U.S. patent application number 14/678165 was filed with the patent office on 2017-06-15 for methods and compositions using fgf23 variant polypeptides.
This patent application is currently assigned to NOVARTIS AG. The applicant listed for this patent is David GLASS, Shou-Ih HU. Invention is credited to David GLASS, Shou-Ih HU.
Application Number | 20170166618 14/678165 |
Document ID | / |
Family ID | 48280855 |
Filed Date | 2017-06-15 |
United States Patent
Application |
20170166618 |
Kind Code |
A1 |
GLASS; David ; et
al. |
June 15, 2017 |
METHODS AND COMPOSITIONS USING FGF23 VARIANT POLYPEPTIDES
Abstract
The present disclosure is directed to methods, kits and
compositions for preventing or treating age-related conditions or
metabolic disorders. The fusion polypeptides of the disclosure
include FGF23 or an active fragment thereof. In one embodiment, the
fusion polypeptide comprises (a) a polypeptide comprising
fibroblast growth factor 23 (FGF23), or a functionally active
variant or derivative thereof, wherein FGF23 has a mutation at one
or more of the positions Q156, C206 and C244; and (b) either a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life, or a
polypeptide comprising at least one extracellular subdomain of a
Klotho protein, or a functionally active variant or derivative
thereof and, optionally (c) a linker. The Klotho fusion proteins
are useful in the treatment and prevention of a variety of
age-related conditions and metabolic disorders. In another
embodiment, the fusion polypeptide comprises a FGF (such as FGF23),
or a functionally active variant or derivative thereof; and a
modified Fc fragment, or a functionally active variant or
derivative thereof In various embodiments of the fusion
polypeptides, FGF23 has mutations which decrease aggregation and
protease-mediated cleavage.
Inventors: |
GLASS; David; (Cambridge,
MA) ; HU; Shou-Ih; (Cambridge, MA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
GLASS; David
HU; Shou-Ih |
Cambridge
Cambridge |
MA
MA |
US
US |
|
|
Assignee: |
NOVARTIS AG
Basel
CH
|
Family ID: |
48280855 |
Appl. No.: |
14/678165 |
Filed: |
April 3, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13743770 |
Jan 17, 2013 |
9139631 |
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14678165 |
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13015159 |
Jan 27, 2011 |
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13743770 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C12N 9/2402 20130101;
C07K 14/50 20130101; A61P 27/02 20180101; A61P 3/04 20180101; C07K
2319/74 20130101; A61K 38/1825 20130101; A61K 38/00 20130101; A61P
21/00 20180101; A61P 3/10 20180101; A61P 9/10 20180101; A61P 25/28
20180101; C07K 2319/30 20130101; A61P 19/10 20180101; A61P 27/12
20180101; A61P 13/12 20180101; A61P 3/00 20180101; A61P 27/16
20180101; A61P 37/00 20180101; C07K 2319/00 20130101; A61P 17/00
20180101; C07K 2319/02 20130101; C12Y 302/01031 20130101; A61P
35/00 20180101 |
International
Class: |
C07K 14/50 20060101
C07K014/50; C12N 9/24 20060101 C12N009/24 |
Claims
1. A fusion polypeptide comprising: (a) a polypeptide comprising
fibroblast growth factor 23 (FGF23), or a functionally active
variant or derivative thereof, wherein FGF23 has a mutation as
described herein; and (b) a fusion partner as described herein, or
a functionally active variant or derivative thereof; and,
optionally (c) a linker.
2. The fusion polypeptide of claim 1, wherein the polypeptide of
(a) is operatively linked to the N-terminus of the polypeptide of
(b).
3. The fusion polypeptide of claim 1, wherein the polypeptide of
(b) is operatively linked to the N-terminus of the polypeptide of
(a).
4. The fusion polypeptide of claim 1, wherein the polypeptide of
(a) and the polypeptide of (b) are connected by a polypeptide
linker.
5. The fusion polypeptide of claim 4, wherein the polypeptide
linker comprises an amino acid sequence selected from the group
consisting of: SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID
NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO:
18.
6. The fusion polypeptide of claim 4, wherein the polypeptide
linker comprises at least 1 and up to about 30 repeats of an amino
acid sequence selected from the group consisting of: SEQ ID NO: 12,
SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID
NO: 17, and SEQ ID NO: 18.
7. The fusion polypeptide of claim 4, wherein the polypeptide of
(a) is connected by a peptide bond to the N-terminus of said
polypeptide linker, and the polypeptide of (b) is connected by a
peptide bond to the C-terminus of said polypeptide linker.
8. The fusion polypeptide of claim 4, wherein the polypeptide of
(a) is connected by a peptide bond to the C-terminus of said
polypeptide linker, and the polypeptide of (b) is connected by a
peptide bond to the N-terminus of said polypeptide linker.
9. The fusion polypeptide of claim 1, wherein the extracellular
subdomain of the Klotho protein is a KL-D1 domain or a KL-D2
domain.
10. The fusion polypeptide of claim 1, wherein the polypeptide of
(a) comprises at least two extracellular subdomains of the Klotho
protein.
11. The fusion polypeptide of claim 10, wherein the at least two
extracellular subdomains of the Klotho protein are at least two
KL-D1 domains in tandem repeats.
12. The fusion polypeptide of claim 10, wherein the at least two
extracellular subdomains of the Klotho protein are at least two
KL-D2 domains in tandem repeats.
13. The fusion polypeptide of claim 10, wherein the at least two
extracellular subdomains of Klotho protein comprise a KL-D1 domain
and a KL-D2 domain.
14. The fusion polypeptide of claim 1, wherein the polypeptide of
(a) is the extracellular domain of the Klotho protein.
15. The fusion polypeptide of claim 1, further comprising a signal
peptide.
16. The fusion polypeptide of claim 15, wherein the signal peptide
is the Klotho signal peptide.
17. The fusion polypeptide of claim 15, wherein the signal peptide
is the IgG signal peptide.
18. The fusion polypeptide of claim 1 that specifically binds to a
fibroblast growth factor receptor.
19. The fusion polypeptide of claim 1, wherein the Klotho protein
is alpha-Klotho.
20. The fusion polypeptide of claim 1, wherein the Klotho protein
is beta-Klotho.
21. The fusion polypeptide of claim 19, wherein the fibroblast
growth factor is fibroblast growth factor-23 (FGF23) or a
fibroblast growth factor-23 variant (R179Q).
22. The fusion polypeptide of claim 20, wherein the fibroblast
growth factor is fibroblast growth factor-19 or fibroblast growth
factor-21.
23. The fusion polypeptide of claim 1 comprising an amino acid
sequence which is 95% or more identical to the amino acid sequence
of SEQ ID NO: 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66,
67, or 68.
24. The fusion polypeptide of claim 1 having the amino acid
sequence of SEQ ID NO: 58, or SEQ ID NO: 68.
25. The fusion polypeptide of claim 1 comprising FcLALA.
26. A pharmaceutical composition comprising the fusion polypeptide
of claim 1 and a pharmaceutically acceptable carrier.
27. A nucleic acid comprising a sequence that encodes the fusion
polypeptide of claim 1.
28. A host cell containing the nucleic acid of claim 27.
29. A vector comprising the nucleic acid of claim 27.
30. A method for treating or preventing an age-related condition, a
metabolic disorder, muscle atrophy, chronic renal disease or
chronic renal failure, hyperphosphatemia, or calcinosis in an
individual, comprising administering to an individual in need
thereof a therapeutically effective dose of a pharmaceutical
composition comprising a fusion polypeptide comprising: (a) a
polypeptide comprising fibroblast growth factor 23 (FGF23), or a
functionally active variant or derivative thereof as described
herein; and (b) a fusion partner as described herein, or a
functionally active variant or derivative thereof; and, optionally
(c) a linker.
31. The method of claim 30, wherein the age-related condition is
selected from the group consisting of sarcopenia, skin atrophy,
muscle wasting, brain atrophy, atherosclerosis, arteriosclerosis,
pulmonary emphysema, osteoporosis, osteoarthritis, immunologic
incompetence, high blood pressure, dementia, Huntington's disease,
Alzheimer's disease, cataracts, age-related macular degeneration,
prostate cancer, stroke, diminished life expectancy, memory loss,
wrinkles, impaired kidney function, and age-related hearing
loss.
32. The method of claim 30, wherein the Klotho protein is alpha
Klotho protein.
33. The method of claim 31, wherein the age-related condition is
muscle wasting, the Klotho protein is alpha Klotho protein, and the
fibroblast growth factor is fibroblast growth factor 23.
Description
[0001] This application is a divisional of U.S. application Ser.
No. 13/743,770, filed Jan. 17, 2013, which is a divisional of U.S.
application Ser. No. 13/015,159, filed Jan. 27, 2011, the contents
of which are incorporated herein by reference in their
entirety.
1. BACKGROUND
[0002] Fibroblast growth factors (FGFs) constitute a family of
homologous polypeptide growth factors expressed in many organisms
(Ornitz and Itoh, Genome Biol. 2: reviews, 3005.1-3005.12 (2001)).
Among vertebrate species, FGFs are highly conserved in both gene
structure and amino-acid sequence, having between 13-71% amino acid
identity with one another. In humans, there are 22 known members of
the FGF family (FGF15 is the mouse ortholog of human FGF19, hence
there is no human FGF15). During early development, FGFs regulate
cell proliferation, migration, and differentiation, but in the
adult organism, FGFs maintain homeostasis, function in tissue
repair, and respond to injury.
[0003] FGFs function as growth factors by binding and thereby
activating cell-surface FGF receptors. FGF receptors (FGFRs) are
tyrosine kinase receptors that activate signal transduction through
autophosphorylation of FGFR, phosphorylation of FRS2 (FGF receptor
substrate 2) and ERK1/2 (extracellular signal-regulated protein
kinase 1/2), and activating Egr-1 (early growth response-1). FGFs
also have a high affinity for heparin sulfate proteoglycans. When
bound to FGFs, heparin sulfate enhances the activation of
FGFRs.
[0004] The alpha-Klotho gene encodes a 130 kDa single pass type I
transmembrane protein with an extracellular domain and a short
cytoplasmic domain. The extracellular domain of alpha-Klotho
protein comprises two subdomains termed, KL-D1 and KL-D2. These two
subdomains share sequence homology to .beta.-glucosidase of
bacteria and plants. The extracellular domain of the alpha-Klotho
protein may be bound to the cell surface by the transmembrane
domain or may be cleaved and released into the extracellular
milieu. Cleavage of the extracellular domain appears to be
facilitated by local low extracellular Ca.sup.2+
concentrations.
[0005] In addition to alpha-Klotho, a homolog of alpha-Klotho,
beta-Klotho, has been identified (Ito et al., Mech. Dev. 98:115-9
(2000)). Beta-Klotho is also a single pass type I transmembrane
protein with extracellular KL-D1 and KL-D2 subdomains.
[0006] Modulation of alpha-Klotho expression has been demonstrated
to produce aging related characteristics in mammals. Mice
homozygous for a loss of function mutation in the alpha-Klotho gene
develop characteristics resembling human aging, including shortened
lifespan, skin atrophy, muscle wasting, arteriosclerosis, pulmonary
emphysema and osteoporosis (Kuro-o et al., Nature, 390:45-51
(1997)). In contrast, overexpression of the alpha-Klotho gene in
mice extends lifespan and increases resistance to oxidative stress
relative to wild-type mice (Kurosu et al., Science 309:1829-1833
(2005); Yamamoto et al., J. Biol. Chem. 280:38029-38034
(2005)).
[0007] Recent studies have demonstrated strikingly similar
biological characteristics between FGF23-deficient mice and
alpha-Klotho-deficient mice (Shimada et al., J. Clin. Invest.
113:561-568 (2004); Yoshida et al. Endocrinology 143:683-689
(2002)), indicating functional crosstalk between FGF23 and
alpha-Klotho. These studies led to the identification of
alpha-Klotho as an obligatory partner of FGF23, in terms of both
binding and signaling through its cognate FGF receptors (Urakawa et
al., Nature 22:1524-6 (2007)). The alpha-Klotho gene is mainly
expressed in kidney, parathyroid gland and choroid plexus. It is
hypothesized that the tissue-specific expression of alpha-Klotho
restricts activation of FGF23 signaling to those tissues.
[0008] Similar to FGF23/alpha-Klotho, beta-Klotho is an obligatory
partner of FGF19 and FGF21, both in binding and in signaling
through their respective cognate FGF receptors (Ogawa et al., Proc.
Natl. Acad. Sci. USA 104:7432-7 (2007); Lin et al., J. Biol Chem.
282:27227-84 (2007); and Wu et al., J. Biol. Chem. 282:29069-72
(2007)). Such studies have also demonstrated the involvement of
beta-Klotho in regulating tissue-specific metabolic activity.
Beta-Klotho was initially shown to act with FGF21 as a cofactor for
regulating carbohydrate and lipid metabolism in adipose tissue.
Beta-Klotho in conjunction with FGF19 regulates bile acid
metabolism in liver, thus explaining elevated bile synthesis in
beta-Klotho deficient mice (Ito et al., J Clin Invest. 2005 August;
115(8):2202-8).
[0009] U.S. Pat. No. 6,579,850 describes polypeptides and
compositions comprising an alpha-Klotho polypeptide. Human and
mouse alpha-Klotho polypeptides are disclosed. The patent also
disclosed that compositions comprising the polypeptides are useful
in treating a syndrome resembling premature aging, treating adult
diseases, and suppressing aging.
[0010] U.S. Pat. No. 7,223,563 describes isolated nucleic acids
encoding the FGF23 polypeptide sequence or recombinant cells
comprising such an isolated nucleic acid. The patent further
relates to methods of diagnosing and treating hypophosphatemic and
hyperphosphatemic disorders, osteoporosis, dermatomyositis, and
coronary artery disease.
[0011] U.S. Pat. No. 7,259,248 describes isolated nucleic acids
encoding the FGF21 polypeptide sequence. The patent further relates
to methods of diagnosing and treating liver disease, conditions
related to thymic function, and methods of treating conditions of
the testis.
2. SUMMARY OF THE INVENTION
[0012] The present disclosure is directed to methods, uses, kits
and compositions for preventing or treating age-related conditions
or metabolic disorders with fusion polypeptides or soluble
polypeptides. The fusion polypeptides of the present disclosure are
formed of a FGF (e.g., FGF23); and either a Klotho protein or an
active fragment thereof (e.g., sKlotho) and/or a Fc fragment (e.g.,
FcLALA); and, optionally, a linker. In some embodiments, the FGF23
is mutated. In some embodiments, the present disclosure provides a
Klotho fusion polypeptide comprising a Klotho protein or an active
fragment thereof and a fibroblast growth factor or an active
fragment thereof. In some embodiments, the fusion polypeptide
comprises a Klotho polypeptide, a FGF (such as FGF23) and a
modified Fc fragment. The Fc fragment can, for example, have
decreased binding to Fc-gamma-receptor and increased serum
half-life. Fusion proteins comprising sKlotho, FGF23 and FcLALA (a
modified Fc fragment having decreased affinity for
Fc-gamma-receptorand/or increased serum half-life) are described in
SEQ ID NOs. 46, 47, 48, and 49. In some embodiments, the fusion
polypeptide or protein comprises a FGF (e.g., FGF23), or a
functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof; and a modified Fc fragment, or a
functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof. Fusion proteins comprising FGF23
and FcLALA are described in SEQ ID NOs. 50, 51, 52 and 53. In some
embodiments, the fusion polypeptide has one or more mutations in
FGF23 which decrease aggregation and/or protease-mediated
cleavage.
[0013] In a first aspect, the disclosure provides a fusion
polypeptide having at least one extracellular subdomain of a Klotho
protein and a fibroblast growth factor or an active fragment
thereof. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity (e.g., decreased Ka
or increased Kd) for Fc-gamma-receptorand/or increased serum
half-life. The Klotho extracellular domain may be derived from
either the alpha or beta Klotho isoforms. Further, although the FGF
component of the Klotho fusion polypeptide is described primarily
with reference to fibroblast growth factor-19, fibroblast growth
factor-21 and fibroblast growth factor-23, it is contemplated that
any of the twenty-three known FGFs can be used in practicing the
disclosure. The reader of the instant application may assume that
each of every combination of alpha or beta extracellular domain
with each human FGF protein or an active fragment thereof are
individually and specifically contemplated.
[0014] According to the present disclosure, the extracellular
domain of the Klotho protein can include one or both of the KL-D1
and KL-D2 domains of a Klotho protein, or a functionally active
variant or derivative (e.g., a variant comprising at least one
conservative amino acid substitution and/or one amino acid
deletion) thereof. In some embodiments, the Klotho fusion
polypeptide of the disclosure has at least two extracellular
subdomains of a Klotho protein, or a functionally active variant or
derivative (e.g., a variant comprising at least one conservative
amino acid substitution and/or one amino acid deletion) thereof.
For example, the at least two extracellular subdomains can be at
least two KL-D1 domains in tandem repeats, at least two KL-D2
domains in tandem repeats, or at least one KL-D1 domain and at
least one KL-D2 domain. In various embodiments, the fusion
polypeptide of the disclosure comprises amino acids 28-292 of the
full length alpha Klotho protein, or amino acids 28-982 (SEQ ID NO:
7). In another embodiment, the fusion polypeptide of the disclosure
comprises amino acids 52-997 of the full length beta Klotho
protein.
[0015] In one embodiment of the present disclosure, the components
of a fusion polypeptide comprise: (a) a polypeptide comprising
fibroblast growth factor 23 (FGF23), or a functionally active
variant or derivative (e.g., a variant comprising at least one
conservative amino acid substitution and/or one amino acid
deletion) thereof, wherein FGF23 has a mutation at one or more of
the positions Q156, C206 and C244; and (b) either a modified Fc
fragment having decreased affinity for Fc-gamma-receptor and/or
increased serum half-life, or a polypeptide comprising at least one
extracellular subdomain of a Klotho protein, or a functionally
active variant or derivative (e.g., a variant comprising at least
one conservative amino acid substitution and/or one amino acid
deletion) thereof; and, optionally (c) a linker. The components can
be, for example, chemically linked or fused in frame by a peptide
bond. They may also linked via a linker. Non-limiting examples of
polypeptide linker are SEQ ID NOs: 11, 12, 13, 14, 15, 16, 17, and
18. Such linkers may comprise at least one and up to about 30
repeats of SEQ ID NOs:11, 12, 13, 14, 15, 16, 17 and 18. In another
non-limiting embodiment, the fusion comprises (2) a FGF or an
active fragment thereof and (3) a modified Fc fragment. The various
components of the fuion can be operatively linked in any order; the
polypeptide (1) can be operatively linked to the N-terminus of the
polypeptide for (2) or (3); the polypeptide for (2) can be
operatively linked to the N-terminus of the polypeptide for (1) or
(3); the polypeptide for (3) can be operatively linked to the
N-terminus of the polypeptide for (1) or (2).
[0016] According to the present disclosure, the extracellular
subdomain of a Klotho protein, the fibroblast growth factor and the
(optional) modified Fc fragment having decreased affinity for
Fc-gamma-receptorand/or increased serum half-life can be
operatively linked to one another in a variety of orientations and
manners. For example, the extracellular subdomain of the Klotho
protein can be operatively linked to the N-terminus of the
fibroblast growth factor or alternatively the fibroblast growth
factor can be operatively linked to the N-terminus of an
extracellular subdomain of the Klotho protein.
[0017] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a sKlotho of a Klotho protein and a linker.
In another embodiment, the present disclosure provides a fusion
polypeptide comprising a sKlotho of the alpha Klotho protein and a
linker. In another embodiment, the present disclosure provides a
fusion polypeptide comprising a sKlotho of the beta Klotho protein
and a linker. In yet another embodiment, the present disclosure
provides a human FGF protein or an active fragment thereof (e.g.,
without signal peptide) and a linker. In one embodiment the
disclosure provides fusion proteins, nucleic acid molecules or
pharmaceutical composition for use in therapy or as medicament for
use in the treatment of a pathological disorder. Pharmaceutical
compositions comprising the fusion proteins of the disclosure and
their uses for treating or preventing age-related conditions or
metabolic disorders are also encompassed by the present disclosure.
In some embodiments, the fusion protein further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptorand/or increased serum half-life.
[0018] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a sKlotho of alpha Klotho protein with
signal peptide fused (directly or indirectly via a linker) to
FGF-23. In another embodiment, the present disclosure provides a
fusion polypeptide comprising a sKlotho of alpha Klotho protein
without signal peptide fused (directly or indirectly via a linker)
to FGF-23. In another embodiment, the present disclosure provides
sKlotho of alpha Klotho protein with signal peptide fused (directly
or indirectly via a linker) to FGF-23 without signal peptide. In
another embodiment, the present disclosure provides a fusion
polypeptide comprising sKlotho of alpha Klotho protein without
signal peptide fused (directly or indirectly via a linker) to
FGF-23 without signal peptide. In some embodiments, the fusion
protein further comprises a modified Fc fragment having decreased
affinity for Fc-gamma-receptorand/or increased serum half-life.
[0019] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a sKlotho of alpha Klotho protein with
signal peptide fused (directly or indirectly via a linker) to
FGF-23 (R179Q) variant. In another embodiment, the present
disclosure provides a fusion polypeptide comprising a sKlotho of
alpha Klotho protein without signal peptide fused (directly or
indirectly via a linker) to FGF-23 (R179Q) variant. In another
embodiment, the present disclosure provides sKlotho of alpha Klotho
protein with signal peptide fused (directly or indirectly via a
linker) to FGF-23 (R179Q) variant without signal peptide. In
another embodiment, the present disclosure provides a fusion
polypeptide comprising sKlotho of alpha Klotho protein without
signal peptide fused (directly or indirectly via a linker) to
FGF-23 (R179Q) variant without signal peptide. In some embodiments,
the fusion protein further comprises a modified Fc fragment having
decreased affinity for Fc-gamma-receptorand/or increased serum
half-life.
[0020] In one embodiment, the present disclosure provides a fusion
polypeptide comprising: (a) a polypeptide comprising fibroblast
growth factor 23 (FGF23), or a functionally active variant or
derivative (e.g., a variant comprising at least one conservative
amino acid substitution and/or one amino acid deletion) thereof,
wherein FGF23 has a mutation at one or more of the positions Q156,
C206 and C244; and (b) either a modified Fc fragment having
decreased affinity for Fc-gamma-receptor and/or increased serum
half-life, or a polypeptide comprising at least one extracellular
subdomain of a Klotho protein, or a functionally active variant or
derivative (e.g., a variant comprising at least one conservative
amino acid substitution and/or one amino acid deletion) thereof;
and, optionally (c) a linker. Such fusion polypeptides are
disclosed in SEQ ID NOs: 54, 55, 56, 57, 58, 59, 60, 61, 62, 63,
64, 65, 66, 67, and 68.
[0021] In one embodiment, the present disclosure provides a fusion
polypeptide comprising (1) sKlotho of alpha Klotho protein with
signal peptide, or a functionally active variant or derivative
(e.g., a variant comprising at least one conservative amino acid
substitution and/or one amino acid deletion) thereof, (2) a linker;
and (3) FGF-23 (R179Q) variant without signal peptide, or a
functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof. In another embodiment, the
present disclosure provides a fusion polypeptide comprising (1)
sKlotho of alpha Klotho protein without signal peptide, or a
functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof; (2) a linker; and (3) FGF-23
(R179Q) variant without signal peptide, or a functionally active
variant or derivative (e.g., a variant comprising at least one
conservative amino acid substitution and/or one amino acid
deletion) thereof. In some embodiments, the fusion polypeptides of
the disclosure are glycosylated. In some embodiments, the fusion
protein further comprises a modified Fc fragment having decreased
affinity for Fc-gamma-receptorand/or increased serum half-life.
[0022] In one embodiment, the present disclosure provides a fusion
polypeptide comprising (1) sKlotho of alpha Klotho protein with
signal peptide (SEQ ID NO: 44 or SEQ ID NO: 45), or a functionally
active variant or derivative (e.g., a variant comprising at least
one conservative amino acid substitution and/or one amino acid
deletion) thereof (2) a linker comprising SEQ ID NO: 11; and (3)
FGF-23 (R179Q) variant without signal peptide (SEQ ID NO: 43), or a
functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof. In another embodiment, the
present disclosure provides a fusion polypeptide comprising (1)
sKlotho of alpha Klotho protein without signal peptide (SEQ ID NO:
7), or a functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof (2) a linker comprising SEQ ID NO:
11; and (3) FGF-23 (R179Q) variant without signal peptide (SEQ ID
NO: 43), or a functionally active variant or derivative (e.g., a
variant comprising at least one conservative amino acid
substitution and/or one amino acid deletion) thereof In one
embodiment, the present disclosure provides a fusion polypeptide
comprising the amino acid sequence of SEQ ID NO: 19, 20, 40, or 41.
In some embodiments, the fusion polypeptides of the disclosure are
glycosylated.
[0023] In one embodiment, the present disclosure provides a fusion
polypeptide comprising sKlotho of alpha Klotho protein with signal
peptide (SEQ ID NO: 44 or SEQ ID NO: 45), or a functionally active
variant or derivative (e.g., a variant comprising at least one
conservative amino acid substitution and/or one amino acid
deletion) thereof; and a linker comprising SEQ ID NO: 11. In
another embodiment, the present disclosure provides a fusion
polypeptide comprising sKlotho of alpha Klotho protein without
signal peptide (SEQ ID NO: 7); and a linker comprising SEQ ID NO:
11. In some embodiments, the fusion polypeptides of the disclosure
are glycosylated. In some embodiments, the fusion protein further
comprises a modified Fc fragment having decreased affinity for
Fc-gamma-receptorand/or increased serum half-life.
[0024] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a human FGF protein or an active fragment
thereof (e.g., without the signal peptide); and a linker comprising
SEQ ID NO: 11. In some embodiments, the fusion polypeptides of the
disclosure are glycosylated. In some embodiments, the fusion
protein further comprises a modified Fc fragment having decreased
affinity for Fc-gamma-receptorand/or increased serum half-life.
[0025] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a human FGF protein (e.g., FGF23) or an
active fragment thereof (e.g., without the signal peptide); a
linker (e.g., a linker comprising SEQ ID NO: 11); and sKlotho (with
or without a signal peptide), or a functionally active variant or
derivative (e.g., a variant comprising at least one conservative
amino acid substitution and/or one amino acid deletion) thereof) or
a Fc-gramma-receptor (e.g., FcLALA); wherein the FGF (e.g., FGF23)
has one or more mutations at these residues: R179, Q156, C206,
and/or C244. In various embodiments, the mutations are R179Q,
Q156A, C2065, and/or C244S. Even though these mutations are
conserved in the human, rhesus, bovine, mouse and rat FGF23,
mutating them does not prevent FGF23 activity. Rather, mutating
these amino acids unexpectedly enhances the qualities of the
proteins, by reducing aggregation, reducing undesired
protease-induced cleavage, and increasing protein production from
cells. In various embodiments, the fusion protein comprising one or
more FGF23 mutation is glycosylated.
[0026] In one embodiment, the present disclosure provides a
pharmaceutical composition (e.g., in an intra-muscular
administering form) comprising (e.g., as a sole pharmaceutically
active ingredient) a fusion polypeptide (e.g., glycosylated or
non-glycosylated) that comprises (1) FGF-23 (R179Q) variant without
signal peptide (SEQ ID NO: 43), or a variant comprising additional
mutations which reduce aggregation and/or protease-mediated
cleavage, or a functionally active variant or derivative (e.g., a
variant comprising at least one conservative amino acid
substitution and/or one amino acid deletion) thereof (2)
optionally, a linker comprising SEQ ID NO: 11; and (3) sKlotho of
alpha Klotho protein with signal peptide (SEQ ID NO: 44 or SEQ ID
NO: 45), or a functionally active variant or derivative (e.g., a
variant comprising at least one conservative amino acid
substitution and/or one amino acid deletion) thereof or a modified
Fc fragment having decreased affinity for Fc-gamma-receptorand/or
increased serum half-life; and uses of the pharmaceutical
composition in therapy or as medicament for the treatment of a
pathological disorder, for example treating and/or preventing
age-related conditions, such as muscular atrophy. In another
embodiment, the present disclosure provides a pharmaceutical
composition (e.g., in an intra-muscular administering form)
comprising (e.g., as a sole pharmaceutically active ingredient) a
fusion polypeptide (e.g., glycosylated or non-glycosylated) that
comprises (1) FGF-23 (R179Q) variant without signal peptide (SEQ ID
NO: 43), or a variant comprising additional mutations which reduce
aggregation and/or protease-mediated cleavage, or a functionally
active variant or derivative (e.g., a variant comprising at least
one conservative amino acid substitution and/or one amino acid
deletion) thereof (2) a linker comprising SEQ ID NO: 11; and (3)
sKlotho of alpha Klotho protein without signal peptide (SEQ ID NO:
7), or a functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof, or a modified Fc fragment having
decreased affinity for Fc-gamma-receptorand/or increased serum
half-life, or a functionally active variant or derivative (e.g., a
variant comprising at least one conservative amino acid
substitution and/or one amino acid deletion) thereof; and uses of
the pharmaceutical composition in therapy or as medicament for the
treatment of a pathological disorder, for example treating and/or
preventing age-related conditions, such as muscular atrophy. In one
embodiment, the present disclosure provides a pharmaceutical
composition (e.g., in an intra-muscular administering form)
comprising (e.g., as a sole pharmaceutically active ingredient) a
fusion polypeptide (e.g., glycosylated or non-glycosylated)
comprising the amino acid sequence of SEQ ID NO: 19, 20, 40, or 41;
and uses of the pharmaceutical composition in therapy or as
medicament for the treatment of a pathological disorder, for
example treating and/or preventing age-related conditions, such as
muscular atrophy.
[0027] In one embodiment, the present disclosure provides a
pharmaceutical composition (e.g., in an intra-muscular
administering form) comprising (e.g., as a sole pharmaceutically
active ingredient) a fusion polypeptide (e.g., glycosylated or
non-glycosylated) that comprises sKlotho of alpha Klotho protein
with signal peptide (SEQ ID NO: 44 or SEQ ID NO: 45); and a linker
comprising SEQ ID NO: 11; and uses of the pharmaceutical
composition for treating and/or preventing age-related conditions,
such as muscular atrophy. In another embodiment, the present
disclosure provides a pharmaceutical composition (e.g., in an
intra-muscular administering form) comprising (e.g., as a sole
pharmaceutically active ingredient) a fusion polypeptide (e.g.,
glycosylated or non-glycosylated) comprising sKlotho of alpha
Klotho protein without signal peptide (SEQ ID NO: 7); and a linker
comprising SEQ ID NO: 11; and uses of the pharmaceutical
composition in therapy or as medicament for the treatment of a
pathological disorder,for example treating and/or preventing
age-related conditions, such as muscular atrophy. In some
embodiments, the fusion protein further comprises a modified Fc
fragment.
[0028] In one embodiment, the present disclosure provides a
pharmaceutical composition comprising (e.g., as a sole
pharmaceutically active ingredient) a fusion polypeptide (e.g.,
glycosylated or non-glycosylated) that comprises a human FGF
protein or an active fragment thereof (e.g., without the signal
peptide); and a linker comprising SEQ ID NO: 11.
[0029] Pharmaceutical compositions comprising the fusion proteins
of the disclosure and their uses in therapy or as medicament for
the treatment of a pathological disorder therapy, for example
treating or preventing age-related conditions (e.g., muscle
atrophy) or metabolic disorders (e.g., diabete) are also
encompassed by the present disclosure.
[0030] In one embodiment, the present disclosure provides a fusion
polypeptide that is at least 85%, at least 90%, at least 91%, at
least 92%, at least 93%, at least 94%, at least 95%, at least 96%,
at least 96%, at least 97%, at least 98%, at least 99% identical to
SEQ ID NO: 19. In another embodiment, the present disclosure
provides a fusion polypeptide that is at least 85%, at least 90%,
at least 91%, at least 92%, at least 93%, at least 94%, at least
95%, at least 96%, at least 96%, at least 97%, at least 98%, at
least 99% identical to SEQ ID NO: 20.
[0031] In one embodiment, the present disclosure provides a fusion
polypeptide that is at least 85%, at least 90%, at least 91%, at
least 92%, at least 93%, at least 94%, at least 95%, at least 96%,
at least 96%, at least 97%, at least 98%, at least 99% identical to
SEQ ID NO: 40. In another embodiment, the present disclosure
provides a fusion polypeptide that is at least 85%, at least 90%,
at least 91%, at least 92%, at least 93%, at least 94%, at least
95%, at least 96%, at least 96%, at least 97%, at least 98%, at
least 99% identical to SEQ ID NO: 41.
[0032] In one embodiment, the present disclosure provides a fusion
polypeptide that is at least 85%, at least 90%, at least 91%, at
least 92%, at least 93%, at least 94%, at least 95%, at least 96%,
at least 96%, at least 97%, at least 98%, at least 99% or 100%
identical to SEQ ID NO: 46. In another embodiment, the present
disclosure provides a fusion polypeptide that is at least 85%, at
least 90%, at least 91%, at least 92%, at least 93%, at least 94%,
at least 95%, at least 96%, at least 96%, at least 97%, at least
98%, at least 99% or 100% identical to SEQ ID NO: 47.
[0033] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 48. In another embodiment, the
present disclosure provides a fusion polypeptide that is at least
85%, at least 90%, at least 91%, at least 92%, at least 93%, at
least 94%, at least 95%, at least 96%, at least 96%, at least 97%,
at least 98%, at least 99% or 100% identical to SEQ ID NO: 49.
[0034] In one embodiment, the present disclosure provides a fusion
polypeptide that is at least 85%, at least 90%, at least 91%, at
least 92%, at least 93%, at least 94%, at least 95%, at least 96%,
at least 96%, at least 97%, at least 98%, at least 99% or 100%
identical to SEQ ID NO: 50. In another embodiment, the present
disclosure provides a fusion polypeptide that is at least 85%, at
least 90%, at least 91%, at least 92%, at least 93%, at least 94%,
at least 95%, at least 96%, at least 96%, at least 97%, at least
98%, at least 99% or 100% identical to SEQ ID NO: 51.
[0035] In one embodiment, the present disclosure provides a fusion
polypeptide that is at least 85%, at least 90%, at least 91%, at
least 92%, at least 93%, at least 94%, at least 95%, at least 96%,
at least 96%, at least 97%, at least 98%, at least 99% or 100%
identical to SEQ ID NO: 52. In another embodiment, the present
disclosure provides a fusion polypeptide that is at least 85%, at
least 90%, at least 91%, at least 92%, at least 93%, at least 94%,
at least 95%, at least 96%, at least 96%, at least 97%, at least
98%, at least 99% or 100% identical to SEQ ID NO: 53.
[0036] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 54.
[0037] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 55.
[0038] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 56.
[0039] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 57.
[0040] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 58.
[0041] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 59.
[0042] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 60.
[0043] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 61.
[0044] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 62.
[0045] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 63.
[0046] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 64.
[0047] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 65.
[0048] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 66.
[0049] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 67.
[0050] In another embodiment, the present disclosure provides a
fusion polypeptide that is at least 85%, at least 90%, at least
91%, at least 92%, at least 93%, at least 94%, at least 95%, at
least 96%, at least 96%, at least 97%, at least 98%, at least 99%
or 100% identical to SEQ ID NO: 68.
[0051] In one embodiment, the present disclosure provides a fusion
polypeptide comprising a sKlotho of beta Klotho protein with signal
peptide fused (directly or indirectly via a linker) to FGF-19 or an
active fragment thereof. In some embodiments, the fusion further
comprises a modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. In another
embodiment, the present disclosure provides a fusion polypeptide
comprising a sKlotho of beta Klotho protein without signal peptide
fused (directly or indirectly via a linker) to FGF-19 or an active
fragment thereof. In another embodiment, the present disclosure
provides a fusion polypeptide comprising a sKlotho of beta Klotho
protein with signal peptide fused (directly or indirectly via a
linker) to FGF-21 or an active fragment thereof. In another
embodiment, the present disclosure provides a fusion polypeptide
comprising a sKlotho of beta Klotho protein without signal peptide
fused (directly or indirectly via a linker) to FGF-21 or an active
fragment thereof.
[0052] The disclosure provides nucleic acid sequences encoding any
of the Klotho fusion polypeptides described herein and host cells
containing the nucleic acids. In some embodiments, the fusion
further comprises a modified Fc fragment having decreased affinity
for Fc-gamma-receptor and/or increased serum half-life.
[0053] The disclosure also provides composition having any of the
Klotho fusion polypeptides contemplated herein. The compositions of
the disclosure can further include heparin. In some embodiments,
the fusion further comprises a modified Fc fragment having
decreased affinity for Fc-gamma-receptor and/or increased serum
half-life.
[0054] The disclosure also provides a method for treating or
preventing an age-related condition in an individual. An individual
(e.g., human) is administered a therapeutically effective dose of a
pharmaceutical composition containing a Klotho fusion polypeptide,
having at least one extracellular subdomain of a Klotho protein
(e.g., alpha Klotho protein) and a fibroblast growth factor or an
active fragment thereof so as to treat or prevent the age-related
condition. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. In particular,
the disclosure provides a method of treating or preventing muscle
wasting comprising administering to an individual (e.g., human) an
therapeutically effective amount of a fusion polypeptide having at
least one extracellular subdomain of an alpha Klotho protein and a
fibroblast growth factor (or an active fragment thereof).
[0055] Additionally, the disclosure provides a method for treating
or preventing a metabolic disorder in an individual. An individual
is administered a therapeutically effective dose of a
pharmaceutical composition containing a fusion polypeptide of the
disclosure, having at least one extracellular subdomain of a Klotho
protein and a fibroblast growth factor (or an active fragment
thereof) so as to treat the metabolic disorder. In some
embodiments, the fusion further comprises a modified Fc fragment
having decreased affinity for Fc-gamma-receptor and/or increased
serum half-life. In particular, a fusion polypeptide of the
disclosure having at least one extracellular subdomain of a
beta-Klotho protein and a fibroblast growth factor 21 is useful for
treating a metabolic disorder.
[0056] Klotho-FGF23 fusion polypeptides of the disclosure can be
used for treating or preventing hyperphosphatemia or calcinosis in
an individual. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. A
pharmacologically effective dose of a pharmaceutical composition
containing the Klotho fusion polypeptide of the disclosure, having
at least one extracellular subdomain of a Klotho protein and a
fibroblast growth factor, is administered to treat or prevent
hyperphosphatemia or calcinosis. In particular, a Klotho fusion
polypeptide of the disclosure having at least one extracellular
subdomain of an alpha Klotho protein and a fibroblast growth factor
23 is useful for treating hyperphosphatemia or calcinosis.
[0057] Klotho-FGF23 fusion polypeptides of the disclosure can be
used for treating or preventing chronic renal disease or chronic
renal failure in an individual. In some embodiments, the fusion
further comprises a modified Fc fragment having decreased affinity
for Fc-gamma-receptor and/or increased serum half-life. A
therapeutically effective dose of a pharmaceutical composition
containing the Klotho fusion polypeptide of the disclosure, having
at least one extracellular subdomain of a Klotho protein (e.g.,
alpha Klotho protein) and a fibroblast growth factor, is
administered to treat or prevent chronic renal disease or chronic
renal failure.
[0058] Klotho-FGF23 fusion polypeptides of the disclosure can be
used for treating or preventing cancer (e.g., breast cancer) in an
individual. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. A
therapeutically effective dose of a pharmaceutical composition
containing the Klotho fusion polypeptide of the disclosure, having
at least one extracellular subdomain of a Klotho protein (e.g.,
alpha Klotho protein) and a fibroblast growth factor, is
administered to treat or prevent cancer or breast cancer.
[0059] The present disclosure provides fusion polypeptides
comprising at least one extracellular subdomain of Klotho protein
and a FGF or an active fragment thereof for use in medicine. In
some embodiments, the fusion further comprises a modified Fc
fragment having decreased affinity for Fc-gamma-receptor and/or
increased serum half-life. In one embodiment, the present
disclosure provides fusion polypeptides comprising at least one
extracellular subdomain of Klotho protein and a FGF or an active
fragment thereof for use in treating or preventing muscle atrophy.
The present disclosure also provides a method of treating or
preventing an age related condition (e.g., muscle atrophy)
comprising administering to an individual in need thereof a
therapeutically effective dose of a pharmaceutical composition
comprising a soluble Klotho protein.
[0060] The disclosure futhermore provides the above described
peptides and fusion polypeptides or pharmaceutical compositions
comprising said peptides for use in therapy, as a medicament or for
use in the treatment of a pathological disorder, for example
age-related condition, metabolic disorder, hyperphosphatemia or
calcinosis, chronic renal disease or chronic renal failure or to
prevent cancer or breast cancer, in an individual. Additonally, the
disclosure further provides use of a polypeptide, nucleic acid or
pharmaceutical composition of the invention in the manifacture of a
medicament for the treatment of a pathological disorder,
particularly for the treatment of the above mentioned disorders,
preferably age related conditions like muscle atrophy.
[0061] The disclosure also includes kits for treating or preventing
an age-related disorder or metabolic disorder in an individual. The
kit includes instructions for use and a purified Klotho fusion
polypeptide having at least one extracellular subdomain of a Klotho
protein and a fibroblast growth factor. In some embodiments, the
fusion further comprises a modified Fc fragment having decreased
affinity for Fc-gamma-receptor and/or increased serum
half-life.
[0062] The disclosure also provides a kit for producing a Klotho
fusion polypeptide of the disclosure. The kit of the disclosure
includes instructions for use and a nucleic acid encoding a Klotho
fusion polypeptide, having at least one extracellular subdomain of
Klotho protein and a fibroblast growth factor. In some embodiments,
the fusion further comprises a modified Fc fragment having
decreased affinity for Fc-gamma-receptor and/or increased serum
half-life.
[0063] In one embodiment of the disclosure, the fusion polypeptide
comprises: (a) a polypeptide comprising a fibroblast growth factor,
or a functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof; and (b) a modified Fc fragment,
or a functionally active variant or derivative (e.g., a variant
comprising at least one conservative amino acid substitution and/or
one amino acid deletion) thereof, having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life
[0064] In one embodiment of the disclosure, the polypeptide of (a)
and the polypeptide of (b) are connected by a polypeptide linker.
The linker can be repeated 1 to 30 times, or more.
[0065] In one embodiment of the disclosure, the polypeptide linker
comprises an amino acid sequence selected from the group consisting
of: SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ
ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18.
[0066] In one embodiment of the disclosure, the polypeptide of (a)
is connected by a peptide bond to the N-terminus of said
polypeptide linker, and the polypeptide of (b) is connected by a
peptide bond to the C-terminus of said polypeptide linker.
[0067] In one embodiment of the disclosure, the fusion polypeptide
further comprises a signal peptide.
[0068] In one embodiment of the disclosure, the signal peptide is
the IgG signal peptide.
[0069] In one embodiment of the disclosure, the fibroblast growth
factor is fibroblast growth factor-23 or a fibroblast growth
factor-23 variant (R179Q).
[0070] In one embodiment of the disclosure, the fibroblast growth
factor is fibroblast growth factor-19 or fibroblast growth
factor-21.
[0071] In one embodiment of the disclosure, fusion polypeptide
comprises an amino acid sequence which is 95% or more identical to
the amino acid sequence of SEQ ID NO: 51, or SEQ ID NO: 53.
[0072] In one embodiment of the disclosure, fusion polypeptide
comprises the amino acid sequence of SEQ ID NO: 51, or SEQ ID NO:
53.
[0073] In one embodiment of the disclosure, fusion polypeptide
comprises FcLALA.
3. BRIEF DESCRIPTION OF THE DRAWINGS
[0074] FIG. 1 illustrates several different embodiments of the
Klotho fusion polypeptides of the disclosure. The represented
fusion polypeptides include one or more Klotho extracellular
subdomains operatively linked to a fibroblast growth factor.
Polypeptides containing one or more Klotho extracellular subdomains
include, for example, an extracellular domain of Klotho (e.g., aa 1
to 982 of human Klotho), or an active fragment of Klotho.
[0075] FIG. 2 illustrates the amino acid and nucleic acid sequences
of several Klotho fusion polypeptides of the disclosure and
components thereof (e.g., Klotho extracellular domain, FGF). Fusion
proteins comprising sKlotho, FGF23 and FcLALA (a modified Fc
fragment having decreased affinity for Fc-gamma-receptor and/or
increased serum half-life) are described in SEQ ID NOs. 46, 47, 48,
and 49. Fusion proteins comprising FGF23 and FcLALA are described
in SEQ ID NOs. 50, 51, 52 and 53.
[0076] FIGS. 3A-3C depict protein expression of an sKlotho-FGF23
fusion protein. FIG. 3A shows that sKlotho-FGF23 fusion protein was
detected in conditioned media by Western blotting with anti-FGF23
antibodies. FIG. 3B shows that sKlotho-FGF23 fusion protein was
detected in conditioned media by SDS-PAGE and Coomassie blue
staining FIG. 3C shows a highly purified sKlotho-FGF23-6.times.His
fusion protein, analyzed by SDS-PAGE and Coomassie blue
staining.
[0077] FIG. 4 illustrates the results of an Egr-1 luciferase assay
comparing the activation level of Egr-1 in cells treated with
conditioned media containing either a
[0078] Klotho fusion polypeptide, a FGF 23 polypeptide only, a
soluble Klotho (sKlotho) polypeptide only, and a soluble Klotho
polypeptide in combination with a FGF 23 polypeptide in the absence
or presence of heparin (20 .mu.g/ml).
[0079] FIGS. 5A-5B depict the results of an Egr-1 luciferase assay
comparing the activation level of Egr-1 in cells treated with
purified Klotho fusion polypeptide, FGF 23 polypeptide, or soluble
Klotho polypeptide in the absence or presence of heparin. FIG. 5A
shows an the results of an experiment comparing the activation
level of Egr-1 in cells treated with FGF 23 alone, sKlotho-His (10
nM or 20 nM) and a combination of FGF 23 and sKlotho-His (10 nM or
20 nM) in the absence or presence of heparin (20 .mu.g/ml). FIG. 5B
shows Egr-1 luciferase reporter activity in cells treated with
sKlotho-FGF23-His fusion (0 nM, 0.6 nM, 1.21 nM, 2.41 nM, 4.83 nM,
9.65 nM, and 19.3 nM).
[0080] FIGS. 6A-6B illustrate the effect of treatment with a
purified sKlotho fusion polypeptide on C2C12 muscle cells. FIG. 6A
shows measurements of myotube diameter in C2C12 muscle cells
treated with either IGF-1 (10 nM), FGF2 (20ng/ml), or a purified
Klotho fusion polypeptide (20 nM), in the absence or presence of
dexamethasone (100 .mu.M). FIG. 6B shows the phosphorylation of
signaling pathway proteins in C2C12 muscle cells by IGF-1 (10 nM),
FGF2 (20 ng/ml), or a purified Klotho fusion polypeptide (20 nM),
in the absence or presence of rapamycin (40 nM).
[0081] FIG. 7 shows activation of EGR-1-luc reporter gene by
sKlotho-FGF23(R179Q)-FcLALA fusion proteins.
[0082] FIG. 8 shows the activation of EGR-1-luc reporter gene by
FGF23(R179Q)-FcLALA proteins.
[0083] FIG. 9 shows the pharmacokinetic profile of FGF23(R179Q) vs
FGF23(R179Q)-FcLALAv2.
[0084] FIG. 10A shows absolute weights of
gastrocnemius-soleus-plantaris (GSP) muscles injected
intramuscularly with sKlotho-FGF23 (KLOFGF) and PBS. FIG. 10B shows
percent weight change (B) of GSP muscles injected intramuscularly
with KLOFGF and PBS.
[0085] FIG. 11. This figure shows activation of EGR-1-luc reporter
gene by FGF23(R179Q)-FcLALA and Q156A, C2065, C244S and C206S/C244S
mutants.
[0086] FIG. 12 shows protein qualities and dimerization of WT
(wild-type), Q156A, C2065, C244S and C206S/C244S mutants of
FGF23(R179Q)-FcLaLa.
4. DETAILED DESCRIPTION
[0087] The present disclosure is directed to methods, kits and
compositions for preventing or treating age-related conditions and
metabolic disorders; and to the use of said compostions in therapy,
as a medicament or for use in the treatment of a pathological
disorder. The fusion polypeptides of the disclosure include a
Klotho protein or active fragment thereof. In some embodiments, the
fusion polypeptides of the disclosure include a Klotho protein or
an active fragment thereof operatively linked to a fibroblast
growth factor polypeptide or active fragment thereof. In some
embodiments, the fusion further comprises a modified Fc fragment
with decreased ability to bind FcRn and/or increased stability in
serum. In another embodiment, the fusion polypeptide comprises a
FGF (e.g., FGF23) and a modified Fc fragment with decreased ability
to bind FcRn and/or increased stability in serum.
[0088] The fusion proteins or sKlotho of the present disclosure are
useful in the treatment and prevention of a variety of age-related
conditions including sarcopenia, skin atrophy, muscle wasting,
brain atrophy, atherosclerosis, arteriosclerosis, pulmonary
emphysema, osteoporosis, osteoarthritis, immunologic incompetence,
high blood pressure, dementia, Huntington's disease, Alzheimer's
disease, cataracts, age-related macular degeneration, prostate
cancer, stroke, diminished life expectancy, memory loss, wrinkles,
impaired kidney function, and age-related hearing loss; and
metabolic disorders including Type II Diabetes, Metabolic Syndrome,
hyperglycemia, and obesity.
[0089] The present disclosure is based at least in part on the
finding that despite the physical constraints (e.g., large size of
both the Klotho and FGF polypeptides) the Klotho-FGF fusion
polypeptides are highly effective in activating an FGF receptor.
This finding is unexpected given that fusion of these two proteins
would likely interfere with the heterodimerization and thus the
activities of the proteins; e.g., the binding domains of the
proteins may be perturbed by the fusion or the proteins may be
mis-oriented spatially if put together in a "cis" formation.
[0090] The fusion polypeptides described herein are advantageous
because they allow the administration of a single therapeutic
protein that has enhanced activity compared to Klotho or FGF
administered alone or together as separate polypeptides. The use of
Klotho and FGF as a single fusion polypeptide rather than as two
separate polypeptides (i.e., a Klotho polypeptide and a separate
FGF polypeptide) is more effective at activating the FGF
receptor.
Definitions
[0091] "Klotho polypeptide", "Klotho protein", or "Klotho" as used
herein, includes active fragments, derivatives, mimetics, variants
and chemically modified compounds or hybrids thereof of wild-type
"Klotho". A Klotho active fragment has the ability to bind to an
FGF polypeptide. Generally, a Klotho active polypeptide contains at
least a Klotho subdomain (e.g., KL-D1 and KL-D2). Wild-type Klotho
has the amino acid sequence as is found in nature. Example Klotho
polypeptides suitable for use with the present disclosure include
alpha-Klotho (SEQ ID NO: 2) and beta-Klotho (SEQ ID NO: 4).
Nucleotide and amino acid sequences of the alpha-Klotho and
beta-Klotho are found in the GenBank database at Accession No.
NM_004795; NP_004786 and NM_175737; NP_783864, respectively. Klotho
polypeptides include those described in U.S. Pat. No. 6,579,850,
the content of which is herein incorporated by reference in its
entirety. The Klotho polypeptides include those from other species
besides humans, including alpha-Klotho from mouse (NP_038851), rat
(NP_112626), rabbit (NP_001075692) and beta-Klotho from mouse
(NP_112457). Species predicted to have alpha-Klotho include
chimpanzee (XP_522655), macaque (XP_001101127), horse
(XP_001495662), cow (XP_001252500), platypus (XP_001510981), and
chicken (XP_417105). Species predicted to have beta-Klotho include
chimpanzee (XP_526550), macaque (XP_001091413), horse
(XP_001495248), dog (XP_536257), rat (XP_001078178), platypus
(XP_001512722), and chicken (XP_423224). The Klotho polypeptides
have an amino acid sequence that is substantially identical to the
amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 4; i.e., at least
70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical
at the amino acid sequences of SEQ ID NO: 2 or SEQ ID NO: 4, or
active fragment thereof.
[0092] "Fusion polypeptide" or "fusion protein", as used herein,
shall mean a polypeptide comprising two or more different
polypeptides or active fragments thereof that are not naturally
present in the same polypeptide. In some embodiments, the two or
more different polypeptides are operatively linked together
covalently, e.g., chemically linked or fused in frame by a peptide
bond. As used herein a "Klotho fusion polypeptide" is a fusion
polypeptide which includes an amino acid sequence from a Klotho
polypeptide or active fragment thereof. A fusion polypeptide can
comprise, as non-limiting examples, Klotho (e.g., sKlotho), FGF
(e.g., FG23), and (optionally) a modified Fc fragment (e.g., a
modified Fc fragment with decreased binding affinity to
FC-gamma-receptor and/or increased serum half-life). Examples of
this type of fusion polypeptide are presented in SEQ ID NOs. 46 to
49. In another embodiment, the fusion proteins comprise FGF (e.g.,
FGF23) and a modified Fc (e.g., FcLALA). Fusion proteins comprising
FGF23 and FcLALA are described in SEQ ID NOs. 50, 51, 52 and 53.
FcLALA is a Fc fragment with a LALA mutation (L234A, L235A), which
triggers ADCC with lowered efficiency, and binds and activates
human complement weakly. Hessell et al. 2007 Nature
449:101-104.
[0093] "Fibroblast growth factor" and "FGF" are used
interchangeably herein and shall refer to polypeptides that
regulate cell proliferation, migration, differentiation,
homeostasis, tissue repair and response to injury in an animal,
including a human subject. FGFs have the ability to bind to a
fibroblast growth factor receptor and regulate its activity,
including autophosphorylation of FGFR, phosphorylation of FRS2 (FGF
receptor substrate 2) and ERK1/2 (extracellular signal-regulated
protein kinase 1/2), and activating Egr-1 (early growth
response-1). The term "FGF" includes active fragments, derivatives,
mimetics, variants and chemically modified compounds or hybrids
thereof of wild-type "FGF", e.g., as known in the art and as
described in U.S. Pat. No. 7,223,563 and U.S. Pat. No. 7,259,248,
the contents of which are incorporated by reference in their
entirety. Wild-type FGF has an amino acid sequence as is found in
nature. Example fibroblast growth factors suitable for use with the
present disclosure include fibroblast growth factor-19 (FGF19; SEQ
ID NO: 31), fibroblast growth factor-21 (FGF21; SEQ ID NO: 33), and
fibroblast growth factor-23 (FGF23; SEQ ID NO: 35). The FGF
polypeptides include those from other species besides humans,
including murine FGFs. Generally, FGF polypeptides have an amino
acid sequence that is substantially identical to the amino acid
sequence of SEQ ID NO: 31, SEQ ID NO: 33 or SEQ ID NO: 35; i.e.,
having an amino acid sequence is which is at least 70%, 75%, 80%,
85%, 90%, 95%, 96%, 97%, 98%, 99% or more or 100% identical to the
amino acid sequences of SEQ ID NO: 31 SEQ ID NO: 33 or SEQ ID NO:
35, or active fragments thereof. Additional non-limiting examples
of FGF, particularly FGF23, are provided at aa 1002-1228 of SEQ ID
NO: 47; aa 1002-1228 of SEQ ID NO: 49; aa 1-251 of SEQ ID NO: 51,
and aa 1-251 of SEQ ID NO: 53; and sequences which are at least
70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more or 100%
identical to these sequences. Nucleotides encoding these sequences
are provided in SEQ ID NOs: 46, 48, 50 and 52.
[0094] The term "FGF", includes active fragments of the full-length
polypeptide. Active FGF fragments that are able to bind to their
corresponding FGF receptors are known in the art and also
contemplated for use in the present disclosure. One skilled in the
art would appreciate, based on the sequences disclosed herein, that
overlapping fragments of the FGFs can be generated using standard
recombinant technology, for example, that described in Sambrook et
al. (1989, Molecular Cloning: A Laboratory Manual, Cold Spring
Harbor Laboratory Press, New York) and Ausubel et al. (1997,
Current Protocols in Molecular Biology, Green & Wiley, New
York). One skilled in the art would appreciate, based on the
disclosure presented herein, that the biological activity of FGF
fragments could be tested by methods well known in the art and
described herein, including binding to the FGF receptor. Similarly,
cell culture models which possess the necessary FGF signal
transduction machinery (i.e. FGF receptor) may be transfected with
FGF fragments and subsequently tested for alterations in FGF
signaling, relative to wild type FGF.
[0095] FGFs are grouped into seven subfamilies based on the
homology of the FGF core homology domain (approximately 120 amino
acids long), which is flanked by N- and C-terminal sequences that
are highly variable in both length and primary sequence,
particularly among different FGF subfamilies (Goetz et al.,
Molecular and Cellular Biology, 2007, Vol. 27, 3417-3428). An FGF
active polypeptide generally contains at least an FGF core homology
domain. In some embodiments, an FGF active polypeptide may contain,
in addition to an FGF core homology domain, flanking sequences
which may confer additional specificity in binding FGF receptors.
FGF19, FGF21, and FGF23 are grouped in the FGF19 subfamily because
the core region of these ligands share high sequence identity
relative to other FGFs (FGF19 v. FGF21: 38% identity; FGF19 v.
FGF23: 36% identity). FGF19 subfamily members act analogously to
signaling molecules of the endocrine system and regulate diverse
physiological processes uncommon to classical FGFs (e.g., FGF19:
energy and bile acid homeostasis; FGF21: glucose and lipid
metabolism; and FGF 23: phosphate and vitamin D homeostasis).
[0096] "Fibroblast growth factor receptor" and "FGFR" as used
herein refer to any one of FGFRs 1-4 known in the art, or splice
variants thereof (e.g., FGFR1c). Example fibroblast growth factor
receptors suitable for use with the present disclosure include
fibroblast growth factor receptor-19 (e.g., FGFR4-beta Klotho),
fibroblast growth factor receptor-21 (e.g., FGFR1c-alpha Klotho),
and fibroblast growth factor receptor-23 (e.g., FGFR1c-alpha
Klotho, FGFR3-alpha Klotho, FGFR4-alpha Klotho).
[0097] "Extracellular domain", as used herein, refers to the
fragment of a transmembrane protein existing outside of a cell
(e.g., not including the intracellular or transmembrane region).
The "extracellular domain of the Klotho protein", "soluble Klotho",
or "sKlotho" (e.g., SEQ ID NO: 7; SEQ ID NO: 39), refers to an
extracellular domain of the Klotho polypeptide that is capable of
binding a fibroblast growth factor, and/or capable of enabling the
binding of a fibroblast growth factor to a fibroblast growth factor
receptor by binding to the fibroblast growth factor. The Klotho
extracellular domain corresponds to amino acid residues 28-982 of
the full length alpha Klotho sequence (SEQ ID NO: 2) and to amino
acid residues 52-997 of the full length beta Klotho sequence (SEQ
ID NO: 4).
[0098] "Extracellular subdomain of Klotho protein" and
"extracellular subdomain of Klotho protein" are used
interchangeably herein and shall refer to a region in the
extracellular domain of the Klotho polypeptide that is capable of
binding a fibroblast growth factor, and/or is capable of enabling
the binding of a fibroblast growth factor to a fibroblast growth
factor receptor by binding to the fibroblast growth factor. In
various embodiments, the fusion comprises a polypeptide comprising
at least one extracellular subdomain of a Klotho protein; a
polypeptide comprising a fibroblast growth factor; and, optionally,
a modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. The Klotho
extracellular domain has two homologous subdomains that are
repeated, i.e., KL-D1 (SEQ ID NO: 5) and KL-D2 (SEQ ID NO: 6).
KL-D1 and KL-D2 correspond respectively to amino acid residues
58-506 and 517-953 of the full length alpha Klotho polypeptide (SEQ
ID NO: 2) and respectively to amino acid residues 77-508 and
571-967 of the full length beta Klotho polypeptide (SEQ ID NO: 4)
and are suitable for use with the present disclosure. Generally, a
polypeptide that contains at least one Klotho subdomain is a Klotho
active polypeptide. The Klotho extracellular subdomain for use with
the polypeptide of the disclosure may be an alpha Klotho or beta
Klotho KL-D1 domain with an amino acid sequence that is
substantially identical to the amino acid sequence of SEQ ID NO: 5
or SEQ ID NO: 37, respectively. Further, the Klotho KL-D1 domain
may have an amino acid sequence that is at least 70%, 75%, 80%,
85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical to the amino
acid sequence of SEQ ID NO: 5 or SEQ ID NO: 37. The Klotho
extracellular subdomain may also be an alpha or beta Klotho
polypeptide KL-D2 domain that is substantially identical to the
amino acid sequence of SEQ ID NO: 6 or SEQ ID NO: 38, respectively.
In a further embodiment, the KL-D2 domain has an amino acid
sequence that is at least at least 70%, 75%, 80%, 85%, 90%, 95%,
96%, 97%, 98%, 99% or more identical to the amino acid sequence of
SEQ ID NO: 6 or SEQ ID NO: 38. In some embodiments, the fusion
comprises at least two extracellular subdomains of the Klotho
protein (e.g., KL-D1 and KL-D2; KL-D1 and KL-D1 in tandem repeats;
KL-D2 and KL-D2 in tandem repeats, etc.).
[0099] "Modified Fc fragment", as used herein, shall mean an Fc
fragment of an antibody comprising a modified sequence. The Fc
fragment is a portion of an antibody comprising the CH2, CH3 and
part of the hinge region. The modified Fc fragment can be derived
from, for example, IgG1, IgG2, IgG3, or IgG4. FcLALA is a modified
Fc fragment with a LALA mutation (L234A, L235A), which triggers
ADCC with lowered efficiency, and binds and activates human
complement weakly. Hessell et al. 2007 Nature 449:101-104.
Additional modifications to the Fc fragment are described in, for
example, U.S. Pat. No. 7,217,798. For example, in various modified
Fc fragments: (a) amino acid residue 250 is glutamic acid and amino
acid residue 428 is phenylalanine; or (b) amino acid residue 250 is
glutamine and amino acid residue 428 is phenylalanine; or (c) amino
acid residue 250 is glutamine and amino acid residue 428 is
leucine. In some embodiments, amino acid residues 250 and 428
differ from the residues present in an unmodified Fc-fusion protein
by amino acid residue 250 being glutamic acid or glutamine and
amino acid residue 428 being leucine or phenylalanine, and wherein
amino acid residues are numbered by the EU numbering system, as
described in U.S. Pat. No. 7,217,798. In some embodiments, the
modified Fc-fusion protein has a higher affinity for FcRn at pH 6.0
than at pH 8.0. Preferably, the modified Fc fragment has decreased
affinity to FcRn and/or increased serum half-life. Non-limiting
examples of modified Fc fragments include that at aa (amino acids)
1234-1459 of SEQ ID NO: 47; aa 1234 to 1450 of SEQ ID NO: 49; aa
257 to 482 of SEQ ID NO: 51; and aa 257 to 473 of SEQ ID NO: 53;
and sequences which are at least 70%, 75%, 80%, 85%, 90%, 95%, 96%,
97%, 98%, 99% or more or 100% identical to these sequences.
Nucleotides encoding these sequences are provided in SEQ ID NOs:
46, 48, 50 and 52.
[0100] "Signal peptide", as used herein, shall mean a peptide chain
(3-60 amino acids long) that directs the post-translational
transport of a protein to the endoplasmic reticulum and may be
cleaved off. Example signal peptides suitable for use with the
present disclosure include the Klotho signal peptide (SEQ ID NO:
19) and the IgG signal peptide (SEQ ID NO: 20). Note that upon
secretion and cleavage by the producer cell line, the signal
peptide (e.g., of the peptides corresponding to SEQ ID NO: 19 and
SEQ ID NO: 20) is cleaved off. Thus, after secretion and cleavage
of the signal peptide by the producer cell lines, the peptide of
SEQ ID NO: 19 would generate the peptide of SEQ ID NO: 41.
[0101] "Linker", as used herein, shall mean a functional group
(e.g., chemical or polypeptide) that covalently attaches two or
more polypeptides or nucleic acids so that they are connected with
one another. As used herein, a "peptide linker" refers to one or
more amino acids used to couple two proteins together (e.g., to
couple the extracellular domain of Klotho and fibroblast growth
factor-23). Peptide linkers suitable for use with the present
disclosure include, but are not limited to, polypeptides with amino
acid sequences represented by SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID
NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14,
SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17 and SEQ ID NO: 18. A
polypeptide linker can comprise at least 1 and up to about 30
repeats of any of these amino acid sequences.
[0102] "Operatively linked", as used herein, shall mean the linking
of two or more biomolecules so that the biological functions,
activities, and/or structure associated with the biomolecules are
at least retained. In reference to polypeptides, the term means
that the linking of two or more polypeptides results in a fusion
polypeptide that retains at least some of the respective individual
activities of each polypeptide component. The two or more
polypeptides may be linked directly or via a linker. In reference
to nucleic acids, the term means that a first polynucleotide is
positioned adjacent to a second polynucleotide that directs
transcription of the first polynucleotide when appropriate
molecules (e.g., transcriptional activator proteins) are bound to
the second polynucleotide.
[0103] "Specifically binds", as used herein, shall refer to the
ability of a first molecule to bind to a target molecule out of
many, different types of molecules to which it may be exposed
because of the ability of the first molecule to adopt a particular
structure conducive to forming non-covalent interactions between
itself and the other target molecule. The first molecule binds to
the target forming a stable complex while there is substantially
less recognition, contact, or complex formation of the first
molecule with any other non-specific molecules.
[0104] "Polypeptide variant" or "protein variant", as used herein,
refers to polypeptides in which one or more amino acids have been
substituted by different amino acids from a reference sequence. It
is well understood in the art that some amino acids may be
substituted by others with broadly similar properties without
changing the nature of the activity of the polypeptide
(conservative substitutions) as described hereinafter. These terms
also encompass polypeptides in which one or more amino acids have
been added or deleted, or replaced with different amino acids,
e.g., protein isoforms. An example variant of fibroblast growth
factor-23 suitable for use with the present disclosure is the
fibroblast growth factor-23 variant (R179Q).
[0105] "Pharmaceutical composition", as used herein, shall mean a
composition containing a compound (e.g., a fusion polypeptide of
the disclosure) that may be administered to treat or prevent a
disease or disorder in an individual.
[0106] "Individual" or "subject", as used herein, shall refer to a
mammal, including, but not limited to, a human or non-human mammal,
such as a bovine, equine, canine, ovine, or feline.
[0107] "Treat", as used herein, shall mean decrease, suppress,
attenuate, diminish, arrest, or stabilize the development or
progression of a disease. In the context of the disclosure, the
administration of the polypeptides of the disclosure may be used to
treat age-related conditions, including sarcopenia, skin atrophy,
muscle wasting, brain atrophy, atherosclerosis, arteriosclerosis,
pulmonary emphysema, osteoporosis, osteoarthritis, immunologic
incompetence, high blood pressure, dementia, Huntington's disease,
Alzheimer's disease, cataracts, age-related macular degeneration,
prostate cancer, stroke, diminished life expectancy, memory loss,
wrinkles, impaired kidney function, and age-related hearing loss;
and metabolic disorders, including Type II Diabetes, Metabolic
Syndrome, hyperglycemia, and obesity.
[0108] "Prevent", as used herein, shall refer to a decrease in the
occurrence of a disorder or decrease in the risk of acquiring a
disorder or its associated symptoms in a subject. In the context of
the disclosure, the administration of the polypeptides of the
disclosure may be used to prevent age-related conditions, including
sarcopenia, skin atrophy, muscle wasting, brain atrophy,
atherosclerosis, arteriosclerosis, pulmonary emphysema,
osteoporosis, osteoarthritis, immunologic incompetence, high blood
pressure, dementia, Huntington's disease, Alzheimer's disease,
cataracts, age-related macular degeneration, prostate cancer,
stroke, diminished life expectancy, memory loss, wrinkles, impaired
kidney function, and age-related hearing loss; and metabolic
disorders, including Type II Diabetes, Metabolic Syndrome,
hyperglycemia, and obesity. The prevention may be complete, e.g.,
the total absence of an age-related condition or metabolic
disorder. The prevention may also be partial, such that the
likelihood of the occurrence of the age-related condition or
metabolic disorder in a subject is less likely to occur than had
the subject not received the present disclosure.
[0109] "Disease", as used herein, shall mean any condition or
disorder that damages or interferes with the normal function of a
cell, tissue, or organ.
[0110] "Age-related condition", as used herein, shall mean any
disease or disorder whose incidence in a population or severity in
an individual correlates with the progression of age. In one
embodiment, the age-related condition is a disease or disorder
whose incidence is at least 1.5 fold higher among human individuals
greater than 60 years of age relative to human individuals between
the ages of 30-40 and in a selected population of greater than
100,000 individuals. Age-related conditions relevant to the present
disclosure include, but are not limited to, sarcopenia, skin
atrophy, muscle wasting, brain atrophy, atherosclerosis,
arteriosclerosis, pulmonary emphysema, osteoporosis,
osteoarthritis, immunologic incompetence, high blood pressure,
dementia, Huntington's disease, Alzheimer's disease, cataracts,
age-related macular degeneration, prostate cancer, stroke,
diminished life expectancy, memory loss, wrinkles, impaired kidney
function, and age-related hearing loss.
[0111] "Metabolic disorder", as used herein, shall mean any disease
or disorder that damages or interferes with normal function in a
cell, tissue, or organ by affecting the production of energy in
cells or the accumulation of toxins in a cell, tissue, organ, or
individual. Metabolic disorders relevant to the present disclosure
include, but are not limited to, Type II Diabetes, Metabolic
Syndrome, hyperglycemia, and obesity.
[0112] An "effective dose" or "effective amount" is an amount
sufficient to effect a beneficial or desired clinical result. In
the context of the disclosure, it is an amount of a Klotho fusion
polypeptide or sKlotho effective to produce the intended
pharmacological, therapeutic or preventive result. A
therapeutically effective dose results in the prevention or
amelioration of the disorder or one or more symptoms of the
disorder, (e.g., an age-related condition or metabolic disorder).
Therapeutically effective doses will vary depending upon the
subject and disease condition being treated, the weight and age of
the subject, the severity of the disease condition, the manner of
administration and the like which can be readily be determined by
one of ordinary skill in the art.
[0113] "Klotho nucleic acid molecule", as used herein is a gene
encoding a Klotho protein. An example human Klotho gene is provided
at GenBank Accession No. NM_004795 (SEQ ID NO: 1). Additional
non-limiting examples of Klotho are provided at aa 1-982 of SEQ ID
NO: 47 and aa 1-982 of SEQ ID NO: 49; and sequences which are at
least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more or
100% identical to these sequences.
[0114] "Fragment", as used herein, refers to a portion of a
polypeptide or nucleic acid molecule. This portion contains,
preferably, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or
more of the entire length of the reference nucleic acid molecule or
polypeptide. A fragment may contain 10, 20, 30, 40, 50, 60, 70, 80,
90, or 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000 or up to
3000 nucleotides or amino acids.
[0115] The term "substantially identical" refers to a polypeptide
or nucleic acid molecule exhibiting at least 50% identity to a
reference amino acid sequence (for example, any one of the amino
acid sequences described herein) or nucleic acid sequence (for
example, any one of the nucleic acid sequences described herein).
Preferably, such a sequence is at least 60%, 70%, 75%, 80% or 85%,
90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical
at the amino acid level or nucleic acid to the sequence used for
comparison.
[0116] The present disclosure is directed to methods, kits and
compositions for preventing or treating age-related conditions and
metabolic disorders; and to the use of said compostions in therapy,
as a medicament or for use in the treatment of a pathological
disorder. In some embodiments, the disclosure provides a fusion
polypeptide having at least one extracellular subdomain of a Klotho
protein. In some embodiments, the fusion polypeptides further
comprise a fibroblast growth factor or an active fragment thereof.
In some embodiments, the fusion further comprises a modified Fc
fragment having decreased affinity for Fc-gamma-receptorand/or
increased serum half-life. In other embodiments, the fusion
comprises an FGF (e.g., FGF19, FGF21, FGF23 or FGF23 variant R179Q)
fused to a modified Fc (e.g., FcLALA). FcLALA is a Fc fragment with
a LALA mutation (L234A, L235A), which triggers ADCC with lowered
efficiency, and binds and activates human complement weakly. The
Klotho extracellular domain may be derived from either the alpha or
beta Klotho isoforms. Further, although the FGF component of the
Klotho fusion polypeptide is described primarily with reference to
fibroblast growth factor-19, fibroblast growth factor-21 and
fibroblast growth factor-23, it is contemplated that any of the
twenty-three known FGFs or an active fragment thereof can be used
in practicing the disclosure.
[0117] The extracellular domain of the Klotho protein can include
one or both of the KL-D1 and KL-D2 domains of a Klotho protein. In
some embodiments, the Klotho fusion polypeptide has at least two
extracellular subdomains of a Klotho protein. For example, the at
least two extracellular subdomains can be at least two KL-D1
domains in tandem repeats, at least two KL-D2 domains in tandem
repeats, or at least one KL-D1 domain and at least one KL-D2
domain.
[0118] The extracellular subdomain of a Klotho protein and the
fibroblast growth factor (or an active fragment thereof) can be
operatively linked to one another in a variety of orientations and
manners. For example, the extracellular subdomain of the Klotho
protein can be operatively linked to the N-terminus of the
fibroblast growth factor or alternatively the fibroblast growth
factor can be operatively linked to the N-terminus of the at least
one extracellular subdomain of the Klotho protein.
[0119] The fusion polypeptide of the disclosure may include one or
both of the Klotho extracellular domains, i.e., KL-D1 (SEQ ID NO:
5) and KL-D2 (SEQ ID NO: 6). KL-D1 and KL-D2 correspond
respectively to amino acid residues 58-506 and 517-953 of the full
length alpha Klotho polypeptide (SEQ ID NO: 2) and to amino acid
residues 77-508 and 571-967 of the full length beta Klotho
polypeptide (SEQ ID NO: 4) and are suitable for use with the
present disclosure. The Klotho fusion polypeptide may have a KL-D1
domain of an alpha Klotho polypeptide having an amino acid sequence
that is substantially identical to the amino acid sequence of SEQ
ID NO: 5 or of a beta Klotho polypeptide having an amino acid
sequence that is substantially identical to the amino acid sequence
of SEQ ID NO: 37. Specifically, the Klotho fusion polypeptide may
have an amino acid sequence that is at least at least 70%, 75%,
80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more identical to SEQ ID
NO: 5 or SEQ ID NO: 37. The Klotho fusion polypeptide may have a
KL-D2 domain of an alpha Klotho polypeptide with an amino acid
sequence that is substantially identical to the amino acid sequence
of SEQ ID NO: 6 or of a beta Klotho polypeptide having an amino
acid sequence that is substantially identical to the amino acid
sequence of SEQ ID NO: 38. Specifically, the Klotho fusion
polypeptide may have an amino acid sequence that is at least at
least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more
identical to SEQ ID NO: 6 or SEQ ID NO: 38, respectively.
[0120] In some embodiments, the Klotho fusion polypeptide of the
disclosure is soluble and is capable of binding to an FGF
receptor.
[0121] The Klotho fusion polypeptides of the disclosure can contain
a polypeptide linker which connects the polypeptide having at least
one extracellular subdomain of a Klotho protein and the fibroblast
growth factor and the (optional) modified Fc fragment. Suitable
linkers are well known in the art and generally contain several Gly
and several Ser residues, e.g., (Gly.sub.4 Ser).sub.3 (SEQ ID NO:
11), Gly.sub.4 Ser polypeptide (SEQ ID NO: 12), Gly (SEQ ID NO:
13), Gly Gly (SEQ ID NO: 14), Gly Ser (SEQ ID NO: 15), Gly.sub.2
Ser (SEQ ID NO: 16), Ala (SEQ ID NO: 17), and Ala Ala (SEQ ID NO:
18). In some embodiments, the linker will have at least 2 and up to
about 30 repeats of an amino acid sequence represented by any one
of SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ
ID NO: 16, SEQ ID NO: 17, or SEQ ID NO: 18.
[0122] When a polypeptide linker is present in the Klotho fusion
polypeptide of the disclosure, the polypeptide having at least one
extracellular subdomain of a Klotho protein may be connected by a
peptide bond to the N-terminus of the linker polypeptide with the
FGF connected by a peptide bond to the C-terminus of the
polypeptide linker. Alternatively, the FGF may be connected by a
peptide bond to the N-terminus of the linker polypeptide with the
polypeptide having at least one extracellular subdomain of Klotho
connected by a peptide bond to the C-terminus of the polypeptide
linker. A chemical linker can also be used to link the two
polypeptides.
[0123] The Klotho fusion polypeptide of the disclosure may include
a signal peptide. Example signal peptides for use with the Klotho
fusion polypeptide include, but are not limited to the Klotho
signal peptide (SEQ ID NO: 8) and the IgG signal peptide (SEQ ID
NO: 9).
[0124] In some embodiments, the disclosure provides a fusion
between a FGF (e.g., FGF19, FGF21, FGF23, or FGF23 variant R179Q)
and a modified Fc (e.g., FcLALA). The fusion can also optionally
comprise linkers between the FGF and Fc portions. The fusion can
also optionally comprise a signal peptide. In various embodiments,
the disclosure encompasses nucleic acids encoding these fusion
polypeptides, vectors comprising these nucleic acids, and host
cells containing these nucleic acids.
4.1. Klotho and Fibroblast Growth Factor Polypeptides
[0125] The Klotho fusion polypeptides of the disclosure are
expected to exhibit biological activities comparable to FGF in
nature, such as binding to an FGF receptor and inducing the
phosphorylation of an FGF receptor, FRS2 (FGF receptor substrate 2)
and ERK1/2 (extracellular signal-regulated protein kinase 1/2) and
activating Egr-1 (early growth response-1) gene. FGF is a secreted
peptide growth factor that binds the FGF receptor. The amino acid
and nucleic acid sequences of FGF are readily available to those of
skill in the art. For example, example nucleotide sequences for
FGF19, FGF21, and FGF23 can be found in the GenBank database at
Accession numbers: NM_005117, NM_019113, and NM_020638,
respectively, and herein as SEQ ID NOs: 30, 32, and 34,
respectively. Example amino sequences for FGF19, FGF21, and FGF23
can be found in the GenBank database at Accession numbers:
NP_005108, NP_061986, and NP_065689, respectively, and herein as
SEQ ID NOs: 31, 35, and 35, respectively. Additionally, FGF may
include one or more alterations which aid in the expression of the
protein, e.g., the FGF23 (R179Q) variant (SEQ ID NO: 36).
[0126] The Klotho protein is a 130 kDa single pass type I
transmembrane protein with an extracellular domain and a short
cytoplasmic domain. The amino acid and nucleic acid sequences of
Klotho are readily available to those of skill in the art. For
example, example nucleotide sequences for alpha-Klotho and
beta-Klotho can be found in the GenBank database at Accession
numbers: NM_004795 and NM_175737, respectively, and herein as SEQ
ID NOs: 7 and 8, respectively. Example amino acid sequences for
alpha-Klotho and beta-Klotho can be found in the GenBank database
at Accession numbers: NP_004786 and NP_783864, respectively, and
herein as SEQ ID NOs: 2 and 4, respectively.
[0127] The Klotho fusion polypeptide of the disclosure can bind to
a fibroblast growth factor receptor and has an alpha-Klotho or
beta-Klotho extracellular domain operatively linked to either
fibroblast growth factor-19 (SEQ ID NO: 31), fibroblast growth
factor-21 (SEQ ID NO: 33), fibroblast growth factor-23 (SEQ ID NO:
35), or variants thereof (which include fibroblast growth factor-23
variant (R179Q) (SEQ ID NO: 36)).
[0128] Specifically, the Klotho fusion polypeptide of the
disclosure may include an alpha-Klotho (SEQ ID NO: 2) which is
operatively coupled to fibroblast growth factor-23 (SEQ ID NO: 35)
or fibroblast growth factor-23 variant (R179Q) (SEQ ID NO: 36).
Additionally, the Klotho fusion polypeptide of the disclosure may
have beta-Klotho (SEQ ID NO: 4), which is operatively coupled to
fibroblast growth factor-19 (SEQ ID NO: 31). The Klotho fusion
polypeptide of the disclosure may include a beta-Klotho (SEQ ID NO:
4), which is operatively coupled to fibroblast growth factor-21
(SEQ ID NO: 33).
[0129] The disclosure includes homologs of the various Klotho and
FGF genes and proteins encoded by those genes. A "homolog," in
reference to a gene refers to a nucleotide sequence that is
substantially identical over at least part of the gene or to its
complementary strand or a part thereof, provided that the
nucleotide sequence encodes a protein that has substantially the
same activity/function as the protein encoded by the gene which it
is a homolog of Homologs of the genes described herein can be
identified by percent identity between amino acid or nucleotide
sequences for putative homologs and the sequences for the genes or
proteins encoded by them (e.g., nucleotide sequences for genes
encoding Klotho and FGF or their complementary strands). Percent
identity may be determined, for example, by visual inspection or by
using various computer programs known in the art or as described
herein. Sequence identity is typically measured using sequence
analysis software (for example, Sequence Analysis Software Package
of the Genetics Computer Group, University of Wisconsin
Biotechnology Center, 1710 University Avenue, Madison, Wis. 53705,
BLAST, BESTFIT, GAP, or PILEUP/PRETTYBOX programs). Such software
matches identical or similar sequences by assigning degrees of
homology to various substitutions, deletions, and/or other
modifications. Conservative amino acid substitutions typically
include substitutions within the following groups: [0130] glycine
and alanine; [0131] valine, isoleucine and leucine; [0132] aspartic
acid, glutamic acid, asparagine and glutamine; [0133] serine and
threonine; [0134] lysine and arginine; and [0135] phenylalanine and
tyrosine.
[0136] Thus, mutating a glycine to alanine would be a conservative
amino acid substititon, as would mutating an alanine to a glycine;
mutating a valine to an isoleucine or leucine would be a
conservative amino acid substation, as would replacing an
isoleucine with valine or leucine, as would replacing leucine with
valine or isoleucine, etc. The disclosure provides variants of all
the amino acid sequences disclosed herein with at least one
conservative amino acid substitution.
[0137] In an example approach to determining the degree of
identity, a BLAST program may be used, with a probability score
between e.sup.-3 and e.sup.-100 indicating a closely related
sequence.
[0138] In one embodiment, the present disclosure provides a fusion
polypeptide of SEQ ID NO: 19.
[0139] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 20.
[0140] In one embodiment, the present disclosure provides a fusion
polypeptide of SEQ ID NO: 40.
[0141] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 41, or a variant thereof
comprising at least one conservative amino acid substition.
[0142] In one embodiment, the present disclosure provides a fusion
polypeptide of SEQ ID NO: 46.
[0143] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 47, or a variant thereof
comprising at least one conservative amino acid substition.
[0144] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 48.
[0145] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 49, or a variant thereof
comprising at least one conservative amino acid substition.
[0146] In one embodiment, the present disclosure provides a fusion
polypeptide of SEQ ID NO: 50.
[0147] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 51, or a variant thereof
comprising at least one conservative amino acid substition.
[0148] In one embodiment, the present disclosure provides a fusion
polypeptide of SEQ ID NO: 52.
[0149] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 53, or a variant thereof
comprising at least one conservative amino acid substition.
[0150] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 54, or a variant thereof
comprising at least one conservative amino acid substition.
[0151] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 55, or a variant thereof
comprising at least one conservative amino acid substition.
[0152] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 56, or a variant thereof
comprising at least one conservative amino acid substition.
[0153] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 57, or a variant thereof
comprising at least one conservative amino acid substition.
[0154] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 58, or a variant thereof
comprising at least one conservative amino acid substition.
[0155] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 59, or a variant thereof
comprising at least one conservative amino acid substition.
[0156] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 60, or a variant thereof
comprising at least one conservative amino acid substition.
[0157] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 61, or a variant thereof
comprising at least one conservative amino acid substition.
[0158] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 62, or a variant thereof
comprising at least one conservative amino acid substition.
[0159] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 63, or a variant thereof
comprising at least one conservative amino acid substition.
[0160] In another embodiment, the present disclosure provides a
fusion polypeptide of
[0161] SEQ ID NO: 64, or a variant thereof comprising at least one
conservative amino acid substition.
[0162] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 65, or a variant thereof
comprising at least one conservative amino acid substition.
[0163] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 66, or a variant thereof
comprising at least one conservative amino acid substition.
[0164] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 67, or a variant thereof
comprising at least one conservative amino acid substition.
[0165] In another embodiment, the present disclosure provides a
fusion polypeptide of SEQ ID NO: 68, or a variant thereof
comprising at least one conservative amino acid substition.
[0166] As used herein, the terms "homology" and "homologous" are
not limited to designate proteins having a theoretical common
genetic ancestor, but includes proteins which may be genetically
unrelated that have, nonetheless, evolved to perform similar
functions and/or have similar structures. Functional homology to
the various proteins described herein also encompasses proteins
that have an activity of the corresponding protein of which it is a
homolog. For proteins to have functional homology, it is not
required that they have significant identity in their amino acid
sequences, but, rather, proteins having functional homology are so
defined by having similar or identical activities. For example,
with respect to a Klotho molecule, the polypeptide should have the
functional characteristics of binding to an FGF polypeptide and
enable the binding of the FGF to an FGFR. With respect to an FGF
molecule, the polypeptide should have the functional
characteristics of binding to an FGFR and causing the activation of
FGFR (e.g., phosphorylation). Assays for assessing FGF binding to
the FGF receptor and/or activation of the FGF signaling pathway are
known in the art and described herein (See Example 2). Assays for
assessing Klotho activity are also known in the art and described
herein (e.g., binding to a FGF polypeptide). Proteins with
structural homology are defined as having analogous tertiary (or
quaternary) structure and do not necessarily require amino acid
identity or nucleic acid identity for the genes encoding them. In
certain circumstances, structural homologs may include proteins
which maintain structural homology only at the active site or
binding site of the protein.
[0167] In addition to structural and functional homology, the
present disclosure further encompasses proteins having amino acid
identity to the various Klotho and FGF amino acid sequences
described herein. To determine the percent identity/homology of two
amino acid sequences, the sequences are aligned for optimal
comparison purposes (e.g., gaps can be introduced in the amino acid
sequence of one protein for optimal alignment with the amino acid
sequence of another protein). The amino acid residues at
corresponding amino acid positions are then compared. When a
position in one sequence is occupied by the same amino acid residue
as the corresponding position in the other, then the molecules are
identical at that position. The percent identity between the two
sequences is a function of the number of identical positions shared
by the sequences (i.e., % identity=# of identical positions/total #
of positions multiplied by 100).
[0168] The amino acid sequences of molecules of the disclosure
described herein have an amino acid sequence which is at least
about 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or more identical
or homologous to an amino acid sequence described herein.
[0169] The nucleic acid sequences of molecules of the disclosure
described herein have a nucleotide sequence which hybridizes to or
is at least about 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or
more identical or homologous to a nucleotide sequence described
herein.
[0170] Nucleic acid molecules appropriate for use in the fusion
polypeptides of the disclosure may have a Klotho or FGF nucleotide
sequence which hybridizes under stringent conditions to the
complement of a nucleic acid molecule encoding Klotho or FGF,
respectively. As used herein, the term "hybridizes under stringent
conditions" is intended to describe conditions for hybridization
and washing under which nucleotide sequences at least about 70%,
80%, 85%, 90% or more homologous to each other typically remain
hybridized to each other. Such stringent conditions are known to
those skilled in the art and can be found in Ausubel et al. Current
Protocols in Molecular Biology, Wiley Interscience, New York
(2001), 6.3.1-6.3.6. A specific, non-limiting example of stringent
hybridization conditions are hybridization in 6.times. sodium
chloride/sodium citrate (SSC) at about 45.degree. C., followed by
one or more washes in 0.2.times.SSC, 0.1% SDS at 50-65.degree.
C.
4.2. Klotho-FGF Fusion Polypeptides of the Disclosure
[0171] In some embodiments of the disclosure, a Klotho fusion
polypeptide has a polypeptide chain having a first polypeptide
sequence of a Klotho polypeptide or an active fragment thereof and
a second polypeptide sequence encoding FGF or an active fragment
thereof. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life.
[0172] The disclosure includes fusion polypeptides which are at
least about 95% or more homologous to an amino acid sequence
presented in SEQ ID NO: 19-28. The amino acid sequence of SEQ ID
NO: 19 encodes a Klotho fusion polypeptide having a Klotho
extracellular domain N-terminally linked to the FGF23 (R179Q)
variant (SEQ ID NO: 36). The amino acid sequence of SEQ ID NO: 20
encodes a Klotho fusion polypeptide having an IgG signal peptide
N-terminally linked to a Klotho extracellular domain lacking a
signal peptide N-terminally linked to the FGF23 (R179Q) variant.
The amino acid sequence of SEQ ID NO: 21 encodes a Klotho fusion
polypeptide having a KL-D1 extracellular subdomain N-terminally
linked to the FGF23 (R179Q) variant. The amino acid sequence of SEQ
ID NO: 22 encodes a Klotho fusion polypeptide having a KL-D2
extracellular subdomain N-terminally linked to the FGF23 (R179Q)
variant. The amino acid sequence of SEQ ID NO: 23 encodes a Klotho
fusion polypeptide having two KL-D1 extracellular subdomains
N-terminally linked to the FGF23 (R179Q) variant. The amino acid
sequence of SEQ ID NO: 24 encodes a Klotho fusion polypeptide
having two KL-D2 extracellular subdomains N-terminally linked to
the FGF23 (R179Q) variant. The amino acid sequence of SEQ ID NO: 25
encodes a Klotho fusion polypeptide having the FGF23 (R179Q)
variant N-terminally linked to a Klotho extracellular domain. The
amino acid sequence of SEQ ID NO: 26 encodes a Klotho fusion
polypeptide having the FGF23 (R179Q) variant N-terminally linked to
a KL-D1 extracellular subdomain. The amino acid sequence of SEQ ID
NO: 27 encodes a Klotho fusion polypeptide having the FGF23 (R179Q)
variant N-terminally linked to a KL-D2 extracellular subdomain. The
amino acid sequence of SEQ ID NO: 28 encodes a Klotho fusion
polypeptide having the FGF23 (R179Q) variant N-terminally linked to
two KL-Dl extracellular subdomains. The amino acid sequence of SEQ
ID NO: 29 encodes a Klotho fusion polypeptide having the FGF23
(R179Q) variant N-terminally linked to two KL-D2 extracellular
subdomains. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life.
[0173] The Klotho fusion polypeptide of the disclosure may include
an amino acid sequence which is at least about 95% identical to the
amino acid sequence set forth in SEQ ID NO: 7. The amino acid
sequence of SEQ ID NO: 7 encodes a Klotho extracellular domain
lacking a signal peptide. In some embodiments, the fusion further
comprises a modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life.
[0174] The subject fusion proteins are described herein and can be
made using methods known in the art. For example, the fusion
polypeptides of the disclosure may be constructed as described in
U.S. Pat. No. 6,194,177. The use of Klotho polypeptides is
described in U.S. Pat. No. 6,579,850. The use of FGF nucleic acid
molecules is described in U.S. Pat. No. 7,223,563.
[0175] In some embodiments, a nucleic acid molecule encoding the
Klotho is cloned by PCR and ligated, in frame, with a nucleic acid
molecule encoding FGF. In some embodiments, the fusion further
comprises a modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. The nucleic
acid encoding the fusion polypeptide is operatively linked to a
promoter to allow for expression. The nucleic acid molecule
encoding the fusion polypeptide is subsequently transfected into a
host cell for expression. The sequence of the final construct can
be confirmed by sequencing.
[0176] When preparing the fusion proteins of the present
disclosure, a nucleic acid molecule encoding an extracellular
subdomain of Klotho will be fused in frame to the nucleic acid
molecule encoding FGF and the (optional) nucleic acid encoding the
modified Fc fragment. Expression of the resulting nucleic acid
molecule results in the extracellular subdomain of Klotho being
fused N-terminal in relation to the FGF polypeptide. Fusions are
also possible in which the extracellular subdomain of Klotho is
fused C-terminal in relation to the FGF polypeptide. Methods for
making fusion proteins are well known in the art.
[0177] The fusion polypeptides of the disclosure have at least two
polypeptides that are covalently linked, in which one polypeptide
comes from one protein sequence or domain, e.g., Klotho, and the
other polypeptide comes from another protein sequence or domain,
e.g., FGF. In some embodiments, the fusion further comprises a
modified Fc fragment having decreased affinity for
Fc-gamma-receptor and/or increased serum half-life. In another
embodiment, the disclosure comprises a FGF fused to a modified Fc
fragment. Klotho and/or FGF and/or the (optional) modified Fc
fragment, of the fusion polypeptides of the disclosure, can be
joined by methods well known to those of skill in the art. These
methods include both chemical and recombinant means.
[0178] Nucleic acids encoding the domains to be incorporated into
the fusion polypeptides of the disclosure can be obtained using
routine techniques in the field of recombinant genetics. Basic
texts disclosing the general methods of use in this disclosure
include Sambrook and Russell, Molecular Cloning, A Laboratory
Manual (3rd ed. 2001); Kriegler, Gene Transfer and Expression: A
Laboratory Manual (1990); and Current Protocols in Molecular
Biology (Ausubel et al., eds., 1994-1999). In nucleic acids
encoding a Klotho fusion polypeptide of the disclosure, the nucleic
acid sequence encoding alpha-Klotho or beta-Klotho, represented by
SEQ ID NO: 1 and SEQ ID NO: 3, respectively, may be used. In
nucleic acids encoding a Klotho fusion polypeptide, the nucleic
acid sequence encoding FGF19, FGF21, or FGF23, represented by SEQ
ID NO: 30, SEQ ID NO: 32 and SEQ ID NO: 34, respectively, may be
used. Nucleic acid sequences of molecules of the disclosure
described herein comprise a nucleotide sequence which hybridizes to
or is at least about 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or
more identical or homologous to SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID
NO: 30, SEQ ID NO: 32, or SEQ ID NO: 34.
[0179] Nucleic acid sequences that encode the various components of
the fusion [Klotho, and/or FGF peptide and/or the (optional)
modified Fc fragment] can be obtained using any of a variety of
methods. For example, the nucleic acid sequences encoding the
polypeptides may be cloned from cDNA and genomic DNA libraries by
hybridization with probes, or isolated using amplification
techniques with oligonucleotide primers. More commonly,
amplification techniques are used to amplify and isolate the Klotho
and FGF sequences using a DNA or RNA template (see, e.g.,
Dieffenfach & Dveksler, PCR Primers: A Laboratory Manual
(1995)). Alternatively, overlapping oligonucleotides can be
produced synthetically and joined to produce one or more of the
domains. Nucleic acids encoding Klotho or FGF can also be isolated
from expression libraries using antibodies as probes.
[0180] According to the present disclosure, the various components
of the fusion [Klotho, and/or, FGF and/or the (optional) modified
Fc fragment] can be linked either directly or via a covalent
linker, including amino acid linkers, such as a polyglycine linker,
or another type of chemical linker, including, carbohydrate
linkers, lipid linkers, fatty acid linkers, polyether linkers, such
as PEG, etc. (See for example, Hermanson, Bioconjugate techniques
(1996)). The polypeptides forming the fusion/fusion polypeptide are
typically linked C-terminus to N-terminus, although they can also
be linked C-terminus to C-terminus, N-terminus to N-terminus, or
N-terminus to C-terminus. One or more polypeptide domains may be
inserted at an internal location within a fusion polypeptide of the
disclosure. The polypeptides of the fusion protein can be in any
order. The fusion polypeptides may be produced by covalently
linking a chain of amino acids from one protein sequence, e.g., an
extracellular subdomain of Klotho, to a chain of amino acids from
another protein sequence, e.g., FGF, by preparing a recombinant
polynucleotide contiguously encoding the fusion protein. The
different chains of amino acids in a fusion protein may be directly
spliced together or may be indirectly spliced together via a
chemical linking group or an amino acid linking group. The amino
acid linking group can be about 200 amino acids or more in length,
or generally 1 to 100 amino acids. In some embodiments, proline
residues are incorporated into the linker to prevent the formation
of significant secondary structural elements by the linker. Linkers
can often be flexible amino acid subsequences that are synthesized
as part of a recombinant fusion protein. Such flexible linkers are
known to persons of skill in the art.
[0181] According to the present disclosure, the amino acid
sequences of the fusion [an extracellular subdomain of Klotho
and/or the FGF and/or the (optional) modified Fc fragment] may be
linked via a peptide linker. Example peptide linkers are well known
in the art and described herein. For example, peptide linkers
generally include several Gly and several Ser residues, such as:
(Gly.sub.4 Ser).sub.3 (SEQ ID NO: 11), Gly.sub.4 Ser polypeptide
(SEQ ID NO: 12), Gly (SEQ ID NO: 13), Gly Gly (SEQ ID NO: 14), Gly
Ser (SEQ ID NO: 15), Gly.sub.2 Ser (SEQ ID NO: 16), Ala (SEQ ID NO:
17), and Ala Ala (SEQ ID NO: 18). Specifically, a peptide linker
for use in a fusion protein of the disclosure may act as a flexible
hinge.
[0182] The signal sequence of Klotho or FGF may be excluded prior
to incorporation of Klotho into a fusion protein of the disclosure.
The signal sequence for Klotho or FGF of the fusion protein may be
included, e.g., the polypeptide represented by SEQ ID NO: 19.
However, such sequences may also be omitted and replaced with the
signal sequence of a different protein, e.g., the IgG signal
sequence (SEQ ID NO: 9). Generally, the pharmaceutical compositions
of the disclosure will contain the mature form of Klotho and
FGF.
[0183] Generally, introns are excluded from either one or both the
Klotho or the FGF moieties prior to incorporation into a fusion
polypeptide.
[0184] The fusion polypeptides of the disclosure may include one or
more polymers covalently attached to one or more reactive amino
acid side chains. By way of example, not limitation, such polymers
include polyethylene glycol (PEG), which can be attached to one or
more free cysteine sulfhydryl residues, thereby blocking the
formation of disulfide bonds and aggregation when the protein is
exposed to oxidizing conditions. In addition, PEGylation of the
fusion polypeptides of the disclosure is expected to provide such
improved properties as increased half-life, solubility, and
protease resistance. The fusion polypeptides of the disclosure may
alternatively be modified by the covalent addition of polymers to
free amino groups such as the lysine epsilon or the N-terminal
amino group. Particular specific cysteines and lysines for covalent
modification will be those not involved in receptor binding,
heparin binding, or in proper protein folding. It will be apparent
to one skilled in the art that the methods for assaying the
biochemical and/or biological activity of the fusion polypeptides
may be employed in order to determine if modification of a
particular amino acid residue affects the activity of the protein
as desired. Other similar suitable modifications are contemplated
and known in the art.
[0185] The disclosure is also directed to the expression of a
fusion polypeptide that is at least about 95% or more homologous to
an amino acid sequence presented in SEQ ID NO: 19-28.
[0186] The present disclosure encompasses a fusion polypeptide
comprising: (a) a polypeptide comprising at least one extracellular
subdomain of a Klotho protein, or a functionally active variant or
derivative thereof; (b) a polypeptide comprising a fibroblast
growth factor, or a functionally active variant or derivative
thereof; and (c) a modified Fc fragment having decreased affinity
for Fc-gamma-receptor and/or increased serum half-life. By "a
functionally active variant or derivative thereof" is meant a
variant or derivative comprising a longer, shorter or altered amino
acid sequence than the corresponding wild-type polypeptide, while
retaining the biological activity. Thus "a functionally active
variant or derivative" of an extracellular subdomain of a Klotho
protein or a fibroblast growth factor comprises fewer, more, or an
altered amino acid sequence than a wild-type extracellular
subdomain of a Klotho protein or a fibroblast growth factor, but
still retains at least one biological activity of the wild-type
polypeptide sequence. A functionally active variant or derivative
of a polypeptide disclosed herein can also comprise the same amino
acid sequence of a polypeptide disclosed herein, but vary in
post-translational modification (e.g., pegylation, methylation
and/or glycosylation), or have additional moieties or elements
added to it. In various embodiments, the variant or derivative of
FGF23 comprises R179Q or does not.
[0187] In one embodiment, a functionally active variant or
derivative polypeptide includes an amino acid sequence at least
about 60% identical to a sequence disclosed herein (e.g., at least
one extracellular domain of a Klotho protein or a fibroblast growth
factor). Preferably, the polypeptide is at least 55%, 60%, 65%,
70%, 75%, 80%, 85%, 90%, 95%, 98%, 99% or more identical to a
sequence disclosed herein.
[0188] As used herein, percent identity of two amino acid sequences
(or of two nucleic acid sequences) is determined using the
algorithm of Karlin and Altschul (PNAS USA 87:2264-2268, 1990),
modified as in Karlin and Altschul, PNAS USA 90:5873-5877, 1993).
Such an algorithm is incorporated into the NBLAST and XBLAST
programs of Altschul et al. (J. Mol. Biol. 215:403-410, 1990).
BLAST nucleotide searches are performed with the NBLAST program,
score=100, wordlength=12. BLAST protein searches are performed with
the XBLAST program, score=50, wordlength=3. To obtain gapped
alignment for comparison purposes GappedBLAST is utilized as
described in Altschul et al. (Nucleic Acids Res. 25:3389-3402,
1997). When utilizing BLAST and GappedBLAST programs the default
parameters of the respective programs (e.g., XBLAST and NBLAST) are
used to obtain nucleotide sequences homologous to a nucleic acid
molecule of the invention.
[0189] Identity or identical means amino acid sequence (or nucleic
acid sequence) similarity and has an art recognized meaning.
Sequences with identity share identical or similar amino acids (or
nucleic acids). Thus, a candidate sequence sharing 85% amino acid
sequence identity with a reference sequence requires that,
following alignment of the candidate sequence with the reference
sequence, 85% of the amino acids in the candidate sequence are
identical to the corresponding amino acids in the reference
sequence, and/or constitute conservative amino acid changes.
[0190] Functionally active variants of a polypeptide disclosed
herein retain substantially the same functional activity of the
original polypeptide or fragment. Naturally occurring functionally
active variants such as allelic variants and species variants and
non-naturally occurring functionally active variants are included
in the invention and can be produced by, for example, mutagenesis
techniques or by direct synthesis.
[0191] A functionally active variant or derivative differs by about
or at least, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,
13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60 or
more amino acid residues from a polypeptide disclosed herein. Where
this comparison requires alignment the sequences are aligned for
maximum homology. The site of variation can occur anywhere in the
polypeptide, as long as activity substantially similar to a
polypeptide disclosed herein.
[0192] Guidance concerning how to make variants and derivatives
with phenotypically silent amino acid substitutions is provided in
Bowie et al., Science, 247:1306-1310 (1990), which teaches that
there are two main strategies for studying the tolerance of an
amino acid sequence to change.
[0193] The first strategy exploits the tolerance of amino acid
substitutions by natural selection during the process of evolution.
By comparing amino acid sequences in different species, the amino
acid positions which have been conserved between species can be
identified. See e.g., FIG. 5. These conserved amino acids are
likely important for protein function. In contrast, the amino acid
positions in which substitutions have been tolerated by natural
selection indicate positions which are not critical for protein
function. Thus, positions tolerating amino acid substitution can be
modified while still maintaining specific binding activity of the
polypeptide.
[0194] The second strategy uses genetic engineering to introduce
amino acid changes at specific positions of a cloned gene to
identify regions critical for protein function. For example,
site-directed mutagenesis or alanine-scanning mutagenesis (the
introduction of single alanine mutations at every residue in the
molecule) can be used (Cunningham et al., Science, 244:1081-1085
(1989)).
[0195] Methods of introducing a mutation into amino acids of a
protein is well known to those skilled in the art. See, e.g.,
Ausubel (ed.), Current Protocols in Molecular Biology, John Wiley
and Sons, Inc. (1994); T. Maniatis, E. F. Fritsch and J. Sambrook,
Molecular Cloning: A Laboratory Manual, Cold Spring Harbor
laboratory, Cold Spring Harbor, N.Y. (1989)). Mutations can also be
introduced using commercially available kits such as
"QuikChange..TM.. Site-Directed Mutagenesis Kit" (Stratagene). The
generation of a polypeptide functionally active variant or
derivative to a polypeptide by replacing an amino acid that does
not influence the function of a polypeptide can be accomplished by
one skilled in the art.
[0196] A variant or derivative can have, for example, one or more
conservative substitutions while still retaining at least one
biological activity. A conservative substitution is one in which an
amino acid is substituted for another amino acid that has similar
properties, such that one skilled in the art of peptide chemistry
would expect the secondary structure and hydropathic nature of the
polypeptide to be substantially unchanged. In general, the
following groups of amino acids represent conservative changes: (1)
ala, pro, gly, glu, asp, gln, asn, ser, thr; (2) cys, ser, tyr,
thr; (3) val, ile, leu, met, ala, phe; (4) lys, arg, his; and (5)
phe, tyr, trp, his.
[0197] Particular example variants and derivatives include, without
limitation, functionally active variants and derivatives of a
polypeptide comprising at least one extracellular subdomain of a
Klotho protein, e.g., a polypeptide comprising at least about 100,
150, 200, 250, 300, 350, 375, 400, or 425 contiguous amino acids of
an extracellular domain of Klotho (e.g., SEQ ID NO: 5 or 6), with
no more than about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,
15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60 or more
amino acid residue differences from the wild-type sequence (as
disclosed in SEQ ID NO: 5 or 6), while retaining at least one
biological activity of the wild-type polypeptide. For example, a
functionally active variant or derivative of a polypeptide
comprising at least one extracellular subdomain of a Klotho protein
comprises a polypeptide comprising at least about 400 contiguous
amino acids of SEQ ID NO: 5 or 6, with no more than about 100 amino
acid residue differences. For example, a functionally active
variant or derivative of a polypeptide comprising at least one
extracellular subdomain of a Klotho protein comprises a polypeptide
comprising at least about 400 contiguous amino acids of SEQ ID NO:
5 or 6, with no more than about 50 amino acid residue differences.
For example, a functionally active variant or derivative of a
polypeptide comprising at least one extracellular subdomain of a
Klotho protein comprises a polypeptide comprising at least about
425 contiguous amino acids of SEQ ID NO: 5 or 6, with no more than
about 25 amino acid residue differences. For example, a
functionally active variant or derivative of a polypeptide
comprising at least one extracellular subdomain of a Klotho protein
comprises a polypeptide comprising at least about 425 contiguous
amino acids of SEQ ID NO: 5 or 6, with no more than about 10 amino
acid residue differences. In another example, a functionally active
variant or derivative of a polypeptide comprising at least one
extracellular subdomain of a Klotho protein comprises a polypeptide
comprising at least about 100, 150, 200, 250, 300, 350, 400, 450,
500, 550, 600, 650, 700, 750, 800, 850, 900, 925, 950 or 982
contiguous amino acids of SEQ ID NO: 7, with no more than about 1,
2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20,
25, 30, 35, 40, 45, 50, 55, 60, 70, 75, 80, 85, 90, 95, 100, 110,
120, 140, 150, 160, 170, 180, 190, or 200 amino acid residue
differences from the wild-type sequence. For example, a
functionally active variant or derivative of a polypeptide
comprising at least one extracellular subdomain of a Klotho protein
comprises a polypeptide comprising at least about 500 contiguous
amino acids of SEQ ID NO: 7, with no more than about 100 amino acid
residue differences. For example, a functionally active variant or
derivative of a polypeptide comprising at least one extracellular
subdomain of a Klotho protein comprises a polypeptide comprising at
least about 600 contiguous amino acids of SEQ ID NO: 7, with no
more than about 100 amino acid residue differences. For example, a
functionally active variant or derivative of a polypeptide
comprising at least one extracellular subdomain of a Klotho protein
comprises a polypeptide comprising at least about 700 contiguous
amino acids of SEQ ID NO: 7, with no more than about 100 amino acid
residue differences. For example, a functionally active variant or
derivative of a polypeptide comprising at least one extracellular
subdomain of a Klotho protein comprises a polypeptide comprising at
least about 800 contiguous amino acids of SEQ ID NO: 7, with no
more than about 100 amino acid residue differences. For example, a
functionally active variant or derivative of a polypeptide
comprising at least one extracellular subdomain of a Klotho protein
comprises a polypeptide comprising at least about 900 contiguous
amino acids of SEQ ID NO: 7, with no more than about 100 amino acid
residue differences. For example, a functionally active variant or
derivative of a polypeptide comprising at least one extracellular
subdomain of a Klotho protein comprises a polypeptide comprising at
least about 900 contiguous amino acids of SEQ ID NO: 7, with no
more than about 50 amino acid residue differences.
[0198] Particular example variants and derivatives include, without
limitation, functionally active variants and derivatives of a
polypeptide comprising a fibroblast growth factor, e.g., a
polypeptide comprising at least about 100, 125, 150, 150, 175, 200,
225, or 250 contiguous amino acids of a fibroblast growth factor,
e.g., FGF19 (SEQ ID NO: 31), FGF21 (SEQ ID NO: 33), or FGF23 (SEQ
ID NO: 35), with no more than about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55,
60 or more amino acid residue differences from the wild-type
sequence (as disclosed in SEQ ID NOs: 31, 33 or 35), while
retaining at least one biological activity of the wild-type
polypeptide. In various embodiments, the variant or derivative can
comprise the R179Q variation or not. For example, a functionally
active variant or derivative of a polypeptide comprising a
fibroblast growth factor comprises a polypeptide comprising at
least about 150 contiguous amino acids of SEQ ID NOs: 31, 33 or 35,
with no more than about 25 amino acid residue differences. For
example, a functionally active variant or derivative of a
polypeptide comprising a fibroblast growth factor comprises a
polypeptide comprising at least about 175 contiguous amino acids of
SEQ ID NOs: 31, 33 or 35, with no more than about 25 amino acid
residue differences. For example, a functionally active variant or
derivative of a polypeptide comprising a fibroblast growth factor
comprises a polypeptide comprising at least about 200 contiguous
amino acids of SEQ ID NOs: 31, 33 or 35, with no more than about 25
amino acid residue differences. For example, a functionally active
variant or derivative of a polypeptide comprising a fibroblast
growth factor comprises a polypeptide comprising at least about 225
contiguous amino acids of SEQ ID NO: 35, with no more than about 50
amino acid residue differences. For example, a functionally active
variant or derivative of a polypeptide comprising a fibroblast
growth factor comprises a polypeptide comprising at least about 225
contiguous amino acids of SEQ ID NO: 35, with no more than about 25
amino acid residue differences.
4.3. Expression of Fusion Polypeptides of the Disclosure
[0199] In order to express the fusion protein of the disclosure,
DNA molecules obtained by any of the methods described herein or
those that are known in the art, can be inserted into appropriate
expression vectors by techniques well known in the art. For
example, a double stranded cDNA can be cloned into a suitable
vector by homopolymeric tailing or by restriction enzyme linking
involving the use of synthetic DNA linkers or by blunt-ended
ligation. DNA ligases are usually used to ligate the DNA molecules
and undesirable joining can be avoided by treatment with alkaline
phosphatase.
[0200] Therefore, the disclosure includes vectors (e.g.,
recombinant plasmids and bacteriophages) that include nucleic acid
molecules (e.g., genes or recombinant nucleic acid molecules
encoding genes) as described herein. The term "recombinant vector"
includes a vector (e.g., plasmid, phage, phasmid, virus, cosmid,
fosmid, or other purified nucleic acid vector) that has been
altered, modified or engineered such that it contains greater,
fewer or different nucleic acid sequences than those included in
the native or natural nucleic acid molecule from which the
recombinant vector was derived. For example, a recombinant vector
may include a nucleotide sequence encoding a Klotho-FGF23 fusion
operatively linked to regulatory sequences, e.g., promoter
sequences, terminator sequences and/or artificial ribosome binding
sites (RBSs), as defined herein. Recombinant vectors which allow
for expression of the genes or nucleic acids included in them are
referred to as "expression vectors."
[0201] For eukaryotic hosts, different transcriptional and
translational regulatory sequences may be employed, depending on
the nature of the host. They may be derived from viral sources,
such as adenovirus, bovine papilloma virus, Simian virus or the
like, where the regulatory signals are associated with a particular
gene which has a high level of expression. Examples include, but
are not limited to, the TK promoter of the Herpes virus, the SV40
early promoter, the yeast ga14 gene promoter, etc. Transcriptional
initiation regulatory signals may be selected which allow for
repression or activation, so that expression of the genes can be
modulated.
[0202] In some of the molecules of the disclosure described herein,
one or more DNA molecules having a nucleotide sequence encoding one
or more polypeptide chains of a fusion polypeptide are operatively
linked to one or more regulatory sequences, which are capable of
integrating the desired DNA molecule into a host cell. Cells which
have been stably transformed by the introduced DNA can be selected,
for example, by introducing one or more markers which allow for
selection of host cells which contain the expression vector. A
selectable marker gene can either be linked directly to a nucleic
acid sequence to be expressed, or be introduced into the same cell
by co-transfection. Additional elements may also be needed for
optimal synthesis of proteins described herein. It would be
apparent to one of ordinary skill in the art which additional
elements to use.
[0203] Factors of importance in selecting a particular plasmid or
viral vector include, but are not limited to, the ease with which
recipient cells that contain the vector are recognized and selected
from those recipient cells which do not contain the vector; the
number of copies of the vector which are desired in a particular
host; and whether it is desirable to be able to "shuttle" the
vector between host cells of different species.
[0204] Once the vector(s) is constructed to include a DNA sequence
for expression, it may be introduced into an appropriate host cell
by one or more of a variety of suitable methods that are known in
the art, including but not limited to, for example, transformation,
transfection, conjugation, protoplast fusion, electroporation,
calcium phosphate-precipitation, direct microinjection, etc.
[0205] Host cells may either be prokaryotic or eukaryotic. Examples
of eukaryotic host cells include, for example, mammalian cells,
such as human, monkey, mouse, and Chinese hamster ovary (CHO)
cells. Such cells facilitate post-translational modifications of
proteins, including, for example, correct folding or glycosylation.
Additionally, yeast cells can also be used to express fusion
polypeptides of the disclosure. Like most mammalian cells, yeast
cells also enable post-translational modifications of proteins,
including, for example, glycosylation. A number of recombinant DNA
strategies exist which utilize strong promoter sequences and high
copy number plasmids that can be utilized for production of
proteins in yeast. Yeast transcription and translation machinery
can recognize leader sequences on cloned mammalian gene products,
thereby enabling the secretion of peptides bearing leader sequences
(i.e., pre-peptides). A particularmethod of high-yield production
of the fusion polypeptides of the disclosure is through the use of
dihydrofolate reductase (DHFR) amplification in DHFR-deficient CHO
cells, by the use of successively increasing levels of methotrexate
as described in U.S. Pat. No. 4,889,803. The polypeptide obtained
may be in a glycosylated form.
[0206] After the introduction of one or more vector(s), host cells
are usually grown in a selective medium, which selects for the
growth of vector-containing cells. Purification of the recombinant
proteins can be carried out by any of the methods known in the art
or described herein, for example, any conventional procedures
involving extraction, precipitation, chromatography and
electrophoresis. A further purification procedure that may be used
for purifying proteins is affinity chromatography using monoclonal
antibodies which bind a target protein. Generally, crude
preparations containing a recombinant protein are passed through a
column on which a suitable monoclonal antibody is immobilized. The
protein usually binds to the column via the specific antibody while
the impurities pass through. After washing the column, the protein
is eluted from the gel by changing pH or ionic strength, for
example.
4.4. Assays for Assessing Fusion Polypeptide Activity
[0207] Assays described herein (See Example 2) and those known in
the art can be used for detecting Klotho or FGF activity of the
fusion polypeptides of the disclosure. Suitable activity assays
include receptor binding assays, cellular proliferation assays and
cell signaling assays. For example, a binding assay which may be
used for determining whether a fusion polypeptide has Klotho or FGF
activity includes, assaying the binding of a fusion polypeptide to
an FGF receptor. FGF receptor binding assays include, but are not
limited to, both competitive and non-competitive assay. For
example, FGF receptor binding can be detected by contacting cells
expressing an FGF receptor with a labeled FGF (for example,
radio-active label) and increasing concentrations of an unlabeled
Klotho-FGF fusion polypeptide. The two ligands that compete for
binding to the same receptor are added to a reaction mixture
containing the cell. The cells are subsequently washed and labeled
FGF is measured. A decrease in the amount of the labeled FGF to its
receptor in the presence of the unlabeled fusion polypeptide is
indicative of binding of the Klotho-FGF fusion polypeptide to the
receptor. Alternatively, the Klotho-FGF fusion polypeptide may be
labeled and direct binding of the fusion polypeptide to the cell is
detected.
[0208] Klotho or FGF activity can also be measured by determining
whether the fusion polypeptide induces a cellular response. For
example, in some embodiments, an assay for detecting the biological
activity of a Klotho-FGF fusion polypeptide involves contacting
cells which express an FGF receptor with a fusion polypeptide,
assaying a cellular response such as, for example, cell
proliferation or Egr-1 activation, myotube diameter in C2C12 cells,
and comparing the cellular response in the presence and absence of
the fusion polypeptide. An increase in the cellular response in the
presence of the fusion polypeptide complex relative to the absence
indicates that the fusion polypeptide has biological activity.
Also, an increase in a downstream signaling event from the receptor
can also be measured as indicia of biological activity (e.g.,
phosphorylation of FGFR, FRS2, ERK1/2, p70S6K etc.).
4.5 Pharmaceutical Compositions and Methods of Treatment
[0209] The disclosure also pertains to pharmaceutical compositions
containing one or more fusion polypeptides of the disclosure and a
pharmaceutically acceptable diluent or carrier. The pharmaceutical
compositions can further include a pharmaceutically effective dose
of heparin. Such pharmaceutical compositions may be included in a
kit or container. Such kit or container may be packaged with
instructions pertaining to the extended in vivo half-life or the in
vitro shelf life of the fusion polypeptides. Optionally associated
with such kit or container(s) can be a notice in the form
prescribed by a governmental agency regulating the manufacture, use
or sale of pharmaceuticals or biological products, which notice
reflects approval by the agency of manufacture, use or sale for
human administration. Such compositions may be used in methods of
treating, preventing, or ameliorating a disease or a disease
symptom (e.g., age-related condition or metabolic disorder) in a
patient, preferably a mammal and most preferably a human, by
administering the pharmaceutical composition to the patient.
[0210] In general, a therapeutically effective amount of a
pharmaceutical composition of the disclosure is from about 0.0001
mg/kg to 0.001 mg/kg; 0.001 mg/kg to about 10 mg/kg body weight or
from about 0.02 mg/kg to about 5 mg/kg body weight. Commonly, a
therapeutically effective amount of a fusion polypeptide is from
about 0.001 mg to about 0.01 mg, about 0.01 mg to about 100 mg, or
from about 100 mg to about 1000 mg, for example. Preferably, a
therapeutically effective amount of a fusion polypeptide is from
about 0.001 mg/kg to 2mg/kg.
[0211] The optimal pharmaceutical formulations for a fusion
polypeptide can be determined by one or ordinary skilled in the art
depending upon the route of administration and desired dosage.
(See, for example, Remington's Pharmaceutical Sciences, 18th Ed.
(1990), Mack Publishing Co., Easton, Pa., the entire disclosure of
which is hereby incorporated by reference).
[0212] The fusion polypeptides of the disclosure may be
administered as a pharmaceutical composition that may be in the
form of a solid, liquid or gas (aerosol). Typical routes of
administration may include, without limitation, oral, topical,
parenteral, sublingual, rectal, vaginal, intradermal and
intranasal. Parenteral administration includes subcutaneous
injections, intravenous, intramuscular, intraperitoneal,
intrapleural, intrasternal injection or infusion techniques.
Preferably, the compositions are administered parenterally. More
preferably, the compositions are administered intravenously.
Pharmaceutical compositions of the disclosure can be formulated so
as to allow a polypeptide of the disclosure to be bioavailable upon
administration of the composition to a subject. Compositions can
take the form of one or more dosage units, where, for example, a
tablet can be a single dosage unit, and a container of a
polypeptide of the disclosure in aerosol form can hold a plurality
of dosage units.
[0213] Materials used in preparing the pharmaceutical compositions
can be non-toxic in the amounts used. It will be evident to those
of ordinary skill in the art that the optimal dosage of the active
ingredient(s) in the pharmaceutical composition will depend on a
variety of factors. Relevant factors include, without limitation,
the type of subject (e.g., human), the overall health of the
subject, the type of age-related condition or metabolic disorder
the subject in need of treatment of, the use of the composition as
part of a multi-drug regimen, the particular form of the
polypeptide of the disclosure, the manner of administration, and
the composition employed.
[0214] The pharmaceutically acceptable carrier or vehicle may be
particulate, so that the compositions are, for example, in tablet
or powder form. The carrier(s) can be liquid, with the compositions
being, for example, an oral syrup or injectable liquid. In
addition, the carrier(s) can be gaseous, so as to provide an
aerosol composition useful in, e.g., inhalatory administration.
[0215] The term "carrier" refers to a diluent, adjuvant or
excipient, with which a polypeptide of the disclosure is
administered. Such pharmaceutical carriers can be liquids, such as
water and oils, including those of petroleum, animal, vegetable or
synthetic origin, such as peanut oil, soybean oil, mineral oil,
sesame oil and the like. The carriers can be saline, gum acacia,
gelatin, starch paste, talc, keratin, colloidal silica, urea, and
the like. In addition, auxiliary, stabilizing, thickening,
lubricating and coloring agents can be used. In one embodiment,
when administered to a subject, the polypeptides of the disclosure
and pharmaceutically acceptable carriers are sterile. Water is a
particular carrier when the polypeptide of the disclosure is
administered intravenously. Saline solutions and aqueous dextrose
and glycerol solutions can also be employed as liquid carriers,
particularly for injectable solutions. Suitable pharmaceutical
carriers also include excipients such as starch, glucose, lactose,
sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium
stearate, glycerol monostearate, talc, sodium chloride, dried skim
milk, glycerol, propylene, glycol, water, ethanol and the like. The
present compositions, if desired, can also contain minor amounts of
wetting or emulsifying agents, or pH buffering agents.
[0216] The composition may be intended for oral administration, and
if so, the composition is preferably in solid or liquid form, where
semi-solid, semi-liquid, suspension and gel forms are included
within the forms considered herein as either solid or liquid.
[0217] As a solid composition for oral administration, the
composition can be formulated into a powder, granule, compressed
tablet, pill, capsule, chewing gum, wafer or the like form. Such a
solid composition typically contains one or more inert diluents. In
addition, one or more of the following can be present: binders such
as ethyl cellulose, carboxymethylcellulose, microcrystalline
cellulose, or gelatin; excipients such as starch, lactose or
dextrins, disintegrating agents such as alginic acid, sodium
alginate, Primogel, corn starch and the like; lubricants such as
magnesium stearate or Sterotex; glidants such as colloidal silicon
dioxide; sweetening agents such as sucrose or saccharin, a
flavoring agent such as peppermint, methyl salicylate or orange
flavoring, and a coloring agent.
[0218] When the pharmaceutical composition is in the form of a
capsule, e.g., a gelatin capsule, it can contain, in addition to
materials of the above type, a liquid carrier such as polyethylene
glycol, cyclodextrin or a fatty oil.
[0219] The pharmaceutical composition can be in the form of a
liquid, e.g., an elixir, syrup, solution, emulsion or suspension.
The liquid can be useful for oral administration or for delivery by
injection. When intended for oral administration, a composition can
contain one or more of a sweetening agent, preservatives,
dye/colorant and flavour enhancer. In a composition for
administration by injection, one or more of a surfactant,
preservative, wetting agent, dispersing agent, suspending agent,
buffer, stabilizer and isotonic agent can also be included.
[0220] The liquid compositions of the disclosure, whether they are
solutions, suspensions or other like form, can also include one or
more of the following: sterile diluents such as water for
injection, saline solution, preferably physiological saline,
Ringer's solution, isotonic sodium chloride, fixed oils such as
synthetic mono or digylcerides which can serve as the solvent or
suspending medium, polyethylene glycols, glycerin, cyclodextrin,
propylene glycol or other solvents; antibacterial agents such as
benzyl alcohol or methyl paraben; antioxidants such as ascorbic
acid or sodium bisulfate; chelating agents such as
ethylenediaminetetraacetic acid; buffers such as acetates, citrates
or phosphates and agents for the adjustment of tonicity such as
sodium chloride or dextrose. A parenteral composition can be
enclosed in an ampoule, a disposable syringe or a multiple-dose
vial made of glass, plastic or other material. Physiological saline
is a particular specific adjuvant. An injectable composition is
preferably sterile.
[0221] The pharmaceutical compositions contain an effective amount
of a compound of the disclosure (e.g., fusion polypeptide) such
that a suitable dosage will be obtained. The pharmaceutical
compositions may contain the known effective amount of the
compounds as currently prescribed for their respective
disorders.
[0222] The route of administration of the polypeptide of the
disclosure used in the prophylactic and/or therapeutic regimens
which will be effective in the prevention, treatment, and/or
management of a age-related condition or metabolic disorder can be
based on the currently prescribed routes of administration for
other therapeutics known in the art. The polypeptides of the
disclosure can be administered by any convenient route, for
example, by infusion or bolus injection, by absorption through
epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and
intestinal mucosa, etc.). Administration can be systemic or local.
Various delivery systems are known, e.g., microparticles,
microcapsules, capsules, etc., and may be useful for administering
a polypeptide of the disclosure. More than one polypeptides of the
disclosure may be administered to a subject. Methods of
administration may include, but are not limited to, oral
administration and parenteral administration; parenteral
administration including, but not limited to, intradermal,
intramuscular, intraperitoneal, intravenous, subcutaneous,
intranasal, epidural, sublingual, intranasal, intracerebral,
intraventricular, intrathecal, intravaginal, transdermal, rectally,
by inhalation, or topically to the ears, nose, eyes, or skin.
[0223] The polypeptides of the disclosure may be administered
parenterally. Specifically, the polypeptides of the disclosure may
be administered intravenously.
[0224] Pulmonary administration can also be employed, e.g., by use
of an inhaler or nebulizer, and formulation with an aerosolizing
agent, or via perfusion in a fluorocarbon or synthetic pulmonary
surfactant. The polypeptides of the disclosure can also be
formulated as a suppository, with traditional binders and carriers
such as triglycerides.
[0225] The polypeptides of the disclosure can be delivered in a
controlled release system. For example, a pump can be used (see
Sefton, CRC Crit. Ref. Biomed. Eng. 1987, 14, 201; Buchwald et al.,
Surgery 1980, 88: 507; Saudek et al., N. Engl. J. Med. 1989, 321:
574). Polymeric materials can also be used for controlled release
of the polypeptides of the disclosure (see Medical Applications of
Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton,
Fla., 1974; Controlled Drug Bioavailability, Drug Product Design
and Performance, Smolen and Ball (eds.), Wiley, New York, 1984;
Ranger and Peppas, J. Macromol. Sci. Rev. Macromol. Chem. 1983, 23,
61; see also Levy et al., Science 1985, 228, 190; During et al.,
Ann. Neurol., 1989, 25, 351; Howard et al., J. Neurosurg., 1989,
71, 105). Specifically, a controlled-release system can be placed
in proximity of the target of the polypeptides of the disclosure,
e.g., the brain, thus requiring only a fraction of the systemic
dose (see, e.g., Goodson, in Medical Applications of Controlled
Release, supra, vol. 2, 1984, pp. 115-138). Other
controlled-release systems discussed in the review by Langer
(Science 1990, 249, 1527-1533) can be used.
[0226] Polymeric materials used to achieve controlled or sustained
release of the polypeptides of the disclosure are disclosed, e.g.,
in U.S. Pat. No. 5,679,377; U.S. Pat. No. 5,916,597; U.S. Pat. No.
5,912,015; U.S. Pat. No. 5,989,463; U.S. Pat. No. 5,128,326; PCT
Publication No. WO 99/15154; and PCT Publication No. WO 99/20253.
Examples of polymers used in sustained release formulations
include, but are not limited to, poly(2-hydroxy ethyl
methacrylate), poly(methyl methacrylate), poly(acrylic acid),
poly(ethylene-co-vinyl acetate), poly(methacrylic acid),
polyglycolides (PLG), polyanhydrides, poly(N-vinyl pyrrolidone),
poly(vinyl alcohol), polyacrylamide, poly(ethylene glycol),
polylactides (PLA), poly(lactide-co-glycolides) (PLGA), and
polyorthoesters. Preferably, the polymer used in a sustained
release formulation is inert, free of leachable impurities, stable
on storage, sterile, and biodegradable.
[0227] In general, a therapeutically effective amount of a
pharmaceutical composition of the disclosure is from about 0.0001
mg/kg to 0.001 mg/kg; 0.001 mg/kg to about 10 mg/kg body weight or
from about 0.02 mg/kg to about 5 mg/kg body weight.
[0228] In other embodiments, the prophylactic and/or therapeutic
regimen involves administering to a patient one or more doses of an
effective amount of a polypeptide of the disclosure, wherein the
dose of an effective amount achieves a plasma level of at least
0.01 .mu.g/mL to at least 400 .mu.g/mL of the polypeptide of the
disclosure.
[0229] A prophylactic and/or therapeutic regimen may involve
administering to a patient a plurality of doses of an effective
amount of a polypeptide of the disclosure, wherein the plurality of
doses maintains a plasma level of at least 0.01 .mu.g/mL, to 400
.mu.g/mL of the polypeptide of the disclosure. The prophylactic
and/or therapeutic regimen may be administered for at least 1 day,
1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7
months, 8 months or 9 months.
[0230] The prophylactic and/or therapeutic regimen may involve
administration of a polypeptide of the disclosure in combination
with one or more additional therapeutics. The recommended dosages
of the one or more therapeutics currently used for the prevention,
treatment, and/or management of an age-related condition or
metabolic disorder can be obtained from any reference in the art
including, but not limited to, Hardman et al., eds., Goodman &
Gilman's The Pharmacological Basis Of Basis Of Therapeutics, 10th
ed., McGraw-Hill, New York, 2001; Physician's Desk Reference
(60.sup.th ed., 2006), which is incorporated herein by reference in
its entirety.
[0231] The disclosure includes methods of treating disorders
wherein agonistic activity of Klotho protein and FGF are desirable.
The disclosure furthermore includes the use of the disclosed
proteins, fusion proteins, nucleic acid molecules or pharmaceutical
composition in therapy or as medicament for the treatment of a
pathological disorder wherein agonistic activity of Klotho protein
and FGF are desirable. Examples of such methods or uses of the
disclosure include, but are not limited to age-related condition or
metabolic disorders.
[0232] The disclosure includes methods for treating or preventing
an age-related condition in an individual; and the use of the
disclosed proteins, fusion proteins, nucleic acid molecules or
pharmaceutical composition in therapy or as medicament for treating
or preventing an age-related condition in an individual. An
individual in need of treatment is administered a pharmacologically
effective dose of a pharmaceutical composition containing a Klotho
fusion polypeptide, having at least one extracellular subdomain of
a Klotho protein and a fibroblast growth factor and an (optional)
modified Fc fragment, so as to treat or prevent the age-related
condition. In some embodiments, the Klotho fusion polypeptide is
co-administered with a pharmacologically effective dose of heparin.
Age-related conditions include sarcopenia, skin atrophy, muscle
wasting, brain atrophy, atherosclerosis, arteriosclerosis,
pulmonary emphysema, osteoporosis, osteoarthritis, immunologic
incompetence, high blood pressure, dementia, Huntington's disease,
Alzheimer's disease, cataracts, age-related macular degeneration,
prostate cancer, stroke, diminished life expectancy, memory loss,
wrinkles, impaired kidney function, and age-related hearing loss.
In some embodiments, the Klotho fusion polypeptide contains at
least one extracellular domain of an alpha Klotho protein. In a
particular embodiment, a Klotho fusion protein containing at least
one extracellular domain of alpha Klotho protein and fibroblast
growth factor 23 is administered to an individual in need of
treatment for muscle wasting.
[0233] The disclosure is also directed to a method for treating or
preventing a metabolic disorder in an individual; and to the use of
the disclosed proteins, fusion proteins, nucleic acid molecules or
pharmaceutical composition in therapy or as medicament for treating
or preventing metabolic disorder in an individual. An individual in
need of treatment is administered a pharmacologically effective
dose of a pharmaceutical composition containing a Klotho fusion
polypeptide, having at least one extracellular subdomain of a
Klotho protein and a fibroblast growth factor so as to treat the
metabolic disorder, and an (optional) modified Fc fragment having
decreased binding to FcRn and/or increased serum half-life and/or
stability. In some embodiments, the Klotho fusion polypeptide is
co-administered with a pharmacologically effective dose of heparin.
The method may be used in the treatment or prevention of Type II
Diabetes, Metabolic Syndrome, hyperglycemia, and obesity. In a
particular embodiment, a Klotho fusion protein containing at least
one extracellular domain of a beta-Klotho protein and fibroblast
growth factor 21 is administered to an individual in need of
treatment for a metabolic disorder.
[0234] The disclosure also provides methods for treating or
preventing hyperphosphatemia or calcinosis in an individual; and
the use of the disclosed proteins, fusion proteins, nucleic acid
molecules or pharmaceutical composition in therapy or as medicament
for treating or preventing hyperphosphatemia or calcinosis in an
individual. An individual in need of treatment is administered a
pharmacologically effective dose of a pharmaceutical composition
containing a Klotho fusion polypeptide, having at least one
extracellular subdomain of a Klotho protein, a fibroblast growth
factor and an (optional) modified Fc fragment so as to treat
hyperphosphatemia or calcinosis. In some embodiments, the Klotho
fusion polypeptide is co-administered with a pharmacologically
effective dose of heparin. In a particular embodiment, a Klotho
fusion protein containing at least one extracellular domain of an
alpha Klotho protein and fibroblast growth factor 23 and an
(optional) modified Fc fragment is administered to an individual in
need of treatment for a hyperphosphatemia or calcinosis.
[0235] The disclosure is also directed to a method for treating or
preventing chronic renal disease or chronic renal failure in an
individual; and to the use of the disclosed proteins, fusion
proteins, nucleic acid molecules or pharmaceutical composition in
therapy or as medicament for treating or preventing chronic renal
disease or chronic renal failure in an individual. An individual in
need of treatment is administered a pharmacologically effective
dose of a pharmaceutical composition containing a Klotho fusion
polypeptide, having at least one extracellular subdomain of a
Klotho protein, a fibroblast growth factor and an (optional)
modified Fc fragment so as to treat chronic renal disease or
chronic renal failure. In some embodiments, the Klotho fusion
polypeptide is co-administered with a pharmacologically effective
dose of heparin. In some embodiments, a Klotho fusion protein
containing at least one extracellular domain of an alpha Klotho
protein is administered to an individual in need of treatment for
chronic renal disease or chronic renal failure.
[0236] The disclosure also includes methods for treating or
preventing cancer in an individual; and the use of the disclosed
proteins, fusion proteins, nucleic acid molecules or pharmaceutical
composition in therapy or as medicament for treating or preventing
cancer in an individual. An individual in need of treatment is
administered a pharmacologically effective dose of a pharmaceutical
composition containing a Klotho fusion polypeptide, having at least
one extracellular subdomain of a Klotho protein, a fibroblast
growth factor and an (optional) modified Fc fragment so as to treat
cancer. The method may be used in the treatment or prevention of
breast cancer. In some embodiments, the Klotho fusion polypeptide
is co-administered with a pharmacologically effective dose of
heparin. In some embodiments, a Klotho fusion protein containing at
least one extracellular domain of an alpha Klotho protein is
administered to an individual in need of treatment for cancer.
[0237] In methods of treating disorders by administering a
pharmaceutical composition containing a Klotho fusion polypeptide;
or when using pharmaceutical composition containing a Klotho fusion
polypeptide in therapy, the Klotho fusion polypeptide and an
(optional) modified Fc fragment has at least one extracellular
subdomain of a Klotho protein and a fibroblast growth factor. In a
particular embodiment, the Klotho fusion protein contains at least
one extracellular domain of a beta Klotho protein and fibroblast
growth factor 21.
[0238] In another embodiment, the fusion comprises a FGF (e.g.,
FGF19, FGF21, FGF23 or FGF23 variant) and a modified Fc fragment
with decreased binding to FcRn and/or increased serum stability.
This type of fusion can be used in various diseases, as described
above, or used to treat or prevent any FGF-related disease known in
the art. The fusion can be administered to an individual in need
thereof
[0239] The fusion polypeptide compositions can be administered
according to any method of administration known to those of skill
in the art and described herein. Particular specific methods of
administration include subcutaneous or intravenous. Other effective
modes of administration are described herein.
4.6. Methods of Treatment and Assays for Assessing Efficacy
[0240] Methods or uses of the disclosure which provide
administering the fusion polypeptides described herein to an
individual can be used to treat a variety of disorders including an
age-related disorder or a metabolic disorder. Without being limited
by any particular theory, fusion polypeptides may be used to treat
disorders in which there is dysregulation of Klotho or FGF. Example
disorders include metabolic disorders and age-related disorders.
For example, both FGF23 or Klotho knock-out mice display a variety
of similar phenotypes including, low physical activity, growth
retardation, muscle wasting, skin atrophy, atherosclerosis, short
life spans, etc. (See Razzaque and Lanske, J. of Endrocrinology,
194:1-10 (2007), which is herein incorporated by reference).
[0241] In particular, fusion polypeptides of the disclosure are
particularly useful in the treatment of aging-related disorders,
including muscle wasting. Without being bound to theory, the
ability of Klotho and FGF23 to control mineral (e.g., phosphate and
calcium) and vitamin D homeostasis may be the means by which these
proteins modulate aging and muscle atrophy.
[0242] On the other hand, fusion polypeptides of the disclosure may
be used for treating a metabolic disorder. For example, beta-Klotho
and FGF19 have been shown to control bile acid homeostasis by
regulating cholesterol 7-.alpha.-hydroxylase (CYP7A1). A
non-limiting example of bile homeostasis disorder is cholestasis.
The beta-Klotho and FGF21 have been shown to induce lipolysis in
adipocytes and, therefore, reduced fat storage and increased
glucose uptake. Non-limiting examples of lipolysis/fat storage
disorders are obesity and associated metabolic and cardiovascular
diseases.
[0243] Based at least in part on the finding that FGF23 is able to
stimulate excretion of phosphate in the urine and thereby reduce
phosphate levels in the serum, Klotho-FGF23 fusion polypeptides of
the disclosure can be used for treating or preventing
hyperphosphatemia or calcinosis in an individual. For example, it
has been shown that a homozygous missense mutation in Klotho
resulting in a deficiency in Klotho in a patient can cause severe
tumoral calcinosis and artery calcification (Ichikawa et al., J.
Clin. Invest. 117:2684-2691 (2007), which is herein incorporated by
reference). An individual is administered a pharmacologically
effective dose of a pharmaceutical composition containing the
Klotho fusion polypeptide, having at least one extracellular
subdomain of a Klotho protein, a fibroblast growth factor and an
(optional) modified Fc fragment so as to treat or prevent
hyperphosphatemia or calcinosis. In particular, a Klotho fusion
polypeptide containing at least one extracellular domain of an
alpha Klotho protein, a fibroblast growth factor and an (optional)
modified Fc fragment is useful for treating hyperphosphatemia or
calcinosis.
[0244] Klotho fusion polypeptides of the disclosure can also be
used for treating or preventing chronic renal disease or chronic
renal failure in an individual. For example, it has been shown that
Klotho expression is reduced in kidney of patients with chronic
renal failure, compared to that in unaffected kidneys (Koh et al.,
Biochem. Biophys. Res. Comm. 280:1015-1020 (2001), which is herein
incorporated by reference). An individual is administered a
pharmacologically effective dose of a pharmaceutical composition
containing the Klotho fusion polypeptide, having at least one
extracellular subdomain of a Klotho protein, a fibroblast growth
factor and an (optional) modified Fc fragment so as to treat or
prevent chronic renal disease or chronic renal failure. In
particular, a Klotho fusion polypeptide containing at least one
extracellular domain of an alpha Klotho protein is useful for
treating chronic renal disease or chronic renal failure.
[0245] Klotho fusion polypeptides of the disclosure can also be
used for treating or preventing cancer in an individual. For
example, it has been shown that Klotho expression is reduced in
breast cancer tissue, compared to normal breast cancer tissue (Wolf
et al., Oncogene (2008) advance online publication, which is herein
incorporated by reference). An individual is administered a
pharmacologically effective dose of a pharmaceutical composition
containing the Klotho fusion polypeptide, having at least one
extracellular subdomain of a Klotho protein, a fibroblast growth
factor and an (optional) modified Fc fragment so as to treat or
prevent cancer or breast cancer. In particular, a Klotho fusion
protein containing at least one extracellular domain of an alpha
Klotho protein is useful for treating cancer or breast cancer.
[0246] Methods for evaluating the efficacy and/or determining the
effective dose of a Klotho fusion polypeptide of the disclosure on
an age-related disorder or metabolic disorder include organismal
based assays, e.g., using a mammal (e.g., a mouse, rat, primate, or
some other non-human), or other animal (e.g., Xenopus, zebrafish,
or an invertebrate such as a fly or nematode). The Klotho fusion
polypeptide can be administered to the organism once or as a
regimen (regular or irregular). A parameter of the organism is then
evaluated, e.g., an age-associated parameter. Klotho fusion
polypeptides that are of interest result in a change in the
parameter relative to a reference, e.g., a parameter of a control
organism. Other parameters (e.g., related to toxicity, clearance,
and pharmacokinetics) can also be evaluated.
[0247] The Klotho fusion polypeptide of the disclosure may be
evaluated using an animal that has a particular disorder, e.g., a
disorder described herein, e.g., an age-related disorder, a
metabolic disorder. These disorders can also provide a sensitized
system in which the test polypeptide's effects on physiology can be
observed. Example disorders include: denervation, disuse atrophy;
metabolic disorders (e.g., disorder of obese and/or diabetic
animals such as db/db mouse and ob/ob mouse); cerebral, liver
ischemia; cisplatin/taxol/vincristine models; various tissue
(xenograph) transplants; transgenic bone models; pain syndromes
(include inflammatory and neuropathic disorders); Paraquat,
genotoxic, and oxidative stress models; and tumor I models.
[0248] For measuring an age-related disorder, the animal model can
be an animal that has an altered phenotype when calorically
restricted. For example, F344 rats provide a useful assay system
for evaluating a Klotho fusion polypeptide. When calorically
restricted, F344 rats have a 0 to 10% incidence of nephropathy.
However, when fed ad libitum, they have a 60 to 100% incidence of
nephropathy.
[0249] To evaluate a Klotho fusion polypeptide of the disclosure,
it is administered to the animal (e.g., an F344 rat or other
suitable animal) and a parameter of the animal is evaluated, e.g.,
after a period of time. The animal can be fed ad libitum or
normally (e.g., not under caloric restriction, although some
parameters can be evaluated under such conditions). Typically, a
cohort of such animals is used for the assay. Generally, a test
polypeptide can be indicated as favorably altering lifespan
regulation in the animal if the test polypeptide affects the
parameter in the direction of the phenotype of a similar animal
subject to caloric restriction. Such test polypeptides may cause at
least some of the lifespan regulatory effects of caloric
restriction, e.g., a subset of such effects, without having to
deprive the organism of caloric intake.
[0250] The parameter to be tested may be an age-associated or
disease associated parameter, e.g., a symptom of the disorder
associated with the animal model. For example, the test polypeptide
can be administered to a SH Rat, and blood pressure is monitored. A
test polypeptide that is favorably indicated can cause an
amelioration of the symptom relative to a similar reference animal
not treated with the polypeptide. Other parameters relevant to a
disorder or to aging can include: antioxidant levels (e.g.,
antioxidant enzyme levels or activity), stress resistance (e.g.,
paraquat resistance), core body temperature, glucose levels,
insulin levels, thyroid-stimulating hormone levels, prolactin
levels, and leutinizing hormone levels.
[0251] To measure the effectiveness of the polypeptides of the
disclosure for treating an age-related disorder, an animal having
decreased Klotho expression may be used, e.g., mouse with a mutant
Klotho; See Kuroo, et al. Nature, 390; 45 (1997) and U.S. Pub. No.
2003/0119910, both of which are herein incorporated by reference in
their entirety. For example, the test polypeptide is administered
to the mutant mouse and age-related parameters are monitored. A
test polypeptide that is favorably indicated can cause an
amelioration of the symptom relative to a similar reference animal
not treated with the polypeptide. A parameter relevant to a
metabolic disorder or to aging can be assessed by measurement of
body weight, examination on the acquisition of reproductive
ability, measurement of blood sugar level, observation of life
span, observation of skin, observation of motor functions such as
walking, and the like. The assessment can also be made by
measurement of thymus weight, observation of the size of calcified
nodules formed on the inner surface of thoracic cavity, and the
like. Further, quantitative determination of mRNA for the Klotho
gene or Klotho protein is also useful for the assessment.
[0252] Still other in vivo models and organismal assays include
evaluating an animal for a metabolic parameter, e.g., a parameter
relevant to an insulin disorder, type II diabetes. Example
metabolic parameters include: glucose concentration, insulin
concentration, and insulin sensitivity.
[0253] Another example system features tumors, e.g., in an animal
model. The tumors can be spontaneous or induced. For example, the
tumors can be developed from cells that have a variety of genetic
constitutions, e.g., they can be p53+ or p53-. It is also possible
to use organisms that an autoimmune disorder, e.g., an NZB mouse,
which is predisposed to SLE. To evaluate features of bone disease,
it is possible, for example, to use an animal that has an
ovariectomy as a model,. e.g., for osteoporosis. Similarly, for
joint disease, the model can be based on adjuvant arthritis (e.g.,
mice can be immunized with cartilage proteoglycans, high mobility
group proteins, streptococcal cell wall material, or collagens);
for kidney disease, kd/kd mice can be used. Animal models of
cognition, particularly learning and memory are also available.
Animal models of diabetes and its complications are also available,
e.g., the streptozotocin model. Canine models can be used, for
example, for evaluating stroke and ischemia.
[0254] In assessing whether a test polypeptide is capable of
altering life span regulation, a number of age-associated
parameters or biomarkers can be monitored or evaluated. Example age
associated parameters include: (i) lifespan of the cell or the
organism; (ii) presence or abundance of a gene transcript or gene
product in the cell or organism that has a biological age dependent
expression pattern; (iii) resistance of the cell or organism to
stress; (iv) one so or more metabolic parameters of the cell or
organism (example parameters include circulating insulin levels,
blood glucose levels; fat content; core body temperature and so
forth); (v) proliferative capacity of the cell or a set of cells
present in the organism; and (vi) physical appearance or behavior
of the cell or organism.
[0255] The term "average lifespan" refers to the average of the age
of death of a cohort of organisms. In some cases, the "average
lifespan" is assessed using a cohort of genetically identical
organisms under controlled environmental conditions. Deaths due to
mishap are discarded. Where average lifespan cannot be determined
(e.g., for humans) under controlled environmental conditions,
reliable statistical information (e.g., from actuarial tables) for
a sufficiently large population can be used as the average
lifespan.
[0256] Characterization of molecular differences between two such
organisms, e.g., one reference organism and one organism treated
with a Klotho fusion polypeptide can reveal a difference in the
physiological state of the organisms. The reference organism and
the treated organism are typically the same chronological age. The
term "chronological age" as used herein refers to time elapsed
since a preselected event, such as conception, a defined
embryological or fetal stage, or, more preferably, birth. A variety
of criteria can be used to determine whether organisms are of the
"same" chronological age for the comparative analysis. Typically,
the degree of accuracy required is a function of the average
lifespan of a wildtype organism. For example, for the nematode C.
elegans, for which the laboratory wildtype strain N2 lives an to
average of about 16 days under some controlled conditions,
organisms of the same age may have lived for the same number of
days. For mice, organism of the same age may have lived for the
same number of weeks or months; for primates or humans, the same
number of years (or within 2, 3, or 5 years); and so forth.
Generally, organisms of the same chronological age may have lived
for an amount of time within 15, 10, 5, 3, 2 or 1% of the average
lifespan of a wildtype organism of that species. Preferably, the
organisms are adult organisms, e.g., the organisms have lived for
at least an amount of time in which the average wildtype organism
has matured to an age at which it is competent to reproduce.
[0257] The organismal screening assay can be performed before the
organisms exhibit overt physical features of aging. For example,
the organisms may be adults that have lived only 10, 30, 40, 50,
60, or 70% of the average lifespan of a wildtype organism of the
same species. Age-associated changes in metabolism, immune
competence, and chromosomal structure have been reported. Any of
these changes can be evaluated, either in a test subject (e.g., for
an organism based assay), or for a patient (e.g., prior, during or
after treatment with a therapeutic described herein.
[0258] A marker associated with caloric restriction can also be
evaluated in a subject organism of a screening assay (or a treated
subject). Although these markers may not be age-associated, they
may be indicative of a physiological state that is altered when the
Klotho pathway is modulated. The marker can be an mRNA or protein
whose abundance changes in calorically restricted animals.
WO01/12851 and U.S. Pat. No. 6,406,853 describe example markers.
Cellular models derived from cells of an animal described herein or
analogous to an animal model described herein can be used for a
cell-based assay.
[0259] Models for evaluating the effect of a test polypeptide on
muscle atrophy include: 1) rat medial gastrocnemius muscle mass
loss resulting from denervation, e.g., by severing the right
sciatic nerve at mid-thigh; 2) rat medial gastrocnemius muscle mass
loss resulting from immobilization, e.g., by fixed the right ankle
joint at 90 degrees of flexion; 3) rat medial gastrocnemius muscle
mass loss resulting from hind limb suspension; (see, e.g., U.S.
2003-0129686); 4) skeletal muscle atrophy resulting from treatment
with the cachectic cytokine, interleukin-1 (IL-1) (R. N. Cooney, S.
R. Kimball, T. C. Vary, Shock 7, 1-16 (1997)); and 5) skeletal
muscle atrophy resulting from treatment with the glucocorticoid,
dexamethasone (A. L. Goldberg, J. Biol. Chem. 244, 3223-9
(1969).)
[0260] Example animal models for AMD include: laser-induced mouse
model simulating exudative (wet) macular degeneration Bora et al.,
Proc. Natl. Acad. Sci. USA., 100:2679-84 (2003); a transgenic mouse
expressing a mutated form of cathepsin D resulting in features
associated with the "geographic atrophy" form of AMD (Rakoczy et
al., Am. J. Pathol., 161:1515-24 (2002)); and a transgenic mouse
over expressing VEGF in the retinal pigment epithelium resulting in
CNV. Schwesinger et al., Am. J. Pathol. 158:1161-72 (2001).
[0261] Example animal models of Parkinson's disease include
primates rendered Parkinsonian by treatment with the dopaminergic
neurotoxin 1-methyl-4 phenyl 1,2,3,6-tetrahydropyridine (MPTP)
(see, e.g., U.S. Patent Publication No. 20030055231 and Wichmann et
al., Ann. N.Y. Acad. Sci., 991:199-213 (2003);
6-hydroxydopamine-lesioned rats (e.g., Lab. Anim. Sci., 49:363-71
(1999)) ; and transgenic invertebrate models (e.g., Lakso et al.,
J. Neurochem. 86:165-72 (2003) and Link, Mech. Ageing Dev.,
122:1639-49 (2001)).
[0262] Example molecular models of Type II diabetes include: a
transgenic mouse having defective Nkx-2.2 or Nkx-6.1; (U.S. Pat.
No. 6,127,598); Zucker Diabetic Fatty fa/fa (ZDF) rat. (U.S. Pat.
No. 6,569,832); and Rhesus monkeys, which spontaneously develop
obesity and subsequently frequently progress to overt type 2
diabetes (Hotta et al., Diabetes, 50:1126-33 (2001); and a
transgenic mouse with a dominant-negative IGF-I receptor
(KR-IGF-IR) having Type 2 diabetes-like insulin resistance.
[0263] Example animal and cellular models for neuropathy include:
vincristine induced sensory-motor neuropathy in mice (U.S. Pat. No.
5,420,112) or rabbits (Ogawa et al., Neurotoxicology, 21:501-11
(2000)); a streptozotocin (STZ)-diabetic rat for study of autonomic
neuropathy (Schmidt et al., Am. J. Pathol., 163:21-8 (2003)); and a
progressive motor neuropathy (pmn) mouse (Martin et al., Genomics,
75:9-16 (2001)).
[0264] Example animal models of hyperphosphatemia or tumoral
calcinosis include Klotho knockout mice and FGF23 knockout mice
(Yoshida et al., Endocrinology 143:683-689 (2002)).
[0265] Example animal models of chronic renal disease or chronic
renal failure include COL4A3+/-mice (Beirowski et al., J. Am. Soc.
Nephrol. 17:1986-1994 (2006)).
[0266] Example animal models of cancer include the transplantation
or implantation of cancer cells or tissue into nude mice, as is
known in the art (Giovanella et al., Adv. Cancer Res. 44:69-120
(1985)). For example, animal models of breast cancer include nude
mice transplanted or implanted with breast cancer cells or tissue
(e.g., Yue et al., Cancer Res. 54:5092-5095 (1994); Glinsky et al.,
Cancer Res. 56:5319-5324 (1996); Visonneau Am. J. Path.
152:1299-1311 (1998)).
[0267] The compositions can be administered to a subject, e.g., an
adult subject, particularly a healthy adult subject or a subject
having an age-related disease. In the latter case, the method can
include evaluating a subject, e.g., to characterize a symptom of an
age-related disease or other disease marker, and thereby
identifying a subject as having a neurodegenerative disease, e.g.,
Alzheimer's or an age-related disease or being pre-disposed to such
a disease.
Skeletal Muscle Atrophy
[0268] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat skeletal muscle atrophy. Muscle atrophy includes
numerous neuromuscular, metabolic, immunological and neurological
disorders and diseases as well as starvation, nutritional
deficiency, metabolic stress, diabetes, aging, muscular dystrophy,
or myopathy. Muscle atrophy occurs during the aging process. Muscle
atrophy also results from reduced use or disuse of the muscle.
Symptoms include a decline in skeletal muscle tissue mass. In human
males, muscle mass declines by one-third between the ages of 50 and
80. Some molecular features of muscle atrophy include the
upregulation of ubiquitin ligases, and the loss of myofibrillar
proteins (Furuno et al., J. Biol. Chem., 265:8550-8557, 1990). The
breakdown of these proteins can be followed, e.g., by measuring
3-methyl-histidine production, which is a specific constituent of
actin, and in certain muscles of myosin (Goodman, Biochem. J.
241:121-12, 1987 and Lowell, et al., Metabolism, 35:1121-112, 1986;
Stein and Schluter, Am. J. Physiol. Endocrinol. Metab. 272:
E688-E696, 1997). Release of creatine kinase (a cell damage marker)
(Jackson, et al., Neurology, 41: 101 104, 1991) can also be
indicative.
Non-Insulin-Dependent Diabetes
[0269] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat Non-insulin-dependent Diabetes. Non-insulin-dependent
Diabetes is also called "adult onset" diabetes and Type 2 diabetes.
Type 2 diabetes also includes "non-obese type 2" and "obese type
2." Type II diabetes can be characterized by (1) reduced
pancreatic-beta-islet-cell secretion of insulin such that less than
necessary amounts of insulin are produced to keep blood glucose
levels in balance and/or (2) "insulin resistance," wherein the body
fails to respond normally to insulin. (U.S. Pat. No. 5,266,561 and
U.S. Pat. No. 6,518,069). For example, glucose-stimulated insulin
levels typically fail to rise above 4.0 nmol/L. (U.S. Pat. No.
5,266,561). Example symptoms of Type II diabetes include:
hyperglycemia while fasting (U.S. Pat. No. 5,266,561); fatigue;
excessive thirst; frequent urination; blurred vision; and an
increased rate of infections. Molecular indications of Type II
diabetes include islet amyloid deposition in the pancreases.
Neuropathy
[0270] Neuropathy can include a central and/or peripheral nerve
dysfunction caused by systemic disease, hereditary condition or
toxic agent affecting motor, sensory, sensorimotor or autonomic
nerves. (see, e.g., US Patent Application No. 20030013771).
Symptoms can vary depending upon the cause of the nerve damage and
the particular types of nerves affected. For example, symptoms of
motor neuropathy include clumsiness in performing physical tasks or
as muscular weakness, exhaustion after minor exertion, difficulty
in standing or walking and attenuation or absence of a
neuromuscular reflex. (U.S. Patent Application No. 20030013771)
symptoms of autonomic neuropathy include constipation, cardiac
irregularities and attenuation of the postural hypotensive reflex.
(U.S. Patent Application No. 20030013771), symptoms of sensory
neuropathy include pain and numbness; tingling in the hands, legs
or feet; and extreme sensitivity to touch, and symptoms of
retinopathy include blurred vision, sudden loss of vision, black
spots, and flashing lights.
Alzheimer's Disease
[0271] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat Alzheimer's Disease (AD). Alzheimer's Disease is a
complex neurodegenerative disease that results in the irreversible
loss of neurons. It provides merely one example of a
neurodegenerative disease that is also an age-related condition.
Clinical hallmarks of Alzheimer's Disease include progressive
impairment in memory, judgment, orientation to physical
surroundings, and language. Neuropathological hallmarks of AD
include region-specific neuronal loss, amyloid plaques, and
neurofibrillary tangles. Amyloid plaques are extracellular plaques
containing the amyloid peptide (also known as Ap, or Ap42), which
is a cleavage product of the, 8-amyloid precursor protein (also
known as APP). Neurofibrillary tangles are insoluble intracellular
aggregates composed of filaments of the abnormally
hyperphosphorylated microtubule-associated protein, taut Amyloid
plaques and neurofibrillary tangles may contribute to secondary
events that lead to neuronal loss by apoptosis (Clark and
Karlawish, Ann. Intern. Med. 138(5):400-410 (2003). For example,
p-amyloid induces caspase-2-dependent apoptosis in cultured neurons
(Troy et al. J Neurosci. 20(4):1386-1392). The deposition of
plaques in viva may trigger apoptosis of proximal neurons in a
similar manner.
[0272] A variety of criteria, including genetic, biochemical,
physiological, and cognitive criteria, can be used to evaluate AD
in a subject. Symptoms and diagnosis of AD are known to medical
practitioners. Some example symptoms and markers of AD are
presented below. Information about these indications and other
indications known to be associated with AD can be used as an
"AD-related parameter." An AD related parameter can include
qualitative or quantitative information. An example of quantitative
information is a numerical value of one or more dimensions, e.g., a
concentration of a protein or a tomographic map. Qualitative
information can include an assessment, e.g., a physician's comments
or a binary ("yes"/"no") and so forth. An AD-related parameter
includes information that indicates that the subject is not
diagnosed with AD or does not have a particular indication of AD,
e.g., a cognitive test result that is not typical of AD or a
genetic APOE polymorphism not associated with AD.
[0273] Progressive cognitive impairment is a hallmark of AD. This
impairment can present as decline in memory, judgment, decision
making, orientation to physical surroundings, and language
(Nussbaum and Ellis, New Eng J. Med. 348(14):1356 35 1364 (2003)).
Exclusion of other forms of dementia can assist in making a
diagnosis of AD. Neuronal death leads to progressive cerebral
atrophy in AD patients. Imaging techniques (e.g., magnetic
resonance imaging, or computer assisted tomography) can be used to
detect AD-associated lesions in the brain and/or brain atrophy.
[0274] AD patients may exhibit biochemical abnormalities that
result from the pathology of the disease. For example, levels of
tan protein in the cerebrospinal fluid is elevated in AD patients
(Andreasen, N. et al. Arch Neurol. 58:349-350 (2001)).
[0275] Levels of amyloid beta 42 (A,B42) peptide can be reduced in
CSF of AD patients. Levels of Ap42 can be increased in the plasma
of AD patients (Ertekein-Taner, N., et al. Science 290:2303 2304
(2000)). Techniques to detect biochemical abnormalities in a sample
from a subject include cellular, immunological, and other
biological methods known in the art. For general guidance, see,
e.g., techniques described in Sambrook & Russell, Molecular
Cloning: A Laboratory Manual, 3r Edition, Cold Spring Harbor
Laboratory, N.Y. (2001), Ausubel et al., Current Protocols in
Molecular Biology (Greene Publishing Associates and Wiley
Interscience, N.Y. (1989), (Harrow, E. and Lane, D. (1988)
Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory
Press, Cold Spring Harbor, N.Y.), and updated editions thereof.
[0276] For example, antibodies, other immunoglobulins, and other
specific binding ligands can be used to detect a biomolecule, e.g.,
a protein or other antigen associated with AD. For example, one or
more specific antibodies can be used to probe a sample. Various
formats are possible, e.g., ELISAs, fluorescence-based assays,
Western blots, and protein arrays. Methods of producing polypeptide
arrays are described in the art, e.g., in De Wildt et al. (2000).
Nature Biotech. 18, 989-994; Lucking et al. (1999). Anal. Biochem.
270, 103-111; Ge, H. (2000). Nucleic Acids Res. 28, e3, I-VII;
MacBeath, G., and Schreiber, S. L. (2000). Science 289, 1760 to 1
763; and WO 99/5 1 773A1.
[0277] In one assay, a non-human animal model of AD (e.g., a mouse
model) is used, e.g., to evaluate a polypeptide or a therapeutic
regimen. For example, U.S. Pat. No. 6,509,515 describes one such
model animal which is naturally able to be used with learning and
memory tests. The animal expresses an amyloid precursor protein
(APP) sequence at a level in brain tissues such that the animal
develops a progressive necrologic disorder within a short period of
time from birth, generally within a year from birth, preferably
within 2 to 6 months, from birth. The APP protein sequence is
introduced into the animal, or an ancestor of the animal, at an
embryonic stage, preferably the one cell, or fertilized oocyte,
stage, and generally not later than about the 8-cell stage. The
zygote or embryo is then developed to term in a pseudo-pregnant as
foster female. The amyloid precursor protein genes are introduced
into an animal embryo so as to be chromosomally incorporated in a
state which results in super endogenous expression of the amyloid
precursor protein and the development of a progressive necrologic
disease in the cortico-limbic areas of the brain, areas of the
brain which are prominently affected in progressive necrologic
disease states such as AD. The gliosis and clinical manifestations
in affected transgenic animals model necrologic disease. The
progressive aspects of the neurologic disease are characterized by
diminished exploratory and/or locomotor behavior and diminished
deoxyglucose uptake/utilization and hypertrophic gliosis in the
cortico-limbic regions of the brain. Further, the changes that are
seen are similar to those that are seen in some aging animals.
Other animal models are also described in U.S. Pat. Nos. 5,387,742;
5,877,399; 6,358,752; and 6,187,992.
Parkinson's Disease
[0278] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat Parkinson's Disease. Parkinson's disease includes
neurodegeneration of dopaminergic neurons in the substantia nigra
resulting in the degeneration of the nigrostriatal dopamine system
that regulates motor function. This pathology, in turn, leads to
motor dysfunctions.(see, e.g., and Lotharius et al., Nat. Rev.
Neurosci., 3:932-42 (2002)). Example motor symptoms include:
akinesia, stooped posture, gait difficulty, postural instability,
catalepsy, muscle rigidity, and tremor. Example non-motor symptoms
include: depression, lack of motivation, passivity, dementia and
gastrointestinal dysfunction (see, e. g., Fahn, Ann. N.Y. Acad.
Sci., 991:1-14 (2003) and Pfeiffer, Lancet Neurol., 2:107-16
(2003)) Parkinson's has been observed in 0.5 to 1 percent of
persons 65 to 69 years of age and 1 to 3 percent among persons 80
years of age and older. (see, e.g., Nussbaum et al., N Engl. J.
Med., 348:1356-64 (2003)). Molecular markers of Parkinson's disease
include reduction in aromatic L amino acid decarboxylase (AADC)
(see, e.g., US App. No. 20020172664); and loss of dopamine content
in the nigrostriatal neurons (see, e.g., Fahn, Ann. N.Y. Acad.
Sci., 991:1-14 (2003) and Lotharius et al., Nat. Rev. Neurosci.,
3:932-42 (2002)). In some familial cases, PD is linked to mutations
in single genes encoding alpha-synuclein and parkin (an E3
ubiquitin ligase) proteins. (e.g., Riess et al., J. Neurol. 250
Suppl 1:13 10 (2003) and Nussbaum et al., N. Engl. J. Med.,
348:1356-64 (2003)). A missense mutation in a neuron-specific
C-terminal ubiquitin hydrolase gene is also associated with
Parkinson's. (e.g., Nussbaum et al., N. Engl. J. Med., 348:1356-64
(2003))
Huntington's Disease
[0279] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat Huntington's Disease. Methods for evaluating the
efficacy and/or determining the effective dose of a Klotho fusion
polypeptide on Huntington's Disease include organismal based
assays, e.g., using a mammal (e.g., a mouse, rat, primate, or some
other non-human), or other animal (e.g., Xenopus, zebrafish, or an
invertebrate such as a fly or nematode). A number of animal model
system for Huntington's disease are available. See, e.g.,
Brouillet, Functional Neurology 15(4): 239-251 (2000); Ona et al.
Nature 399: 263-267 (1999), Bates et al. Hum Mol Genet.
6(10):1633-7 (1997); Hansson et al. J. of Neurochemistry 78:
694-703; and Rubinsztein, D. C., Trends in Genetics, Vol. 1S, No.
4, pp. 202-209 (a review on various animal and non-human models of
HD).
[0280] An example of such an animal model is the transgenic mouse
strain is the R6/2 line (Mangiarini et al. Cell 87: 493-506
(1996)). The R6/2 mice are transgenic Huntington's disease mice,
which over-express exon 1 of the human HD gene (under the control
of the endogenous promoter). The exon 1 of the R6/2 human HD gene
has an expanded CAG/polyglutamine repeat lengths (150 CAG repeats
on average). These mice develop a progressive, ultimately fatal
neurological disease with many features of human Huntington's
disease. Abnormal aggregates, constituted in part by the N terminal
part of Huntingtin (encoded by HD exon 1), are observed in R6/2
mice, both 45 in the cytoplasm and nuclei of cells (Davies et al.
Cell 90: 537-548 (1997)). For example, the human Huntingtin protein
in the transgenic animal is encoded by a gene that includes at
least 55 CAG repeats and more preferably about 150 CAG repeats.
These transgenic animals can develop a Huntington's disease-like
phenotype.
[0281] These transgenic mice are characterized by reduced weight
gain, reduced lifespan and motor impairment characterized by
abnormal gait, resting tremor, hindlimb clasping and hyperactivity
from 8 to 10 weeks after birth (for example the R6/2 strain; see
Mangiarini et al. Cell 87: 493-506 (1996)). The phenotype worsens
progressively toward hypokinesia. The brains of these transgenic
mice also demonstrate neurochemical and histological abnormalities,
such as changes in neurotransmitter receptors (glutamate,
dopaminergic), decreased concentration of N-acetylaspartate (a
marker of neuronal integrity) and reduced striatum and brain size.
Accordingly, evaluating can include assessing parameters related to
neurotransmitter levels, neurotransmitter receptor levels, brain
size and striatum size. In addition, abnormal aggregates containing
the transgenic part of or full-length human Huntingtin protein are
present in the brain tissue of these animals (e.g., the R6/2
transgenic mouse strain). See, e.g., Mangiarini et al. Cell 87:
493-506 (1996), Davies et al. Cell 90: 537-548 (1997), Brouillet,
Functional Neurology 15(4): 239-251 (2000) and Cha et al. Proc.
Natl. Acad. Sci. USA 95: 6480-6485 (1998).
[0282] To test the effect of the test polypeptide or known
polypeptide described in the application in an animal model,
different concentrations of test polypeptide are administered to
the transgenic animal, for example by injecting the test
polypeptide into circulation of the animal. A Huntington's
disease-like symptom may be evaluated in the animal. The
progression of the Huntington's disease-like symptoms, e.g., as
described above for the mouse model, is then monitored to determine
whether treatment with the test polypeptide results in reduction or
delay of symptoms. In another assay, disaggregation of the
Huntingtin protein aggregates in these animals is monitored. The
animal can then be sacrificed and brain slices are obtained. The
brain slices are then analyzed for the presence of aggregates
containing the transgenic human Huntingtin protein, a portion
thereof, or a fusion protein comprising human Huntingtin protein,
or a portion thereof. This analysis can includes, for example,
staining the slices of brain tissue with anti-Huntingtin antibody
and adding a secondary antibody conjugated with FITC which
recognizes the anti-Huntington's antibody (e.g., the
anti-Huntingtin antibody is mouse anti-human antibody and the
secondary antibody is specific for human antibody) and visualizing
the protein aggregates by fluorescent microscopy.
[0283] A variety of methods are available to evaluate and/or
monitor Huntington's disease. A variety of clinical symptoms and
indicia for the disease are known. Huntington's disease causes a
movement disorder, psychiatric difficulties and cognitive changes.
The degree, age of onset, and manifestation of these symptoms can
vary. The movement disorder can include quick, random, dance-like
movements called chorea.
[0284] Example motor evaluations include: ocular pursuit, saccade
initiation, saccade velocity, dysarthria, tongue protrusion, finger
tap ability, pronate/supinate, a lo fist-hand-palm sequence,
rigidity of arms, bradykinesia, maximal dystonia (trunk, upper and
lower extremities), maximal chorea (e.g., trunk, face, upper and
lower extremities), gait, tandem walking, and retropulsion. An
example treatment can cause a change in the Total Motor Score 4
(TMS-4), a subscale of the UHDRS, e.g., over a one-year period.
Cancer
[0285] Methods or uses of the disclosure which provide
administering the Klotho fusion polypeptide to an individual can be
used to treat cacner. Cancer includes any disease that is caused by
or results in inappropriately high levels of cell division,
inappropriately low levels of apoptosis, or both. Examples of
cancers include, without limitation, leukemias (e.g., acute
leukemia, acute lymphocytic leukemia, acute myelocytic leukemia,
acute myeloblastic leukemia, acute promyelocytic leukemia, acute
myelomonocytic leukemia, acute monocytic leukemia, acute
erythroleukemia, chronic leukemia, chronic myelocytic leukemia,
chronic lymphocytic leukemia), polycythemia vera, lymphoma
(Hodgkin's disease, non-Hodgkin's disease), Waldenstrom's
macroglobulinemia, heavy chain disease, and solid tumors such as
sarcomas and carcinomas (e.g., fibrosarcoma, myxosarcoma,
liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma,
angiosarcoma, endotheliosarcoma, lymphangiosarcoma,
lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's
tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma,
pancreatic cancer, breast cancer, ovarian cancer, prostate cancer,
squamous cell carcinoma, basal cell carcinoma, adenocarcinoma,
sweat gland carcinoma, sebaceous gland carcinoma, papillary
carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary
carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma,
nile duct carcinoma, choriocarcinoma, seminoma, embryonal
carcinoma, Wilm's tumor, cervical cancer, uterine cancer,
testicular cancer, lung carcinoma, small cell lung carcinoma,
bladder carcinoma, epithelial carcinoma, glioma, astrocytoma,
medulloblastoma, craniopharyngioma, ependymoma, pinealoma,
hemangioblastoma, acoustic neuroma, oligodenroglioma, schwannoma,
meningioma, melanoma, neuroblastoma, and retinoblastoma).
Lymphoproliferative disorders are also considered to be
proliferative diseases.
[0286] All patents, patent applications, and published references
cited herein are hereby incorporated by reference in their
entirety. While this disclosure has been particularly shown and
described with references to embodiments thereof, it will be
understood by those skilled in the art that various changes in form
and details may be made therein without departing from the scope of
the disclosure encompassed by the appended claims.
5. EXAMPLES
Example 1
Expression and Purification of Klotho Fusion Polypeptides
[0287] Expression of the Klotho Fusion Polypeptide
[0288] The polypeptides of the disclosure were made by transiently
transfecting HEK293T cells with an expression vector encoding a
Klotho fusion polypeptide having the extracellular domain of alpha
Klotho and the FGF23 (R179Q) variant. Conditioned media containing
expressed polypeptides were generated by transient transfection of
the respective expression plasmids for Klotho, FGF23, and the
Klotho-FGF23(R179Q) fusion protein. The transfections were
performed in 6-well plates using Lipofectamine 2000 (Invitrogen,
Cat #11668-019). Five hours after transfection, the transfection
mix was replaced with 3 ml DMEM plus 1% FBS. Conditioned media were
collected 72 hours after the addition of 3 ml DMEM plus 1% FBS.
Samples of conditioned medium from various transiently transfected
HEK293T cells were separated by SDS-polyacrylamide gel
electrophoresis (SDS-PAGE) and analyzed by Western blot (FIG. 3A)
or stained with Coomassie blue (FIG. 3B).
[0289] SDS-polyacrylamide gel electrophoresis was performed on
various samples (lane 1, Control; lane 2, FGF23; lane 3, sKlotho;
lanes 4-6, sKlotho-FGF23). Coomassie blue staining revealed the
expression of a high, >180 kDa band (FIG. 3B, indicated by arrow
on the right) that was not present in lanes 1-3, which contained
samples that had not been transfected with the vector encoding the
Klotho fusion polypeptide. The quality of the Klotho fusion
polypeptide secreted into the media was evaluated by Western blot
(FIG. 3A). An anti-FGF23 rat monoclonal IgG2A (R&D Systems,
Cat#MAB26291) was used as the primary antibody to detect the Klotho
fusion polypeptides by Western blot. The Western blot confirmed
that the additional bands observed in the Coomassie stained gels
were Klotho fusion polypeptides. The Western blot confirmed that
the Klotho fusion polypeptides had the expected molecular weight
for the Klotho fusion polypeptide. This analysis shows the
expression of the Klotho-FGF23(R179Q) fusion protein.
[0290] Purification of the Klotho Fusion Polypeptide
[0291] The polypeptides of the disclosure were purified from
conditioned media from a culture of HEK293T cells transiently
transfected with an expression vector encoding a Klotho fusion
polypeptide having the extracellular domain of alpha Klotho and the
FGF23 R179Q variant. To generate conditioned medium, an expression
vector encoding sKlotho-FGF23-6.times.His was transfected (500
.mu.g DNA in 18 ml of OptiMEM 1 (GIBCO, Cat #11058) mixed with 18
ml of 2 .mu.g/ml polyethlinimine (PEI) into HEK293 cells grown in
suspension in expression medium (464 ml of HEK293T cells at
10.sup.6 cells/ml in Freestype 293 expression medium (GIBCO, Cat
#12338)). After transfection, the culture was allowed to grow (120
hours; 37.degree. C. in a 5% CO.sub.2 incubator; shaking at 125
rpm). At the end of incubation, conditioned medium was harvested by
centrifugation (1000 rpm for five minutes). The conditioned medium
was then applied to a nickel-agarose column. The
sKlotho-FGF23-6.times.His bound tightly to the column and was
eluted with 50 mM imidazole. The resulting purified material was
then dialyzed in PBS to remove imidazole. A sample of the purified
sKlotho-FGF23-6.times.His was separated by SDS-PAGE (lane 1,
purified sKlotho-FGF23-6.times.His; lane 2, molecular weight
marker) and analyzed by staining with Coomassie blue (FIG. 3C). The
stained SDS-PAGE gel confirmed that the purified
sKlotho-FGF23-6.times.His had the expected molecular weight. The
inability to detect bands corresponding to proteins other than
full-length sKlotho-FGF23-6.times.His in the lane loaded with the
purified material also showed that the sKlotho-FGF23-6.times.His
was purified.
Example 2
In Vitro Assay Assessing the Activity of the Klotho Fusion
Polypeptide
[0292] Egr-1-luciferase
[0293] The biological activity of the expressed alpha Klotho fusion
polypeptide was tested in Egr-1-luciferase reporter assays. Binding
of the Klotho fusion polypeptide to the FGF23 receptor resulted in
the downstream activation of Egr-1 and the expression of a
luciferase reporter regulated by the Egr-1 promoter. The
Egr-1-luciferase reporter gene was constructed based on that
reported by Urakawa et al. (Nature, 2006, Vol 444, 770-774).
HEK293T cells seeded in 48-well poly-D-lysine plate were
transfected with the Egr-1-luciferase reporter gene together with a
transfection normalization reporter gene (Renilla luciferase). Five
hours after transfection of the Egr-1 luciferase reporter gene, the
transfection mix was replaced with 3 ml DMEM plus 1% FBS.
Conditioned media were collected 72 hours after the addition of 3
ml DMEM plus 1% FBS. Five hours later, the transfection mix was
replaced with a sample to be tested for activity. In initial
experiments, 50% conditioned medium (alone or containing Klotho,
FGF23, Klotho and FGF23, and the Klotho-FGF23(R179Q) fusion
protein) and 50% DMEM with 1% FBS in the presence or absence of 20
.mu.g/ml heparin (Sigma, Cat#H8537; dissolved in DMEM as 2 mg/ml
stock) were tested in the Egr-1-luciferase reporter assays (FIG.
4). Further experiments used defined quantities of the purified
polypeptides (FIGS. 5A and 5B). Cells were lysed 20 hours later in
passive lysis buffer (Promega, Cat #E194A) and luciferase
activities were determined using Dual-Glo Luciferase Assay System
(Promega, Cat #E2940).
[0294] In initial experiments, Klotho fusion polypeptide activity
was demonstrated in unfractionated conditioned medium. Using the
Egr-1-luciferase reporter gene (FIG. 4) these experiments
quantified the fold changes in the expression of the luciferase
reporter. Conditioned medium containing a combination of FGF23 and
the extracellular domain of Klotho protein activated
Egr-1-luciferase, but conditioned medium containing only FGF23 or
conditioned medium containing only the extracellular domain of
Klotho, did not activate Egr-1-luciferase. Conditioned medium
containing the fusion protein sKlotho-FGF23(R179Q) activated the
Egr-1-luciferase reporter gene in contrast to conditioned media
containing either FGF23 or Klotho alone. In these experiments,
conditioned medium containing the fusion protein
sKlotho-FGF23(R179Q) activated the Egr-1-luciferase reporter gene
significantly better than conditioned medium containing a
combination of FGF23 and Klotho. In the presence of heparin, the
inductions by conditioned medium containing the fusion protein
sKlotho-FGF23(R179Q) and the conditioned medium containing a
combination of FGF23 and Klotho were significantly enhanced. Table
1 lists the relative expression of various FGF-Klotho fusion
polypeptides in conditioned medium and the relative activity of the
unfractionated conditioned medium corresponding to the various
FGF-Klotho fusion polypeptides in Egr-1-luciferase reporter
assays.
TABLE-US-00001 TABLE 1 Expression and Activities of sKlotho-FGF23
fusion variants sKlotho-FGF23 fusion Activity in Egr-1- constructs
Expression luc reporter gene 1 sKlotho-FGF23 good yes 2 IgG
sp-sKlotho-FGF23 good yes 3 sKL-D1-FGF23 good no 4 sKL-D2-FGF23 no
n.a. 5 s(KL-D1)2-FGF23 good no 6 sKL-D1/D2-FGF23 no n.a. 7
ssKlotho(.DELTA.N-26)-FGF23 poor no* 8
sKLD1-D2(.DELTA.692-965)-FGF23 poor no* 9
sKL-D1-D2(.DELTA.507-798)-FGF23 poor no* 10 FGF23-sKlotho poor no*
*lack of activity may be the result of low expression
[0295] Egr-1-luciferase reporter assays were also performed using
defined quantities of proteins purified from the conditioned
medium, using the purification procedure as described in Example 1.
Consistent with previous results using unfractionated conditioned
medium containing the expressed polypeptides, treatment with a
combination of purified FGF23 and sKlotho resulted in luciferase
reporter activity, but treatment with purified FGF23 alone did not
(FIG. 5A). The luciferase reporter activity from the combination of
purified FGF23 and sKlotho was further dependent on the dose of
purified sKlotho, and the effect could be enhanced by the presence
of heparin (20 .mu.g/ml). An effect of the
sKlotho-FGF23-6.times.His fusion polypeptide on luciferase activity
could be detected at concentrations as low as about 1.21 nM (1.2
fold change) and at least up to about 19.3 nM (2.4 fold change) in
Egr-1-luciferase reporter assays (FIG. 5B). The activity of the
sKlotho-FGF23-6.times.His fusion polypeptide on luciferase activity
was significantly enhanced in the presence of heparin (20
.mu.g/ml). In the presence of heparin, the effect of the
sKlotho-FGF23-6.times.His fusion polypeptide on luciferase activity
could be detected at a concentration as low as about 0.6 nM (2.0
fold change). The result showed that purified
sKlotho-FGF23-6.times.His dose-dependently induced the EGR-1-luc
reporter gene, and that treatment with
sKlotho-FGF23-6.times.His.
Example 3
In Vitro Assay Assessing the Effect of the Klotho Fusion
Polypeptide on Muscle Cells
[0296] The biological effect of the expressed Klotho fusion
polypeptide was tested on C2C12 myoblasts. Treatment of C2C12
myoblasts with IGF-1, FGF2, or sKlotho-FGF23 resulted in myotube
growth and phosphorylation of signaling proteins. C2C12 myoblasts
were seeded at a density of 40,000 cells/well in 6-well
poly-D-lysine and fibronectin coated plates in growth medium (3
parts DMEM and 1 part F12), 10% FBS, 1% Glut; 1% P/S; 1% Linolic
acid; 0.1% ITS: [insulin (10 mg/ml), transferrin (5.5 mg/ml), and
selenium (5 ng/ml)]. After myoblasts reached confluence (3 days),
medium was changed into differentiation medium (DMED with 2% horse
serum; 1% Glut; 1% P/S).
[0297] For the myotube diameter experiments, three days after
confluent media was changed into differentiation medium, cells were
treated with IGF-1 (10 nM), FGF2 (20 ng/ml) or sKlotho-FGF23 (20
nM) in the absence or presence of dexamethasone (100 .mu.M) for 24
hours in differentiation medium. At the end of treatment, cells
were fixed with glutaraldehyde (5% in PBS) and multiple fluorescent
images were collected. Myotube diameter was measured using the
Pipeline Pilot program to determine hypertrophy or atrophy.
[0298] For the signaling protein phosphorylation,experiments, three
days after confluent media was changed into differentiation medium,
cells were starved for four hours with DMEM without FBS and then
treated with IGF-1 (10 nM), FGF2 (20 ng/ml) or sKlotho-FGF23 (20
nM) in the absence or presence of Rapamycin (40 nM) for 30 min.
Cells were lysed in RIPA buffer in the presence of protease and
phosphatase inhibitors. Western blot analysis was carried out and
membranes were probed with different antibodies as indicated in the
figure and developed on X-ray films, which were scanned.
[0299] The results of this study showed that sKlotho-FGF23 resulted
in an increase in myotube diameter compared to the control and
induced C2C12 myotube hypertrophy similar to results for IGF-1 and
FGF2 (FIG. 5A). In addition, treatment with sKlotho-FGF23, IGF-1,
and FGF2 could partially reverse myotube atrophy induced by
dexamethasone, based on measurements of myotube diameter. No
difference was observed between sKlotho-FGF23 and FGF2 on myotube
morphology (measured by thickness of the myotubes) in the absence
or presence of dexamethasone. The trophic effects of sKlotho-FGF23,
IGF-1, and FGF2 were statistically significant.
[0300] Consistent with the effects on C2C12 myotubes, sKlotho-FGF23
fusion protein signaling led to the phosphorylation of p70S6K and
ERK, but not AKT or FoxO, in C2C12 myotubes (FIG. 5B). The effect
of sKlotho-FGF23 on signaling was similar to that of FGF2, but was
distinct from that of IGF-1. The extent of ERK phosphorylation by
sKlotho-FGF23 was observed to be less than that of IGF-1 or FGF2.
The phosphorylation of p70S6K by sKlotho-FGF23 was rapamycin
sensitive. In the experiments involving C2C12 cells, heparin was
not required to activate signaling. These results show that a
sKlotho-FGF23 fusion polypeptide activated signaling in C2C12
myotubes.
Example 4
Fusion Polypeptides Comprising sKlotho, FGF23 and FcLALA
[0301] Various fusion polypeptides are constructed using sKlotho,
FGF23, and a modified Fc fragment of an antibody. These modified Fc
molecules have altered (decreased) binding to FcRn and thus
increased serum half-life. They also have modified bioavailability
and alterered transport to mucosal surfaces and other targets in
the body. In this example, the FGF23 and sKlotho are fused to
FcLALA, which is described in U.S. Pat. No. 7,217,798 and Hessell
et al. 2007 Nature 449:101-104, Intervening between the various
components of these fusion polypeptides are linkers, as described
in Lode et al. 1998 Proc. Natl. Acad. Sci. USA 95: 2475-2480. These
fusions are inserted into constructs, e.g., pcDNA3.1 (Invitrogen,
Carlsbad, Calif.), and expressed in HEK293 cells.
[0302] A. sKlotho-FGF23-FcLALA v1
[0303] A fusion is constructed which comprises: sKlotho, a linker,
FGF23, another linker, and FcLALA. This embodiment, designated
sKlotho-FGF23-FcLALA v1, is presented in SEQ ID NOs: 46 and 47,
below.
[0304] The nucleotide sequence of sKlotho-FGF23-FcLALA v1 (wherein
initiation ATG as 1) is presented as SEQ ID NO: 46.
[0305] The amino acid sequence of sKlotho-FGF23-FcLALA v1 is
presented below as SEQ ID NO: 47.
[0306] In this sequence, the various components of the fusion are
as follows:
[0307] sKlotho: 1-982; Linker1: 983-1001; FGF23: 1002-1228; Linker
2; 1229-1233; FcLALA: 1234-1459.
[0308] B. sKlotho-FGF23-FcLALA v2
[0309] A fusion is constructed which comprises: sKlotho, a linker,
FGF23, another linker, and FcLALA. This embodiment is designated
sKlotho-FGF23-FcLALA v2 and presented as SEQ ID NOs: 48 and 49,
below.
[0310] The nucleotide sequence of sKlotho-FGF23-FcLALA v2 (wherein
initiation ATG as 1) is presented as SEQ ID NO: 48.
[0311] The amino acid sequence of sKlotho-FGF23-FcLALA v2 is
presented below as SEQ ID NO: 49.
[0312] In this sequence, the various components of the fusion are
as follows:
[0313] sKlotho: (aa or amino acids) 1-982; Linker 1: 983-1001;
FGF23: 1002-1228; Linker 2; 1229-1233; FcLALA: 1234-1450.
[0314] Other fusion polypeptides can be constructed by combining in
various combinations the FGF, Klotho, modified Fc fragments, and
(optionally) linker sequences, and variants and derivatives
thereof, as described herein or known in the art.
Example 5
Fusion Polypeptides Comprising FGF23 and FcLALA
[0315] Various fusion polypeptides are constructed using FGF23, and
a modified Fc fragment of an antibody, as described in U.S. Pat.
No. 7,217,798. These modified Fc molecules have altered (decreased)
binding to FcRn and thus increased serum half-life. They also have
modified bioavailability and alterered transport to mucosal
surfaces and other targets in the body. In this example, FGF23 is
fused to FcLALA, Intervening between the various components of
these fusion polypeptides are linkers, as described in Lode et al.
1998 Proc. Natl. Acad. Sci. USA 95: 2475-2480. These fusions are
inserted constructs, e.g., pcDNA3.1 (Invitrogen, Carlsbad, Calif.),
and expressed in HEK293 cells.
[0316] C. FGF23-FcLALA v1
[0317] A fusion is constructed which comprises: FGF23, a linker,
and FcLALA. This construct is designated FGF23-FcLALA v1 and
presented below as SEQ ID NOs: 50 and 51.
[0318] The nucleotide sequence of FGF23-FcLALA v1 (wherein
initiation ATG as 1) is presented below as SEQ ID NO: 50.
[0319] The amino acid sequence of FGF23(R179Q)-FcLALAv1 is
presented below as SEQ ID NO: 51.
[0320] In this sequence, the various components of the fusion are
as follows:
[0321] FGF23: (aa) 1-251; Linker: 252-256; FcLALA: 257-482.
[0322] D. FGF23-FcLALA v2
[0323] A fusion is constructed which comprises: FGF23-FcLALA v2,
which comprises FGF23 and FcLALA.
[0324] The nucleotide sequence of FGF23-FcLALA v2 (wherein
initiation ATG as 1) is presented below as SEQ ID NO: 52.
[0325] The amino acid sequence of FGF23(R179Q)-FcLALAv2 is
presented below as SEQ ID NO: 53.
[0326] In this sequence, the various components of the fusion are
as follows:
[0327] FGF23: 1-251; Linker: 252-256; FcLALA: 257-473.
[0328] Other fusion polypeptides can be constructed by combining in
various combinations the FGF sequences, modified Fc fragments, and
(optionally) linkers, and variants and derivatives thereof, as
described herein or known in the art.
[0329] E. Activation of Egr-1-luc reporter gene by
sKlotho-FGF23(R179Q)-FcLALA fusion proteins; activation of
Egr-1-luc reporter gene by FGF23(R179Q)-FcLALA proteins; and
pharmacokinetic profile of FGF23(R179Q) vs FGF23(R179Q)-FcLaLav2
are determined.
[0330] FIG. 7 shows the activation of Egr-1-luc reporter gene by
sKlotho-FGF23(R179Q)-FcLALA fusion proteins. HEK293T cells are
transiently transfected with the Egr-1-luc reporter gene and
incubated with the indicated conditioned media in the absence or
presence of 20 .mu.g/ml heparin. Luciferase activities are then
determined 18 hours later. The result shows that
sklotho-FGF23-FcLALA fusion proteins induces the reporter gene
activity. These inductions are significantly enhanced in the
presence of heparin. sKF-Fcv1: sKlotho-FGF23-FcLALAv1; sKF-Fcv2:
sKlotho-FGF23-FcLALAv2
[0331] FIG. 8 shows the activation of Egr-1-luc reporter gene by
FGF23(R179Q)-FcLALA proteins. HEK293T cells are transiently
transfected with the Egr-1-luc reporter gene together with the
full-length transmembrane form of Klotho and incubated with the
indicated 30% conditioned media. Luciferase activities are then
determined 18 hours later. The results show that FGF23-FcLALA
fusion proteins induce the reporter gene activity in a similar
manner as the FGF23.
[0332] FIG. 9 shows the pharmacokinetic profile of FGF23(R179Q) vs
FGF23(R179Q)-FcLALAv2. Four mice per group are injected
subcutaneously with FGF23(R179Q)-6.times.His or
FGF23(R179Q)-FcLALAv2 at 2 mg/kg. At the indicated times, serum
samples are collected and analyzed for FGF23 by ELISA.
FGF23(R179Q)-FcLALA concentration in serum remains elevated at the
24 hr time point, while FGF23(R179Q)-6.times.His is back to basal
level. This results indicate that with the addition of FcLALA, the
in vivo half-life of FGF23(R179Q) is significantly improved.
Example 6
In Vivo Efficacy of sKlotho-FGF23 Fusion in Enhancing Muscle Growth
After Dexamethasone-Induced Muscle Atrophy
[0333] Experimental data shows that intramuscular injection of
sKlotho-FGF23 significantly enhanced growth of muscle mass after
dexamethasone-induced muscle atrophy. In this experiment, the
peptide corresponding to that of SEQ ID NO: 41 is used.
[0334] FIG. 10A shows absolute weights of the
gastrocnemius-soleus-plantaris (GSP) muscles showing that
intramuscular injection of sKlotho-FGF23 (KLOFGF) significantly
enhanced regrowth of muscle mass after dexamethasone (DEX)-induced
muscle atrophy compared with intramuscular injection of sKlotho
(sKLO) or phosphate buffered saline (PBS). FIG. 10B shows percent
weight change (B) of the gastrocnemius-soleus-plantaris (GSP)
muscles showing that intramuscular injection of sKlotho-FGF23
(KLOFGF) significantly enhanced regrowth of muscle mass after
dexamethasone (DEX)-induced muscle atrophy compared with
intramuscular injection of sKlotho (sKLO) or phosphate buffered
saline (PBS).
[0335] Eighty male C57BL/6 mice, aged 15 weeks, are randomized by
body weight into 8 groups each of 10 mice. Four groups receive
water without DEX (W21d) while the other four receive DEX in
drinking water at 2.4 mg/kg/day for three weeks (D21d). After the
three weeks, DEX treatment is stopped and one W21d and one D21d
group is immediately sacrificed to establish the degree of muscle
atrophy induced by the DEX treatment. The remaining three groups of
W21d or D21d mice are allowed to recover for another 14 days (R14d)
during which period they receive an intramuscular injection of
2.times.50 .mu.l of PBS, sKlotho-FGF23 (KLOFGF; 1.6 mg/ml), or
sKlotho (sKLO; 1.6 mg/ml), respectively, every other day into the
right gastrocnemius-soleus-plantaris muscle complex. The mice are
sacrificed 24 h after the last intramuscular injection and the
muscle weights determined and expressed as absolute weight (A) or
percent change compared to the W21d+PBS group.
[0336] These data show the in vivo efficacy of sKlotho-FGF23 fusion
in enhancing muscle growth after dexamethasone-induced muscle
atrophy.
Example 7
Additional Mutations in the FGF23 Portion of Fusion Proteins which
Reduce Aggregation, Reduce Undesired Protease-Induced Cleavage, and
Increase Production
[0337] Several mutations are investigated within the FGF23 portion
of sKlotho-FGF23 and FGF23-FcLaLa fusion polypeptides. These
include Q156, C206 and C244 (wherein the number is based on the
FGF23 amino acid sequence). Example individual mutations include
Q156A, C2065 and C244S, and mutations at any of these sites can be
combined with a mutation at R179 (e.g., R179Q). Example sequences
are provided in SEQ ID NO: 54 to 68 of FIG. 2.
[0338] C206 and C244 are suspected to be involved in dimerization;
and Q156 is a site identified by the inventors as a protease
sensitive site. Mutating these amino acids to any other amino acid
enhances the qualities of the proteins, by reducing aggregation,
reducing undesired protease-induced cleavage, and increasing
protein production from cells, without interfering with FGF23
activity. This is an unexpected result, as these three positions
are conserved in the FGF23 proteins found in human, rhesus, bovine,
mouse and rat. This conservation is shown below in the comparison
between SEQ ID NOs: 69, 70, 71, 72 and 73, with the Q156, C206 and
C244 in bold, underlined font.
TABLE-US-00002 hFGF23
MLGARLRLWVCALCSVCSMSVLRAYPNASPLLGSSWGGLIHLYTATARNSYHLQIHKNGH rhesus
MLGARLRLWVCALCSVCSMSVIRAYPNASPLLGSSWGGLIHLYTATARNSYHLQIHKNGH bovine
MLGARLGLWVCTLSCV-----VQAYPNSSPLLGSSWGGLTHLYTATARNSYHLQIHGDGH mouse
MLGTCLRLLVGVLCTVCSLGTARAYPDTSPLLGSNWGSLTHLYTATARTSYHLQIHRDGH rat
MLGACLRLLVGALCTVCSLGTARAYSDTSPLLGSNWGSLTHLYTATARNSYHLQIHRDGH hFGF23
VDGAPHQTIYSALMIRSEDAGFVVITGVMSRRYLCMDFRGNIFGSHYFDPENCRFQHQTL rhesus
VDGAPHQTIYSALMIRSEDAGFVVITGVMSRRYLCMDFRGNIFGSHYFNPENCRFRHWTL bovine
VDGSPQQTVYSALMIRSEDAGFVVITGVMSRRYLCMDFTGNIFGSHHFSPESCRFRQRTL mouse
VDGTPHQTIYSALMITSEDAGSVVITGAMTRRFLCMDLHGNIFGSLHFSPENCKFRQWTL rat
VDGTPHQTIYSALMITSEDAGSVVIIGAMTRRFLCMDLRGNIFGSYHFSPENCRFRQWTL hFGF23
ENGYDVYHSPQYHFLVSLGRAKRAFLPGMNPPPYSQFLSRRNEIPLIHFNTPI-PRRHTR rhesus
ENGYDVYHSPQHHFLVSLGRAKRAFLPGMNPPPYSQFLSRRNEIPLIHFNTPR-PRRHTR bovine
ENGYDVYHSPQHRFLVSLGRAKRAFLPGTNPPPYAQFLSRRNEIPLPHFAATARPRRHTR mouse
ENGYDVYLSQKHHYLVSLGRAKRIFQPGTNPPPFSQFLARRNEVPLLHFYTVR-PRRHTR rat
ENGYDVYLSPKHHYLVSLGRSKRIFQPGTNPPPFSQFLARRNEVPLLHFYTAR-PRRHTR hFGF23
SAEDDSERDPLNVLKPRARMTPAPASCSQELPSAEDNSPMASDPLGVVRGGRVNTHAGGT rhesus
SAEDDSERDPLNVLKPRARMTPAPASCSQELPSAEDNSPVASDPLGVVRGGRVNTHAGGT bovine
SAHDSG--DPLSVLKPRARATPVPAACSQELPSAEDSGPAASDPLGVLRGHRLDVRAGSA mouse
SAEDPPERDPLNVLKPRPRATPVPVSCSRELPSAEEGGPAASDPLGVLRRGRGDARGGAG rat
SAEDPPERDPLNVLKPRPRATPIPVSCSRELPSAEEGGPAASDPLGVLRRGRGDARRGAG hFGF23
GPEGCRPFAKFI (SEQ ID NO: 69) rhesus GPEACRPFPKFI (SEQ ID NO: 70)
bovine GAERCRPFPGFA (SEQ ID NO: 71) mouse GADRCRPFPRFV (SEQ ID NO:
72) rat GTDRCRPFPRFV (SEQ ID NO: 73)
[0339] The fact that these three mutations do not prevent FGF23
activity is shown in FIG. 11. This figure shows activation of
Egr-1-luc reporter gene by FGF23(R179Q)-FcLALA and Q156A, C206S,
C244S and C206S/C244S mutants.
[0340] HEK293T cells are transiently transfected with the EGR-1-luc
reporter gene together with the full-length transmembrane form of
Klotho and indicated FGF23-FcLaLa mutants. Luciferase activities
are then determined 18 hours later. The results show that C206S,
C244S, C206S/C244S (three independent clones) and Q156A (three
independent clones) mutants are equally effective as FGF23-FcLALA
fusion proteins in activating EGR-1-Luc reporter gene activity.
[0341] Data showing that mutating C244 and C206 alter dimerization
and aggregation of FGF23 is shown in FIG. 12. This figure shows
protein qualities of WT, Q156A, C206S, C244S and C206S/C244S
mutants of FGF23(R179Q)-FcLaLa. Conditioned medium from HEK293T
cells transient transfected with the indicated FGF23-FcLaLa
expression vectors are analyzed by Western blot using an FGF23
antibody. The result shows that C206S/C244S mutation prevents
protein dimerization and Q156A mutation has reduced proteolytic
fragments.
[0342] Thus, surprisingly, even though these Q156, C206 and C244
residues are conserved across species, they can mutated without
reducing FGF23 activity and can enhance the qualities of the
protein by reducing aggregation and cleavage and by improving
production.
[0343] Unless defined otherwise, the technical and scientific terms
used herein have the same meaning as that usually understood by a
specialist familiar with the field to which the disclosure
belongs.
[0344] Unless indicated otherwise, all methods, steps, techniques
and manipulations that are not specifically described in detail can
be performed and have been performed in a manner known per se, as
will be clear to the skilled person. Reference is for example again
made to the standard handbooks and the general background art
mentioned herein and to the further references cited therein.
[0345] Claims to the invention are non-limiting and are provided
below.
[0346] Although particular embodiments and claims have been
disclosed herein in detail, this has been done by way of example
for purposes of illustration only, and is not intended to be
limiting with respect to the scope of the appended claims, or the
scope of subject matter of claims of any corresponding future
application. In particular, it is contemplated by the inventors
that various substitutions, alterations, and modifications may be
made to the disclosure without departing from the spirit and scope
of the disclosure as defined by the claims. The choice of nucleic
acid starting material or clone of interest is believed to be a
matter of routine for a person of ordinary skill in the art with
knowledge of the embodiments described herein. Other aspects,
advantages, and modifications considered to be within the scope of
the following claims. Redrafting of claim scope in later filed
corresponding applications may be due to limitations by the patent
laws of various countries and should not be interpreted as giving
up subject matter of the claims.
TABLE-US-00003 SEQUENCE LISTING (FIG 2) Human Klotho nucleic acid
sequence (NM_004795) (SEQ ID NO: 1) Protein coding region: 9-3047 1
cgcgcagcat gcccgccagc gccccgccgc gccgcccgcg gccgccgccg ccgtcgctgt
61 cgctgctgct ggtgctgctg ggcctgggcg gccgccgcct gcgtgcggag
ccgggcgacg 121 gcgcgcagac ctgggcccgt ttctcgcggc ctcctgcccc
cgaggccgcg ggcctcttcc 181 agggcacctt ccccgacggc ttcctctggg
ccgtgggcag cgccgcctac cagaccgagg 241 gcggctggca gcagcacggc
aagggtgcgt ccatctggga tacgttcacc caccaccccc 301 tggcaccccc
gggagactcc cggaacgcca gtctgccgtt gggcgccccg tcgccgctgc 361
agcccgccac cggggacgta gccagcgaca gctacaacaa cgtcttccgc gacacggagg
421 cgctgcgcga gctcggggtc actcactacc gcttctccat ctcgtgggcg
cgagtgctcc 481 ccaatggcag cgcgggcgtc cccaaccgcg aggggctgcg
ctactaccgg cgcctgctgg 541 agcggctgcg ggagctgggc gtgcagcccg
tggtcaccct gtaccactgg gacctgcccc 601 agcgcctgca ggacgcctac
ggcggctggg ccaaccgcgc cctggccgac cacttcaggg 661 attacgcgga
gctctgcttc cgccacttcg gcggtcaggt caagtactgg atcaccatcg 721
acaaccccta cgtggtggcc tggcacggct acgccaccgg gcgcctggcc cccggcatcc
781 ggggcagccc gcggctcggg tacctggtgg cgcacaacct cctcctggct
catgccaaag 841 tctggcatct ctacaatact tctttccgtc ccactcaggg
aggtcaggtg tccattgccc 901 taagctctca ctggatcaat cctcgaagaa
tgaccgacca cagcatcaaa gaatgtcaaa 961 aatctctgga ctttgtacta
ggttggtttg ccaaacccgt atttattgat ggtgactatc 1021 ccgagagcat
gaagaataac ctttcatcta ttctgcctga ttttactgaa tctgagaaaa 1081
agttcatcaa aggaactgct gacttttttg ctctttgctt tggacccacc ttgagttttc
1141 aacttttgga ccctcacatg aagttccgcc aattggaatc tcccaacctg
aggcaactgc 1201 tttcctggat tgaccttgaa tttaaccatc ctcaaatatt
tattgtggaa aatggctggt 1261 ttgtctcagg gaccaccaag agagatgatg
ccaaatatat gtattacctc aaaaagttca 1321 tcatggaaac cttaaaagcc
atcaagctgg atggggtgga tgtcatcggg tataccgcat 1381 ggtccctcat
ggatggtttc gagtggcaca gaggttacag catcaggcgt ggactcttct 1441
atgttgactt tctaagccag gacaagatgt tgttgccaaa gtcttcagcc ttgttctacc
1501 aaaagctgat agagaaaaat ggcttccctc ctttacctga aaatcagccc
ctagaaggga 1561 catttccctg tgactttgct tggggagttg ttgacaacta
cattcaagta gataccactc 1621 tgtctcagtt taccgacctg aatgtttacc
tgtgggatgt ccaccacagt aaaaggctta 1681 ttaaagtgga tggggttgtg
accaagaaga ggaaatccta ctgtgttgac tttgctgcca 1741 tccagcccca
gatcgcttta ctccaggaaa tgcacgttac acattttcgc ttctccctgg 1801
actgggccct gattctccct ctgggtaacc agtcccaggt gaaccacacc atcctgcagt
1861 actatcgctg catggccagc gagcttgtcc gtgtcaacat caccccagtg
gtggccctgt 1921 ggcagcctat ggccccgaac caaggactgc cgcgcctcct
ggccaggcag ggcgcctggg 1981 agaaccccta cactgccctg gcctttgcag
agtatgcccg actgtgcttt caagagctcg 2041 gccatcacgt caagctttgg
ataacgatga atgagccgta tacaaggaat atgacataca 2101 gtgctggcca
caaccttctg aaggcccatg ccctggcttg gcatgtgtac aatgaaaagt 2161
ttaggcatgc tcagaatggg aaaatatcca tagccttgca ggctgattgg atagaacctg
2221 cctgcccttt ctcccaaaag gacaaagagg tggccgagag agttttggaa
tttgacattg 2281 gctggctggc tgagcccatt ttcggctctg gagattatcc
atgggtgatg agggactggc 2341 tgaaccaaag aaacaatttt cttcttcctt
atttcactga agatgaaaaa aagctaatcc 2401 agggtacctt tgactttttg
gctttaagcc attataccac catccttgta gactcagaaa 2461 aagaagatcc
aataaaatac aatgattacc tagaagtgca agaaatgacc gacatcacgt 2521
ggctcaactc ccccagtcag gtggcggtag tgccctgggg gttgcgcaaa gtgctgaact
2581 ggctgaagtt caagtacgga gacctcccca tgtacataat atccaacgga
atcgatgacg 2641 ggctgcatgc tgaggacgac cagctgaggg tgtattatat
gcagaattac ataaacgaag 2701 ctctcaaagc ccacatactg gatggtatca
atctttgcgg atactttgct tattcgttta 2761 acgaccgcac agctccgagg
tttggcctct atcgttatgc tgcagatcag tttgagccca 2821 aggcatccat
gaaacattac aggaaaatta ttgacagcaa tggtttcccg ggcccagaaa 2881
ctctggaaag attttgtcca gaagaattca ccgtgtgtac tgagtgcagt ttttttcaca
2941 cccgaaagtc tttactggct ttcatagctt ttctattttt tgcttctatt
atttctctct 3001 cccttatatt ttactactcg aagaaaggca gaagaagtta
caaatagttc tgaacatttt 3061 tctattcatt cattttgaaa taattatgca
gacacatcag ctgttaacca tttgcacctc 3121 taagtgttgt gaaactgtaa
atttcataca tttgacttct agaaaacatt tttgtggctt 3181 atgacagagg
ttttgaaatg ggcataggtg atcgtaaaat attgaataat gcgaatagtg 3241
cctgaatttg ttctcttttt gggtgattaa aaaactgaca ggcactataa tttctgtaac
3301 acactaacaa aagcatgaaa aataggaacc acaccaatgc aacatttgtg
cagaaatttg 3361 aatgacaaga ttaggaatat tttcttctgc acccacttct
aaatttaatg tttttctgga 3421 agtagtaatt gcaagagttc gaatagaaag
ttatgtacca agtaaccatt tctcagctgc 3481 cataataatg cctagtggct
tcccctctgt caaatctagt ttcctatgga aaagaagatg 3541 gcagatacag
gagagacgac agagggtcct aggctggaat gttcctttcg aaagcaatgc 3601
ttctatcaaa tactagtatt aatttatgta tctggttaat gacatacttg gagagcaaat
3661 tatggaaatg tgtattttat atgatttttg aggtcctgtc taaaccctgt
gtccctgagg 3721 gatctgtctc actggcatct tgttgagggc cttgcacata
ggaaactttt gataagtatc 3781 tgcggaaaaa caaacatgaa tcctgtgata
ttgggctctt caggaagcat aaagcaattg 3841 tgaaatacag tataccgcag
tggctctagg tggaggaaag gaggaaaaag tgcttattat 3901 gtgcaacatt
atgattaatc tgattataca ccatttttga gcagatcttg gaatgaatga 3961
catgaccttt ccctagagaa taaggatgaa ataatcactc attctatgaa cagtgacact
4021 actttctatt ctttagctgt actgtaattt ctttgagttg atagttttac
aaattcttaa 4081 taggttcaaa agcaatctgg tctgaataac actggatttg
tttctgtgat ctctgaggtc 4141 tattttatgt ttttgctgct acttctgtgg
aagtagcttt gaactagttt tactttgaac 4201 tttcacgctg aaacatgcta
gtgatatcta gaaagggcta attaggtctc atcctttaat 4261 gccccttaaa
taagtcttgc tgattttcag acagggaagt ctctctatta cactggagct 4321
gttttataga taagtcaata ttgtatcagg caagataaac caatgtcata acaggcattg
4381 ccaacctcac tgacacaggg tcatagtgta taataatata ctgtactata
taatatatca 4441 tctttagagg tatgattttt tcatgaaaga taagcttttg
gtaatattca ttttaaagtg 4501 gacttattaa aattggatgc tagagaatca
agtttatttt atgtatatat ttttctgatt 4561 ataagagtaa tatatgttca
ttgtaaaaat ttttaaaaca cagaaactat atgcaaagaa 4621 aaaataaaaa
ttatctataa tctcagaacc cagaaatagc cactattaac atttcctacg 4681
tattttattt tacatagatc atattgtata tagttagtat ctttattaat ttttattatg
4741 aaactttcct ttgtcattat tagtcttcaa aagcatgatt tttaatagtt
gttgagtatt 4801 ccaccacagg aatgtatcac aacttaaccg ttcccgtttg
ttagactagt ttcttattaa 4861 tgttgatgaa tgttgtttaa aaataatttt
gttgctacat ttactttaat ttccttgact 4921 gtaaagagaa gtaattttgc
tccttgataa agtattatat taataataaa tctgcctgca 4981 actttttgcc
ttctttcata atc Klotho amino acid sequence (NP_004786) (SEQ ID NO:
2) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA
PEAAGLFQGT 61 FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP
PGDSRNASLP LGAPSPLQPA 121 TGDVASDSYN NVFRDTEALR ELGVTHYRFS
ISWARVLPNG SAGVPNREGL RYYRRLLERL 181 RELGVQPVVT LYHWDLPQRL
QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP 241 YVVAWHGYAT
GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301
HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP DFTESEKKFI
361 KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW IDLEFNHPQI
FIVENGWFVS 421 GTTKRDDAKY MYYLKKFIME TLKAIKLDGV DVIGYTAWSL
MDGFEWHRGY SIRRGLFYVD 481 FLSQDKMLLP KSSALFYQKL IEKNGFPPLP
ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ 541 FTDLNVYLWD VHHSKRLIKV
DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ
VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL LARQGAWENP 661
YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG HNLLKAHALA WHVYNEKFRH
721 AQNGKISIAL QADWIEPACP FSQKDKEVAE RVLEFDIGWL AEPIFGSGDY
PWVMRDWLNQ 781 RNNFLLPYFT EDEKKLIQGT FDFLALSHYT TILVDSEKED
PIKYNDYLEV QEMTDITWLN 841 SPSQVAVVPW GLRKVLNWLK FKYGDLPMYI
ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR
TAPRFGLYRY AADQFEPKAS MKHYRKIIDS NGFPGPETLE 961 RFCPEEFTVC
TECSFFHTRK SLLAFIAFLF FASIISLSLI FYYSKKGRRS YK beta-Klotho nucleic
acid sequence (NM_175737) (SEQ ID NO: 3) Protein coding region:
98-3232 1 atcctcagtc tcccagttca agctaatcat tgacagagct ttacaatcac
aagcttttac 61 tgaagctttg ataagacagt ccagcagttg gtggcaaatg
aagccaggct gtgcggcagg 121 atctccaggg aatgaatgga ttttcttcag
cactgatgaa ataaccacac gctataggaa 181 tacaatgtcc aacgggggat
tgcaaagatc tgtcatcctg tcagcactta ttctgctacg 241 agctgttact
ggattctctg gagatggaag agctatatgg tctaaaaatc ctaattttac 301
tccggtaaat gaaagtcagc tgtttctcta tgacactttc cctaaaaact ttttctgggg
361 tattgggact ggagcattgc aagtggaagg gagttggaag aaggatggaa
aaggaccttc 421 tatatgggat catttcatcc acacacacct taaaaatgtc
agcagcacga atggttccag 481 tgacagttat atttttctgg aaaaagactt
atcagccctg gattttatag gagtttcttt 541 ttatcaattt tcaatttcct
ggccaaggct tttccccgat ggaatagtaa cagttgccaa 601 cgcaaaaggt
ctgcagtact acagtactct tctggacgct ctagtgctta gaaacattga 661
acctatagtt actttatacc actgggattt gcctttggca ctacaagaaa aatatggggg
721 gtggaaaaat gataccataa tagatatctt caatgactat gccacatact
gtttccagat 781 gtttggggac cgtgtcaaat attggattac aattcacaac
ccatatctag tggcttggca 841 tgggtatggg acaggtatgc atgcccctgg
agagaaggga aatttagcag ctgtctacac 901 tgtgggacac aacttgatca
aggctcactc gaaagtttgg cataactaca acacacattt 961 ccgcccacat
cagaagggtt ggttatcgat cacgttggga tctcattgga tcgagccaaa 1021
ccggtcggaa aacacgatgg atatattcaa atgtcaacaa tccatggttt ctgtgcttgg
1081 atggtttgcc aaccctatcc atggggatgg cgactatcca gaggggatga
gaaagaagtt 1141 gttctccgtt ctacccattt tctctgaagc agagaagcat
gagatgagag gcacagctga 1201 tttctttgcc ttttcttttg gacccaacaa
cttcaagccc ctaaacacca tggctaaaat 1261 gggacaaaat gtttcactta
atttaagaga agcgctgaac tggattaaac tggaatacaa 1321 caaccctcga
atcttgattg ctgagaatgg ctggttcaca gacagtcgtg tgaaaacaga 1381
agacaccacg gccatctaca tgatgaagaa tttcctcagc caggtgcttc aagcaataag
1441 gttagatgaa atacgagtgt ttggttatac tgcctggtct ctcctggatg
gctttgaatg 1501 gcaggatgct tacaccatcc gccgaggatt attttatgtg
gattttaaca gtaaacagaa 1561 agagcggaaa cctaagtctt cagcacacta
ctacaaacag atcatacgag aaaatggttt 1621 ttctttaaaa gagtccacgc
cagatgtgca gggccagttt ccctgtgact tctcctgggg 1681 tgtcactgaa
tctgttctta agcccgagtc tgtggcttcg tccccacagt tcagcgatcc 1741
tcatctgtac gtgtggaacg ccactggcaa cagactgttg caccgagtgg aaggggtgag
1801 gctgaaaaca cgacccgctc aatgcacaga ttttgtaaac atcaaaaaac
aacttgagat 1861 gttggcaaga atgaaagtca cccactaccg gtttgctctg
gattgggcct cggtccttcc 1921 cactggcaac ctgtccgcgg tgaaccgaca
ggccctgagg tactacaggt gcgtggtcag 1981 tgaggggctg aagcttggca
tctccgcgat ggtcaccctg tattatccga cccacgccca 2041 cctaggcctc
cccgagcctc tgttgcatgc cgacgggtgg ctgaacccat cgacggccga 2101
ggccttccag gcctacgctg ggctgtgctt ccaggagctg ggggacctgg tgaagctctg
2161 gatcaccatc aacgagccta accggctaag tgacatctac aaccgctctg
gcaacgacac 2221 ctacggggcg gcgcacaacc tgctggtggc ccacgccctg
gcctggcgcc tctacgaccg 2281 gcagttcagg ccctcacagc gcggggccgt
gtcgctgtcg ctgcacgcgg actgggcgga 2341 acccgccaac ccctatgctg
actcgcactg gagggcggcc gagcgcttcc tgcagttcga 2401 gatcgcctgg
ttcgccgagc cgctcttcaa gaccggggac taccccgcgg ccatgaggga 2461
atacattgcc tccaagcacc gacgggggct ttccagctcg gccctgccgc gcctcaccga
2521 ggccgaaagg aggctgctca agggcacggt cgacttctgc gcgctcaacc
acttcaccac 2581 taggttcgtg atgcacgagc agctggccgg cagccgctac
gactcggaca gggacatcca 2641 gtttctgcag gacatcaccc gcctgagctc
ccccacgcgc ctggctgtga ttccctgggg 2701 ggtgcgcaag ctgctgcggt
gggtccggag gaactacggc gacatggaca tttacatcac 2761 cgccagtggc
atcgacgacc aggctctgga ggatgaccgg ctccggaagt actacctagg 2821
gaagtacctt caggaggtgc tgaaagcata cctgattgat aaagtcagaa tcaaaggcta
2881 ttatgcattc aaactggctg aagagaaatc taaacccaga tttggattct
tcacatctga 2941 ttttaaagct aaatcctcaa tacaatttta caacaaagtg
atcagcagca ggggcttccc 3001 ttttgagaac agtagttcta gatgcagtca
gacccaagaa aatacagagt gcactgtctg 3061 cttattcctt gtgcagaaga
aaccactgat attcctgggt tgttgcttct tctccaccct 3121 ggttctactc
ttatcaattg ccatttttca aaggcagaag agaagaaagt tttggaaagc 3181
aaaaaactta caacacatac cattaaagaa aggcaagaga gttgttagct aaactgatct
3241 gtctgcatga tagacagttt aaaaattcat cccagttcc beta-Klotho amino
acid sequence (NP_783864) (SEQ ID NO: 4) 1 mkpgcaagsp gnewiffstd
eittryrntm sngglqrsvi lsalillrav tgfsgdgrai 61 wsknpnftpv
nesqlflydt fpknffwgig tgalqvegsw kkdgkgpsiw dhfihthlkn 121
vsstngssds yiflekdlsa ldfigvsfyq fsiswprlfp dgivtvanak glqyystlld
181 alvlrniepi vtlyhwdlpl alqekyggwk ndtiidifnd yatycfqmfg
drvkywitih 241 npylvawhgy gtgmhapgek gnlaavytvg hnlikahskv
whnynthfrp hqkgwlsitl 301 gshwiepnrs entmdifkcq qsmvsvlgwf
anpihgdgdy pegmrkklfs vlpifseaek 361 hemrgtadff afsfgpnnfk
pintmakmgq nvslnlreal nwikleynnp riliaengwf 421 tdsrvktedt
taiymmknfl sqvlqairld eirvfgytaw slldgfewqd aytirrglfy 481
vdfnskqker kpkssahyyk qiirengfsl kestpdvqgq fpcdfswgvt esvlkpesva
541 sspqfsdphl yvwnatgnrl lhrvegvrlk trpaqctdfv nikkqlemla
rmkvthyrfa 601 ldwasvlptg nlsavnrqal ryyrcvvseg lklgisamvt
lyypthahlg lpepllhadg 661 wlnpstaeaf qayaglcfqe lgdlvklwit
inepnrlsdi ynrsgndtyg aahnllvaha 721 lawrlydrqf rpsqrgaysl
slhadwaepa npyadshwra aerflqfeia wfaeplfktg 781 dypaamreyi
askhrrglss salprlteae rrllkgtvdf calnhfttrf vmheqlagsr 841
ydsdrdiqfl qditrlsspt rlavipwgvr kllrwvrrny gdmdiyitas giddqaledd
901 rlrkyylgky lqevlkayli dkvrikgyya fklaeekskp rfgfftsdfk
akssiqfynk 961 vissrgfpfe nsssrcsqtq entectvclf lvqkkplifl
gccffstivl llsialfqrq 1021 krrkfwkakn lqhiplkkgk rvvs Human Klotho
domain 1 (KL-D1) amino acid sequence (SEQ ID NO: 5) 58 qgt 61
fpdgflwavg saayqteggw qqhgkgasiw dtfthhplap pgdsrnaslp lgapsplqpa
121 tgdvasdsyn nvfrdtealr elgvthyrfs iswarvlpng sagvpnregl
ryyrrllerl 181 relgvqpvvt lyhwdlpqrl qdayggwanr aladhfrdya
elcfrhfggq vkywitidnp 241 yvvawhgyat grlapgirgs prlgylvahn
lllahakvwh lyntsfrptq ggqvsialss 301 hwinprrmtd hsikecqksl
dfvlgwfakp vfidgdypes mknnlssilp dftesekkfl 361 kgtadffalc
fgptlsfqll dphmkfrqle spnlrqllsw idlefnhpql fivengwfvs 421
gttkrddaky myylkkfime tlkalkldgv dvigytawsl mdgfewhrgy sirrglfyvd
481 flsqdkmllp kssalfyqkl lekngf Human Klotho domain 2 (KL-D2)
amino acid sequence (SEQ ID NO: 6) 517 gtfp cdfawgvvdn yiqvdttlsq
541 ftdlnvylwd vhhskrlikv dgvvtkkrks ycvdfaalqp qiallqemhv
thfrfsldwa 601 111plgnqsq vnhtilqyyr cmaselvrvn itpvvalwqp
mapnqglprl larqgawenp 661 ytalafaeya rlcfgelghh vklwitmnep
ytrnmtysag hnllkahala whvynekfrh 721 aqngkisial qadwiepacp
fsqkdkevae rvlefdigwl aeplfgsgdy pwvmrdwlnq 781 rnnfllpyft
edekkliqgt fdflalshyt tilvdseked pikyndylev qemtditwln 841
spsqvavvpw glrkvinwlk fkygdlpmyi isngiddglh aeddqlrvyy mqnyinealk
901 ahildginlc gyfaysfndr taprfglyry aadqfepkas mkhyrkilds ngf
Klotho extracellular domain (without signal peptide) amino acid
sequence (SEQ ID NO: 7) 28 epgdgaq twarfsrppa peaaglfqgt 61
fpdgflwavg saayqteggw qqhgkgasiw dtfthhplap pgdsrnaslp lgapsplqpa
121 tgdvasdsyn nvfrdtealr elgvthyrfs iswarvlpng sagvpnregl
ryyrrllerl 181 relgvqpvvt lyhwdlpqrl qdayggwanr aladhfrdya
elcfrhfggq vkywitidnp 241 yvvawhgyat grlapgirgs prlgylvahn
lllahakvwh lyntsfrptq ggqvsialss 301 hwinprrmtd hsikecqksl
dfvlgwfakp vfidgdypes mknnlssilp dftesekkfl 361 kgtadffalc
fgptlsfqll dphmkfrqle spnlrqllsw idlefnhpql fivengwfvs 421
gttkrddaky myylkkfime tlkalkldgv dvigytawsl mdgfewhrgy sirrglfyvd
481 flsqdkmllp kssalfyqkl lekngfpplp enqplegtfp cdfawgvvdn
yiqvdttlsq 541 ftdlnvylwd vhhskrlikv dgvvtkkrks ycvdfaalqp
qiallqemhv thfrfsldwa 601 lllplgnqsq vnhtilqyyr cmaselvrvn
itpvvalwqp mapnqglprl larqgawenp 661 ytalafaeya rlcfgelghh
vklwitmnep ytrnmtysag hnllkahala whvynekfrh 721 aqngkisial
qadwiepacp fsqkdkevae rvlefdigwl aeplfgsgdy pwvmrdwlnq 781
rnnfllpyft edekkliqgt fdflalshyt tilvdseked pikyndylev qemtditwln
841 spsqvavvpw glrkvinwlk fkygdlpmyi isngiddglh aeddqlrvyy
mqnyinealk 901 ahildginlc gyfaysfndr taprfglyry aadqfepkas
mkhyrkilds ngfpgpetle 961 rfcpeeftvc tecsffhtrk sl Klotho signal
peptide amino acid sequence (SEQ ID NO: 8) 1 mpasapprrp rppppslsll
lvllglggrr lra IgG signal peptide amino acid sequence (SEQ ID NO:
9) 1 msvltqvlal lllwltgtrc rrlra (Gly.sub.4 Ser).sub.3 polypeptide
linker nucleic acid sequence (SEQ ID NO: 10) 1 ggaggtggag
gttcaggagg tggaggttca ggaggtggag gttca (Gly.sub.4 Ser).sub.3
polypeptide linker amino acid sequence (SEQ ID NO: 11) 1 GGGGSGGGGS
GGGGS (Gly.sub.4 Ser) polypeptide linker amino acid sequence (SEQ
ID NO: 12) 1 GGGGS (Gly) polypeptide linker amino acid sequence
(SEQ ID NO: 13) 1 G (Gly Gly) polypeptide linker amino acid
sequence (SEQ ID NO: 14)
1 GG (Gly Ser) polypeptide linker amino acid sequence (SEQ ID NO:
15) 1 GS (Gly.sub.2 Ser) polypeptide linker amino acid sequence
(SEQ ID NO: 16) 1 GGS (Ala) polypeptide linker amino acid sequence
(SEQ ID NO: 17) 1 A (Ala Ala) polypeptide linker amino acid
sequence (SEQ ID NO: 18) 1 AA Klotho signal peptide-Klotho
extracellular domain-FGF23 (R179Q) amino acid sequence (SEQ ID NO:
19) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51
PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101
PGDSRNASLP LGAPSPLQPA TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151
ISWARVLPNG SAGVPNREGL RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201
QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251
GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301
HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351
DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401
IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451
DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501
IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551
VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601
LILPLGNQSQ VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL 651
LARQGAWENP YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701
HNLLKAHALA WHVYNEKFRH AQNGKISIAL QADWIEPACP FSQKDKEVAE 751
RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801
FDFLALSHYT TILVDSEKED PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851
GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901
AHILDGINLC GYFAYSFNDR TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951
NGFPGPETLE RFCPEEFTVC TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001
KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051
MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101
YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151
PRRHTQSAED DSERDPLNVL KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201
LGVVRGGRVN THAGGTGPEG CRPFAKFI* IgG signal peptide-Klotho
extracellular domain-FGF23 (R179Q) amino acid sequence (SEQ ID NO:
20) 1 MSVLTQVLAL LLLWLTGLGG RRLRAEPGDG AQTWARFSRP PAPEAAGLFQ 51
GTFPDGFLWA VGSAAYQTEG GWQQHGKGAS IWDTFTHHPL APPGDSRNAS 101
LPLGAPSPLQ PATGDVASDS YNNVFRDTEA LRELGVTHYR FSISWARVLP 151
NGSAGVPNRE GLRYYRRLLE RLRELGVQPV VTLYHWDLPQ RLQDAYGGWA 201
NRALADHFRD YAELCFRHFG GQVKYWITID NPYVVAWHGY ATGRLAPGIR 251
GSPRLGYLVA HNLLLAHAKV WHLYNTSFRP TQGGQVSIAL SSHWINPRPM 301
TDHSIKECQK SLDFVLGWFA KPVFIDGDYP ESMKNNLSSI LPDFTESEKK 351
FIKGTADFFA LCFGPTLSFQ LLDPHMKFRQ LESPNLRQLL SWIDLEFNHP 401
QIFIVENGWF VSGTTKRDDA KYMYYLKKFI METLKAIKLD GVDVIGYTAW 451
SLMDGFEWHR GYSIRRGLFY VDFLSQDKML LPKSSALFYQ KLIEKNGFPP 501
LPENQPLEGT FPCDFAWGVV DNYIQVDTTL SQFTDLNVYL WDVHHSKRLI 551
KVDGVVTKKR KSYCVDFAAI QPQIALLQEM HVTHFRFSLD WALILPLGNQ 601
SQVNHTILQY YRCMASELVR VNITPVVALW QPMAPNQGLP RLLARQGAWE 651
NPYTALAFAE YARLCFQELG HHVKLWITMN EPYTRNMTYS AGHNLLKAHA 701
LAWHVYNEKF RHAQNGKISI ALQADWIEPA CPFSQKDKEV AERVLEFDIG 751
WLAEPIFGSG DYPWVMRDWL NQRNNFLLPY FTEDEKKLIQ GTFDFLALSH 801
YTTILVDSEK EDPIKYNDYL EVQEMTDITW LNSPSQVAVV PWGLRKVLNW 851
LKFKYGDLPM YIISNGIDDG LHAEDDQLRV YYMQNYINEA LKAHILDGIN 901
LCGYFAYSFN DRTAPRFGLY RYAADQFEPK ASMKHYRKII DSNGFPGPET 951
LERFCPEEFT VCTECSFFHT RKSLGSGGGG SGGGGSGGGG SLKYPNASPL 1001
LGSSWGGLIH LYTATARNSY HLQIHKNGHV DGAPHQTIYS ALMIRSEDAG 1051
FVVITGVMSR RYLCMDFRGN IFGSHYFDPE NCRFQHQTLE NGYDVYHSPQ 1101
YHFLVSLGRA KRAFLPGMNP PPYSQFLSRR NEIPLIHFNT PIPRRHTQSA 1151
EDDSERDPLN VLKPRARMTP APASCSQELP SAEDNSPMAS DPLGVVRGGR 1201
VNTHAGGTGP EGCRPFAKFI * KL-D1-FGF23 (R179Q) amino acid sequence
(SEQ ID NO: 21) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ
TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW QQHGKGASIW
DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA TGDVASDSYN NVFRDTEALR
ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL RYYRRLLERL RELGVQPVVT
LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP
YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ
GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES
MKNNLSSILP 351 DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE
SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME
TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP
KSSALFYQKL 501 IEKNGFPPLP ENQPLEGSGG GGSGGGGSGG GGSLKYPNAS
PLLGSSWGGL 551 IHLYTATARN SYHLQIHKNG HVDGAPHQTI YSALMIRSED
AGFVVITGVM 601 SRRYLCMDFR GNIFGSHYFD PENCRFQHQT LENGYDVYHS
PQYHFLVSLG 651 RAKRAFLPGM NPPPYSQFLS RRNEIPLIHF NTPIPRRHTQ
SAEDDSERDP 701 LNVLKPRAPM TPAPASCSQE LPSAEDNSPM ASDPLGVVRG
GRVNTHAGGT 751 GPEGCRPFAK FI* KL-D2-FGF23 (R179Q) amino acid
sequence (SEQ ID NO: 22) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR
LPLPENQPLE GTFPCDFAWG 51 VVDNYIQVDT TLSQFTDLNV YLWDVHHSKR
LIKVDGVVTK KRKSYCVDFA 101 AIQPQIALLQ EMHVTHFRFS LDWALILPLG
NQSQVNHTIL QYYRCMASEL 151 VRVNITPVVA LWQPMAPNQG LPRLLARQGA
WENPYTALAF AEYARLCFQE 201 LGHHVKLWIT MNEPYTRNMT YSAGHNLLKA
HALAWHVYNE KFRHAQNGKI 251 SIALQADWIE PACPFSQKDK EVAERVLEFD
IGWLAEPIFG SGDYPWVMRD 301 WLNQRNNFLL PYFTEDEKKL IQGTFDFLAL
SHYTTILVDS EKEDPIKYND 351 YLEVQEMTDI TWLNSPSQVA VVPWGLRKVL
NWLKFKYGDL PMYIISNGID 401 DGLHAEDDQL RVYYMQNYIN EALKAHILDG
INLCGYFAYS FNDRTAPRFG 451 LYRYAADQFE PKASMKHYRK IIDSNGFPGP
ETLERFCPEE FTVCTECSFF 501 HTRKSLGSGG GGSGGGGSGG GGSLKYPNAS
PLLGSSWGGL IHLYTATARN 551 SYHLQIHKNG HVDGAPHQTI YSALMIRSED
AGFVVITGVM SRRYLCMDFR 601 GNIFGSHYFD PENCRFQHQT LENGYDVYHS
PQYHFLVSLG RAKRAFLPGM 651 NPPPYSQFLS RRNEIPLIHF NTPIPRRHTQ
SAEDDSERDP LNVLKPRARM 701 TPAPASCSQE LPSAEDNSPM ASDPLGVVRG
GRVNTHAGGT GPEGCRPFAK 751 FI* (KL-D1)2-FGF23 (R179Q) amino acid
sequence (SEQ ID NO: 23) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR
LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW
QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA TGDVASDSYN
NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL RYYRRLLERL
RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ
VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH
LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL DFVLGWFAKP
VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC FGPTLSFQLL
DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS GTTKRDDAKY
MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD
FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGSGT FPDGFLWAVG
SAAYQTEGGW QQHGKGASIW 551 DTFTHHPLAP PGDSRNASLP LGAPSPLQPA
TGDVASDSYN NVFRDTEALR 601 ELGVTHYRFS ISWARVLPNG SAGVPNREGL
RYYRRLLERL RELGVQPVVT 651 LYHWDLPQRL QDAYGGWANR ALADHFRDYA
ELCFRHFGGQ VKYWITIDNP 701 YVVAWHGYAT GRLAPGIRGS PRLGYLVAHN
LLLAHAKVWH LYNTSFRPTQ 751 GGQVSIALSS HWINPRRMTD HSIKECQKSL
DFVLGWFAKP VFIDGDYPES 801 MKNNLSSILP DFTESEKKFI KGTADFFALC
FGPTLSFQLL DPHMKFRQLE 851 SPNLRQLLSW IDLEFNHPQI FIVENGWFVS
GTTKRDDAKY MYYLKKFIME 901 TLKAIKLDGV DVIGYTAWSL MDGFEWHRGY
SIRRGLFYVD FLSQDKMLLP 951 KSSALFYQKL IEKNGFPEFG SGGGGSGGGG
SGGGGSLKYP NASPLLGSSW 1001 GGLIHLYTAT ARNSYHLQIH KNGHVDGAPH
QTIYSALMIR SEDAGFVVIT 1051 GVMSRRYLCM DFRGNIFGSH YFDPENCRFQ
HQTLENGYDV YHSPQYHFLV 1101 SLGRAKRAFL PGMNPPPYSQ FLSRRNEIPL
IHFNTPIPRR HTQSAEDDSE 1151 RDPLNVLKPR ARMTPAPASC SQELPSAEDN
SPMASDPLGV VRGGRVNTHA 1201 GGTGPEGCRP FAKFI* (KL-D2).sub.2-FGF23
(R179Q) amino acid sequence (SEQ ID NO: 24) 1 MPASAPPRRP RPPPPSLSLL
LVLLGLGGRR LPLPENQPLE GTFPCDFAWG 51 VVDNYIQVDT TLSQFTDLNV
YLWDVHHSKR LIKVDGVVTK KRKSYCVDFA 101 AIQPQIALLQ EMHVTHFRFS
LDWALILPLG NQSQVNHTIL QYYRCMASEL 151 VRVNITPVVA LWQPMAPNQG
LPRLLARQGA WENPYTALAF AEYARLCFQE 201 LGHHVKLWIT MNEPYTRNMT
YSAGHNLLKA HALAWHVYNE KFRHAQNGKI 251 SIALQADWIE PACPFSQKDK
EVAERVLEFD IGWLAEPIFG SGDYPWVMRD 301 WLNQRNNFLL PYFTEDEKKL
IQGTFDFLAL SHYTTILVDS EKEDPIKYND 351 YLEVQEMTDI TWLNSPSQVA
VVPWGLRKVL NWLKFKYGDL PMYIISNGID 401 DGLHAEDDQL RVYYMQNYIN
EALKAHILDG INLCGYFAYS FNDRTAPRFG 451 LYRYAADQFE PKASMKHYRK
IIDSNGFPGP ETLERFCPEE FTVCTECSFF 501 HTRKSLGTFP CDFAWGVVDN
YIQVDTTLSQ FTDLNVYLWD VHHSKRLIKV 551 DGVVTKKRKS YCVDFAAIQP
QIALLQEMHV THFRFSLDWA LILPLGNQSQ 601 VNHTILQYYR CMASELVRVN
ITPVVALWQP MAPNQGLPRL LARQGAWENP 651 YTALAFAEYA RLCFQELGHH
VKLWITMNEP YTRNMTYSAG HNLLKAHALA 701 WHVYNEKFRH AQNGKISIAL
QADWIEPACP FSQKDKEVAE RVLEFDIGWL 751 AEPIFGSGDY PWVMRDWLNQ
RNNFLLPYFT EDEKKLIQGT FDFLALSHYT 801 TILVDSEKED PIKYNDYLEV
QEMTDITWLN SPSQVAVVPW GLRKVLNWLK 851 FKYGDLPMYI ISNGIDDGLH
AEDDQLRVYY MQNYINEALK AHILDGINLC 901 GYFAYSFNDR TAPRFGLYRY
AADQFEPKAS MKHYRKIIDS NGFGSGGGGS 951 GGGGSGGGGS LKYPNASPLL
GSSWGGLIHL YTATARNSYH LQIHKNGHVD 1001 GAPHQTIYSA LMIRSEDAGF
VVITGVMSRR YLCMDFRGNI FGSHYFDPEN 1051 CRFQHQTLEN GYDVYHSPQY
HFLVSLGRAK RAFLPGMNPP PYSQFLSRRN 1101 EIPLIHFNTP IPRRHTQSAE
DDSERDPLNV LKPRARMTPA PASCSQELPS 1151 AEDNSPMASD PLGVVRGGRV
NTHAGGTGPE GCRPFAKFI* FGF23 (R179Q)-Klotho extracellular domain
amino acid sequence (SEQ ID NO: 25) 1 MLGARLRLWV CALCSVCSMS
VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY
SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL
ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN
TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IGSGGGGSGG GGSGGGGSLK
EPGDGAQTWA RFSRPPAPEA AGLFQGTFPD 301 GFLWAVGSAA YQTEGGWQQH
GKGASIWDTF THHPLAPPGD SRNASLPLGA 351 PSPLQPATGD VASDSYNNVF
RDTEALRELG VTHYRFSISW ARVLPNGSAG 401 VPNREGLRYY RRLLERLREL
GVQPVVTLYH WDLPQRLQDA YGGWANRALA 451 DHFRDYAELC FRHFGGQVKY
WITIDNPYVV AWHGYATGRL APGIRGSPRL 501 GYLVAHNLLL AHAKVWHLYN
TSFRPTQGGQ VSIALSSHWI NPRRMTDHSI 551 KECQKSLDFV LGWFAKPVFI
DGDYPESMKN NLSSILPDFT ESEKKFIKGT 601 ADFFALCFGP TLSFQLLDPH
MKFRQLESPN LRQLLSWIDL EFNHPQIFIV 651 ENGWFVSGTT KRDDAKYMYY
LKKFIMETLK AIKLDGVDVI GYTAWSLMDG 701 FEWHRGYSIR RGLFYVDFLS
QDKMLLPKSS ALFYQKLIEK NGFPPLPENQ 751 PLEGTFPCDF AWGVVDNYIQ
VDTTLSQFTD LNVYLWDVHH SKRLIKVDGV 801 VTKKRKSYCV DFAAIQPQIA
LLQEMHVTHF RFSLDWALIL PLGNQSQVNH 851 TILQYYRCMA SELVRVNITP
VVALWQPMAP NQGLPRLLAR QGAWENPYTA 901 LAFAEYARLC FQELGHHVKL
WITMNEPYTR NMTYSAGHNL LKAHALAWHV 951 YNEKFRHAQN GKISIALQAD
WIEPACPFSQ KDKEVAERVL EFDIGWLAEP 1001 IFGSGDYPWV MRDWLNQRNN
FLLPYFTEDE KKLIQGTFDF LALSHYTTIL 1051 VDSEKEDPIK YNDYLEVQEM
TDITWLNSPS QVAVVPWGLR KVLNWLKFKY 1101 GDLPMYIISN GIDDGLHAED
DQLRVYYMQN YINEALKAHI LDGINLCGYF 1151 AYSFNDRTAP RFGLYRYAAD
QFEPKASMKH YRKIIDSNGF PGPETLERFC 1201 PEEFTVCTEC SFFHTRKSL* FGF23
(R179Q)-KL-D1 amino acid sequence (SEQ ID NO: 26) 1 MLGARLRLWV
CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP
ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR
RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL
PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IQGTFPDGFL
WAVGSAAYQT EGGWQQHGKG ASIWDTFTHH PLAPPGDSRN 301 ASLPLGAPSP
LQPATGDVAS DSYNNVFRDT EALRELGVTH YRFSISWARV 351 LPNGSAGVPN
REGLRYYRRL LERLRELGVQ PVVTLYHWDL PQRLQDAYGG 401 WANRALADHF
RDYAELCFRH FGGQVKYWIT IDNPYVVAWH GYATGRLAPG 451 IRGSPRLGYL
VAHNLLLAHA KVWHLYNTSF RPTQGGQVSI ALSSHWINPR 501 RMTDHSIKEC
QKSLDFVLGW FAKPVFIDGD YPESMKNNLS SILPDFTESE 551 KKFIKGTADF
FALCFGPTLS FQLLDPHMKF RQLESPNLRQ LLSWIDLEFN 601 HPQIFIVENG
WFVSGTTKRD DAKYMYYLKK FIMETLKAIK LDGVDVIGYT 651 AWSLMDGFEW
HRGYSIRRGL FYVDFLSQDK MLLPKSSALF YQKLIEKNGF 652 * FGF23
(R179Q)-KL-D2 amino acid sequence (SEQ ID NO: 27) 1 MLGARLRLWV
CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP
ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR
RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL
PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IGTFPCDFAW
GVVDNYIQVD TTLSQFTDLN VYLWDVHHSK RLIKVDGVVT 301 KKRKSYCVDF
AAIQPQIALL QEMHVTHFRF SLDWALILPL GNQSQVNHTI 351 LQYYRCMASE
LVRVNITPVV ALWQPMAPNQ GLPRLLARQG AWENPYTALA 401 FAEYARLCFQ
ELGHHVKLWI TMNEPYTRNM TYSAGHNLLK AHALAWHVYN 451 EKFRHAQNGK
ISIALQADWI EPACPFSQKD KEVAERVLEF DIGWLAEPIF 501 GSGDYPWVMR
DWLNQRNNFL LPYFTEDEKK LIQGTFDFLA LSHYTTILVD 551 SEKEDPIKYN
DYLEVQEMTD ITWLNSPSQV AVVPWGLRKV LNWLKFKYGD 601 LPMYIISNGI
DDGLHAEDDQ LRVYYMQNYI NEALKAHILD GINLCGYFAY 651 SFNDRTAPRF
GLYRYAADQF EPKASMKHYR KIIDSNGF* FGF23 (R179Q)-(KL-D1).sub.2 amino
acid sequence (SEQ ID NO: 28) 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS
AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG PEGCRPFAKF 251 IQGTFPDGFL WAVGSAAYQT EGGWQQHGKG
ASIWDTFTHH PLAPPGDSRN 301 ASLPLGAPSP LQPATGDVAS DSYNNVFRDT
EALRELGVTH YRFSISWARV 351 LPNGSAGVPN REGLRYYRRL LERLRELGVQ
PVVTLYHWDL PQRLQDAYGG 401 WANRALADHF RDYAELCFRH FGGQVKYWIT
IDNPYVVAWH GYATGRLAPG 451 IRGSPRLGYL VAHNLLLAHA KVWHLYNTSF
RPTQGGQVSI ALSSHWINPR 501 RMTDHSIKEC QKSLDFVLGW FAKPVFIDGD
YPESMKNNLS SILPDFTESE 551 KKFIKGTADF FALCFGPTLS FQLLDPHMKF
RQLESPNLRQ LLSWIDLEFN 601 HPQIFIVENG WFVSGTTKRD DAKYMYYLKK
FIMETLKAIK LDGVDVIGYT 651 AWSLMDGFEW HRGYSIRRGL FYVDFLSQDK
MLLPKSSALF YQKLIEKNGF 701 QGTFPDGFLW AVGSAAYQTE GGWQQHGKGA
SIWDTFTHHP LAPPGDSRNA 751 SLPLGAPSPL QPATGDVASD SYNNVFRDTE
ALRELGVTHY RFSISWARVL 801 PNGSAGVPNR EGLRYYRRLL ERLRELGVQP
VVTLYHWDLP QRLQDAYGGW 851 ANRALADHFR DYAELCFRHF GGQVKYWITI
DNPYVVAWHG YATGRLAPGI 901 RGSPRLGYLV AHNLLLAHAK VWHLYNTSFR
PTQGGQVSIA LSSHWINPRR 951 MTDHSIKECQ KSLDFVLGWF AKPVFIDGDY
PESMKNNLSS ILPDFTESEK 1001 KFIKGTADFF ALCFGPTLSF QLLDPHMKFR
QLESPNLRQL LSWIDLEFNH 1051 PQIFIVENGW FVSGTTKRDD AKYMYYLKKF
IMETLKAIKL DGVDVIGYTA 1101 WSLMDGFEWH RGYSIRRGLF YVDFLSQDKM
LLPKSSALFY QKLIEKNGF* FGF23 (R179Q)-(KL-D2).sub.2 amino acid
sequence (SEQ ID NO: 29) 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS
AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG PEGCRPFAKF
251 IGTFPCDFAW GVVDNYIQVD TTLSQFTDLN VYLWDVHHSK RLIKVDGVVT 301
KKRKSYCVDF AAIQPQIALL QEMHVTHFRF SLDWALILPL GNQSQVNHTI 351
LQYYRCMASE LVRVNITPVV ALWQPMAPNQ GLPRLLARQG AWENPYTALA 401
FAEYARLCFQ ELGHHVKLWI TMNEPYTRNM TYSAGHNLLK AHALAWHVYN 451
EKFRHAQNGK ISIALQADWI EPACPFSQKD KEVAERVLEF DIGWLAEPIF 501
GSGDYPWVMR DWLNQRNNFL LPYFTEDEKK LIQGTFDFLA LSHYTTILVD 551
SEKEDPIKYN DYLEVQEMTD ITWLNSPSQV AVVPWGLRKV LNWLKFKYGD 601
LPMYIISNGI DDGLHAEDDQ LRVYYMQNYI NEALKAHILD GINLCGYFAY 651
SFNDRTAPRF GLYRYAADQF EPKASMKHYR KIIDSNGFGT FPCDFAWGVV 701
DNYIQVDTTL SQFTDLNVYL WDVHHSKRLI KVDGVVTKKR KSYCVDFAAI 751
QPQIALLQEM HVTHFRFSLD WALILPLGNQ SQVNHTILQY YRCMASELVR 801
VNITPVVALW QPMAPNQGLP RLLARQGAWE NPYTALAFAE YARLCFQELG 851
HHVKLWITMN EPYTRNMTYS AGHNLLKAHA LAWHVYNEKF RHAQNGKISI 901
ALQADWIEPA CPFSQKDKEV AERVLEFDIG WLAEPIFGSG DYPWVMRDWL 951
NQRNNFLLPY FTEDEKKLIQ GTFDFLALSH YTTILVDSEK EDPIKYNDYL 1001
EVQEMTDITW LNSPSQVAVV PWGLRKVLNW LKFKYGDLPM YIISNGIDDG 1051
LHAEDDQLRV YYMQNYINEA LKAHILDGIN LCGYFAYSFN DRTAPRFGLY 1101
RYAADQFEPK ASMKHYRKII DSNGF* FGF19 nucleic acid sequence
(NM_005117) (SEQ ID NO: 30) Protein coding region (464-1114) 1
gctcccagcc aagaacctcg gggccgctgc gcggtgggga ggagttcccc gaaacccggc
61 cgctaagcga ggcctcctcc tcccgcagat ccgaacggcc tgggcggggt
caccccggct 121 gggacaagaa gccgccgcct gcctgcccgg gcccggggag
ggggctgggg ctggggccgg 181 aggcggggtg tgagtgggtg tgtgcggggg
gcggaggctt gatgcaatcc cgataagaaa 241 tgctcgggtg tcttgggcac
ctacccgtgg ggcccgtaag gcgctactat ataaggctgc 301 cggcccggag
ccgccgcgcc gtcagagcag gagcgctgcg tccaggatct agggccacga 361
ccatcccaac ccggcactca cagccccgca gcgcatcccg gtcgccgccc agcctcccgc
421 acccccatcg ccggagctgc gccgagagcc ccagggaggt gccatgcgga
gcgggtgtgt 481 ggtggtccac gtatggatcc tggccggcct ctggctggcc
gtggccgggc gccccctcgc 541 cttctcggac gcggggcccc acgtgcacta
cggctggggc gaccccatcc gcctgcggca 601 cctgtacacc tccggccccc
acgggctctc cagctgcttc ctgcgcatcc gtgccgacgg 661 cgtcgtggac
tgcgcgcggg gccagagcgc gcacagtttg ctggagatca aggcagtcgc 721
tctgcggacc gtggccatca agggcgtgca cagcgtgcgg tacctctgca tgggcgccga
781 cggcaagatg caggggctgc ttcagtactc ggaggaagac tgtgctttcg
aggaggagat 841 ccgcccagat ggctacaatg tgtaccgatc cgagaagcac
cgcctcccgg tctccctgag 901 cagtgccaaa cagcggcagc tgtacaagaa
cagaggcttt cttccactct ctcatttcct 961 gcccatgctg cccatggtcc
cagaggagcc tgaggacctc aggggccact tggaatctga 1021 catgttctct
tcgcccctgg agaccgacag catggaccca tttgggcttg tcaccggact 1081
ggaggccgtg aggagtccca gctttgagaa gtaactgaga ccatgcccgg gcctcttcac
1141 tgctgccagg ggctgtggta cctgcagcgt gggggacgtg cttctacaag
aacagtcctg 1201 agtccacgtt ctgtttagct ttaggaagaa acatctagaa
gttgtacata ttcagagttt 1261 tccattggca gtgccagttt ctagccaata
gacttgtctg atcataacat tgtaagcctg 1321 tagcttgccc agctgctgcc
tgggccccca ttctgctccc tcgaggttgc tggacaagct 1381 gctgcactgt
ctcagttctg cttgaatacc tccatcgatg gggaactcac ttcctttgga 1441
aaaattctta tgtcaagctg aaattctcta attttttctc atcacttccc caggagcagc
1501 cagaagacag gcagtagttt taatttcagg aacaggtgat ccactctgta
aaacagcagg 1561 taaatttcac tcaaccccat gtgggaattg atctatatct
ctacttccag ggaccatttg 1621 cccttcccaa atccctccag gccagaactg
actggagcag gcatggccca ccaggcttca 1681 ggagtagggg aagcctggag
ccccactcca gccctgggac aacttgagaa ttccccctga 1741 ggccagttct
gtcatggatg ctgtcctgag aataacttgc tgtcccggtg tcacctgctt 1801
ccatctccca gcccaccagc cctctgccca cctcacatgc ctccccatgg attggggcct
1861 cccaggcccc ccaccttatg tcaacctgca cttcttgttc aaaaatcagg
aaaagaaaag 1921 atttgaagac cccaagtctt gtcaataact tgctgtgtgg
aagcagcggg ggaagaccta 1981 gaaccctttc cccagcactt ggttttccaa
catgatattt atgagtaatt tattttgata 2041 tgtacatctc ttattttctt
acattattta tgcccccaaa ttatatttat gtatgtaagt 2101 gaggtttgtt
ttgtatatta aaatggagtt tgtttgtaaa aaaaaaaaaa aaaaaaa FGF19 amino
acid sequence (NP_005108) (SEQ ID NO: 31) 1 MRSGCVVVHV WILAGLWLAV
AGRPLAFSDA GPHVHYGWGD PIRLRHLYTS GPHGLSSCFL 61 RIRADGVVDC
ARGQSAHSLL EIKAVALRTV AIKGVHSVRY LCMGADGKMQ GLLQYSEEDC 121
AFEEEIRPDG YNVYRSEKHR LPVSLSSAKQ RQLYKNRGFL PLSHFLPMLP MVPEEPEDLR
181 GHLESDMFSS PLETDSMDPF GLVTGLEAVR SPSFEK FGF21 nucleic acid
sequence (NM_019113) (SEQ ID NO: 32) Protein coding region 151-780
1 CTGTCAGCTG AGGATCCAGC CGAAAGAGGA GCCAGGCACT CAGGCCACCT GAGTCTACTC
61 ACCTGGACAA CTGGAATCTG GCACCAATTC TAAACCACTC AGCTTCTCCG
AGCTCACACC 121 CCGGAGATCA CCTGAGGACC CGAGCCATTG ATGGACTCGG
ACGAGACCGG GTTCGAGCAC 181 TCAGGACTGT GGGTTTCTGT GCTGGCTGGT
CTTCTGCTGG GAGCCTGCCA GGCACACCCC 241 ATCCCTGACT CCAGTCCTCT
CCTGCAATTC GGGGGCCAAG TCCGGCAGCG GTACCTCTAC 301 ACAGATGATG
CCCAGCAGAC AGAAGCCCAC CTGGAGATCA GGGAGGATGG GACGGTGGGG 361
GGCGCTGCTG ACCAGAGCCC CGAAAGTCTC CTGCAGCTGA AAGCCTTGAA GCCGGGAGTT
421 ATTCAAATCT TGGGAGTCAA GACATCCAGG TTCCTGTGCC AGCGGCCAGA
TGGGGCCCTG 481 TATGGATCGC TCCACTTTGA CCCTGAGGCC TGCAGCTTCC
GGGAGCTGCT TCTTGAGGAC 541 GGATACAATG TTTACCAGTC CGAAGCCCAC
GGCCTCCCGC TGCACCTGCC AGGGAACAAG 601 TCCCCACACC GGGACCCTGC
ACCCCGAGGA CCAGCTCGCT TCCTGCCACT ACCAGGCCTG 661 CCCCCCGCAC
TCCCGGAGCC ACCCGGAATC CTGGCCCCCC AGCCCCCCGA TGTGGGCTCC 721
TCGGACCCTC TGAGCATGGT GGGACCTTCC CAGGGCCGAA GCCCCAGCTA CGCTTCCTGA
781 AGCCAGAGGC TGTTTACTAT GACATCTCCT CTTTATTTAT TAGGTTATTT
ATCTTATTTA 841 TTTTTTTATT TTTCTTACTT GAGATAATAA AGAGTTCCAG
AGGAGAAAAA AAAAAAAAAA 901 AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA
AAAAAAAAAA FGF21 amino acid sequence (NP_061986) (SEQ ID NO: 33) 1
MDSDETGFEH SGLWVSVLAG LLLGACQAHP IPDSSPLLQF GGQVRQRYLY TDDAQQTEAH
61 LEIREDGTVG GAADQSPESL LQLKALKPGV IQILGVKTSR FLCQRPDGAL
YGSLHFDPEA 121 CSFRELLLED GYNVYQSEAH GLPLHLPGNK SPHRDPAPRG
PARFLPLPGL PPALPEPPGI 181 LAPQPPDVGS SDPLSMVGPS QGRSPSYAS FGF23
nucleic acid sequence (NM_020638) (SEQ ID NO: 34) Protein coding
region 147-902 1 cggcaaaaag gagggaatcc agtctaggat cctcacacca
gctacttgca agggagaagg 61 aaaaggccag taaggcctgg gccaggagag
tcccgacagg agtgtcaggt ttcaatctca 121 gcaccagcca ctcagagcag
ggcacgatgt tgggggcccg cctcaggctc tgggtctgtg 181 ccttgtgcag
cgtctgcagc atgagcgtcc tcagagccta tcccaatgcc tccccactgc 241
tcggctccag ctggggtggc ctgatccacc tgtacacagc cacagccagg aacagctacc
301 acctgcagat ccacaagaat ggccatgtgg atggcgcacc ccatcagacc
atctacagtg 361 ccctgatgat cagatcagag gatgctggct ttgtggtgat
tacaggtgtg atgagcagaa 421 gatacctctg catggatttc agaggcaaca
tttttggatc acactatttc gacccggaga 481 actgcaggtt ccaacaccag
acgctggaaa acgggtacga cgtctaccac tctcctcagt 541 atcacttcct
ggtcagtctg ggccgggcga agagagcctt cctgccaggc atgaacccac 601
ccccgtactc ccagttcctg tcccggagga acgagatccc cctaattcac ttcaacaccc
661 ccataccacg gcggcacacc cggagcgccg aggacgactc ggagcgggac
cccctgaacg 721 tgctgaagcc ccgggcccgg atgaccccgg ccccggcctc
ctgttcacag gagctcccga 781 gcgccgagga caacagcccg atggccagtg
acccattagg ggtggtcagg ggcggtcgag 841 tgaacacgca cgctggggga
acgggcccgg aaggctgccg ccccttcgcc aagttcatct 901 agggtcgctg
gaagggcacc ctctttaacc catccctcag caaacgcagc tcttcccaag 961
gaccaggtcc cttgacgttc cgaggatggg aaaggtgaca ggggcatgta tggaatttgc
1021 tgcttctctg gggtcccttc cacaggaggt cctgtgagaa ccaacctttg
aggcccaagt 1081 catggggttt caccgccttc ctcactccat atagaacacc
tttcccaata ggaaacccca 1141 acaggtaaac tagaaatttc cccttcatga
aggtagagag aaggggtctc tcccaacata 1201 tttctcttcc ttgtgcctct
cctctttatc acttttaagc ataaaaaaaa aaaaaaaaaa 1261 aaaaaaaaaa
aaaagcagtg ggttcctgag ctcaagactt tgaaggtgta gggaagagga 1321
aatcggagat cccagaagct tctccactgc cctatgcatt tatgttagat gccccgatcc
1381 cactggcatt tgagtgtgca aaccttgaca ttaacagctg aatggggcaa
gttgatgaaa 1441 acactacttt caagccttcg ttcttccttg agcatctctg
gggaagagct gtcaaaagac 1501 tggtggtagg ctggtgaaaa cttgacagct
agacttgatg cttgctgaaa tgaggcagga 1561 atcataatag aaaactcagc
ctccctacag ggtgagcacc ttctgtctcg ctgtctccct 1621 ctgtgcagcc
acagccagag ggcccagaat ggccccactc tgttcccaag cagttcatga 1681
tacagcctca ccttttggcc ccatctctgg tttttgaaaa tttggtctaa ggaataaata
1741 gcttttacac tggctcacga aaatctgccc tgctagaatt tgcttttcaa
aatggaaata 1801 aattccaact ctcctaagag gcatttaatt aaggctctac
ttccaggttg agtaggaatc 1861 cattctgaac aaactacaaa aatgtgactg
ggaagggggc tttgagagac tgggactgct 1921 ctgggttagg ttttctgtgg
actgaaaaat cgtgtccttt tctctaaatg aagtggcatc 1981 aaggactcag
ggggaaagaa atcaggggac atgttataga agttatgaaa agacaaccac 2041
atggtcaggc tcttgtctgt ggtctctagg gctctgcagc agcagtggct cttcgattag
2101 ttaaaactct cctaggctga cacatctggg tctcaatccc cttggaaatt
cttggtgcat 2161 taaatgaagc cttaccccat tactgcggtt cttcctgtaa
gggggctcca ttttcctccc 2221 tctctttaaa tgaccaccta aaggacagta
tattaacaag caaagtcgat tcaacaacag 2281 cttcttccca gtcacttttt
tttttctcac tgccatcaca tactaacctt atactttgat 2341 ctattctttt
tggttatgag agaaatgttg ggcaactgtt tttacctgat ggttttaagc 2401
tgaacttgaa ggactggttc ctattctgaa acagtaaaac tatgtataat agtatatagc
2461 catgcatggc aaatatttta atatttctgt tttcatttcc tgttggaaat
attatcctgc 2521 ataatagcta ttggaggctc ctcagtgaaa gatcccaaaa
ggattttggt ggaaaactag 2581 ttgtaatctc acaaactcaa cactaccatc
aggggttttc tttatggcaa agccaaaata 2641 gctcctacaa tttcttatat
ccctcgtcat gtggcagtat ttatttattt atttggaagt 2701 ttgcctatcc
ttctatattt atagatattt ataaaaatgt aacccctttt tcctttcttc 2761
tgtttaaaat aaaaataaaa tttatctcag cttctgttag cttatcctct ttgtagtact
2821 acttaaaagc atgtcggaat ataagaataa aaaggattat gggaggggaa
cattagggaa 2881 atccagagaa ggcaaaattg aaaaaaagat tttagaattt
taaaattttc aaagatttct 2941 tccattcata aggagactca atgattttaa
ttgatctaga cagaattatt taagttttat 3001 caatattgga tttctggt FGF23
amino acid sequence (NP_065689) (SEQ ID NO: 35) 1 MLGARLRLWV
CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS YHLQIHKNGH 61
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG NIFGSHYFDP ENCRFQHQTL
121 ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN PPPYSQFLSR RNEIPLIHFN
TPIPRRHTRS 181 AEDDSERDPL NVLKPRARMT PAPASCSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG 241 PEGCRPFAKF I FGF23 (R179Q) amino acid
sequence (SEQ ID NO: 36) 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS YHLQIHKNGH 61 VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG NIFGSHYFDP ENCRFQHQTL 121 ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS 181
AEDDSERDPL NVLKPRARMT PAPASCSQEL PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG
241 PEGCRPFAKF I Human beta-Klotho domain 1 (b-KL-D1) amino acid
sequence (SEQ ID NO: 37) 77 ydt fpknffwgig tgalqvegsw kkdgkgpsiw
dhfihthlkn 121 vsstngssds yiflekdlsa ldfigvsfyq fsiswprlfp
dgivtvanak glqyystlld 181 alvirniepi vtlyhwdlpl alqekyggwk
ndtildifnd yatycfqmfg drvkywitih 241 npylvawhgy gtgmhapgek
gnlaavytvg hnlikahskv whnynthfrp hqkgwlsitl 301 gshwiepnrs
entmdifkcq qsmvsvlgwf anpihgdgdy pegmrkklfs vlpifseaek 361
hemrgtadff afsfgpnnfk pintmakmgq nvslnlreal nwikleynnp rillaengwf
421 tdsrvktedt talymmknfl sqvlqairld eirvfgytaw slldgfewqd
aytirrglfy 481 vdfnskqker kpkssahyyk qiirengf Human beta-Klotho
domain 2 (b-KL-D2) amino acid sequence (SEQ ID NO: 38) 571
trpaqctdfv nikkqlemla rmkvthyrfa 601 ldwasvlptg nlsavnrqal
ryyrcvvseg lklgisamvt lyypthahlg lpepllhadg 661 wlnpstaeaf
qayaglcfqe lgdlvklwit inepnrlsdi ynrsgndtyg aahnllvaha 721
lawrlydrqf rpsqrgaysl slhadwaepa npyadshwra aerflqfela wfaeplfktg
781 dypaamreyi askhrrglss salprlteae rrllkgtvdf calnhfttrf
vmheqlagsr 841 ydsdrdiqfl qditrlsspt rlavipwgvr kllrwvrrny
gdmdlyitas giddqaledd 901 rlrkyylgky lqevlkayll dkvrikgyya
fklaeekskp rfgfftsdfk akssiqfynk 961 vissrgf Beta-Klotho
extracellular domain (without signal peptide) amino acid sequence
(SEQ ID NO: 39) 52 gfsgdgral 61 wsknpnftpv nesqlflydt fpknffwgig
tgalqvegsw kkdgkgpsiw dhfihthlkn 121 vsstngssds yiflekdlsa
ldfigvsfyq fsiswprlfp dgivtvanak glqyystlld 181 alvirniepi
vtlyhwdlpl alqekyggwk ndtildifnd yatycfqmfg drvkywitih 241
npylvawhgy gtgmhapgek gnlaavytvg hnlikahskv whnynthfrp hqkgwlsitl
301 gshwiepnrs entmdifkcq qsmvsvlgwf anpihgdgdy pegmrkklfs
vlpifseaek 361 hemrgtadff afsfgpnnfk pintmakmgq nvslnlreal
nwikleynnp rillaengwf 421 tdsrvktedt talymmknfl sqvlqairld
eirvfgytaw slldgfewqd aytirrglfy 481 vdfnskqker kpkssahyyk
qiirengfsl kestpdvqgq fpcdfswgvt esvlkpesva 541 sspqfsdphl
yvwnatgnrl lhrvegvrlk trpaqctdfv nikkqlemla rmkvthyrfa 601
ldwasvlptg nlsavnrqal ryyrcvvseg lklgisamvt lyypthahlg lpepllhadg
661 wlnpstaeaf qayaglcfqe lgdlvklwit inepnrlsdi ynrsgndtyg
aahnllvaha 721 lawrlydrqf rpsqrgaysl slhadwaepa npyadshwra
aerflqfela wfaeplfktg 781 dypaamreyi askhrrglss salprlteae
rrllkgtvdf calnhfttrf vmheqlagsr 841 ydsdrdiqfl qditrlsspt
rlavipwgvr kllrwvrrny gdmdlyitas giddqaledd 901 rlrkyylgky
lqevlkayll dkvrikgyya fklaeekskp rfgfftsdfk akssiqfynk 961
vissrgfpfe nsssrcsqtq entectvclf lvqkkpl sKlotho without signal
peptide-FGF23 amino acid sequence (without signal peptide)(SEQ ID
NO: 40) EPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW
QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA TGDVASDSYN
NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL RYYRRLLERL
RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ
VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH
LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL DFVLGWFAKP
VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC FGPTLSFQLL
DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS GTTKRDDAKY
MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD
FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN
YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP
QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR CMASELVRVN
ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA RLCFQELGHH
VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH AQNGKISIAL
QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ
RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED PIKYNDYLEV
QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH
AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR TAPRFGLYRY
AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC TECSFFHTRK
SLGSGGGGSG GGGSGGGGSL 1001 KYPNASPLLG SSWGGLIHLY TATARNSYHL
QIHKNGHVDG APHQTIYSAL 1051 MIRSEDAGFV VITGVMSRRY LCMDFRGNIF
GSHYFDPENC RFQHQTLENG 1101 YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP
YSQFLSRRNE IPLIHFNTPI 1151 PRRHTRSAED DSERDPLNVL KPRARMTPAP
ASCSQELPSA EDNSPMASDP 1201 LGVVRGGRVN THAGGTGPEG CRPFAKFI* sKlotho
without signal peptide-FGF23 (R179Q) (without signal peptide) amino
acid sequence (SEQ ID NO: 41) EPGDGAQ TWARFSRPPA 51 PEAAGLFQGT
FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP
LGAPSPLQPA TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG
SAGVPNREGL RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR
ALADHFRDYA ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS
PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD
HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI
KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI
FIVENGWFVS GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL
MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP
ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV
DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ
VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP
YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA
WHVYNEKFRH AQNGKISIAL QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL
AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT
TILVDSEKED PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK
FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901 AHILDGINLC
GYFAYSFNDR TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE
RFCPEEFTVC TECSFFHTRK SLGSGGGGSG GGGSGGGGSL
1001 KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051
MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101
YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151
PRRHTQSAED DSERDPLNVL KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201
LGVVRGGRVN THAGGTGPEG CRPFAKFI* FGF23 without signal peptide (SEQ
ID NO: 42) YPNASP LLGSSWGGLI HLYTATARNS YHLQIHKNGH 61 VDGAPHQTIY
SALMIRSEDA GFVVITGVMS RRYLCMDFRG NIFGSHYFDP ENCRFQHQTL 121
ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN PPPYSQFLSR RNEIPLIHFN TPIPRRHTRS
181 AEDDSERDPL NVLKPRARMT PAPASCSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG 241 PEGCRPFAKF I FGF23(R179Q) without signal peptide
(SEQ ID NO: 43) YPNASP LLGSSWGGLI HLYTATARNS YHLQIHKNGH 61
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG NIFGSHYFDP ENCRFQHQTL
121 ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN PPPYSQFLSR RNEIPLIHFN
TPIPRRHTQS 181 AEDDSERDPL NVLKPRARMT PAPASCSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG 241 PEGCRPFAKF I sKlotho with Klotho signal
peptide (SEQ ID NO: 44) 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR
LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW
QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA TGDVASDSYN
NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL RYYRRLLERL
RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ
VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH
LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL DFVLGWFAKP
VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC FGPTLSFQLL
DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS GTTKRDDAKY
MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD
FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN
YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP
QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR CMASELVRVN
ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA RLCFQELGHH
VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH AQNGKISIAL
QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ
RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED PIKYNDYLEV
QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH
AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR TAPRFGLYRY
AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC TECSFFHTRK SL
sKlotho with IgG Signal peptide (SEQ ID NO: 45) 1 MSVLTQVLAL
LLLWLTGLGG RRLRAEPGDG AQTWARFSRP PAPEAAGLFQ 51 GTFPDGFLWA
VGSAAYQTEG GWQQHGKGAS IWDTFTHHPL APPGDSRNAS 101 LPLGAPSPLQ
PATGDVASDS YNNVFRDTEA LRELGVTHYR FSISWARVLP 151 NGSAGVPNRE
GLRYYRRLLE RLRELGVQPV VTLYHWDLPQ RLQDAYGGWA 201 NRALADHFRD
YAELCFRHFG GQVKYWITID NPYVVAWHGY ATGRLAPGIR 251 GSPRLGYLVA
HNLLLAHAKV WHLYNTSFRP TQGGQVSIAL SSHWINPRPM 301 TDHSIKECQK
SLDFVLGWFA KPVFIDGDYP ESMKNNLSSI LPDFTESEKK 351 FIKGTADFFA
LCFGPTLSFQ LLDPHMKFRQ LESPNLRQLL SWIDLEFNHP 401 QIFIVENGWF
VSGTTKRDDA KYMYYLKKFI METLKAIKLD GVDVIGYTAW 451 SLMDGFEWHR
GYSIRRGLFY VDFLSQDKML LPKSSALFYQ KLIEKNGFPP 501 LPENQPLEGT
FPCDFAWGVV DNYIQVDTTL SQFTDLNVYL WDVHHSKRLI 551 KVDGVVTKKR
KSYCVDFAAI QPQIALLQEM HVTHFRFSLD WALILPLGNQ 601 SQVNHTILQY
YRCMASELVR VNITPVVALW QPMAPNQGLP RLLARQGAWE 651 NPYTALAFAE
YARLCFQELG HHVKLWITMN EPYTRNMTYS AGHNLLKAHA 701 LAWHVYNEKF
RHAQNGKISI ALQADWIEPA CPFSQKDKEV AERVLEFDIG 751 WLAEPIFGSG
DYPWVMRDWL NQRNNFLLPY FTEDEKKLIQ GTFDFLALSH 801 YTTILVDSEK
EDPIKYNDYL EVQEMTDITW LNSPSQVAVV PWGLRKVLNW 851 LKFKYGDLPM
YIISNGIDDG LHAEDDQLRV YYMQNYINEA LKAHILDGIN 901 LCGYFAYSFN
DRTAPRFGLY RYAADQFEPK ASMKHYRKII DSNGFPGPET 951 LERFCPEEFT
VCTECSFFHT RKSL* sKlotho-FGF23-FcLALA v1 (SEQ ID NO: 46) 1
ATGCCCGCCA GCGCCCCGCC GCGCCGCCCG CGGCCGCCGC CGCCGTCGCT GTCGCTGCTG
61 CTGGTGCTGC TGGGCCTGGG CGGCCGCCGC CTGCGTGCGG AGCCGGGCGA
CGGCGCGCAG 121 ACCTGGGCCC GTTTCTCGCG GCCTCCTGCC CCCGAGGCCG
CGGGCCTCTT CCAGGGCACC 181 TTCCCCGACG GCTTCCTCTG GGCCGTGGGC
AGCGCCGCCT ACCAGACCGA GGGCGGCTGG 241 CAGCAGCACG GCAAGGGTGC
GTCCATCTGG GATACGTTCA CCCACCACCC CCTGGCACCC 301 CCGGGAGACT
CCCGGAACGC CAGTCTGCCG TTGGGCGCCC CGTCGCCGCT GCAGCCCGCC 361
ACCGGGGACG TAGCCAGCGA CAGCTACAAC AACGTCTTCC GCGACACGGA GGCGCTGCGC
421 GAGCTCGGGG TCACTCACTA CCGCTTCTCC ATCTCGTGGG CGCGAGTGCT
CCCCAATGGC 481 AGCGCGGGCG TCCCCAACCG CGAGGGGCTG CGCTACTACC
GGCGCCTGCT GGAGCGGCTG 541 CGGGAGCTGG GCGTGCAGCC CGTGGTCACC
CTGTACCACT GGGACCTGCC CCAGCGCCTG 601 CAGGACGCCT ACGGCGGCTG
GGCCAACCGC GCCCTGGCCG ACCACTTCAG GGATTACGCG 661 GAGCTCTGCT
TCCGCCACTT CGGCGGTCAG GTCAAGTACT GGATCACCAT CGACAACCCC 721
TACGTGGTGG CCTGGCACGG CTACGCCACC GGGCGCCTGG CCCCCGGCAT CCGGGGCAGC
781 CCGCGGCTCG GGTACCTGGT GGCGCACAAC CTCCTCCTGG CTCATGCCAA
AGTCTGGCAT 841 CTCTACAATA CTTCTTTCCG TCCCACTCAG GGAGGTCAGG
TGTCCATTGC CCTAAGCTCT 901 CACTGGATCA ATCCTCGAAG AATGACCGAC
CACAGCATCA AAGAATGTCA AAAATCTCTG 961 GACTTTGTAC TAGGTTGGTT
TGCCAAACCC GTATTTATTG ATGGTGACTA TCCCGAGAGC 1021 ATGAAGAATA
ACCTTTCATC TATTCTGCCT GATTTTACTG AATCTGAGAA AAAGTTCATC 1081
AAAGGAACTG CTGACTTTTT TGCTCTTTGC TTTGGACCCA CCTTGAGTTT TCAACTTTTG
1141 GACCCTCACA TGAAGTTCCG CCAATTGGAA TCTCCCAACC TGAGGCAACT
GCTTTCCTGG 1201 ATTGACCTTG AATTTAACCA TCCTCAAATA TTTATTGTGG
AAAATGGCTG GTTTGTCTCA 1261 GGGACCACCA AGAGAGATGA TGCCAAATAT
ATGTATTACC TCAAAAAGTT CATCATGGAA 1321 ACCTTAAAAG CCATCAAGCT
GGATGGGGTG GATGTCATCG GGTATACCGC ATGGTCCCTC 1381 ATGGATGGTT
TCGAGTGGCA CAGAGGTTAC AGCATCAGGC GTGGACTCTT CTATGTTGAC 1441
TTTCTAAGCC AGGACAAGAT GTTGTTGCCA AAGTCTTCAG CCTTGTTCTA CCAAAAGCTG
1501 ATAGAGAAAA ATGGCTTCCC TCCTTTACCT GAAAATCAGC CCCTAGAAGG
GACATTTCCC 1561 TGTGACTTTG CTTGGGGAGT TGTTGACAAC TACATTCAAG
TAGATACCAC TCTGTCTCAG 1621 TTTACCGACC TGAATGTTTA CCTGTGGGAT
GTCCACCACA GTAAAAGGCT TATTAAAGTG 1681 GATGGGGTTG TGACCAAGAA
GAGGAAATCC TACTGTGTTG ACTTTGCTGC CATCCAGCCC 1741 CAGATCGCTT
TACTCCAGGA AATGCACGTT ACACATTTTC GCTTCTCCCT GGACTGGGCC 1801
CTGATTCTCC CTCTGGGTAA CCAGTCCCAG GTGAACCACA CCATCCTGCA GTACTATCGC
1861 TGCATGGCCA GCGAGCTTGT CCGTGTCAAC ATCACCCCAG TGGTGGCCCT
GTGGCAGCCT 1921 ATGGCCCCGA ACCAAGGACT GCCGCGCCTC CTGGCCAGGC
AGGGCGCCTG GGAGAACCCC 1981 TACACTGCCC TGGCCTTTGC AGAGTATGCC
CGACTGTGCT TTCAAGAGCT CGGCCATCAC 2041 GTCAAGCTTT GGATAACGAT
GAATGAGCCG TATACAAGGA ATATGACATA CAGTGCTGGC 2101 CACAACCTTC
TGAAGGCCCA TGCCCTGGCT TGGCATGTGT ACAATGAAAA GTTTAGGCAT 2161
GCTCAGAATG GGAAAATATC CATAGCCTTG CAGGCTGATT GGATAGAACC TGCCTGCCCT
2221 TTCTCCCAAA AGGACAAAGA GGTGGCCGAG AGAGTTTTGG AATTTGACAT
TGGCTGGCTG 2281 GCTGAGCCCA TTTTCGGCTC TGGAGATTAT CCATGGGTGA
TGAGGGACTG GCTGAACCAA 2341 AGAAACAATT TTCTTCTTCC TTATTTCACT
GAAGATGAAA AAAAGCTAAT CCAGGGTACC 2401 TTTGACTTTT TGGCTTTAAG
CCATTATACC ACCATCCTTG TAGACTCAGA AAAAGAAGAT 2461 CCAATAAAAT
ACAATGATTA CCTAGAAGTG CAAGAAATGA CCGACATCAC GTGGCTCAAC 2521
TCCCCCAGTC AGGTGGCGGT AGTGCCCTGG GGGTTGCGCA AAGTGCTGAA CTGGCTGAAG
2581 TTCAAGTACG GAGACCTCCC CATGTACATA ATATCCAACG GAATCGATGA
CGGGCTGCAT 2641 GCTGAGGACG ACCAGCTGAG GGTGTATTAT ATGCAGAATT
ACATAAACGA AGCTCTCAAA 2701 GCCCACATAC TGGATGGTAT CAATCTTTGC
GGATACTTTG CTTATTCGTT TAACGACCGC 2761 ACAGCTCCGA GGTTTGGCCT
CTATCGTTAT GCTGCAGATC AGTTTGAGCC CAAGGCATCC 2821 ATGAAACATT
ACAGGAAAAT TATTGACAGC AATGGTTTCC CGGGCCCAGA AACTCTGGAA 2881
AGATTTTGTC CAGAAGAATT CACCGTGTGT ACTGAGTGCA GTTTTTTTCA CACCCGAAAG
2941 TCTTTAGGAT CCGGAGGTGG AGGTTCAGGA GGTGGAGGTT CAGGAGGTGG
AGGTTCACTT 3001 AAGTATCCCA ATGCCTCCCC ACTGCTCGGC TCCAGCTGGG
GTGGCCTGAT CCACCTGTAC 3061 ACAGCCACAG CCAGGAACAG CTACCACCTG
CAGATCCACA AGAATGGCCA TGTGGATGGC 3121 GCACCCCATC AGACCATCTA
CAGTGCCCTG ATGATCAGAT CAGAGGATGC TGGCTTTGTG 3181 GTGATTACAG
GTGTGATGAG CAGAAGATAC CTCTGCATGG ATTTCAGAGG CAACATTTTT 3241
GGATCACACT ATTTCGACCC GGAGAACTGC AGGTTCCAAC ACCAGACGCT GGAAAACGGG
3301 TACGACGTCT ACCACTCTCC TCAGTATCAC TTCCTGGTCA GTCTGGGCCG
GGCGAAGAGA 3361 GCCTTCCTGC CAGGCATGAA CCCACCCCCG TACTCCCAGT
TCCTGTCCCG GAGGAACGAG 3421 ATCCCCCTAA TTCACTTCAA CACCCCCATA
CCACGGCGGC ACACCCAGAG CGCCGAGGAC 3481 GACTCGGAGC GGGACCCCCT
GAACGTGCTG AAGCCCCGGG CCCGGATGAC CCCGGCCCCG 3541 GCCTCCTGTT
CACAGGAGCT CCCGAGCGCC GAGGACAACA GCCCGATGGC CAGTGACCCA 3601
TTAGGGGTGG TCAGGGGCGG TCGAGTGAAC ACGCACGCTG GGGGAACGGG CCCGGAAGGC
3661 TGCCGCCCCT TCGCCAAGTT CATCGGAGGT GGAGGTTCAA AAACCCACAC
GTGTCCTCCT 3721 TGTCCTGCCC CAGAAGCAGC AGGTGGTCCA TCAGTTTTTC
TTTTCCCTCC CAAACCCAAG 3781 GATACGCTGA TGATCTCTCG CACGCCTGAG
GTGACATGCG TCGTAGTAGA CGTGAGCCAC 3841 GAAGATCCCG AGGTGAAGTT
CAATTGGTAT GTGGACGGAG TAGAAGTGCA TAACGCGAAA 3901 ACTAAGCCGC
GCGAGGAACA ATATAACAGT ACTTACAGGG TGGTATCCGT GCTCACAGTC 3961
CTGCACCAGG ACTGGCTGAA CGGTAAGGAA TACAAGTGCA AAGTAAGCAA CAAGGCACTT
4021 CCCGCGCCTA TTGAGAAAAC AATCTCCAAG GCGAAGGGAC AACCAAGAGA
ACCTCAGGTT 4081 TACACTCTCC CGCCTTCCAG GGAAGAGATG ACCAAAAATC
AAGTTTCCCT GACTTGCCTC 4141 GTCAAAGGAT TCTACCCTTC CGACATTGCT
GTTGAATGGG AAAGCAATGG ACAACCAGAG 4201 AACAACTACA AGACAACACC
CCCGGTGCTG GATAGTGACG GATCTTTCTT TCTCTACTCA 4261 AAGCTGACCG
TGGATAAGTC CAGGTGGCAG CAGGGAAACG TGTTTTCCTG CTCTGTCATG 4321
CATGAAGCGC TGCATAATCA CTATACCCAG AAGTCTCTGA GCTTGAGCCC AGGCAAGTAA
sKlotho-FGF23-FcLALA v1 (SEQ ID NO: 47) 1 MPASAPPRRP RPPPPSLSLL
LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG
SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA
TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL
RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA
ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN
LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL
DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC
FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS
GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY
SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP
CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS
YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR
CMASELVRVN ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA
RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH
AQNGKISIAL QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY
PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED
PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI
ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR
TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC
TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001 KYPNASPLLG SSWGGLIHLY
TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051 MIRSEDAGFV VITGVMSRRY
LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101 YDVYHSPQYH FLVSLGRAKR
AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151 PRRHTQSAED DSERDPLNVL
KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201 LGVVRGGRVN THAGGTGPEG
CRPFAKFIGG GGSKTHTCPP CPAPEAAGGP 1251 SVFLFPPKPK DTLMISRTPE
VTCVVVDVSH EDPEVKFNWY VDGVEVHNAK 1301 TKPREEQYNS TYRVVSVLTV
LHQDWLNGKE YKCKVSNKAL PAPIEKTISK 1351 AKGQPREPQV YTLPPSREEM
TKNQVSLTCL VKGFYPSDIA VEWESNGQPE 1401 NNYKTTPPVL DSDGSFFLYS
KLTVDKSRWQ QGNVFSCSVM HEALHNHYTQ 1451 KSLSLSPGK*
sKlotho-FGF23-FcLALA v2 (SEQ ID NO: 48) 1 ATGCCCGCCA GCGCCCCGCC
GCGCCGCCCG CGGCCGCCGC CGCCGTCGCT GTCGCTGCTG 61 CTGGTGCTGC
TGGGCCTGGG CGGCCGCCGC CTGCGTGCGG AGCCGGGCGA CGGCGCGCAG 121
ACCTGGGCCC GTTTCTCGCG GCCTCCTGCC CCCGAGGCCG CGGGCCTCTT CCAGGGCACC
181 TTCCCCGACG GCTTCCTCTG GGCCGTGGGC AGCGCCGCCT ACCAGACCGA
GGGCGGCTGG 241 CAGCAGCACG GCAAGGGTGC GTCCATCTGG GATACGTTCA
CCCACCACCC CCTGGCACCC 301 CCGGGAGACT CCCGGAACGC CAGTCTGCCG
TTGGGCGCCC CGTCGCCGCT GCAGCCCGCC 361 ACCGGGGACG TAGCCAGCGA
CAGCTACAAC AACGTCTTCC GCGACACGGA GGCGCTGCGC 421 GAGCTCGGGG
TCACTCACTA CCGCTTCTCC ATCTCGTGGG CGCGAGTGCT CCCCAATGGC 481
AGCGCGGGCG TCCCCAACCG CGAGGGGCTG CGCTACTACC GGCGCCTGCT GGAGCGGCTG
541 CGGGAGCTGG GCGTGCAGCC CGTGGTCACC CTGTACCACT GGGACCTGCC
CCAGCGCCTG 601 CAGGACGCCT ACGGCGGCTG GGCCAACCGC GCCCTGGCCG
ACCACTTCAG GGATTACGCG 661 GAGCTCTGCT TCCGCCACTT CGGCGGTCAG
GTCAAGTACT GGATCACCAT CGACAACCCC 721 TACGTGGTGG CCTGGCACGG
CTACGCCACC GGGCGCCTGG CCCCCGGCAT CCGGGGCAGC 781 CCGCGGCTCG
GGTACCTGGT GGCGCACAAC CTCCTCCTGG CTCATGCCAA AGTCTGGCAT 841
CTCTACAATA CTTCTTTCCG TCCCACTCAG GGAGGTCAGG TGTCCATTGC CCTAAGCTCT
901 CACTGGATCA ATCCTCGAAG AATGACCGAC CACAGCATCA AAGAATGTCA
AAAATCTCTG 961 GACTTTGTAC TAGGTTGGTT TGCCAAACCC GTATTTATTG
ATGGTGACTA TCCCGAGAGC 1021 ATGAAGAATA ACCTTTCATC TATTCTGCCT
GATTTTACTG AATCTGAGAA AAAGTTCATC 1081 AAAGGAACTG CTGACTTTTT
TGCTCTTTGC TTTGGACCCA CCTTGAGTTT TCAACTTTTG 1141 GACCCTCACA
TGAAGTTCCG CCAATTGGAA TCTCCCAACC TGAGGCAACT GCTTTCCTGG 1201
ATTGACCTTG AATTTAACCA TCCTCAAATA TTTATTGTGG AAAATGGCTG GTTTGTCTCA
1261 GGGACCACCA AGAGAGATGA TGCCAAATAT ATGTATTACC TCAAAAAGTT
CATCATGGAA 1321 ACCTTAAAAG CCATCAAGCT GGATGGGGTG GATGTCATCG
GGTATACCGC ATGGTCCCTC 1381 ATGGATGGTT TCGAGTGGCA CAGAGGTTAC
AGCATCAGGC GTGGACTCTT CTATGTTGAC 1441 TTTCTAAGCC AGGACAAGAT
GTTGTTGCCA AAGTCTTCAG CCTTGTTCTA CCAAAAGCTG 1501 ATAGAGAAAA
ATGGCTTCCC TCCTTTACCT GAAAATCAGC CCCTAGAAGG GACATTTCCC 1561
TGTGACTTTG CTTGGGGAGT TGTTGACAAC TACATTCAAG TAGATACCAC TCTGTCTCAG
1621 TTTACCGACC TGAATGTTTA CCTGTGGGAT GTCCACCACA GTAAAAGGCT
TATTAAAGTG 1681 GATGGGGTTG TGACCAAGAA GAGGAAATCC TACTGTGTTG
ACTTTGCTGC CATCCAGCCC 1741 CAGATCGCTT TACTCCAGGA AATGCACGTT
ACACATTTTC GCTTCTCCCT GGACTGGGCC 1801 CTGATTCTCC CTCTGGGTAA
CCAGTCCCAG GTGAACCACA CCATCCTGCA GTACTATCGC 1861 TGCATGGCCA
GCGAGCTTGT CCGTGTCAAC ATCACCCCAG TGGTGGCCCT GTGGCAGCCT 1921
ATGGCCCCGA ACCAAGGACT GCCGCGCCTC CTGGCCAGGC AGGGCGCCTG GGAGAACCCC
1981 TACACTGCCC TGGCCTTTGC AGAGTATGCC CGACTGTGCT TTCAAGAGCT
CGGCCATCAC 2041 GTCAAGCTTT GGATAACGAT GAATGAGCCG TATACAAGGA
ATATGACATA CAGTGCTGGC 2101 CACAACCTTC TGAAGGCCCA TGCCCTGGCT
TGGCATGTGT ACAATGAAAA GTTTAGGCAT 2161 GCTCAGAATG GGAAAATATC
CATAGCCTTG CAGGCTGATT GGATAGAACC TGCCTGCCCT 2221 TTCTCCCAAA
AGGACAAAGA GGTGGCCGAG AGAGTTTTGG AATTTGACAT TGGCTGGCTG 2281
GCTGAGCCCA TTTTCGGCTC TGGAGATTAT CCATGGGTGA TGAGGGACTG GCTGAACCAA
2341 AGAAACAATT TTCTTCTTCC TTATTTCACT GAAGATGAAA AAAAGCTAAT
CCAGGGTACC 2401 TTTGACTTTT TGGCTTTAAG CCATTATACC ACCATCCTTG
TAGACTCAGA AAAAGAAGAT 2461 CCAATAAAAT ACAATGATTA CCTAGAAGTG
CAAGAAATGA CCGACATCAC GTGGCTCAAC 2521 TCCCCCAGTC AGGTGGCGGT
AGTGCCCTGG GGGTTGCGCA AAGTGCTGAA CTGGCTGAAG 2581 TTCAAGTACG
GAGACCTCCC CATGTACATA ATATCCAACG GAATCGATGA CGGGCTGCAT 2641
GCTGAGGACG ACCAGCTGAG GGTGTATTAT ATGCAGAATT ACATAAACGA AGCTCTCAAA
2701 GCCCACATAC TGGATGGTAT CAATCTTTGC GGATACTTTG CTTATTCGTT
TAACGACCGC 2761 ACAGCTCCGA GGTTTGGCCT CTATCGTTAT GCTGCAGATC
AGTTTGAGCC CAAGGCATCC 2821 ATGAAACATT ACAGGAAAAT TATTGACAGC
AATGGTTTCC CGGGCCCAGA AACTCTGGAA 2881 AGATTTTGTC CAGAAGAATT
CACCGTGTGT ACTGAGTGCA GTTTTTTTCA CACCCGAAAG 2941 TCTTTAGGAT
CCGGAGGTGG AGGTTCAGGA GGTGGAGGTT CAGGAGGTGG AGGTTCACTT 3001
AAGTATCCCA ATGCCTCCCC ACTGCTCGGC TCCAGCTGGG GTGGCCTGAT CCACCTGTAC
3061 ACAGCCACAG CCAGGAACAG CTACCACCTG CAGATCCACA AGAATGGCCA
TGTGGATGGC 3121 GCACCCCATC AGACCATCTA CAGTGCCCTG ATGATCAGAT
CAGAGGATGC TGGCTTTGTG 3181 GTGATTACAG GTGTGATGAG CAGAAGATAC
CTCTGCATGG ATTTCAGAGG CAACATTTTT 3241 GGATCACACT ATTTCGACCC
GGAGAACTGC AGGTTCCAAC ACCAGACGCT GGAAAACGGG 3301 TACGACGTCT
ACCACTCTCC TCAGTATCAC TTCCTGGTCA GTCTGGGCCG GGCGAAGAGA 3361
GCCTTCCTGC CAGGCATGAA CCCACCCCCG TACTCCCAGT TCCTGTCCCG GAGGAACGAG
3421 ATCCCCCTAA TTCACTTCAA CACCCCCATA CCACGGCGGC ACACCCAGAG
CGCCGAGGAC 3481 GACTCGGAGC GGGACCCCCT GAACGTGCTG AAGCCCCGGG
CCCGGATGAC CCCGGCCCCG 3541 GCCTCCTGTT CACAGGAGCT CCCGAGCGCC
GAGGACAACA GCCCGATGGC CAGTGACCCA 3601 TTAGGGGTGG TCAGGGGCGG
TCGAGTGAAC ACGCACGCTG GGGGAACGGG CCCGGAAGGC 3661 TGCCGCCCCT
TCGCCAAGTT CATCGGAGGT GGAGGTTCAG CCCCAGAAGC AGCAGGTGGT 3721
CCATCAGTTT TTCTTTTCCC TCCCAAACCC AAGGATACGC TGATGATCTC TCGCACGCCT
3781 GAGGTGACAT GCGTCGTAGT AGACGTGAGC CACGAAGATC CCGAGGTGAA
GTTCAATTGG 3841 TATGTGGACG GAGTAGAAGT GCATAACGCG AAAACTAAGC
CGCGCGAGGA ACAATATAAC 3901 AGTACTTACA GGGTGGTATC CGTGCTCACA
GTCCTGCACC AGGACTGGCT GAACGGTAAG 3961 GAATACAAGT GCAAAGTAAG
CAACAAGGCA CTTCCCGCGC CTATTGAGAA AACAATCTCC 4021 AAGGCGAAGG
GACAACCAAG AGAACCTCAG GTTTACACTC TCCCGCCTTC CAGGGAAGAG 4081
ATGACCAAAA ATCAAGTTTC CCTGACTTGC CTCGTCAAAG GATTCTACCC TTCCGACATT
4141 GCTGTTGAAT GGGAAAGCAA TGGACAACCA GAGAACAACT ACAAGACAAC
ACCCCCGGTG 4201 CTGGATAGTG ACGGATCTTT CTTTCTCTAC TCAAAGCTGA
CCGTGGATAA GTCCAGGTGG 4261 CAGCAGGGAA ACGTGTTTTC CTGCTCTGTC
ATGCATGAAG CGCTGCATAA TCACTATACC 4321 CAGAAGTCTC TGAGCTTGAG
CCCAGGCAAG TAA sKlotho-FGF23-FcLALA v2 (SEQ ID NO: 49) 1 MPASAPPRRP
RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT
FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP
LGAPSPLQPA TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG
SAGVPNREGL RYYRRLLERL RELGVQPVVT LYHWDLPQRL
201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251
GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301
HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351
DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401
IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451
DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501
IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551
VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601
LILPLGNQSQ VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL 651
LARQGAWENP YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701
HNLLKAHALA WHVYNEKFRH AQNGKISIAL QADWIEPACP FSQKDKEVAE 751
RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801
FDFLALSHYT TILVDSEKED PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851
GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901
AHILDGINLC GYFAYSFNDR TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951
NGFPGPETLE RFCPEEFTVC TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001
KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051
MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101
YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151
PRRHTQSAED DSERDPLNVL KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201
LGVVRGGRVN THAGGTGPEG CRPFAKFIGG GGSAPEAAGG PSVFLFPPKP 1251
KDTLMISRTP EVTCVVVDVS HEDPEVKFNW YVDGVEVHNA KTKPREEQYN 1301
STYRVVSVLT VLHQDWLNGK EYKCKVSNKA LPAPIEKTIS KAKGQPREPQ 1351
VYTLPPSREE MTKNQVSLTC LVKGFYPSDI AVEWESNGQP ENNYKTTPPV 1401
LDSDGSFFLY SKLTVDKSRW QQGNVFSCSV MHEALHNHYT QKSLSLSPGK 1451 *
FGF23-FcLALA v1 (SEQ ID NO: 50) 1 ATGTTGGGGG CCCGCCTCAG GCTCTGGGTC
TGTGCCTTGT GCAGCGTCTG CAGCATGAGC 61 GTCCTCAGAG CCTATCCCAA
TGCCTCCCCA CTGCTCGGCT CCAGCTGGGG TGGCCTGATC 121 CACCTGTACA
CAGCCACAGC CAGGAACAGC TACCACCTGC AGATCCACAA GAATGGCCAT 181
GTGGATGGCG CACCCCATCA GACCATCTAC AGTGCCCTGA TGATCAGATC AGAGGATGCT
241 GGCTTTGTGG TGATTACAGG TGTGATGAGC AGAAGATACC TCTGCATGGA
TTTCAGAGGC 301 AACATTTTTG GATCACACTA TTTCGACCCG GAGAACTGCA
GGTTCCAACA CCAGACGCTG 361 GAAAACGGGT ACGACGTCTA CCACTCTCCT
CAGTATCACT TCCTGGTCAG TCTGGGCCGG 421 GCGAAGAGAG CCTTCCTGCC
AGGCATGAAC CCACCCCCGT ACTCCCAGTT CCTGTCCCGG 481 AGGAACGAGA
TCCCCCTAAT TCACTTCAAC ACCCCCATAC CACGGCGGCA CACCCAGAGC 541
GCCGAGGACG ACTCGGAGCG GGACCCCCTG AACGTGCTGA AGCCCCGGGC CCGGATGACC
601 CCGGCCCCGG CCTCCTGTTC ACAGGAGCTC CCGAGCGCCG AGGACAACAG
CCCGATGGCC 661 AGTGACCCAT TAGGGGTGGT CAGGGGCGGT CGAGTGAACA
CGCACGCTGG GGGAACGGGC 721 CCGGAAGGCT GCCGCCCCTT CGCCAAGTTC
ATCGGAGGTG GAGGTTCAAA AACCCACACG 781 TGTCCTCCTT GTCCTGCCCC
AGAAGCAGCA GGTGGTCCAT CAGTTTTTCT TTTCCCTCCC 841 AAACCCAAGG
ATACGCTGAT GATCTCTCGC ACGCCTGAGG TGACATGCGT CGTAGTAGAC 901
GTGAGCCACG AAGATCCCGA GGTGAAGTTC AATTGGTATG TGGACGGAGT AGAAGTGCAT
961 AACGCGAAAA CTAAGCCGCG CGAGGAACAA TATAACAGTA CTTACAGGGT
GGTATCCGTG 1021 CTCACAGTCC TGCACCAGGA CTGGCTGAAC GGTAAGGAAT
ACAAGTGCAA AGTAAGCAAC 1081 AAGGCACTTC CCGCGCCTAT TGAGAAAACA
ATCTCCAAGG CGAAGGGACA ACCAAGAGAA 1141 CCTCAGGTTT ACACTCTCCC
GCCTTCCAGG GAAGAGATGA CCAAAAATCA AGTTTCCCTG 1201 ACTTGCCTCG
TCAAAGGATT CTACCCTTCC GACATTGCTG TTGAATGGGA AAGCAATGGA 1261
CAACCAGAGA ACAACTACAA GACAACACCC CCGGTGCTGG ATAGTGACGG ATCTTTCTTT
1321 CTCTACTCAA AGCTGACCGT GGATAAGTCC AGGTGGCAGC AGGGAAACGT
GTTTTCCTGC 1381 TCTGTCATGC ATGAAGCGCT GCATAATCAC TATACCCAGA
AGTCTCTGAG CTTGAGCCCA 1441 GGCAAGTAA FGF23(R179Q)-FcLALAv1 (SEQ ID
NO: 51) 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51
YHLQIHKNGH VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101
NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151
PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201
PAPASCSQEL PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251
IGGGGSKTHT CPPCPAPEAA GGPSVFLFPP KPKDTLMISR TPEVTCVVVD 301
VSHEDPEVKF NWYVDGVEVH NAKTKPREEQ YNSTYRVVSV LTVLHQDWLN 351
GKEYKCKVSN KALPAPIEKT ISKAKGQPRE PQVYTLPPSR EEMTKNQVSL 401
TCLVKGFYPS DIAVEWESNG QPENNYKTTP PVLDSDGSFF LYSKLTVDKS 451
RWQQGNVFSC SVMHEALHNH YTQKSLSLSP GK* FGF23-FcLALA v2 (SEQ ID NO:
52) 1 ATGTTGGGGG CCCGCCTCAG GCTCTGGGTC TGTGCCTTGT GCAGCGTCTG
CAGCATGAGC 61 GTCCTCAGAG CCTATCCCAA TGCCTCCCCA CTGCTCGGCT
CCAGCTGGGG TGGCCTGATC 121 CACCTGTACA CAGCCACAGC CAGGAACAGC
TACCACCTGC AGATCCACAA GAATGGCCAT 181 GTGGATGGCG CACCCCATCA
GACCATCTAC AGTGCCCTGA TGATCAGATC AGAGGATGCT 241 GGCTTTGTGG
TGATTACAGG TGTGATGAGC AGAAGATACC TCTGCATGGA TTTCAGAGGC 301
AACATTTTTG GATCACACTA TTTCGACCCG GAGAACTGCA GGTTCCAACA CCAGACGCTG
361 GAAAACGGGT ACGACGTCTA CCACTCTCCT CAGTATCACT TCCTGGTCAG
TCTGGGCCGG 421 GCGAAGAGAG CCTTCCTGCC AGGCATGAAC CCACCCCCGT
ACTCCCAGTT CCTGTCCCGG 481 AGGAACGAGA TCCCCCTAAT TCACTTCAAC
ACCCCCATAC CACGGCGGCA CACCCAGAGC 541 GCCGAGGACG ACTCGGAGCG
GGACCCCCTG AACGTGCTGA AGCCCCGGGC CCGGATGACC 601 CCGGCCCCGG
CCTCCTGTTC ACAGGAGCTC CCGAGCGCCG AGGACAACAG CCCGATGGCC 661
AGTGACCCAT TAGGGGTGGT CAGGGGCGGT CGAGTGAACA CGCACGCTGG GGGAACGGGC
721 CCGGAAGGCT GCCGCCCCTT CGCCAAGTTC ATCGGAGGTG GAGGTTCAGC
CCCAGAAGCA 781 GCAGGTGGTC CATCAGTTTT TCTTTTCCCT CCCAAACCCA
AGGATACGCT GATGATCTCT 841 CGCACGCCTG AGGTGACATG CGTCGTAGTA
GACGTGAGCC ACGAAGATCC CGAGGTGAAG 901 TTCAATTGGT ATGTGGACGG
AGTAGAAGTG CATAACGCGA AAACTAAGCC GCGCGAGGAA 961 CAATATAACA
GTACTTACAG GGTGGTATCC GTGCTCACAG TCCTGCACCA GGACTGGCTG 1021
AACGGTAAGG AATACAAGTG CAAAGTAAGC AACAAGGCAC TTCCCGCGCC TATTGAGAAA
1081 ACAATCTCCA AGGCGAAGGG ACAACCAAGA GAACCTCAGG TTTACACTCT
CCCGCCTTCC 1141 AGGGAAGAGA TGACCAAAAA TCAAGTTTCC CTGACTTGCC
TCGTCAAAGG ATTCTACCCT 1201 TCCGACATTG CTGTTGAATG GGAAAGCAAT
GGACAACCAG AGAACAACTA CAAGACAACA 1261 CCCCCGGTGC TGGATAGTGA
CGGATCTTTC TTTCTCTACT CAAAGCTGAC CGTGGATAAG 1321 TCCAGGTGGC
AGCAGGGAAA CGTGTTTTCC TGCTCTGTCA TGCATGAAGC GCTGCATAAT 1381
CACTATACCC AGAAGTCTCT GAGCTTGAGC CCAGGCAAGT AA
FGF23(R179Q)-FcLALAv2 (SEQ ID NO: 53) 1 MLGARLRLWV CALCSVCSMS
VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY
SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL
ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN
TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IGGGGSAPEA AGGPSVFLFP
PKPKDTLMIS RTPEVTCVVV DVSHEDPEVK 301 FNWYVDGVEV HNAKTKPREE
QYNSTYRVVS VLTVLHQDWL NGKEYKCKVS 351 NKALPAPIEK TISKAKGQPR
EPQVYTLPPS REEMTKNQVS LTCLVKGFYP 401 SDIAVEWESN GQPENNYKTT
PPVLDSDGSF FLYSKLTVDK SRWQQGNVFS 451 CSVMHEALHN HYTQKSLSLS PGK*
Amino acid sequence of sKlotho-FGF23 (R1156Q, C1183S) (SEQ ID NO:
54) sKlotho: aa [amino acid] 1-982; Linker1: aa 983-1001; FGF23: aa
1002-1228 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA
51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101
PGDSRNASLP LGAPSPLQPA TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151
ISWARVLPNG SAGVPNREGL RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201
QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251
GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301
HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351
DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401
IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451
DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501
IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551
VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601
LILPLGNQSQ VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL 651
LARQGAWENP YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701
HNLLKAHALA WHVYNEKFRH AQNGKISIAL QADWIEPACP FSQKDKEVAE 751
RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801
FDFLALSHYT TILVDSEKED PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851
GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901
AHILDGINLC GYFAYSFNDR TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951
NGFPGPETLE RFCPEEFTVC TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001
KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051
MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101
YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151
PRRHTQSAED DSERDPLNVL KPRARMTPAP ASSSQELPSA EDNSPMASDP 1201
LGVVRGGRVN THAGGTGPEG CRPFAKFI* Amino acid sequence of
sKlotho-FGF23 (R1156Q, C1221S) (SEQ ID NO: 55) sKlotho: 1-982;
Linker1: 983-1001; FGF23: 1002-1228; 1 MPASAPPRRP RPPPPSLSLL
LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG
SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA
TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL
RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA
ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN
LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL
DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC
FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS
GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY
SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP
CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS
YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR
CMASELVRVN ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA
RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH
AQNGKISIAL QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY
PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED
PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI
ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR
TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC
TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001 KYPNASPLLG SSWGGLIHLY
TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051 MIRSEDAGFV VITGVMSRRY
LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101 YDVYHSPQYH FLVSLGRAKR
AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151 PRRHTQSAED DSERDPLNVL
KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201 LGVVRGGRVN THAGGTGPEG
SRPFAKFI* Amino acid sequence of sKlotho-FGF23 (R1156Q, Q1133A)
(SEQ ID NO: 56) sKlotho: 1-982; Linker1: 983-1001; FGF23: 1002-1228
1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51
PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101
PGDSRNASLP LGAPSPLQPA TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151
ISWARVLPNG SAGVPNREGL RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201
QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251
GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301
HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351
DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401
IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451
DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501
IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551
VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601
LILPLGNQSQ VNHTILQYYR CMASELVRVN ITPVVALWQP MAPNQGLPRL 651
LARQGAWENP YTALAFAEYA RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701
HNLLKAHALA WHVYNEKFRH AQNGKISIAL QADWIEPACP FSQKDKEVAE 751
RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801
FDFLALSHYT TILVDSEKED PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851
GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901
AHILDGINLC GYFAYSFNDR TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951
NGFPGPETLE RFCPEEFTVC TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001
KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051
MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101
YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSAFLSRRNE IPLIHFNTPI 1151
PRRHTQSAED DSERDPLNVL KPRARMTPAP ASCSQELPSA EDNSPMASDP 1201
LGVVRGGRVN THAGGTGPEG CRPFAKFI* Amino acid sequence of
sKlotho-FGF23 (R1156Q, C1183S, C1221S) (SEQ ID NO: 57) sKlotho:
1-982; Linker1: 983-1001; FGF23: 1002-1228 1 MPASAPPRRP RPPPPSLSLL
LVLLGLGGRR LRAEPGDGAQ TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG
SAAYQTEGGW QQHGKGASIW DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA
TGDVASDSYN NVFRDTEALR ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL
RYYRRLLERL RELGVQPVVT LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA
ELCFRHFGGQ VKYWITIDNP YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN
LLLAHAKVWH LYNTSFRPTQ GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL
DFVLGWFAKP VFIDGDYPES MKNNLSSILP 351 DFTESEKKFI KGTADFFALC
FGPTLSFQLL DPHMKFRQLE SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS
GTTKRDDAKY MYYLKKFIME TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY
SIRRGLFYVD FLSQDKMLLP KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP
CDFAWGVVDN YIQVDTTLSQ FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS
YCVDFAAIQP QIALLQEMHV THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR
CMASELVRVN ITPVVALWQP MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA
RLCFQELGHH VKLWITMNEP YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH
AQNGKISIAL QADWIEPACP FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY
PWVMRDWLNQ RNNFLLPYFT EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED
PIKYNDYLEV QEMTDITWLN SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI
ISNGIDDGLH AEDDQLRVYY MQNYINEALK 901 AHILDGINLC GYFAYSFNDR
TAPRFGLYRY AADQFEPKAS MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC
TECSFFHTRK SLGSGGGGSG GGGSGGGGSL 1001 KYPNASPLLG SSWGGLIHLY
TATARNSYHL QIHKNGHVDG APHQTIYSAL 1051 MIRSEDAGFV VITGVMSRRY
LCMDFRGNIF GSHYFDPENC RFQHQTLENG 1101 YDVYHSPQYH FLVSLGRAKR
AFLPGMNPPP YSQFLSRRNE IPLIHFNTPI 1151 PRRHTQSAED DSERDPLNVL
KPRARMTPAP ASSSQELPSA EDNSPMASDP 1201 LGVVRGGRVN THAGGTGPEG
SRPFAKFI* Amino acid sequence of sKlotho-FGF23 (R1156Q, C1183S,
C1221S, Q1133A) (SEQ ID NO: 58) sKlotho: 1-982; Linker1: 983-1001;
FGF23: 1002-1228 1 MPASAPPRRP RPPPPSLSLL LVLLGLGGRR LRAEPGDGAQ
TWARFSRPPA 51 PEAAGLFQGT FPDGFLWAVG SAAYQTEGGW QQHGKGASIW
DTFTHHPLAP 101 PGDSRNASLP LGAPSPLQPA TGDVASDSYN NVFRDTEALR
ELGVTHYRFS 151 ISWARVLPNG SAGVPNREGL RYYRRLLERL RELGVQPVVT
LYHWDLPQRL 201 QDAYGGWANR ALADHFRDYA ELCFRHFGGQ VKYWITIDNP
YVVAWHGYAT 251 GRLAPGIRGS PRLGYLVAHN LLLAHAKVWH LYNTSFRPTQ
GGQVSIALSS 301 HWINPRRMTD HSIKECQKSL DFVLGWFAKP VFIDGDYPES
MKNNLSSILP 351 DFTESEKKFI KGTADFFALC FGPTLSFQLL DPHMKFRQLE
SPNLRQLLSW 401 IDLEFNHPQI FIVENGWFVS GTTKRDDAKY MYYLKKFIME
TLKAIKLDGV 451 DVIGYTAWSL MDGFEWHRGY SIRRGLFYVD FLSQDKMLLP
KSSALFYQKL 501 IEKNGFPPLP ENQPLEGTFP CDFAWGVVDN YIQVDTTLSQ
FTDLNVYLWD 551 VHHSKRLIKV DGVVTKKRKS YCVDFAAIQP QIALLQEMHV
THFRFSLDWA 601 LILPLGNQSQ VNHTILQYYR CMASELVRVN ITPVVALWQP
MAPNQGLPRL 651 LARQGAWENP YTALAFAEYA RLCFQELGHH VKLWITMNEP
YTRNMTYSAG 701 HNLLKAHALA WHVYNEKFRH AQNGKISIAL QADWIEPACP
FSQKDKEVAE 751 RVLEFDIGWL AEPIFGSGDY PWVMRDWLNQ RNNFLLPYFT
EDEKKLIQGT 801 FDFLALSHYT TILVDSEKED PIKYNDYLEV QEMTDITWLN
SPSQVAVVPW 851 GLRKVLNWLK FKYGDLPMYI ISNGIDDGLH AEDDQLRVYY
MQNYINEALK 901 AHILDGINLC GYFAYSFNDR TAPRFGLYRY AADQFEPKAS
MKHYRKIIDS 951 NGFPGPETLE RFCPEEFTVC TECSFFHTRK SLGSGGGGSG
GGGSGGGGSL 1001 KYPNASPLLG SSWGGLIHLY TATARNSYHL QIHKNGHVDG
APHQTIYSAL 1051 MIRSEDAGFV VITGVMSRRY LCMDFRGNIF GSHYFDPENC
RFQHQTLENG 1101 YDVYHSPQYH FLVSLGRAKR AFLPGMNPPP YSAFLSRRNE
IPLIHFNTPI 1151 PRRHTQSAED DSERDPLNVL KPRARMTPAP ASSSQELPSA
EDNSPMASDP 1201 LGVVRGGRVN THAGGTGPEG SRPFAKFI* Amino acid sequence
of FGF23(R179Q; C206S)-FcLALAv1 (SEQ ID NO: 59) FGF23: 1-251;
Linker: 252-256; FcLALA: 257-482 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN
151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201
PAPASSSQEL PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251
IGGGGSKTHT CPPCPAPEAA GGPSVFLFPP KPKDTLMISR TPEVTCVVVD 301
VSHEDPEVKF NWYVDGVEVH NAKTKPREEQ YNSTYRVVSV LTVLHQDWLN 351
GKEYKCKVSN KALPAPIEKT ISKAKGQPRE PQVYTLPPSR EEMTKNQVSL 401
TCLVKGFYPS DIAVEWESNG QPENNYKTTP PVLDSDGSFF LYSKLTVDKS 451
RWQQGNVFSC SVMHEALHNH YTQKSLSLSP GK* Amino acid sequence of
FGF23(R179Q, C244S)-FcLALAv1 (SEQ ID NO: 60) FGF23: 1-251; Linker:
252-256; FcLALA: 257-482 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS
AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG PEGSRPFAKF 251 IGGGGSKTHT CPPCPAPEAA GGPSVFLFPP
KPKDTLMISR TPEVTCVVVD 301 VSHEDPEVKF NWYVDGVEVH NAKTKPREEQ
YNSTYRVVSV LTVLHQDWLN 351 GKEYKCKVSN KALPAPIEKT ISKAKGQPRE
PQVYTLPPSR EEMTKNQVSL 401 TCLVKGFYPS DIAVEWESNG QPENNYKTTP
PVLDSDGSFF LYSKLTVDKS 451 RWQQGNVFSC SVMHEALHNH YTQKSLSLSP GK*
Amino acid sequence of FGF23(R179Q, Q156A)-FcLALAv1 (SEQ ID NO: 61)
FGF23: 1-251; Linker: 252-256; FcLALA: 257-482 1 MLGARLRLWV
CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP
ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSAFLSR
RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL
PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IGGGGSKTHT
CPPCPAPEAA GGPSVFLFPP KPKDTLMISR TPEVTCVVVD 301 VSHEDPEVKF
NWYVDGVEVH NAKTKPREEQ YNSTYRVVSV LTVLHQDWLN 351 GKEYKCKVSN
KALPAPIEKT ISKAKGQPRE PQVYTLPPSR EEMTKNQVSL 401 TCLVKGFYPS
DIAVEWESNG QPENNYKTTP PVLDSDGSFF LYSKLTVDKS 451 RWQQGNVFSC
SVMHEALHNH YTQKSLSLSP GK* Amino acid sequence of FGF23(R179Q,
C206S, C2445)-FcLALAv1 (SEQ ID NO: 62) FGF23: 1-251; Linker:
252-256; FcLALA: 257-482 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS
AEDDSERDPL NVLKPRARMT 201 PAPASSSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG PEGSRPFAKF 251 IGGGGSKTHT CPPCPAPEAA GGPSVFLFPP
KPKDTLMISR TPEVTCVVVD 301 VSHEDPEVKF NWYVDGVEVH NAKTKPREEQ
YNSTYRVVSV LTVLHQDWLN 351 GKEYKCKVSN KALPAPIEKT ISKAKGQPRE
PQVYTLPPSR EEMTKNQVSL 401 TCLVKGFYPS DIAVEWESNG QPENNYKTTP
PVLDSDGSFF LYSKLTVDKS 451 RWQQGNVFSC SVMHEALHNH YTQKSLSLSP GK*
Amino acid sequence of FGF23(R179Q, C206S, C244S, Q156A)-FcLALAv1
(SEQ ID NO: 63) FGF23: 1-251; Linker: 252-256; FcLALA: 257-482 1
MLGARLRLWV CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51
YHLQIHKNGH VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101
NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151
PPPYSAFLSR RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201
PAPASSSQEL PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGSRPFAKF 251
IGGGGSKTHT CPPCPAPEAA GGPSVFLFPP KPKDTLMISR TPEVTCVVVD 301
VSHEDPEVKF NWYVDGVEVH NAKTKPREEQ YNSTYRVVSV LTVLHQDWLN 351
GKEYKCKVSN KALPAPIEKT ISKAKGQPRE PQVYTLPPSR EEMTKNQVSL 401
TCLVKGFYPS DIAVEWESNG QPENNYKTTP PVLDSDGSFF LYSKLTVDKS 451
RWQQGNVFSC SVMHEALHNH YTQKSLSLSP GK* Amino acid sequence of
FGF23(R179Q, C206S)-FcLALAv2 (SEQ ID NO: 64) FGF23: 1-251; Linker:
252-256; FcLALA: 257-473 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP
LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA
GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP
QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS
AEDDSERDPL NVLKPRARMT 201 PAPASSSQEL PSAEDNSPMA SDPLGVVRGG
RVNTHAGGTG PEGCRPFAKF 251 IGGGGSAPEA AGGPSVFLFP PKPKDTLMIS
RTPEVTCVVV DVSHEDPEVK 301 FNWYVDGVEV HNAKTKPREE QYNSTYRVVS
VLTVLHQDWL NGKEYKCKVS 351 NKALPAPIEK TISKAKGQPR EPQVYTLPPS
REEMTKNQVS LTCLVKGFYP 401 SDIAVEWESN GQPENNYKTT PPVLDSDGSF
FLYSKLTVDK SRWQQGNVFS 451 CSVMHEALHN HYTQKSLSLS PGK* Amino acid
sequence of FGF23(R179Q,C244S)-FcLALAv2 (SEQ ID NO: 65) FGF23:
1-251; Linker: 252-256; FcLALA: 257-473 1 MLGARLRLWV CALCSVCSMS
VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY
SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL
ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN
TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG PEGSRPFAKF 251 IGGGGSAPEA AGGPSVFLFP
PKPKDTLMIS RTPEVTCVVV DVSHEDPEVK 301 FNWYVDGVEV HNAKTKPREE
QYNSTYRVVS VLTVLHQDWL NGKEYKCKVS 351 NKALPAPIEK TISKAKGQPR
EPQVYTLPPS REEMTKNQVS LTCLVKGFYP 401 SDIAVEWESN GQPENNYKTT
PPVLDSDGSF FLYSKLTVDK SRWQQGNVFS 451 CSVMHEALHN HYTQKSLSLS PGK*
Amino acid sequence of FGF23(R179Q,Q156A)-FcLALAv2 (SEQ ID NO: 66)
FGF23: 1-251; Linker: 252-256; FcLALA: 257-473 1 MLGARLRLWV
CALCSVCSMS VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH
VDGAPHQTIY SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP
ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSAFLSR
RNEIPLIHFN TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASCSQEL
PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG PEGCRPFAKF 251 IGGGGSAPEA
AGGPSVFLFP PKPKDTLMIS RTPEVTCVVV DVSHEDPEVK 301 FNWYVDGVEV
HNAKTKPREE QYNSTYRVVS VLTVLHQDWL NGKEYKCKVS 351 NKALPAPIEK
TISKAKGQPR EPQVYTLPPS REEMTKNQVS LTCLVKGFYP 401 SDIAVEWESN
GQPENNYKTT PPVLDSDGSF FLYSKLTVDK SRWQQGNVFS 451 CSVMHEALHN
HYTQKSLSLS PGK* Amino acid sequence of FGF23(R179Q, C206S,
C244S)-FcLALAv2 (SEQ ID NO: 67) FGF23: 1-251; Linker: 252-256;
FcLALA: 257-473 1 MLGARLRLWV CALCSVCSMS VLRAYPNASP LLGSSWGGLI
HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY SALMIRSEDA GFVVITGVMS
RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL ENGYDVYHSP QYHFLVSLGR
AKRAFLPGMN 151 PPPYSQFLSR RNEIPLIHFN TPIPRRHTQS AEDDSERDPL
NVLKPRARMT 201 PAPASSSQEL PSAEDNSPMA SDPLGVVRGG RVNTHAGGTG
PEGSRPFAKF 251 IGGGGSAPEA AGGPSVFLFP PKPKDTLMIS RTPEVTCVVV
DVSHEDPEVK 301 FNWYVDGVEV HNAKTKPREE QYNSTYRVVS VLTVLHQDWL
NGKEYKCKVS 351 NKALPAPIEK TISKAKGQPR EPQVYTLPPS REEMTKNQVS
LTCLVKGFYP 401 SDIAVEWESN GQPENNYKTT PPVLDSDGSF FLYSKLTVDK
SRWQQGNVFS 451 CSVMHEALHN HYTQKSLSLS PGK* Amino acid sequence of
FGF23(R179Q, C206S, C244S, Q156A)-FcLALAv2 (SEQ ID NO: 68) FGF23:
1-251; Linker: 252-256; FcLALA: 257-473 1 MLGARLRLWV CALCSVCSMS
VLRAYPNASP LLGSSWGGLI HLYTATARNS 51 YHLQIHKNGH VDGAPHQTIY
SALMIRSEDA GFVVITGVMS RRYLCMDFRG 101 NIFGSHYFDP ENCRFQHQTL
ENGYDVYHSP QYHFLVSLGR AKRAFLPGMN 151 PPPYSAFLSR RNEIPLIHFN
TPIPRRHTQS AEDDSERDPL NVLKPRARMT 201 PAPASSSQEL PSAEDNSPMA
SDPLGVVRGG RVNTHAGGTG PEGSRPFAKF 251 IGGGGSAPEA AGGPSVFLFP
PKPKDTLMIS RTPEVTCVVV DVSHEDPEVK 301 FNWYVDGVEV HNAKTKPREE
QYNSTYRVVS VLTVLHQDWL NGKEYKCKVS 351 NKALPAPIEK TISKAKGQPR
EPQVYTLPPS REEMTKNQVS LTCLVKGFYP 401 SDIAVEWESN GQPENNYKTT
PPVLDSDGSF FLYSKLTVDK SRWQQGNVFS 451 CSVMHEALHN HYTQKSLSLS PGK*
Sequence CWU 1
1
7415003DNAHomo sapiens 1cgcgcagcat gcccgccagc gccccgccgc gccgcccgcg
gccgccgccg ccgtcgctgt 60cgctgctgct ggtgctgctg ggcctgggcg gccgccgcct
gcgtgcggag ccgggcgacg 120gcgcgcagac ctgggcccgt ttctcgcggc
ctcctgcccc cgaggccgcg ggcctcttcc 180agggcacctt ccccgacggc
ttcctctggg ccgtgggcag cgccgcctac cagaccgagg 240gcggctggca
gcagcacggc aagggtgcgt ccatctggga tacgttcacc caccaccccc
300tggcaccccc gggagactcc cggaacgcca gtctgccgtt gggcgccccg
tcgccgctgc 360agcccgccac cggggacgta gccagcgaca gctacaacaa
cgtcttccgc gacacggagg 420cgctgcgcga gctcggggtc actcactacc
gcttctccat ctcgtgggcg cgagtgctcc 480ccaatggcag cgcgggcgtc
cccaaccgcg aggggctgcg ctactaccgg cgcctgctgg 540agcggctgcg
ggagctgggc gtgcagcccg tggtcaccct gtaccactgg gacctgcccc
600agcgcctgca ggacgcctac ggcggctggg ccaaccgcgc cctggccgac
cacttcaggg 660attacgcgga gctctgcttc cgccacttcg gcggtcaggt
caagtactgg atcaccatcg 720acaaccccta cgtggtggcc tggcacggct
acgccaccgg gcgcctggcc cccggcatcc 780ggggcagccc gcggctcggg
tacctggtgg cgcacaacct cctcctggct catgccaaag 840tctggcatct
ctacaatact tctttccgtc ccactcaggg aggtcaggtg tccattgccc
900taagctctca ctggatcaat cctcgaagaa tgaccgacca cagcatcaaa
gaatgtcaaa 960aatctctgga ctttgtacta ggttggtttg ccaaacccgt
atttattgat ggtgactatc 1020ccgagagcat gaagaataac ctttcatcta
ttctgcctga ttttactgaa tctgagaaaa 1080agttcatcaa aggaactgct
gacttttttg ctctttgctt tggacccacc ttgagttttc 1140aacttttgga
ccctcacatg aagttccgcc aattggaatc tcccaacctg aggcaactgc
1200tttcctggat tgaccttgaa tttaaccatc ctcaaatatt tattgtggaa
aatggctggt 1260ttgtctcagg gaccaccaag agagatgatg ccaaatatat
gtattacctc aaaaagttca 1320tcatggaaac cttaaaagcc atcaagctgg
atggggtgga tgtcatcggg tataccgcat 1380ggtccctcat ggatggtttc
gagtggcaca gaggttacag catcaggcgt ggactcttct 1440atgttgactt
tctaagccag gacaagatgt tgttgccaaa gtcttcagcc ttgttctacc
1500aaaagctgat agagaaaaat ggcttccctc ctttacctga aaatcagccc
ctagaaggga 1560catttccctg tgactttgct tggggagttg ttgacaacta
cattcaagta gataccactc 1620tgtctcagtt taccgacctg aatgtttacc
tgtgggatgt ccaccacagt aaaaggctta 1680ttaaagtgga tggggttgtg
accaagaaga ggaaatccta ctgtgttgac tttgctgcca 1740tccagcccca
gatcgcttta ctccaggaaa tgcacgttac acattttcgc ttctccctgg
1800actgggccct gattctccct ctgggtaacc agtcccaggt gaaccacacc
atcctgcagt 1860actatcgctg catggccagc gagcttgtcc gtgtcaacat
caccccagtg gtggccctgt 1920ggcagcctat ggccccgaac caaggactgc
cgcgcctcct ggccaggcag ggcgcctggg 1980agaaccccta cactgccctg
gcctttgcag agtatgcccg actgtgcttt caagagctcg 2040gccatcacgt
caagctttgg ataacgatga atgagccgta tacaaggaat atgacataca
2100gtgctggcca caaccttctg aaggcccatg ccctggcttg gcatgtgtac
aatgaaaagt 2160ttaggcatgc tcagaatggg aaaatatcca tagccttgca
ggctgattgg atagaacctg 2220cctgcccttt ctcccaaaag gacaaagagg
tggccgagag agttttggaa tttgacattg 2280gctggctggc tgagcccatt
ttcggctctg gagattatcc atgggtgatg agggactggc 2340tgaaccaaag
aaacaatttt cttcttcctt atttcactga agatgaaaaa aagctaatcc
2400agggtacctt tgactttttg gctttaagcc attataccac catccttgta
gactcagaaa 2460aagaagatcc aataaaatac aatgattacc tagaagtgca
agaaatgacc gacatcacgt 2520ggctcaactc ccccagtcag gtggcggtag
tgccctgggg gttgcgcaaa gtgctgaact 2580ggctgaagtt caagtacgga
gacctcccca tgtacataat atccaacgga atcgatgacg 2640ggctgcatgc
tgaggacgac cagctgaggg tgtattatat gcagaattac ataaacgaag
2700ctctcaaagc ccacatactg gatggtatca atctttgcgg atactttgct
tattcgttta 2760acgaccgcac agctccgagg tttggcctct atcgttatgc
tgcagatcag tttgagccca 2820aggcatccat gaaacattac aggaaaatta
ttgacagcaa tggtttcccg ggcccagaaa 2880ctctggaaag attttgtcca
gaagaattca ccgtgtgtac tgagtgcagt ttttttcaca 2940cccgaaagtc
tttactggct ttcatagctt ttctattttt tgcttctatt atttctctct
3000cccttatatt ttactactcg aagaaaggca gaagaagtta caaatagttc
tgaacatttt 3060tctattcatt cattttgaaa taattatgca gacacatcag
ctgttaacca tttgcacctc 3120taagtgttgt gaaactgtaa atttcataca
tttgacttct agaaaacatt tttgtggctt 3180atgacagagg ttttgaaatg
ggcataggtg atcgtaaaat attgaataat gcgaatagtg 3240cctgaatttg
ttctcttttt gggtgattaa aaaactgaca ggcactataa tttctgtaac
3300acactaacaa aagcatgaaa aataggaacc acaccaatgc aacatttgtg
cagaaatttg 3360aatgacaaga ttaggaatat tttcttctgc acccacttct
aaatttaatg tttttctgga 3420agtagtaatt gcaagagttc gaatagaaag
ttatgtacca agtaaccatt tctcagctgc 3480cataataatg cctagtggct
tcccctctgt caaatctagt ttcctatgga aaagaagatg 3540gcagatacag
gagagacgac agagggtcct aggctggaat gttcctttcg aaagcaatgc
3600ttctatcaaa tactagtatt aatttatgta tctggttaat gacatacttg
gagagcaaat 3660tatggaaatg tgtattttat atgatttttg aggtcctgtc
taaaccctgt gtccctgagg 3720gatctgtctc actggcatct tgttgagggc
cttgcacata ggaaactttt gataagtatc 3780tgcggaaaaa caaacatgaa
tcctgtgata ttgggctctt caggaagcat aaagcaattg 3840tgaaatacag
tataccgcag tggctctagg tggaggaaag gaggaaaaag tgcttattat
3900gtgcaacatt atgattaatc tgattataca ccatttttga gcagatcttg
gaatgaatga 3960catgaccttt ccctagagaa taaggatgaa ataatcactc
attctatgaa cagtgacact 4020actttctatt ctttagctgt actgtaattt
ctttgagttg atagttttac aaattcttaa 4080taggttcaaa agcaatctgg
tctgaataac actggatttg tttctgtgat ctctgaggtc 4140tattttatgt
ttttgctgct acttctgtgg aagtagcttt gaactagttt tactttgaac
4200tttcacgctg aaacatgcta gtgatatcta gaaagggcta attaggtctc
atcctttaat 4260gccccttaaa taagtcttgc tgattttcag acagggaagt
ctctctatta cactggagct 4320gttttataga taagtcaata ttgtatcagg
caagataaac caatgtcata acaggcattg 4380ccaacctcac tgacacaggg
tcatagtgta taataatata ctgtactata taatatatca 4440tctttagagg
tatgattttt tcatgaaaga taagcttttg gtaatattca ttttaaagtg
4500gacttattaa aattggatgc tagagaatca agtttatttt atgtatatat
ttttctgatt 4560ataagagtaa tatatgttca ttgtaaaaat ttttaaaaca
cagaaactat atgcaaagaa 4620aaaataaaaa ttatctataa tctcagaacc
cagaaatagc cactattaac atttcctacg 4680tattttattt tacatagatc
atattgtata tagttagtat ctttattaat ttttattatg 4740aaactttcct
ttgtcattat tagtcttcaa aagcatgatt tttaatagtt gttgagtatt
4800ccaccacagg aatgtatcac aacttaaccg ttcccgtttg ttagactagt
ttcttattaa 4860tgttgatgaa tgttgtttaa aaataatttt gttgctacat
ttactttaat ttccttgact 4920gtaaagagaa gtaattttgc tccttgataa
agtattatat taataataaa tctgcctgca 4980actttttgcc ttctttcata atc
500321012PRTHomo sapiens 2Met Pro Ala Ser Ala Pro Pro Arg Arg Pro
Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu Val Leu Leu
Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro Gly Asp Gly
Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro Ala Pro Glu
Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55 60 Phe Leu
Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp 65 70 75 80
Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His His 85
90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu
Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val Ala
Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr Glu Ala Leu
Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser Ile Ser Trp
Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly Val Pro Asn
Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu Glu Arg Leu
Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185 190 His Trp
Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala 195 200 205
Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe 210
215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn
Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg
Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu Gly Tyr Leu
Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys Val Trp His
Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly Gly Gln Val
Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro Arg Arg Met
Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310 315 320 Asp
Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp 325 330
335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe
340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe
Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu
Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser Pro Asn Leu
Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu Phe Asn His
Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe Val Ser Gly
Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430 Tyr Leu Lys
Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435 440 445 Gly
Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe 450 455
460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp
465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro Lys Ser Ser
Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn Gly Phe Pro
Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Thr Phe Pro Cys
Asp Phe Ala Trp Gly Val Val 515 520 525 Asp Asn Tyr Ile Gln Val Asp
Thr Thr Leu Ser Gln Phe Thr Asp Leu 530 535 540 Asn Val Tyr Leu Trp
Asp Val His His Ser Lys Arg Leu Ile Lys Val 545 550 555 560 Asp Gly
Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala 565 570 575
Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val Thr His 580
585 590 Phe Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn
Gln 595 600 605 Ser Gln Val Asn His Thr Ile Leu Gln Tyr Tyr Arg Cys
Met Ala Ser 610 615 620 Glu Leu Val Arg Val Asn Ile Thr Pro Val Val
Ala Leu Trp Gln Pro 625 630 635 640 Met Ala Pro Asn Gln Gly Leu Pro
Arg Leu Leu Ala Arg Gln Gly Ala 645 650 655 Trp Glu Asn Pro Tyr Thr
Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660 665 670 Cys Phe Gln Glu
Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn 675 680 685 Glu Pro
Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu 690 695 700
Lys Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe Arg His 705
710 715 720 Ala Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp
Ile Glu 725 730 735 Pro Ala Cys Pro Phe Ser Gln Lys Asp Lys Glu Val
Ala Glu Arg Val 740 745 750 Leu Glu Phe Asp Ile Gly Trp Leu Ala Glu
Pro Ile Phe Gly Ser Gly 755 760 765 Asp Tyr Pro Trp Val Met Arg Asp
Trp Leu Asn Gln Arg Asn Asn Phe 770 775 780 Leu Leu Pro Tyr Phe Thr
Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr 785 790 795 800 Phe Asp Phe
Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser 805 810 815 Glu
Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln Glu 820 825
830 Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val Ala Val Val
835 840 845 Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu Lys Phe Lys
Tyr Gly 850 855 860 Asp Leu Pro Met Tyr Ile Ile Ser Asn Gly Ile Asp
Asp Gly Leu His 865 870 875 880 Ala Glu Asp Asp Gln Leu Arg Val Tyr
Tyr Met Gln Asn Tyr Ile Asn 885 890 895 Glu Ala Leu Lys Ala His Ile
Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900 905 910 Phe Ala Tyr Ser Phe
Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr 915 920 925 Arg Tyr Ala
Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr 930 935 940 Arg
Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu Glu 945 950
955 960 Arg Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu Cys Ser Phe
Phe 965 970 975 His Thr Arg Lys Ser Leu Leu Ala Phe Ile Ala Phe Leu
Phe Phe Ala 980 985 990 Ser Ile Ile Ser Leu Ser Leu Ile Phe Tyr Tyr
Ser Lys Lys Gly Arg 995 1000 1005 Arg Ser Tyr Lys 1010 33279DNAHomo
sapiens 3atcctcagtc tcccagttca agctaatcat tgacagagct ttacaatcac
aagcttttac 60tgaagctttg ataagacagt ccagcagttg gtggcaaatg aagccaggct
gtgcggcagg 120atctccaggg aatgaatgga ttttcttcag cactgatgaa
ataaccacac gctataggaa 180tacaatgtcc aacgggggat tgcaaagatc
tgtcatcctg tcagcactta ttctgctacg 240agctgttact ggattctctg
gagatggaag agctatatgg tctaaaaatc ctaattttac 300tccggtaaat
gaaagtcagc tgtttctcta tgacactttc cctaaaaact ttttctgggg
360tattgggact ggagcattgc aagtggaagg gagttggaag aaggatggaa
aaggaccttc 420tatatgggat catttcatcc acacacacct taaaaatgtc
agcagcacga atggttccag 480tgacagttat atttttctgg aaaaagactt
atcagccctg gattttatag gagtttcttt 540ttatcaattt tcaatttcct
ggccaaggct tttccccgat ggaatagtaa cagttgccaa 600cgcaaaaggt
ctgcagtact acagtactct tctggacgct ctagtgctta gaaacattga
660acctatagtt actttatacc actgggattt gcctttggca ctacaagaaa
aatatggggg 720gtggaaaaat gataccataa tagatatctt caatgactat
gccacatact gtttccagat 780gtttggggac cgtgtcaaat attggattac
aattcacaac ccatatctag tggcttggca 840tgggtatggg acaggtatgc
atgcccctgg agagaaggga aatttagcag ctgtctacac 900tgtgggacac
aacttgatca aggctcactc gaaagtttgg cataactaca acacacattt
960ccgcccacat cagaagggtt ggttatcgat cacgttggga tctcattgga
tcgagccaaa 1020ccggtcggaa aacacgatgg atatattcaa atgtcaacaa
tccatggttt ctgtgcttgg 1080atggtttgcc aaccctatcc atggggatgg
cgactatcca gaggggatga gaaagaagtt 1140gttctccgtt ctacccattt
tctctgaagc agagaagcat gagatgagag gcacagctga 1200tttctttgcc
ttttcttttg gacccaacaa cttcaagccc ctaaacacca tggctaaaat
1260gggacaaaat gtttcactta atttaagaga agcgctgaac tggattaaac
tggaatacaa 1320caaccctcga atcttgattg ctgagaatgg ctggttcaca
gacagtcgtg tgaaaacaga 1380agacaccacg gccatctaca tgatgaagaa
tttcctcagc caggtgcttc aagcaataag 1440gttagatgaa atacgagtgt
ttggttatac tgcctggtct ctcctggatg gctttgaatg 1500gcaggatgct
tacaccatcc gccgaggatt attttatgtg gattttaaca gtaaacagaa
1560agagcggaaa cctaagtctt cagcacacta ctacaaacag atcatacgag
aaaatggttt 1620ttctttaaaa gagtccacgc cagatgtgca gggccagttt
ccctgtgact tctcctgggg 1680tgtcactgaa tctgttctta agcccgagtc
tgtggcttcg tccccacagt tcagcgatcc 1740tcatctgtac gtgtggaacg
ccactggcaa cagactgttg caccgagtgg aaggggtgag 1800gctgaaaaca
cgacccgctc aatgcacaga ttttgtaaac atcaaaaaac aacttgagat
1860gttggcaaga atgaaagtca cccactaccg gtttgctctg gattgggcct
cggtccttcc 1920cactggcaac ctgtccgcgg tgaaccgaca ggccctgagg
tactacaggt gcgtggtcag 1980tgaggggctg aagcttggca tctccgcgat
ggtcaccctg tattatccga cccacgccca 2040cctaggcctc cccgagcctc
tgttgcatgc cgacgggtgg ctgaacccat cgacggccga 2100ggccttccag
gcctacgctg ggctgtgctt ccaggagctg ggggacctgg tgaagctctg
2160gatcaccatc aacgagccta accggctaag tgacatctac aaccgctctg
gcaacgacac 2220ctacggggcg gcgcacaacc tgctggtggc ccacgccctg
gcctggcgcc tctacgaccg 2280gcagttcagg ccctcacagc gcggggccgt
gtcgctgtcg ctgcacgcgg actgggcgga 2340acccgccaac ccctatgctg
actcgcactg gagggcggcc gagcgcttcc tgcagttcga 2400gatcgcctgg
ttcgccgagc cgctcttcaa gaccggggac taccccgcgg ccatgaggga
2460atacattgcc tccaagcacc gacgggggct ttccagctcg gccctgccgc
gcctcaccga 2520ggccgaaagg aggctgctca agggcacggt cgacttctgc
gcgctcaacc acttcaccac 2580taggttcgtg atgcacgagc agctggccgg
cagccgctac gactcggaca gggacatcca 2640gtttctgcag gacatcaccc
gcctgagctc ccccacgcgc ctggctgtga ttccctgggg 2700ggtgcgcaag
ctgctgcggt gggtccggag gaactacggc gacatggaca tttacatcac
2760cgccagtggc atcgacgacc aggctctgga ggatgaccgg ctccggaagt
actacctagg 2820gaagtacctt caggaggtgc tgaaagcata cctgattgat
aaagtcagaa tcaaaggcta 2880ttatgcattc aaactggctg aagagaaatc
taaacccaga tttggattct tcacatctga 2940ttttaaagct aaatcctcaa
tacaatttta caacaaagtg atcagcagca ggggcttccc 3000ttttgagaac
agtagttcta gatgcagtca gacccaagaa aatacagagt gcactgtctg
3060cttattcctt gtgcagaaga aaccactgat attcctgggt tgttgcttct
tctccaccct 3120ggttctactc ttatcaattg ccatttttca aaggcagaag
agaagaaagt tttggaaagc 3180aaaaaactta caacacatac cattaaagaa
aggcaagaga gttgttagct aaactgatct 3240gtctgcatga tagacagttt
aaaaattcat cccagttcc 327941044PRTHomo sapiens 4Met Lys Pro Gly Cys
Ala Ala Gly Ser
Pro Gly Asn Glu Trp Ile Phe 1 5 10 15 Phe Ser Thr Asp Glu Ile Thr
Thr Arg Tyr Arg Asn Thr Met Ser Asn 20 25 30 Gly Gly Leu Gln Arg
Ser Val Ile Leu Ser Ala Leu Ile Leu Leu Arg 35 40 45 Ala Val Thr
Gly Phe Ser Gly Asp Gly Arg Ala Ile Trp Ser Lys Asn 50 55 60 Pro
Asn Phe Thr Pro Val Asn Glu Ser Gln Leu Phe Leu Tyr Asp Thr 65 70
75 80 Phe Pro Lys Asn Phe Phe Trp Gly Ile Gly Thr Gly Ala Leu Gln
Val 85 90 95 Glu Gly Ser Trp Lys Lys Asp Gly Lys Gly Pro Ser Ile
Trp Asp His 100 105 110 Phe Ile His Thr His Leu Lys Asn Val Ser Ser
Thr Asn Gly Ser Ser 115 120 125 Asp Ser Tyr Ile Phe Leu Glu Lys Asp
Leu Ser Ala Leu Asp Phe Ile 130 135 140 Gly Val Ser Phe Tyr Gln Phe
Ser Ile Ser Trp Pro Arg Leu Phe Pro 145 150 155 160 Asp Gly Ile Val
Thr Val Ala Asn Ala Lys Gly Leu Gln Tyr Tyr Ser 165 170 175 Thr Leu
Leu Asp Ala Leu Val Leu Arg Asn Ile Glu Pro Ile Val Thr 180 185 190
Leu Tyr His Trp Asp Leu Pro Leu Ala Leu Gln Glu Lys Tyr Gly Gly 195
200 205 Trp Lys Asn Asp Thr Ile Ile Asp Ile Phe Asn Asp Tyr Ala Thr
Tyr 210 215 220 Cys Phe Gln Met Phe Gly Asp Arg Val Lys Tyr Trp Ile
Thr Ile His 225 230 235 240 Asn Pro Tyr Leu Val Ala Trp His Gly Tyr
Gly Thr Gly Met His Ala 245 250 255 Pro Gly Glu Lys Gly Asn Leu Ala
Ala Val Tyr Thr Val Gly His Asn 260 265 270 Leu Ile Lys Ala His Ser
Lys Val Trp His Asn Tyr Asn Thr His Phe 275 280 285 Arg Pro His Gln
Lys Gly Trp Leu Ser Ile Thr Leu Gly Ser His Trp 290 295 300 Ile Glu
Pro Asn Arg Ser Glu Asn Thr Met Asp Ile Phe Lys Cys Gln 305 310 315
320 Gln Ser Met Val Ser Val Leu Gly Trp Phe Ala Asn Pro Ile His Gly
325 330 335 Asp Gly Asp Tyr Pro Glu Gly Met Arg Lys Lys Leu Phe Ser
Val Leu 340 345 350 Pro Ile Phe Ser Glu Ala Glu Lys His Glu Met Arg
Gly Thr Ala Asp 355 360 365 Phe Phe Ala Phe Ser Phe Gly Pro Asn Asn
Phe Lys Pro Leu Asn Thr 370 375 380 Met Ala Lys Met Gly Gln Asn Val
Ser Leu Asn Leu Arg Glu Ala Leu 385 390 395 400 Asn Trp Ile Lys Leu
Glu Tyr Asn Asn Pro Arg Ile Leu Ile Ala Glu 405 410 415 Asn Gly Trp
Phe Thr Asp Ser Arg Val Lys Thr Glu Asp Thr Thr Ala 420 425 430 Ile
Tyr Met Met Lys Asn Phe Leu Ser Gln Val Leu Gln Ala Ile Arg 435 440
445 Leu Asp Glu Ile Arg Val Phe Gly Tyr Thr Ala Trp Ser Leu Leu Asp
450 455 460 Gly Phe Glu Trp Gln Asp Ala Tyr Thr Ile Arg Arg Gly Leu
Phe Tyr 465 470 475 480 Val Asp Phe Asn Ser Lys Gln Lys Glu Arg Lys
Pro Lys Ser Ser Ala 485 490 495 His Tyr Tyr Lys Gln Ile Ile Arg Glu
Asn Gly Phe Ser Leu Lys Glu 500 505 510 Ser Thr Pro Asp Val Gln Gly
Gln Phe Pro Cys Asp Phe Ser Trp Gly 515 520 525 Val Thr Glu Ser Val
Leu Lys Pro Glu Ser Val Ala Ser Ser Pro Gln 530 535 540 Phe Ser Asp
Pro His Leu Tyr Val Trp Asn Ala Thr Gly Asn Arg Leu 545 550 555 560
Leu His Arg Val Glu Gly Val Arg Leu Lys Thr Arg Pro Ala Gln Cys 565
570 575 Thr Asp Phe Val Asn Ile Lys Lys Gln Leu Glu Met Leu Ala Arg
Met 580 585 590 Lys Val Thr His Tyr Arg Phe Ala Leu Asp Trp Ala Ser
Val Leu Pro 595 600 605 Thr Gly Asn Leu Ser Ala Val Asn Arg Gln Ala
Leu Arg Tyr Tyr Arg 610 615 620 Cys Val Val Ser Glu Gly Leu Lys Leu
Gly Ile Ser Ala Met Val Thr 625 630 635 640 Leu Tyr Tyr Pro Thr His
Ala His Leu Gly Leu Pro Glu Pro Leu Leu 645 650 655 His Ala Asp Gly
Trp Leu Asn Pro Ser Thr Ala Glu Ala Phe Gln Ala 660 665 670 Tyr Ala
Gly Leu Cys Phe Gln Glu Leu Gly Asp Leu Val Lys Leu Trp 675 680 685
Ile Thr Ile Asn Glu Pro Asn Arg Leu Ser Asp Ile Tyr Asn Arg Ser 690
695 700 Gly Asn Asp Thr Tyr Gly Ala Ala His Asn Leu Leu Val Ala His
Ala 705 710 715 720 Leu Ala Trp Arg Leu Tyr Asp Arg Gln Phe Arg Pro
Ser Gln Arg Gly 725 730 735 Ala Val Ser Leu Ser Leu His Ala Asp Trp
Ala Glu Pro Ala Asn Pro 740 745 750 Tyr Ala Asp Ser His Trp Arg Ala
Ala Glu Arg Phe Leu Gln Phe Glu 755 760 765 Ile Ala Trp Phe Ala Glu
Pro Leu Phe Lys Thr Gly Asp Tyr Pro Ala 770 775 780 Ala Met Arg Glu
Tyr Ile Ala Ser Lys His Arg Arg Gly Leu Ser Ser 785 790 795 800 Ser
Ala Leu Pro Arg Leu Thr Glu Ala Glu Arg Arg Leu Leu Lys Gly 805 810
815 Thr Val Asp Phe Cys Ala Leu Asn His Phe Thr Thr Arg Phe Val Met
820 825 830 His Glu Gln Leu Ala Gly Ser Arg Tyr Asp Ser Asp Arg Asp
Ile Gln 835 840 845 Phe Leu Gln Asp Ile Thr Arg Leu Ser Ser Pro Thr
Arg Leu Ala Val 850 855 860 Ile Pro Trp Gly Val Arg Lys Leu Leu Arg
Trp Val Arg Arg Asn Tyr 865 870 875 880 Gly Asp Met Asp Ile Tyr Ile
Thr Ala Ser Gly Ile Asp Asp Gln Ala 885 890 895 Leu Glu Asp Asp Arg
Leu Arg Lys Tyr Tyr Leu Gly Lys Tyr Leu Gln 900 905 910 Glu Val Leu
Lys Ala Tyr Leu Ile Asp Lys Val Arg Ile Lys Gly Tyr 915 920 925 Tyr
Ala Phe Lys Leu Ala Glu Glu Lys Ser Lys Pro Arg Phe Gly Phe 930 935
940 Phe Thr Ser Asp Phe Lys Ala Lys Ser Ser Ile Gln Phe Tyr Asn Lys
945 950 955 960 Val Ile Ser Ser Arg Gly Phe Pro Phe Glu Asn Ser Ser
Ser Arg Cys 965 970 975 Ser Gln Thr Gln Glu Asn Thr Glu Cys Thr Val
Cys Leu Phe Leu Val 980 985 990 Gln Lys Lys Pro Leu Ile Phe Leu Gly
Cys Cys Phe Phe Ser Thr Leu 995 1000 1005 Val Leu Leu Leu Ser Ile
Ala Ile Phe Gln Arg Gln Lys Arg Arg 1010 1015 1020 Lys Phe Trp Lys
Ala Lys Asn Leu Gln His Ile Pro Leu Lys Lys 1025 1030 1035 Gly Lys
Arg Val Val Ser 1040 5449PRTHomo sapiens 5Gln Gly Thr Phe Pro Asp
Gly Phe Leu Trp Ala Val Gly Ser Ala Ala 1 5 10 15 Tyr Gln Thr Glu
Gly Gly Trp Gln Gln His Gly Lys Gly Ala Ser Ile 20 25 30 Trp Asp
Thr Phe Thr His His Pro Leu Ala Pro Pro Gly Asp Ser Arg 35 40 45
Asn Ala Ser Leu Pro Leu Gly Ala Pro Ser Pro Leu Gln Pro Ala Thr 50
55 60 Gly Asp Val Ala Ser Asp Ser Tyr Asn Asn Val Phe Arg Asp Thr
Glu 65 70 75 80 Ala Leu Arg Glu Leu Gly Val Thr His Tyr Arg Phe Ser
Ile Ser Trp 85 90 95 Ala Arg Val Leu Pro Asn Gly Ser Ala Gly Val
Pro Asn Arg Glu Gly 100 105 110 Leu Arg Tyr Tyr Arg Arg Leu Leu Glu
Arg Leu Arg Glu Leu Gly Val 115 120 125 Gln Pro Val Val Thr Leu Tyr
His Trp Asp Leu Pro Gln Arg Leu Gln 130 135 140 Asp Ala Tyr Gly Gly
Trp Ala Asn Arg Ala Leu Ala Asp His Phe Arg 145 150 155 160 Asp Tyr
Ala Glu Leu Cys Phe Arg His Phe Gly Gly Gln Val Lys Tyr 165 170 175
Trp Ile Thr Ile Asp Asn Pro Tyr Val Val Ala Trp His Gly Tyr Ala 180
185 190 Thr Gly Arg Leu Ala Pro Gly Ile Arg Gly Ser Pro Arg Leu Gly
Tyr 195 200 205 Leu Val Ala His Asn Leu Leu Leu Ala His Ala Lys Val
Trp His Leu 210 215 220 Tyr Asn Thr Ser Phe Arg Pro Thr Gln Gly Gly
Gln Val Ser Ile Ala 225 230 235 240 Leu Ser Ser His Trp Ile Asn Pro
Arg Arg Met Thr Asp His Ser Ile 245 250 255 Lys Glu Cys Gln Lys Ser
Leu Asp Phe Val Leu Gly Trp Phe Ala Lys 260 265 270 Pro Val Phe Ile
Asp Gly Asp Tyr Pro Glu Ser Met Lys Asn Asn Leu 275 280 285 Ser Ser
Ile Leu Pro Asp Phe Thr Glu Ser Glu Lys Lys Phe Ile Lys 290 295 300
Gly Thr Ala Asp Phe Phe Ala Leu Cys Phe Gly Pro Thr Leu Ser Phe 305
310 315 320 Gln Leu Leu Asp Pro His Met Lys Phe Arg Gln Leu Glu Ser
Pro Asn 325 330 335 Leu Arg Gln Leu Leu Ser Trp Ile Asp Leu Glu Phe
Asn His Pro Gln 340 345 350 Ile Phe Ile Val Glu Asn Gly Trp Phe Val
Ser Gly Thr Thr Lys Arg 355 360 365 Asp Asp Ala Lys Tyr Met Tyr Tyr
Leu Lys Lys Phe Ile Met Glu Thr 370 375 380 Leu Lys Ala Ile Lys Leu
Asp Gly Val Asp Val Ile Gly Tyr Thr Ala 385 390 395 400 Trp Ser Leu
Met Asp Gly Phe Glu Trp His Arg Gly Tyr Ser Ile Arg 405 410 415 Arg
Gly Leu Phe Tyr Val Asp Phe Leu Ser Gln Asp Lys Met Leu Leu 420 425
430 Pro Lys Ser Ser Ala Leu Phe Tyr Gln Lys Leu Ile Glu Lys Asn Gly
435 440 445 Phe 6437PRTHomo sapiens 6Gly Thr Phe Pro Cys Asp Phe
Ala Trp Gly Val Val Asp Asn Tyr Ile 1 5 10 15 Gln Val Asp Thr Thr
Leu Ser Gln Phe Thr Asp Leu Asn Val Tyr Leu 20 25 30 Trp Asp Val
His His Ser Lys Arg Leu Ile Lys Val Asp Gly Val Val 35 40 45 Thr
Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala Ala Ile Gln Pro 50 55
60 Gln Ile Ala Leu Leu Gln Glu Met His Val Thr His Phe Arg Phe Ser
65 70 75 80 Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln Ser Gln
Val Asn 85 90 95 His Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser
Glu Leu Val Arg 100 105 110 Val Asn Ile Thr Pro Val Val Ala Leu Trp
Gln Pro Met Ala Pro Asn 115 120 125 Gln Gly Leu Pro Arg Leu Leu Ala
Arg Gln Gly Ala Trp Glu Asn Pro 130 135 140 Tyr Thr Ala Leu Ala Phe
Ala Glu Tyr Ala Arg Leu Cys Phe Gln Glu 145 150 155 160 Leu Gly His
His Val Lys Leu Trp Ile Thr Met Asn Glu Pro Tyr Thr 165 170 175 Arg
Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu Lys Ala His Ala 180 185
190 Leu Ala Trp His Val Tyr Asn Glu Lys Phe Arg His Ala Gln Asn Gly
195 200 205 Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu Pro Ala
Cys Pro 210 215 220 Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val
Leu Glu Phe Asp 225 230 235 240 Ile Gly Trp Leu Ala Glu Pro Ile Phe
Gly Ser Gly Asp Tyr Pro Trp 245 250 255 Val Met Arg Asp Trp Leu Asn
Gln Arg Asn Asn Phe Leu Leu Pro Tyr 260 265 270 Phe Thr Glu Asp Glu
Lys Lys Leu Ile Gln Gly Thr Phe Asp Phe Leu 275 280 285 Ala Leu Ser
His Tyr Thr Thr Ile Leu Val Asp Ser Glu Lys Glu Asp 290 295 300 Pro
Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln Glu Met Thr Asp Ile 305 310
315 320 Thr Trp Leu Asn Ser Pro Ser Gln Val Ala Val Val Pro Trp Gly
Leu 325 330 335 Arg Lys Val Leu Asn Trp Leu Lys Phe Lys Tyr Gly Asp
Leu Pro Met 340 345 350 Tyr Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu
His Ala Glu Asp Asp 355 360 365 Gln Leu Arg Val Tyr Tyr Met Gln Asn
Tyr Ile Asn Glu Ala Leu Lys 370 375 380 Ala His Ile Leu Asp Gly Ile
Asn Leu Cys Gly Tyr Phe Ala Tyr Ser 385 390 395 400 Phe Asn Asp Arg
Thr Ala Pro Arg Phe Gly Leu Tyr Arg Tyr Ala Ala 405 410 415 Asp Gln
Phe Glu Pro Lys Ala Ser Met Lys His Tyr Arg Lys Ile Ile 420 425 430
Asp Ser Asn Gly Phe 435 7 949PRTHomo sapiens 7Glu Pro Gly Asp Gly
Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro Pro 1 5 10 15 Ala Pro Glu
Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly Phe 20 25 30 Leu
Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp Gln 35 40
45 Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His His Pro
50 55 60 Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu
Gly Ala 65 70 75 80 Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val Ala
Ser Asp Ser Tyr 85 90 95 Asn Asn Val Phe Arg Asp Thr Glu Ala Leu
Arg Glu Leu Gly Val Thr 100 105 110 His Tyr Arg Phe Ser Ile Ser Trp
Ala Arg Val Leu Pro Asn Gly Ser 115 120 125 Ala Gly Val Pro Asn Arg
Glu Gly Leu Arg Tyr Tyr Arg Arg Leu Leu 130 135 140 Glu Arg Leu Arg
Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr His 145 150 155 160 Trp
Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala Asn 165 170
175 Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe Arg
180 185 190 His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn
Pro Tyr 195 200 205 Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu
Ala Pro Gly Ile 210 215 220 Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val
Ala His Asn Leu Leu Leu 225 230 235 240 Ala His Ala Lys Val Trp His
Leu Tyr Asn Thr Ser Phe Arg Pro Thr 245 250 255 Gln Gly Gly Gln Val
Ser Ile Ala Leu Ser Ser His Trp Ile Asn Pro 260 265 270 Arg Arg Met
Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu Asp 275 280 285 Phe
Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp Tyr 290 295
300 Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe Thr
305 310 315 320 Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe
Phe Ala Leu 325 330 335 Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu
Asp Pro His Met Lys 340 345 350 Phe Arg Gln Leu Glu Ser Pro Asn Leu
Arg Gln Leu Leu Ser Trp Ile 355 360 365 Asp
Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly Trp 370 375
380 Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr Tyr
385 390 395 400 Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys
Leu Asp Gly 405 410 415 Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu
Met Asp Gly Phe Glu 420 425 430 Trp His Arg Gly Tyr Ser Ile Arg Arg
Gly Leu Phe Tyr Val Asp Phe 435 440 445 Leu Ser Gln Asp Lys Met Leu
Leu Pro Lys Ser Ser Ala Leu Phe Tyr 450 455 460 Gln Lys Leu Ile Glu
Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn Gln 465 470 475 480 Pro Leu
Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val Asp 485 490 495
Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu Asn 500
505 510 Val Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val
Asp 515 520 525 Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp
Phe Ala Ala 530 535 540 Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met
His Val Thr His Phe 545 550 555 560 Arg Phe Ser Leu Asp Trp Ala Leu
Ile Leu Pro Leu Gly Asn Gln Ser 565 570 575 Gln Val Asn His Thr Ile
Leu Gln Tyr Tyr Arg Cys Met Ala Ser Glu 580 585 590 Leu Val Arg Val
Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro Met 595 600 605 Ala Pro
Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala Trp 610 615 620
Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu Cys 625
630 635 640 Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met
Asn Glu 645 650 655 Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His
Asn Leu Leu Lys 660 665 670 Ala His Ala Leu Ala Trp His Val Tyr Asn
Glu Lys Phe Arg His Ala 675 680 685 Gln Asn Gly Lys Ile Ser Ile Ala
Leu Gln Ala Asp Trp Ile Glu Pro 690 695 700 Ala Cys Pro Phe Ser Gln
Lys Asp Lys Glu Val Ala Glu Arg Val Leu 705 710 715 720 Glu Phe Asp
Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly Asp 725 730 735 Tyr
Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe Leu 740 745
750 Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr Phe
755 760 765 Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp
Ser Glu 770 775 780 Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu
Val Gln Glu Met 785 790 795 800 Thr Asp Ile Thr Trp Leu Asn Ser Pro
Ser Gln Val Ala Val Val Pro 805 810 815 Trp Gly Leu Arg Lys Val Leu
Asn Trp Leu Lys Phe Lys Tyr Gly Asp 820 825 830 Leu Pro Met Tyr Ile
Ile Ser Asn Gly Ile Asp Asp Gly Leu His Ala 835 840 845 Glu Asp Asp
Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn Glu 850 855 860 Ala
Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr Phe 865 870
875 880 Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr
Arg 885 890 895 Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys
His Tyr Arg 900 905 910 Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro
Glu Thr Leu Glu Arg 915 920 925 Phe Cys Pro Glu Glu Phe Thr Val Cys
Thr Glu Cys Ser Phe Phe His 930 935 940 Thr Arg Lys Ser Leu 945
833PRTHomo sapiens 8Met Pro Ala Ser Ala Pro Pro Arg Arg Pro Arg Pro
Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu Val Leu Leu Gly Leu
Gly Gly Arg Arg Leu Arg 20 25 30 Ala 925PRTHomo sapiens 9Met Ser
Val Leu Thr Gln Val Leu Ala Leu Leu Leu Leu Trp Leu Thr 1 5 10 15
Gly Thr Arg Cys Arg Arg Leu Arg Ala 20 25 1045DNAHomo sapiens
10ggaggtggag gttcaggagg tggaggttca ggaggtggag gttca 451115PRTHomo
sapiens 11Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser 1 5 10 15 125PRTHomo sapiens 12Gly Gly Gly Gly Ser 1 5
131PRTHomo sapiens 13Gly 1 142PRTHomo sapiens 14Gly Gly 1
152PRTHomo sapiens 15Gly Ser 1 163PRTHomo sapiens 16Gly Gly Ser 1
171PRTHomo sapiens 17Ala 1 182PRTHomo sapiens 18Ala Ala 1
191228PRTHomo sapiens 19Met Pro Ala Ser Ala Pro Pro Arg Arg Pro Arg
Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu Val Leu Leu Gly
Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro Gly Asp Gly Ala
Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro Ala Pro Glu Ala
Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55 60 Phe Leu Trp
Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp 65 70 75 80 Gln
Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His His 85 90
95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu Gly
100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val Ala Ser
Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr Glu Ala Leu Arg
Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser Ile Ser Trp Ala
Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly Val Pro Asn Arg
Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu Glu Arg Leu Arg
Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185 190 His Trp Asp
Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala 195 200 205 Asn
Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe 210 215
220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro
225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu
Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val
Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys Val Trp His Leu
Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly Gly Gln Val Ser
Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro Arg Arg Met Thr
Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310 315 320 Asp Phe
Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp 325 330 335
Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe 340
345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe
Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp
Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser Pro Asn Leu Arg
Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu Phe Asn His Pro
Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe Val Ser Gly Thr
Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430 Tyr Leu Lys Lys
Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435 440 445 Gly Val
Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe 450 455 460
Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp 465
470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro Lys Ser Ser Ala
Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn Gly Phe Pro Pro
Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Thr Phe Pro Cys Asp
Phe Ala Trp Gly Val Val 515 520 525 Asp Asn Tyr Ile Gln Val Asp Thr
Thr Leu Ser Gln Phe Thr Asp Leu 530 535 540 Asn Val Tyr Leu Trp Asp
Val His His Ser Lys Arg Leu Ile Lys Val 545 550 555 560 Asp Gly Val
Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala 565 570 575 Ala
Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val Thr His 580 585
590 Phe Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln
595 600 605 Ser Gln Val Asn His Thr Ile Leu Gln Tyr Tyr Arg Cys Met
Ala Ser 610 615 620 Glu Leu Val Arg Val Asn Ile Thr Pro Val Val Ala
Leu Trp Gln Pro 625 630 635 640 Met Ala Pro Asn Gln Gly Leu Pro Arg
Leu Leu Ala Arg Gln Gly Ala 645 650 655 Trp Glu Asn Pro Tyr Thr Ala
Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660 665 670 Cys Phe Gln Glu Leu
Gly His His Val Lys Leu Trp Ile Thr Met Asn 675 680 685 Glu Pro Tyr
Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu 690 695 700 Lys
Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe Arg His 705 710
715 720 Ala Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp Ile
Glu 725 730 735 Pro Ala Cys Pro Phe Ser Gln Lys Asp Lys Glu Val Ala
Glu Arg Val 740 745 750 Leu Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro
Ile Phe Gly Ser Gly 755 760 765 Asp Tyr Pro Trp Val Met Arg Asp Trp
Leu Asn Gln Arg Asn Asn Phe 770 775 780 Leu Leu Pro Tyr Phe Thr Glu
Asp Glu Lys Lys Leu Ile Gln Gly Thr 785 790 795 800 Phe Asp Phe Leu
Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser 805 810 815 Glu Lys
Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln Glu 820 825 830
Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val Ala Val Val 835
840 845 Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu Lys Phe Lys Tyr
Gly 850 855 860 Asp Leu Pro Met Tyr Ile Ile Ser Asn Gly Ile Asp Asp
Gly Leu His 865 870 875 880 Ala Glu Asp Asp Gln Leu Arg Val Tyr Tyr
Met Gln Asn Tyr Ile Asn 885 890 895 Glu Ala Leu Lys Ala His Ile Leu
Asp Gly Ile Asn Leu Cys Gly Tyr 900 905 910 Phe Ala Tyr Ser Phe Asn
Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr 915 920 925 Arg Tyr Ala Ala
Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr 930 935 940 Arg Lys
Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu Glu 945 950 955
960 Arg Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu Cys Ser Phe Phe
965 970 975 His Thr Arg Lys Ser Leu Gly Ser Gly Gly Gly Gly Ser Gly
Gly Gly 980 985 990 Gly Ser Gly Gly Gly Gly Ser Leu Lys Tyr Pro Asn
Ala Ser Pro Leu 995 1000 1005 Leu Gly Ser Ser Trp Gly Gly Leu Ile
His Leu Tyr Thr Ala Thr 1010 1015 1020 Ala Arg Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val 1025 1030 1035 Asp Gly Ala Pro His
Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg 1040 1045 1050 Ser Glu Asp
Ala Gly Phe Val Val Ile Thr Gly Val Met Ser Arg 1055 1060 1065 Arg
Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly Ser His 1070 1075
1080 Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His Gln Thr Leu Glu
1085 1090 1095 Asn Gly Tyr Asp Val Tyr His Ser Pro Gln Tyr His Phe
Leu Val 1100 1105 1110 Ser Leu Gly Arg Ala Lys Arg Ala Phe Leu Pro
Gly Met Asn Pro 1115 1120 1125 Pro Pro Tyr Ser Gln Phe Leu Ser Arg
Arg Asn Glu Ile Pro Leu 1130 1135 1140 Ile His Phe Asn Thr Pro Ile
Pro Arg Arg His Thr Gln Ser Ala 1145 1150 1155 Glu Asp Asp Ser Glu
Arg Asp Pro Leu Asn Val Leu Lys Pro Arg 1160 1165 1170 Ala Arg Met
Thr Pro Ala Pro Ala Ser Cys Ser Gln Glu Leu Pro 1175 1180 1185 Ser
Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu Gly Val 1190 1195
1200 Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly Pro
1205 1210 1215 Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile 1220 1225 20
1220PRTHomo sapiens 20Met Ser Val Leu Thr Gln Val Leu Ala Leu Leu
Leu Leu Trp Leu Thr 1 5 10 15 Gly Leu Gly Gly Arg Arg Leu Arg Ala
Glu Pro Gly Asp Gly Ala Gln 20 25 30 Thr Trp Ala Arg Phe Ser Arg
Pro Pro Ala Pro Glu Ala Ala Gly Leu 35 40 45 Phe Gln Gly Thr Phe
Pro Asp Gly Phe Leu Trp Ala Val Gly Ser Ala 50 55 60 Ala Tyr Gln
Thr Glu Gly Gly Trp Gln Gln His Gly Lys Gly Ala Ser 65 70 75 80 Ile
Trp Asp Thr Phe Thr His His Pro Leu Ala Pro Pro Gly Asp Ser 85 90
95 Arg Asn Ala Ser Leu Pro Leu Gly Ala Pro Ser Pro Leu Gln Pro Ala
100 105 110 Thr Gly Asp Val Ala Ser Asp Ser Tyr Asn Asn Val Phe Arg
Asp Thr 115 120 125 Glu Ala Leu Arg Glu Leu Gly Val Thr His Tyr Arg
Phe Ser Ile Ser 130 135 140 Trp Ala Arg Val Leu Pro Asn Gly Ser Ala
Gly Val Pro Asn Arg Glu 145 150 155 160 Gly Leu Arg Tyr Tyr Arg Arg
Leu Leu Glu Arg Leu Arg Glu Leu Gly 165 170 175 Val Gln Pro Val Val
Thr Leu Tyr His Trp Asp Leu Pro Gln Arg Leu 180 185 190 Gln Asp Ala
Tyr Gly Gly Trp Ala Asn Arg Ala Leu Ala Asp His Phe 195 200 205 Arg
Asp Tyr Ala Glu Leu Cys Phe Arg His Phe Gly Gly Gln Val Lys 210 215
220 Tyr Trp Ile Thr Ile Asp Asn Pro Tyr Val Val Ala Trp His Gly Tyr
225 230 235 240 Ala Thr Gly Arg Leu Ala Pro Gly Ile Arg Gly Ser Pro
Arg Leu Gly 245 250 255 Tyr Leu Val Ala His Asn Leu Leu Leu Ala His
Ala Lys Val Trp His 260 265 270 Leu Tyr Asn Thr Ser Phe Arg Pro Thr
Gln Gly Gly Gln Val Ser Ile 275 280 285 Ala Leu Ser Ser His Trp Ile
Asn Pro Arg Arg Met Thr Asp His Ser 290 295 300 Ile Lys Glu Cys Gln
Lys Ser Leu Asp Phe Val Leu Gly Trp Phe Ala 305 310 315 320 Lys Pro
Val Phe Ile Asp Gly Asp Tyr Pro Glu Ser Met Lys Asn Asn 325 330 335
Leu Ser Ser Ile Leu Pro Asp Phe Thr Glu Ser Glu Lys Lys Phe Ile 340
345 350 Lys Gly
Thr Ala Asp Phe Phe Ala Leu Cys Phe Gly Pro Thr Leu Ser 355 360 365
Phe Gln Leu Leu Asp Pro His Met Lys Phe Arg Gln Leu Glu Ser Pro 370
375 380 Asn Leu Arg Gln Leu Leu Ser Trp Ile Asp Leu Glu Phe Asn His
Pro 385 390 395 400 Gln Ile Phe Ile Val Glu Asn Gly Trp Phe Val Ser
Gly Thr Thr Lys 405 410 415 Arg Asp Asp Ala Lys Tyr Met Tyr Tyr Leu
Lys Lys Phe Ile Met Glu 420 425 430 Thr Leu Lys Ala Ile Lys Leu Asp
Gly Val Asp Val Ile Gly Tyr Thr 435 440 445 Ala Trp Ser Leu Met Asp
Gly Phe Glu Trp His Arg Gly Tyr Ser Ile 450 455 460 Arg Arg Gly Leu
Phe Tyr Val Asp Phe Leu Ser Gln Asp Lys Met Leu 465 470 475 480 Leu
Pro Lys Ser Ser Ala Leu Phe Tyr Gln Lys Leu Ile Glu Lys Asn 485 490
495 Gly Phe Pro Pro Leu Pro Glu Asn Gln Pro Leu Glu Gly Thr Phe Pro
500 505 510 Cys Asp Phe Ala Trp Gly Val Val Asp Asn Tyr Ile Gln Val
Asp Thr 515 520 525 Thr Leu Ser Gln Phe Thr Asp Leu Asn Val Tyr Leu
Trp Asp Val His 530 535 540 His Ser Lys Arg Leu Ile Lys Val Asp Gly
Val Val Thr Lys Lys Arg 545 550 555 560 Lys Ser Tyr Cys Val Asp Phe
Ala Ala Ile Gln Pro Gln Ile Ala Leu 565 570 575 Leu Gln Glu Met His
Val Thr His Phe Arg Phe Ser Leu Asp Trp Ala 580 585 590 Leu Ile Leu
Pro Leu Gly Asn Gln Ser Gln Val Asn His Thr Ile Leu 595 600 605 Gln
Tyr Tyr Arg Cys Met Ala Ser Glu Leu Val Arg Val Asn Ile Thr 610 615
620 Pro Val Val Ala Leu Trp Gln Pro Met Ala Pro Asn Gln Gly Leu Pro
625 630 635 640 Arg Leu Leu Ala Arg Gln Gly Ala Trp Glu Asn Pro Tyr
Thr Ala Leu 645 650 655 Ala Phe Ala Glu Tyr Ala Arg Leu Cys Phe Gln
Glu Leu Gly His His 660 665 670 Val Lys Leu Trp Ile Thr Met Asn Glu
Pro Tyr Thr Arg Asn Met Thr 675 680 685 Tyr Ser Ala Gly His Asn Leu
Leu Lys Ala His Ala Leu Ala Trp His 690 695 700 Val Tyr Asn Glu Lys
Phe Arg His Ala Gln Asn Gly Lys Ile Ser Ile 705 710 715 720 Ala Leu
Gln Ala Asp Trp Ile Glu Pro Ala Cys Pro Phe Ser Gln Lys 725 730 735
Asp Lys Glu Val Ala Glu Arg Val Leu Glu Phe Asp Ile Gly Trp Leu 740
745 750 Ala Glu Pro Ile Phe Gly Ser Gly Asp Tyr Pro Trp Val Met Arg
Asp 755 760 765 Trp Leu Asn Gln Arg Asn Asn Phe Leu Leu Pro Tyr Phe
Thr Glu Asp 770 775 780 Glu Lys Lys Leu Ile Gln Gly Thr Phe Asp Phe
Leu Ala Leu Ser His 785 790 795 800 Tyr Thr Thr Ile Leu Val Asp Ser
Glu Lys Glu Asp Pro Ile Lys Tyr 805 810 815 Asn Asp Tyr Leu Glu Val
Gln Glu Met Thr Asp Ile Thr Trp Leu Asn 820 825 830 Ser Pro Ser Gln
Val Ala Val Val Pro Trp Gly Leu Arg Lys Val Leu 835 840 845 Asn Trp
Leu Lys Phe Lys Tyr Gly Asp Leu Pro Met Tyr Ile Ile Ser 850 855 860
Asn Gly Ile Asp Asp Gly Leu His Ala Glu Asp Asp Gln Leu Arg Val 865
870 875 880 Tyr Tyr Met Gln Asn Tyr Ile Asn Glu Ala Leu Lys Ala His
Ile Leu 885 890 895 Asp Gly Ile Asn Leu Cys Gly Tyr Phe Ala Tyr Ser
Phe Asn Asp Arg 900 905 910 Thr Ala Pro Arg Phe Gly Leu Tyr Arg Tyr
Ala Ala Asp Gln Phe Glu 915 920 925 Pro Lys Ala Ser Met Lys His Tyr
Arg Lys Ile Ile Asp Ser Asn Gly 930 935 940 Phe Pro Gly Pro Glu Thr
Leu Glu Arg Phe Cys Pro Glu Glu Phe Thr 945 950 955 960 Val Cys Thr
Glu Cys Ser Phe Phe His Thr Arg Lys Ser Leu Gly Ser 965 970 975 Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Leu 980 985
990 Lys Tyr Pro Asn Ala Ser Pro Leu Leu Gly Ser Ser Trp Gly Gly Leu
995 1000 1005 Ile His Leu Tyr Thr Ala Thr Ala Arg Asn Ser Tyr His
Leu Gln 1010 1015 1020 Ile His Lys Asn Gly His Val Asp Gly Ala Pro
His Gln Thr Ile 1025 1030 1035 Tyr Ser Ala Leu Met Ile Arg Ser Glu
Asp Ala Gly Phe Val Val 1040 1045 1050 Ile Thr Gly Val Met Ser Arg
Arg Tyr Leu Cys Met Asp Phe Arg 1055 1060 1065 Gly Asn Ile Phe Gly
Ser His Tyr Phe Asp Pro Glu Asn Cys Arg 1070 1075 1080 Phe Gln His
Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His Ser 1085 1090 1095 Pro
Gln Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 1100 1105
1110 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser
1115 1120 1125 Arg Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro
Ile Pro 1130 1135 1140 Arg Arg His Thr Gln Ser Ala Glu Asp Asp Ser
Glu Arg Asp Pro 1145 1150 1155 Leu Asn Val Leu Lys Pro Arg Ala Arg
Met Thr Pro Ala Pro Ala 1160 1165 1170 Ser Cys Ser Gln Glu Leu Pro
Ser Ala Glu Asp Asn Ser Pro Met 1175 1180 1185 Ala Ser Asp Pro Leu
Gly Val Val Arg Gly Gly Arg Val Asn Thr 1190 1195 1200 His Ala Gly
Gly Thr Gly Pro Glu Gly Cys Arg Pro Phe Ala Lys 1205 1210 1215 Phe
Ile 1220 21762PRTHomo sapiens 21Met Pro Ala Ser Ala Pro Pro Arg Arg
Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu Val Leu
Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro Gly Asp
Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro Ala Pro
Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55 60 Phe
Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp 65 70
75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His
His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu
Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp
Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr Glu
Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser Ile
Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly Val
Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu Glu
Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185 190
His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala 195
200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys
Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile
Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala Thr
Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu Gly
Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys Val
Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly Gly
Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro Arg
Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310 315
320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp
325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro
Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala
Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe Gln
Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser Pro
Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu Phe
Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe Val
Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430 Tyr
Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435 440
445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe
450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr
Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro Lys
Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn Gly
Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Ser Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser
Leu Lys Tyr Pro Asn Ala Ser Pro Leu Leu Gly 530 535 540 Ser Ser Trp
Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg Asn 545 550 555 560
Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala Pro 565
570 575 His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala
Gly 580 585 590 Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu
Cys Met Asp 595 600 605 Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe
Asp Pro Glu Asn Cys 610 615 620 Arg Phe Gln His Gln Thr Leu Glu Asn
Gly Tyr Asp Val Tyr His Ser 625 630 635 640 Pro Gln Tyr His Phe Leu
Val Ser Leu Gly Arg Ala Lys Arg Ala Phe 645 650 655 Leu Pro Gly Met
Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg Arg 660 665 670 Asn Glu
Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg His 675 680 685
Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu 690
695 700 Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln
Glu 705 710 715 720 Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu Gly 725 730 735 Val Val Arg Gly Gly Arg Val Asn Thr His
Ala Gly Gly Thr Gly Pro 740 745 750 Glu Gly Cys Arg Pro Phe Ala Lys
Phe Ile 755 760 22752PRTHomo sapiens 22Met Pro Ala Ser Ala Pro Pro
Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu
Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Pro 20 25 30 Leu Pro Glu
Asn Gln Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala 35 40 45 Trp
Gly Val Val Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln 50 55
60 Phe Thr Asp Leu Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg
65 70 75 80 Leu Ile Lys Val Asp Gly Val Val Thr Lys Lys Arg Lys Ser
Tyr Cys 85 90 95 Val Asp Phe Ala Ala Ile Gln Pro Gln Ile Ala Leu
Leu Gln Glu Met 100 105 110 His Val Thr His Phe Arg Phe Ser Leu Asp
Trp Ala Leu Ile Leu Pro 115 120 125 Leu Gly Asn Gln Ser Gln Val Asn
His Thr Ile Leu Gln Tyr Tyr Arg 130 135 140 Cys Met Ala Ser Glu Leu
Val Arg Val Asn Ile Thr Pro Val Val Ala 145 150 155 160 Leu Trp Gln
Pro Met Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala 165 170 175 Arg
Gln Gly Ala Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu 180 185
190 Tyr Ala Arg Leu Cys Phe Gln Glu Leu Gly His His Val Lys Leu Trp
195 200 205 Ile Thr Met Asn Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser
Ala Gly 210 215 220 His Asn Leu Leu Lys Ala His Ala Leu Ala Trp His
Val Tyr Asn Glu 225 230 235 240 Lys Phe Arg His Ala Gln Asn Gly Lys
Ile Ser Ile Ala Leu Gln Ala 245 250 255 Asp Trp Ile Glu Pro Ala Cys
Pro Phe Ser Gln Lys Asp Lys Glu Val 260 265 270 Ala Glu Arg Val Leu
Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile 275 280 285 Phe Gly Ser
Gly Asp Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln 290 295 300 Arg
Asn Asn Phe Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu 305 310
315 320 Ile Gln Gly Thr Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr
Ile 325 330 335 Leu Val Asp Ser Glu Lys Glu Asp Pro Ile Lys Tyr Asn
Asp Tyr Leu 340 345 350 Glu Val Gln Glu Met Thr Asp Ile Thr Trp Leu
Asn Ser Pro Ser Gln 355 360 365 Val Ala Val Val Pro Trp Gly Leu Arg
Lys Val Leu Asn Trp Leu Lys 370 375 380 Phe Lys Tyr Gly Asp Leu Pro
Met Tyr Ile Ile Ser Asn Gly Ile Asp 385 390 395 400 Asp Gly Leu His
Ala Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln 405 410 415 Asn Tyr
Ile Asn Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn 420 425 430
Leu Cys Gly Tyr Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg 435
440 445 Phe Gly Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala
Ser 450 455 460 Met Lys His Tyr Arg Lys Ile Ile Asp Ser Asn Gly Phe
Pro Gly Pro 465 470 475 480 Glu Thr Leu Glu Arg Phe Cys Pro Glu Glu
Phe Thr Val Cys Thr Glu 485 490 495 Cys Ser Phe Phe His Thr Arg Lys
Ser Leu Gly Ser Gly Gly Gly Gly 500 505 510 Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser Leu Lys Tyr Pro Asn 515 520 525 Ala Ser Pro Leu
Leu Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr 530 535 540 Thr Ala
Thr Ala Arg Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly 545 550 555
560 His Val Asp Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile
565 570 575 Arg Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val Met
Ser Arg 580 585 590 Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe
Gly Ser His Tyr 595 600 605 Phe Asp Pro Glu Asn Cys Arg Phe Gln His
Gln Thr Leu Glu Asn Gly 610 615 620 Tyr Asp Val Tyr His Ser Pro Gln
Tyr His Phe Leu Val Ser Leu Gly 625 630 635 640 Arg Ala Lys Arg Ala
Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser 645 650 655 Gln Phe Leu
Ser Arg Arg Asn Glu
Ile Pro Leu Ile His Phe Asn Thr 660 665 670 Pro Ile Pro Arg Arg His
Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg 675 680 685 Asp Pro Leu Asn
Val Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro 690 695 700 Ala Ser
Cys Ser Gln Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met 705 710 715
720 Ala Ser Asp Pro Leu Gly Val Val Arg Gly Gly Arg Val Asn Thr His
725 730 735 Ala Gly Gly Thr Gly Pro Glu Gly Cys Arg Pro Phe Ala Lys
Phe Ile 740 745 750 231215PRTHomo sapiens 23Met Pro Ala Ser Ala Pro
Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu
Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu
Pro Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45
Pro Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50
55 60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly
Trp 65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe
Thr His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala
Ser Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr
Gly Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp
Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe
Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala
Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175
Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180
185 190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp
Ala 195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu
Leu Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile
Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr
Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg
Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala
Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln
Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300
Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305
310 315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp
Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile
Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly
Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser
Phe Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu
Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu
Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp
Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425
430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp
435 440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp
Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu
Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu
Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys
Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly
Ser Gly Thr Phe Pro Asp Gly Phe Leu Trp Ala 515 520 525 Val Gly Ser
Ala Ala Tyr Gln Thr Glu Gly Gly Trp Gln Gln His Gly 530 535 540 Lys
Gly Ala Ser Ile Trp Asp Thr Phe Thr His His Pro Leu Ala Pro 545 550
555 560 Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu Gly Ala Pro Ser
Pro 565 570 575 Leu Gln Pro Ala Thr Gly Asp Val Ala Ser Asp Ser Tyr
Asn Asn Val 580 585 590 Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly
Val Thr His Tyr Arg 595 600 605 Phe Ser Ile Ser Trp Ala Arg Val Leu
Pro Asn Gly Ser Ala Gly Val 610 615 620 Pro Asn Arg Glu Gly Leu Arg
Tyr Tyr Arg Arg Leu Leu Glu Arg Leu 625 630 635 640 Arg Glu Leu Gly
Val Gln Pro Val Val Thr Leu Tyr His Trp Asp Leu 645 650 655 Pro Gln
Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala Asn Arg Ala Leu 660 665 670
Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe Arg His Phe Gly 675
680 685 Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro Tyr Val Val
Ala 690 695 700 Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly Ile
Arg Gly Ser 705 710 715 720 Pro Arg Leu Gly Tyr Leu Val Ala His Asn
Leu Leu Leu Ala His Ala 725 730 735 Lys Val Trp His Leu Tyr Asn Thr
Ser Phe Arg Pro Thr Gln Gly Gly 740 745 750 Gln Val Ser Ile Ala Leu
Ser Ser His Trp Ile Asn Pro Arg Arg Met 755 760 765 Thr Asp His Ser
Ile Lys Glu Cys Gln Lys Ser Leu Asp Phe Val Leu 770 775 780 Gly Trp
Phe Ala Lys Pro Val Phe Ile Asp Gly Asp Tyr Pro Glu Ser 785 790 795
800 Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe Thr Glu Ser Glu
805 810 815 Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala Leu Cys
Phe Gly 820 825 830 Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met
Lys Phe Arg Gln 835 840 845 Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu
Ser Trp Ile Asp Leu Glu 850 855 860 Phe Asn His Pro Gln Ile Phe Ile
Val Glu Asn Gly Trp Phe Val Ser 865 870 875 880 Gly Thr Thr Lys Arg
Asp Asp Ala Lys Tyr Met Tyr Tyr Leu Lys Lys 885 890 895 Phe Ile Met
Glu Thr Leu Lys Ala Ile Lys Leu Asp Gly Val Asp Val 900 905 910 Ile
Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe Glu Trp His Arg 915 920
925 Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp Phe Leu Ser Gln
930 935 940 Asp Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe Tyr Gln
Lys Leu 945 950 955 960 Ile Glu Lys Asn Gly Phe Pro Glu Phe Gly Ser
Gly Gly Gly Gly Ser 965 970 975 Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser Leu Lys Tyr Pro Asn Ala 980 985 990 Ser Pro Leu Leu Gly Ser Ser
Trp Gly Gly Leu Ile His Leu Tyr Thr 995 1000 1005 Ala Thr Ala Arg
Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly 1010 1015 1020 His Val
Asp Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met 1025 1030 1035
Ile Arg Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val Met 1040
1045 1050 Ser Arg Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe
Gly 1055 1060 1065 Ser His Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln
His Gln Thr 1070 1075 1080 Leu Glu Asn Gly Tyr Asp Val Tyr His Ser
Pro Gln Tyr His Phe 1085 1090 1095 Leu Val Ser Leu Gly Arg Ala Lys
Arg Ala Phe Leu Pro Gly Met 1100 1105 1110 Asn Pro Pro Pro Tyr Ser
Gln Phe Leu Ser Arg Arg Asn Glu Ile 1115 1120 1125 Pro Leu Ile His
Phe Asn Thr Pro Ile Pro Arg Arg His Thr Gln 1130 1135 1140 Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys 1145 1150 1155
Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln Glu 1160
1165 1170 Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro
Leu 1175 1180 1185 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala
Gly Gly Thr 1190 1195 1200 Gly Pro Glu Gly Cys Arg Pro Phe Ala Lys
Phe Ile 1205 1210 1215 241189PRTHomo sapiens 24Met Pro Ala Ser Ala
Pro Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu
Leu Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Pro 20 25 30 Leu
Pro Glu Asn Gln Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala 35 40
45 Trp Gly Val Val Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln
50 55 60 Phe Thr Asp Leu Asn Val Tyr Leu Trp Asp Val His His Ser
Lys Arg 65 70 75 80 Leu Ile Lys Val Asp Gly Val Val Thr Lys Lys Arg
Lys Ser Tyr Cys 85 90 95 Val Asp Phe Ala Ala Ile Gln Pro Gln Ile
Ala Leu Leu Gln Glu Met 100 105 110 His Val Thr His Phe Arg Phe Ser
Leu Asp Trp Ala Leu Ile Leu Pro 115 120 125 Leu Gly Asn Gln Ser Gln
Val Asn His Thr Ile Leu Gln Tyr Tyr Arg 130 135 140 Cys Met Ala Ser
Glu Leu Val Arg Val Asn Ile Thr Pro Val Val Ala 145 150 155 160 Leu
Trp Gln Pro Met Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala 165 170
175 Arg Gln Gly Ala Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu
180 185 190 Tyr Ala Arg Leu Cys Phe Gln Glu Leu Gly His His Val Lys
Leu Trp 195 200 205 Ile Thr Met Asn Glu Pro Tyr Thr Arg Asn Met Thr
Tyr Ser Ala Gly 210 215 220 His Asn Leu Leu Lys Ala His Ala Leu Ala
Trp His Val Tyr Asn Glu 225 230 235 240 Lys Phe Arg His Ala Gln Asn
Gly Lys Ile Ser Ile Ala Leu Gln Ala 245 250 255 Asp Trp Ile Glu Pro
Ala Cys Pro Phe Ser Gln Lys Asp Lys Glu Val 260 265 270 Ala Glu Arg
Val Leu Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile 275 280 285 Phe
Gly Ser Gly Asp Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln 290 295
300 Arg Asn Asn Phe Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu
305 310 315 320 Ile Gln Gly Thr Phe Asp Phe Leu Ala Leu Ser His Tyr
Thr Thr Ile 325 330 335 Leu Val Asp Ser Glu Lys Glu Asp Pro Ile Lys
Tyr Asn Asp Tyr Leu 340 345 350 Glu Val Gln Glu Met Thr Asp Ile Thr
Trp Leu Asn Ser Pro Ser Gln 355 360 365 Val Ala Val Val Pro Trp Gly
Leu Arg Lys Val Leu Asn Trp Leu Lys 370 375 380 Phe Lys Tyr Gly Asp
Leu Pro Met Tyr Ile Ile Ser Asn Gly Ile Asp 385 390 395 400 Asp Gly
Leu His Ala Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln 405 410 415
Asn Tyr Ile Asn Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn 420
425 430 Leu Cys Gly Tyr Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro
Arg 435 440 445 Phe Gly Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu Pro
Lys Ala Ser 450 455 460 Met Lys His Tyr Arg Lys Ile Ile Asp Ser Asn
Gly Phe Pro Gly Pro 465 470 475 480 Glu Thr Leu Glu Arg Phe Cys Pro
Glu Glu Phe Thr Val Cys Thr Glu 485 490 495 Cys Ser Phe Phe His Thr
Arg Lys Ser Leu Gly Thr Phe Pro Cys Asp 500 505 510 Phe Ala Trp Gly
Val Val Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu 515 520 525 Ser Gln
Phe Thr Asp Leu Asn Val Tyr Leu Trp Asp Val His His Ser 530 535 540
Lys Arg Leu Ile Lys Val Asp Gly Val Val Thr Lys Lys Arg Lys Ser 545
550 555 560 Tyr Cys Val Asp Phe Ala Ala Ile Gln Pro Gln Ile Ala Leu
Leu Gln 565 570 575 Glu Met His Val Thr His Phe Arg Phe Ser Leu Asp
Trp Ala Leu Ile 580 585 590 Leu Pro Leu Gly Asn Gln Ser Gln Val Asn
His Thr Ile Leu Gln Tyr 595 600 605 Tyr Arg Cys Met Ala Ser Glu Leu
Val Arg Val Asn Ile Thr Pro Val 610 615 620 Val Ala Leu Trp Gln Pro
Met Ala Pro Asn Gln Gly Leu Pro Arg Leu 625 630 635 640 Leu Ala Arg
Gln Gly Ala Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe 645 650 655 Ala
Glu Tyr Ala Arg Leu Cys Phe Gln Glu Leu Gly His His Val Lys 660 665
670 Leu Trp Ile Thr Met Asn Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser
675 680 685 Ala Gly His Asn Leu Leu Lys Ala His Ala Leu Ala Trp His
Val Tyr 690 695 700 Asn Glu Lys Phe Arg His Ala Gln Asn Gly Lys Ile
Ser Ile Ala Leu 705 710 715 720 Gln Ala Asp Trp Ile Glu Pro Ala Cys
Pro Phe Ser Gln Lys Asp Lys 725 730 735 Glu Val Ala Glu Arg Val Leu
Glu Phe Asp Ile Gly Trp Leu Ala Glu 740 745 750 Pro Ile Phe Gly Ser
Gly Asp Tyr Pro Trp Val Met Arg Asp Trp Leu 755 760 765 Asn Gln Arg
Asn Asn Phe Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys 770 775 780 Lys
Leu Ile Gln Gly Thr Phe Asp Phe Leu Ala Leu Ser His Tyr Thr 785 790
795 800 Thr Ile Leu Val Asp Ser Glu Lys Glu Asp Pro Ile Lys Tyr Asn
Asp 805 810 815 Tyr Leu Glu Val Gln Glu Met Thr Asp Ile Thr Trp Leu
Asn Ser Pro 820 825 830 Ser Gln Val Ala Val Val Pro Trp Gly Leu Arg
Lys Val Leu Asn Trp 835 840 845 Leu Lys Phe Lys Tyr Gly Asp Leu Pro
Met Tyr Ile Ile Ser Asn Gly 850 855 860 Ile Asp Asp Gly Leu His Ala
Glu Asp Asp Gln Leu Arg Val Tyr Tyr 865 870 875 880 Met Gln Asn Tyr
Ile Asn Glu Ala Leu Lys Ala His Ile Leu Asp Gly 885 890 895 Ile Asn
Leu Cys Gly Tyr Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala 900 905 910
Pro Arg Phe Gly Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys 915
920 925 Ala Ser Met Lys His Tyr Arg Lys Ile Ile Asp Ser Asn Gly Phe
Gly 930 935 940 Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser 945 950 955 960 Leu Lys Tyr Pro Asn Ala Ser Pro Leu Leu
Gly Ser Ser Trp Gly Gly 965 970 975 Leu Ile His Leu Tyr Thr Ala Thr
Ala Arg Asn Ser Tyr His
Leu Gln 980 985 990 Ile His Lys Asn Gly His Val Asp Gly Ala Pro His
Gln Thr Ile Tyr 995 1000 1005 Ser Ala Leu Met Ile Arg Ser Glu Asp
Ala Gly Phe Val Val Ile 1010 1015 1020 Thr Gly Val Met Ser Arg Arg
Tyr Leu Cys Met Asp Phe Arg Gly 1025 1030 1035 Asn Ile Phe Gly Ser
His Tyr Phe Asp Pro Glu Asn Cys Arg Phe 1040 1045 1050 Gln His Gln
Thr Leu Glu Asn Gly Tyr Asp Val Tyr His Ser Pro 1055 1060 1065 Gln
Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala Phe 1070 1075
1080 Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg
1085 1090 1095 Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile
Pro Arg 1100 1105 1110 Arg His Thr Gln Ser Ala Glu Asp Asp Ser Glu
Arg Asp Pro Leu 1115 1120 1125 Asn Val Leu Lys Pro Arg Ala Arg Met
Thr Pro Ala Pro Ala Ser 1130 1135 1140 Cys Ser Gln Glu Leu Pro Ser
Ala Glu Asp Asn Ser Pro Met Ala 1145 1150 1155 Ser Asp Pro Leu Gly
Val Val Arg Gly Gly Arg Val Asn Thr His 1160 1165 1170 Ala Gly Gly
Thr Gly Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe 1175 1180 1185 Ile
251219PRTHomo sapiens 25Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gly Ser
Gly Gly Gly 245 250 255 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser Leu Lys Glu Pro 260 265 270 Gly Asp Gly Ala Gln Thr Trp Ala Arg
Phe Ser Arg Pro Pro Ala Pro 275 280 285 Glu Ala Ala Gly Leu Phe Gln
Gly Thr Phe Pro Asp Gly Phe Leu Trp 290 295 300 Ala Val Gly Ser Ala
Ala Tyr Gln Thr Glu Gly Gly Trp Gln Gln His 305 310 315 320 Gly Lys
Gly Ala Ser Ile Trp Asp Thr Phe Thr His His Pro Leu Ala 325 330 335
Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu Gly Ala Pro Ser 340
345 350 Pro Leu Gln Pro Ala Thr Gly Asp Val Ala Ser Asp Ser Tyr Asn
Asn 355 360 365 Val Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val
Thr His Tyr 370 375 380 Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro
Asn Gly Ser Ala Gly 385 390 395 400 Val Pro Asn Arg Glu Gly Leu Arg
Tyr Tyr Arg Arg Leu Leu Glu Arg 405 410 415 Leu Arg Glu Leu Gly Val
Gln Pro Val Val Thr Leu Tyr His Trp Asp 420 425 430 Leu Pro Gln Arg
Leu Gln Asp Ala Tyr Gly Gly Trp Ala Asn Arg Ala 435 440 445 Leu Ala
Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe Arg His Phe 450 455 460
Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro Tyr Val Val 465
470 475 480 Ala Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly Ile
Arg Gly 485 490 495 Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu
Leu Leu Ala His 500 505 510 Ala Lys Val Trp His Leu Tyr Asn Thr Ser
Phe Arg Pro Thr Gln Gly 515 520 525 Gly Gln Val Ser Ile Ala Leu Ser
Ser His Trp Ile Asn Pro Arg Arg 530 535 540 Met Thr Asp His Ser Ile
Lys Glu Cys Gln Lys Ser Leu Asp Phe Val 545 550 555 560 Leu Gly Trp
Phe Ala Lys Pro Val Phe Ile Asp Gly Asp Tyr Pro Glu 565 570 575 Ser
Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe Thr Glu Ser 580 585
590 Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala Leu Cys Phe
595 600 605 Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met Lys
Phe Arg 610 615 620 Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser
Trp Ile Asp Leu 625 630 635 640 Glu Phe Asn His Pro Gln Ile Phe Ile
Val Glu Asn Gly Trp Phe Val 645 650 655 Ser Gly Thr Thr Lys Arg Asp
Asp Ala Lys Tyr Met Tyr Tyr Leu Lys 660 665 670 Lys Phe Ile Met Glu
Thr Leu Lys Ala Ile Lys Leu Asp Gly Val Asp 675 680 685 Val Ile Gly
Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe Glu Trp His 690 695 700 Arg
Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp Phe Leu Ser 705 710
715 720 Gln Asp Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe Tyr Gln
Lys 725 730 735 Leu Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn
Gln Pro Leu 740 745 750 Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly
Val Val Asp Asn Tyr 755 760 765 Ile Gln Val Asp Thr Thr Leu Ser Gln
Phe Thr Asp Leu Asn Val Tyr 770 775 780 Leu Trp Asp Val His His Ser
Lys Arg Leu Ile Lys Val Asp Gly Val 785 790 795 800 Val Thr Lys Lys
Arg Lys Ser Tyr Cys Val Asp Phe Ala Ala Ile Gln 805 810 815 Pro Gln
Ile Ala Leu Leu Gln Glu Met His Val Thr His Phe Arg Phe 820 825 830
Ser Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln Ser Gln Val 835
840 845 Asn His Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser Glu Leu
Val 850 855 860 Arg Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro
Met Ala Pro 865 870 875 880 Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg
Gln Gly Ala Trp Glu Asn 885 890 895 Pro Tyr Thr Ala Leu Ala Phe Ala
Glu Tyr Ala Arg Leu Cys Phe Gln 900 905 910 Glu Leu Gly His His Val
Lys Leu Trp Ile Thr Met Asn Glu Pro Tyr 915 920 925 Thr Arg Asn Met
Thr Tyr Ser Ala Gly His Asn Leu Leu Lys Ala His 930 935 940 Ala Leu
Ala Trp His Val Tyr Asn Glu Lys Phe Arg His Ala Gln Asn 945 950 955
960 Gly Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu Pro Ala Cys
965 970 975 Pro Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val Leu
Glu Phe 980 985 990 Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser
Gly Asp Tyr Pro 995 1000 1005 Trp Val Met Arg Asp Trp Leu Asn Gln
Arg Asn Asn Phe Leu Leu 1010 1015 1020 Pro Tyr Phe Thr Glu Asp Glu
Lys Lys Leu Ile Gln Gly Thr Phe 1025 1030 1035 Asp Phe Leu Ala Leu
Ser His Tyr Thr Thr Ile Leu Val Asp Ser 1040 1045 1050 Glu Lys Glu
Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln 1055 1060 1065 Glu
Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val Ala 1070 1075
1080 Val Val Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu Lys Phe
1085 1090 1095 Lys Tyr Gly Asp Leu Pro Met Tyr Ile Ile Ser Asn Gly
Ile Asp 1100 1105 1110 Asp Gly Leu His Ala Glu Asp Asp Gln Leu Arg
Val Tyr Tyr Met 1115 1120 1125 Gln Asn Tyr Ile Asn Glu Ala Leu Lys
Ala His Ile Leu Asp Gly 1130 1135 1140 Ile Asn Leu Cys Gly Tyr Phe
Ala Tyr Ser Phe Asn Asp Arg Thr 1145 1150 1155 Ala Pro Arg Phe Gly
Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu 1160 1165 1170 Pro Lys Ala
Ser Met Lys His Tyr Arg Lys Ile Ile Asp Ser Asn 1175 1180 1185 Gly
Phe Pro Gly Pro Glu Thr Leu Glu Arg Phe Cys Pro Glu Glu 1190 1195
1200 Phe Thr Val Cys Thr Glu Cys Ser Phe Phe His Thr Arg Lys Ser
1205 1210 1215 Leu 26700PRTHomo sapiens 26Met Leu Gly Ala Arg Leu
Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser
Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser
Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45
Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50
55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp
Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr
Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr
Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu
Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu
Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met
Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn
Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175
His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180
185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser
Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His
Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala
Lys Phe Ile Gln Gly Thr Phe Pro 245 250 255 Asp Gly Phe Leu Trp Ala
Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly 260 265 270 Gly Trp Gln Gln
His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr 275 280 285 His His
Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro 290 295 300
Leu Gly Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val Ala Ser 305
310 315 320 Asp Ser Tyr Asn Asn Val Phe Arg Asp Thr Glu Ala Leu Arg
Glu Leu 325 330 335 Gly Val Thr His Tyr Arg Phe Ser Ile Ser Trp Ala
Arg Val Leu Pro 340 345 350 Asn Gly Ser Ala Gly Val Pro Asn Arg Glu
Gly Leu Arg Tyr Tyr Arg 355 360 365 Arg Leu Leu Glu Arg Leu Arg Glu
Leu Gly Val Gln Pro Val Val Thr 370 375 380 Leu Tyr His Trp Asp Leu
Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly 385 390 395 400 Trp Ala Asn
Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu 405 410 415 Cys
Phe Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp 420 425
430 Asn Pro Tyr Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu Ala
435 440 445 Pro Gly Ile Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val Ala
His Asn 450 455 460 Leu Leu Leu Ala His Ala Lys Val Trp His Leu Tyr
Asn Thr Ser Phe 465 470 475 480 Arg Pro Thr Gln Gly Gly Gln Val Ser
Ile Ala Leu Ser Ser His Trp 485 490 495 Ile Asn Pro Arg Arg Met Thr
Asp His Ser Ile Lys Glu Cys Gln Lys 500 505 510 Ser Leu Asp Phe Val
Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp 515 520 525 Gly Asp Tyr
Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro 530 535 540 Asp
Phe Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe 545 550
555 560 Phe Ala Leu Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp
Pro 565 570 575 His Met Lys Phe Arg Gln Leu Glu Ser Pro Asn Leu Arg
Gln Leu Leu 580 585 590 Ser Trp Ile Asp Leu Glu Phe Asn His Pro Gln
Ile Phe Ile Val Glu 595 600 605 Asn Gly Trp Phe Val Ser Gly Thr Thr
Lys Arg Asp Asp Ala Lys Tyr 610 615 620 Met Tyr Tyr Leu Lys Lys Phe
Ile Met Glu Thr Leu Lys Ala Ile Lys 625 630 635 640 Leu Asp Gly Val
Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp 645 650 655 Gly Phe
Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr 660 665 670
Val Asp Phe Leu Ser Gln Asp Lys Met Leu Leu Pro Lys Ser Ser Ala 675
680 685 Leu Phe Tyr Gln Lys Leu Ile Glu Lys Asn Gly Phe 690 695 700
27688PRTHomo sapiens 27Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln
Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe
Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp
Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala
Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro
Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly
Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230
235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gly Thr Phe Pro
Cys 245 250 255 Asp Phe Ala Trp Gly Val Val Asp Asn Tyr Ile Gln Val
Asp Thr Thr 260 265 270 Leu Ser Gln Phe Thr Asp Leu Asn Val Tyr Leu
Trp Asp Val His His 275 280 285 Ser Lys Arg Leu Ile Lys Val Asp Gly
Val Val Thr Lys Lys Arg Lys 290 295 300 Ser Tyr Cys Val Asp Phe Ala
Ala Ile Gln Pro Gln Ile Ala Leu Leu 305 310 315 320 Gln Glu Met His
Val Thr His Phe Arg Phe Ser Leu Asp Trp Ala Leu 325 330 335 Ile Leu
Pro Leu Gly Asn Gln Ser Gln Val Asn His Thr Ile Leu Gln 340 345 350
Tyr Tyr Arg Cys Met Ala Ser Glu Leu Val Arg Val Asn Ile Thr Pro 355
360 365 Val Val Ala Leu Trp Gln Pro Met Ala Pro Asn Gln Gly Leu Pro
Arg 370 375 380 Leu Leu Ala Arg Gln Gly Ala Trp Glu Asn Pro Tyr Thr
Ala Leu Ala 385 390 395 400 Phe Ala Glu Tyr Ala Arg Leu Cys Phe Gln
Glu Leu Gly His His Val 405 410 415 Lys Leu Trp Ile Thr Met Asn Glu
Pro Tyr Thr Arg Asn Met Thr Tyr 420 425 430 Ser Ala Gly His Asn Leu
Leu Lys Ala His Ala Leu Ala Trp His Val 435 440 445 Tyr Asn Glu Lys
Phe Arg His Ala Gln Asn Gly Lys Ile Ser Ile Ala 450 455 460 Leu Gln
Ala Asp Trp Ile Glu Pro Ala Cys Pro Phe Ser Gln Lys Asp 465 470 475
480 Lys Glu Val Ala Glu Arg Val Leu Glu Phe Asp Ile Gly Trp Leu Ala
485 490 495 Glu Pro Ile Phe Gly Ser Gly Asp Tyr Pro Trp Val Met Arg
Asp Trp 500 505 510 Leu Asn Gln Arg Asn Asn Phe Leu Leu Pro Tyr Phe
Thr Glu Asp Glu 515 520 525 Lys Lys Leu Ile Gln Gly Thr Phe Asp Phe
Leu Ala Leu Ser His Tyr 530 535 540 Thr Thr Ile Leu Val Asp Ser Glu
Lys Glu Asp Pro Ile Lys Tyr Asn 545 550 555 560 Asp Tyr Leu Glu Val
Gln Glu Met Thr Asp Ile Thr Trp Leu Asn Ser 565 570 575 Pro Ser Gln
Val Ala Val Val Pro Trp Gly Leu Arg Lys Val Leu Asn 580 585 590 Trp
Leu Lys Phe Lys Tyr Gly Asp Leu Pro Met Tyr Ile Ile Ser Asn 595 600
605 Gly Ile Asp Asp Gly Leu His Ala Glu Asp Asp Gln Leu Arg Val Tyr
610 615 620 Tyr Met Gln Asn Tyr Ile Asn Glu Ala Leu Lys Ala His Ile
Leu Asp 625 630 635 640 Gly Ile Asn Leu Cys Gly Tyr Phe Ala Tyr Ser
Phe Asn Asp Arg Thr 645 650 655 Ala Pro Arg Phe Gly Leu Tyr Arg Tyr
Ala Ala Asp Gln Phe Glu Pro 660 665 670 Lys Ala Ser Met Lys His Tyr
Arg Lys Ile Ile Asp Ser Asn Gly Phe 675 680 685 28 1149PRTHomo
sapiens 28Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys
Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala
Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu
Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His
Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln Thr Ile Tyr
Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly Phe Val Val
Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90 95 Asp Phe
Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn 100 105 110
Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His 115
120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg
Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe
Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe Asn
Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp Asp
Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala Arg
Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro Ser
Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly Val
Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230 235
240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gln Gly Thr Phe Pro
245 250 255 Asp Gly Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr
Glu Gly 260 265 270 Gly Trp Gln Gln His Gly Lys Gly Ala Ser Ile Trp
Asp Thr Phe Thr 275 280 285 His His Pro Leu Ala Pro Pro Gly Asp Ser
Arg Asn Ala Ser Leu Pro 290 295 300 Leu Gly Ala Pro Ser Pro Leu Gln
Pro Ala Thr Gly Asp Val Ala Ser 305 310 315 320 Asp Ser Tyr Asn Asn
Val Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu 325 330 335 Gly Val Thr
His Tyr Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro 340 345 350 Asn
Gly Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg 355 360
365 Arg Leu Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr
370 375 380 Leu Tyr His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr
Gly Gly 385 390 395 400 Trp Ala Asn Arg Ala Leu Ala Asp His Phe Arg
Asp Tyr Ala Glu Leu 405 410 415 Cys Phe Arg His Phe Gly Gly Gln Val
Lys Tyr Trp Ile Thr Ile Asp 420 425 430 Asn Pro Tyr Val Val Ala Trp
His Gly Tyr Ala Thr Gly Arg Leu Ala 435 440 445 Pro Gly Ile Arg Gly
Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn 450 455 460 Leu Leu Leu
Ala His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe 465 470 475 480
Arg Pro Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp 485
490 495 Ile Asn Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln
Lys 500 505 510 Ser Leu Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val
Phe Ile Asp 515 520 525 Gly Asp Tyr Pro Glu Ser Met Lys Asn Asn Leu
Ser Ser Ile Leu Pro 530 535 540 Asp Phe Thr Glu Ser Glu Lys Lys Phe
Ile Lys Gly Thr Ala Asp Phe 545 550 555 560 Phe Ala Leu Cys Phe Gly
Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro 565 570 575 His Met Lys Phe
Arg Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu 580 585 590 Ser Trp
Ile Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu 595 600 605
Asn Gly Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr 610
615 620 Met Tyr Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile
Lys 625 630 635 640 Leu Asp Gly Val Asp Val Ile Gly Tyr Thr Ala Trp
Ser Leu Met Asp 645 650 655 Gly Phe Glu Trp His Arg Gly Tyr Ser Ile
Arg Arg Gly Leu Phe Tyr 660 665 670 Val Asp Phe Leu Ser Gln Asp Lys
Met Leu Leu Pro Lys Ser Ser Ala 675 680 685 Leu Phe Tyr Gln Lys Leu
Ile Glu Lys Asn Gly Phe Gln Gly Thr Phe 690 695 700 Pro Asp Gly Phe
Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu 705 710 715 720 Gly
Gly Trp Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe 725 730
735 Thr His His Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu
740 745 750 Pro Leu Gly Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp
Val Ala 755 760 765 Ser Asp Ser Tyr Asn Asn Val Phe Arg Asp Thr Glu
Ala Leu Arg Glu 770 775 780 Leu Gly Val Thr His Tyr Arg Phe Ser Ile
Ser Trp Ala Arg Val Leu 785 790 795 800 Pro Asn Gly Ser Ala Gly Val
Pro Asn Arg Glu Gly Leu Arg Tyr Tyr 805 810 815 Arg Arg Leu Leu Glu
Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val 820 825 830 Thr Leu Tyr
His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly 835 840 845 Gly
Trp Ala Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu 850 855
860 Leu Cys Phe Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile
865 870 875 880 Asp Asn Pro Tyr Val Val Ala Trp His Gly Tyr Ala Thr
Gly Arg Leu 885 890 895 Ala Pro Gly Ile Arg Gly Ser Pro Arg Leu Gly
Tyr Leu Val Ala His 900 905 910 Asn Leu Leu Leu Ala His Ala Lys Val
Trp His Leu Tyr Asn Thr Ser 915 920 925 Phe Arg Pro Thr Gln Gly Gly
Gln Val Ser Ile Ala Leu Ser Ser His 930 935 940 Trp Ile Asn Pro Arg
Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln 945 950 955 960 Lys Ser
Leu Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile 965 970 975
Asp Gly Asp Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu 980
985 990 Pro Asp Phe Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala
Asp 995 1000 1005 Phe Phe Ala Leu Cys Phe Gly Pro Thr Leu Ser Phe
Gln Leu Leu 1010 1015 1020 Asp Pro His Met Lys Phe Arg Gln Leu Glu
Ser Pro Asn Leu Arg 1025 1030 1035 Gln Leu Leu Ser Trp Ile Asp Leu
Glu Phe Asn His Pro Gln Ile 1040 1045 1050 Phe Ile Val Glu Asn Gly
Trp Phe Val Ser Gly Thr Thr Lys Arg 1055 1060 1065 Asp Asp Ala Lys
Tyr Met Tyr Tyr Leu Lys Lys Phe Ile Met Glu 1070 1075 1080 Thr Leu
Lys Ala Ile Lys Leu Asp Gly Val Asp Val Ile Gly Tyr 1085 1090 1095
Thr Ala Trp Ser Leu Met Asp Gly Phe Glu Trp His Arg Gly Tyr 1100
1105 1110 Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp Phe Leu Ser Gln
Asp 1115 1120 1125 Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe Tyr
Gln Lys Leu 1130 1135 1140 Ile Glu Lys Asn Gly Phe 1145 29
1125PRTHomo sapiens 29Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gly Thr
Phe Pro Cys 245 250 255 Asp Phe Ala Trp Gly Val Val Asp Asn Tyr Ile
Gln Val Asp Thr Thr 260 265 270 Leu Ser Gln Phe Thr Asp Leu Asn Val
Tyr Leu Trp Asp Val His His 275 280 285 Ser Lys Arg Leu Ile Lys Val
Asp Gly Val Val Thr Lys Lys Arg Lys 290 295 300 Ser Tyr Cys Val Asp
Phe Ala Ala Ile Gln Pro Gln Ile Ala Leu Leu 305 310 315 320 Gln Glu
Met His Val Thr His Phe Arg Phe Ser Leu Asp Trp Ala Leu 325 330 335
Ile Leu Pro Leu Gly Asn Gln Ser Gln Val Asn His Thr Ile Leu Gln 340
345 350 Tyr Tyr Arg Cys Met Ala Ser Glu Leu Val Arg Val Asn Ile Thr
Pro 355 360 365 Val Val Ala Leu Trp Gln Pro Met Ala Pro Asn Gln Gly
Leu Pro Arg 370 375 380 Leu Leu Ala Arg Gln Gly Ala Trp Glu Asn Pro
Tyr Thr Ala Leu Ala 385 390 395 400 Phe Ala Glu Tyr Ala Arg Leu Cys
Phe Gln Glu Leu Gly His His Val 405 410 415 Lys Leu Trp Ile Thr Met
Asn Glu Pro Tyr Thr Arg Asn Met Thr Tyr 420 425 430 Ser Ala Gly His
Asn Leu Leu Lys Ala His Ala Leu Ala Trp His Val 435 440 445 Tyr Asn
Glu Lys Phe Arg His Ala Gln Asn Gly Lys Ile Ser Ile Ala 450 455 460
Leu Gln Ala Asp Trp Ile Glu Pro Ala Cys Pro Phe Ser Gln Lys Asp 465
470 475 480 Lys Glu Val Ala Glu Arg Val Leu Glu Phe Asp Ile Gly Trp
Leu Ala 485 490 495 Glu Pro Ile Phe Gly Ser Gly Asp Tyr Pro Trp Val
Met Arg Asp Trp 500 505 510 Leu Asn Gln Arg Asn Asn Phe Leu Leu Pro
Tyr Phe Thr Glu Asp Glu 515 520 525 Lys Lys Leu Ile Gln Gly Thr Phe
Asp Phe Leu Ala Leu Ser His Tyr 530 535 540 Thr Thr Ile Leu Val Asp
Ser Glu Lys Glu Asp Pro Ile Lys Tyr Asn 545 550 555 560 Asp Tyr Leu
Glu Val Gln Glu Met Thr Asp Ile Thr Trp Leu Asn Ser 565 570 575 Pro
Ser Gln Val Ala Val Val Pro Trp Gly Leu Arg Lys Val Leu Asn 580 585
590 Trp Leu Lys Phe Lys Tyr Gly Asp Leu Pro Met Tyr Ile Ile Ser Asn
595 600
605 Gly Ile Asp Asp Gly Leu His Ala Glu Asp Asp Gln Leu Arg Val Tyr
610 615 620 Tyr Met Gln Asn Tyr Ile Asn Glu Ala Leu Lys Ala His Ile
Leu Asp 625 630 635 640 Gly Ile Asn Leu Cys Gly Tyr Phe Ala Tyr Ser
Phe Asn Asp Arg Thr 645 650 655 Ala Pro Arg Phe Gly Leu Tyr Arg Tyr
Ala Ala Asp Gln Phe Glu Pro 660 665 670 Lys Ala Ser Met Lys His Tyr
Arg Lys Ile Ile Asp Ser Asn Gly Phe 675 680 685 Gly Thr Phe Pro Cys
Asp Phe Ala Trp Gly Val Val Asp Asn Tyr Ile 690 695 700 Gln Val Asp
Thr Thr Leu Ser Gln Phe Thr Asp Leu Asn Val Tyr Leu 705 710 715 720
Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val Asp Gly Val Val 725
730 735 Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala Ala Ile Gln
Pro 740 745 750 Gln Ile Ala Leu Leu Gln Glu Met His Val Thr His Phe
Arg Phe Ser 755 760 765 Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn
Gln Ser Gln Val Asn 770 775 780 His Thr Ile Leu Gln Tyr Tyr Arg Cys
Met Ala Ser Glu Leu Val Arg 785 790 795 800 Val Asn Ile Thr Pro Val
Val Ala Leu Trp Gln Pro Met Ala Pro Asn 805 810 815 Gln Gly Leu Pro
Arg Leu Leu Ala Arg Gln Gly Ala Trp Glu Asn Pro 820 825 830 Tyr Thr
Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu Cys Phe Gln Glu 835 840 845
Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn Glu Pro Tyr Thr 850
855 860 Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu Lys Ala His
Ala 865 870 875 880 Leu Ala Trp His Val Tyr Asn Glu Lys Phe Arg His
Ala Gln Asn Gly 885 890 895 Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp
Ile Glu Pro Ala Cys Pro 900 905 910 Phe Ser Gln Lys Asp Lys Glu Val
Ala Glu Arg Val Leu Glu Phe Asp 915 920 925 Ile Gly Trp Leu Ala Glu
Pro Ile Phe Gly Ser Gly Asp Tyr Pro Trp 930 935 940 Val Met Arg Asp
Trp Leu Asn Gln Arg Asn Asn Phe Leu Leu Pro Tyr 945 950 955 960 Phe
Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr Phe Asp Phe Leu 965 970
975 Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser Glu Lys Glu Asp
980 985 990 Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln Glu Met Thr
Asp Ile 995 1000 1005 Thr Trp Leu Asn Ser Pro Ser Gln Val Ala Val
Val Pro Trp Gly 1010 1015 1020 Leu Arg Lys Val Leu Asn Trp Leu Lys
Phe Lys Tyr Gly Asp Leu 1025 1030 1035 Pro Met Tyr Ile Ile Ser Asn
Gly Ile Asp Asp Gly Leu His Ala 1040 1045 1050 Glu Asp Asp Gln Leu
Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 1055 1060 1065 Glu Ala Leu
Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly 1070 1075 1080 Tyr
Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly 1085 1090
1095 Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met
1100 1105 1110 Lys His Tyr Arg Lys Ile Ile Asp Ser Asn Gly Phe 1115
1120 1125 30 2157DNAHomo sapiens 30gctcccagcc aagaacctcg gggccgctgc
gcggtgggga ggagttcccc gaaacccggc 60cgctaagcga ggcctcctcc tcccgcagat
ccgaacggcc tgggcggggt caccccggct 120gggacaagaa gccgccgcct
gcctgcccgg gcccggggag ggggctgggg ctggggccgg 180aggcggggtg
tgagtgggtg tgtgcggggg gcggaggctt gatgcaatcc cgataagaaa
240tgctcgggtg tcttgggcac ctacccgtgg ggcccgtaag gcgctactat
ataaggctgc 300cggcccggag ccgccgcgcc gtcagagcag gagcgctgcg
tccaggatct agggccacga 360ccatcccaac ccggcactca cagccccgca
gcgcatcccg gtcgccgccc agcctcccgc 420acccccatcg ccggagctgc
gccgagagcc ccagggaggt gccatgcgga gcgggtgtgt 480ggtggtccac
gtatggatcc tggccggcct ctggctggcc gtggccgggc gccccctcgc
540cttctcggac gcggggcccc acgtgcacta cggctggggc gaccccatcc
gcctgcggca 600cctgtacacc tccggccccc acgggctctc cagctgcttc
ctgcgcatcc gtgccgacgg 660cgtcgtggac tgcgcgcggg gccagagcgc
gcacagtttg ctggagatca aggcagtcgc 720tctgcggacc gtggccatca
agggcgtgca cagcgtgcgg tacctctgca tgggcgccga 780cggcaagatg
caggggctgc ttcagtactc ggaggaagac tgtgctttcg aggaggagat
840ccgcccagat ggctacaatg tgtaccgatc cgagaagcac cgcctcccgg
tctccctgag 900cagtgccaaa cagcggcagc tgtacaagaa cagaggcttt
cttccactct ctcatttcct 960gcccatgctg cccatggtcc cagaggagcc
tgaggacctc aggggccact tggaatctga 1020catgttctct tcgcccctgg
agaccgacag catggaccca tttgggcttg tcaccggact 1080ggaggccgtg
aggagtccca gctttgagaa gtaactgaga ccatgcccgg gcctcttcac
1140tgctgccagg ggctgtggta cctgcagcgt gggggacgtg cttctacaag
aacagtcctg 1200agtccacgtt ctgtttagct ttaggaagaa acatctagaa
gttgtacata ttcagagttt 1260tccattggca gtgccagttt ctagccaata
gacttgtctg atcataacat tgtaagcctg 1320tagcttgccc agctgctgcc
tgggccccca ttctgctccc tcgaggttgc tggacaagct 1380gctgcactgt
ctcagttctg cttgaatacc tccatcgatg gggaactcac ttcctttgga
1440aaaattctta tgtcaagctg aaattctcta attttttctc atcacttccc
caggagcagc 1500cagaagacag gcagtagttt taatttcagg aacaggtgat
ccactctgta aaacagcagg 1560taaatttcac tcaaccccat gtgggaattg
atctatatct ctacttccag ggaccatttg 1620cccttcccaa atccctccag
gccagaactg actggagcag gcatggccca ccaggcttca 1680ggagtagggg
aagcctggag ccccactcca gccctgggac aacttgagaa ttccccctga
1740ggccagttct gtcatggatg ctgtcctgag aataacttgc tgtcccggtg
tcacctgctt 1800ccatctccca gcccaccagc cctctgccca cctcacatgc
ctccccatgg attggggcct 1860cccaggcccc ccaccttatg tcaacctgca
cttcttgttc aaaaatcagg aaaagaaaag 1920atttgaagac cccaagtctt
gtcaataact tgctgtgtgg aagcagcggg ggaagaccta 1980gaaccctttc
cccagcactt ggttttccaa catgatattt atgagtaatt tattttgata
2040tgtacatctc ttattttctt acattattta tgcccccaaa ttatatttat
gtatgtaagt 2100gaggtttgtt ttgtatatta aaatggagtt tgtttgtaaa
aaaaaaaaaa aaaaaaa 215731216PRTHomo sapiens 31Met Arg Ser Gly Cys
Val Val Val His Val Trp Ile Leu Ala Gly Leu 1 5 10 15 Trp Leu Ala
Val Ala Gly Arg Pro Leu Ala Phe Ser Asp Ala Gly Pro 20 25 30 His
Val His Tyr Gly Trp Gly Asp Pro Ile Arg Leu Arg His Leu Tyr 35 40
45 Thr Ser Gly Pro His Gly Leu Ser Ser Cys Phe Leu Arg Ile Arg Ala
50 55 60 Asp Gly Val Val Asp Cys Ala Arg Gly Gln Ser Ala His Ser
Leu Leu 65 70 75 80 Glu Ile Lys Ala Val Ala Leu Arg Thr Val Ala Ile
Lys Gly Val His 85 90 95 Ser Val Arg Tyr Leu Cys Met Gly Ala Asp
Gly Lys Met Gln Gly Leu 100 105 110 Leu Gln Tyr Ser Glu Glu Asp Cys
Ala Phe Glu Glu Glu Ile Arg Pro 115 120 125 Asp Gly Tyr Asn Val Tyr
Arg Ser Glu Lys His Arg Leu Pro Val Ser 130 135 140 Leu Ser Ser Ala
Lys Gln Arg Gln Leu Tyr Lys Asn Arg Gly Phe Leu 145 150 155 160 Pro
Leu Ser His Phe Leu Pro Met Leu Pro Met Val Pro Glu Glu Pro 165 170
175 Glu Asp Leu Arg Gly His Leu Glu Ser Asp Met Phe Ser Ser Pro Leu
180 185 190 Glu Thr Asp Ser Met Asp Pro Phe Gly Leu Val Thr Gly Leu
Glu Ala 195 200 205 Val Arg Ser Pro Ser Phe Glu Lys 210 215
32940DNAHomo sapiens 32ctgtcagctg aggatccagc cgaaagagga gccaggcact
caggccacct gagtctactc 60acctggacaa ctggaatctg gcaccaattc taaaccactc
agcttctccg agctcacacc 120ccggagatca cctgaggacc cgagccattg
atggactcgg acgagaccgg gttcgagcac 180tcaggactgt gggtttctgt
gctggctggt cttctgctgg gagcctgcca ggcacacccc 240atccctgact
ccagtcctct cctgcaattc gggggccaag tccggcagcg gtacctctac
300acagatgatg cccagcagac agaagcccac ctggagatca gggaggatgg
gacggtgggg 360ggcgctgctg accagagccc cgaaagtctc ctgcagctga
aagccttgaa gccgggagtt 420attcaaatct tgggagtcaa gacatccagg
ttcctgtgcc agcggccaga tggggccctg 480tatggatcgc tccactttga
ccctgaggcc tgcagcttcc gggagctgct tcttgaggac 540ggatacaatg
tttaccagtc cgaagcccac ggcctcccgc tgcacctgcc agggaacaag
600tccccacacc gggaccctgc accccgagga ccagctcgct tcctgccact
accaggcctg 660ccccccgcac tcccggagcc acccggaatc ctggcccccc
agccccccga tgtgggctcc 720tcggaccctc tgagcatggt gggaccttcc
cagggccgaa gccccagcta cgcttcctga 780agccagaggc tgtttactat
gacatctcct ctttatttat taggttattt atcttattta 840tttttttatt
tttcttactt gagataataa agagttccag aggagaaaaa aaaaaaaaaa
900aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 94033209PRTHomo
sapiens 33Met Asp Ser Asp Glu Thr Gly Phe Glu His Ser Gly Leu Trp
Val Ser 1 5 10 15 Val Leu Ala Gly Leu Leu Leu Gly Ala Cys Gln Ala
His Pro Ile Pro 20 25 30 Asp Ser Ser Pro Leu Leu Gln Phe Gly Gly
Gln Val Arg Gln Arg Tyr 35 40 45 Leu Tyr Thr Asp Asp Ala Gln Gln
Thr Glu Ala His Leu Glu Ile Arg 50 55 60 Glu Asp Gly Thr Val Gly
Gly Ala Ala Asp Gln Ser Pro Glu Ser Leu 65 70 75 80 Leu Gln Leu Lys
Ala Leu Lys Pro Gly Val Ile Gln Ile Leu Gly Val 85 90 95 Lys Thr
Ser Arg Phe Leu Cys Gln Arg Pro Asp Gly Ala Leu Tyr Gly 100 105 110
Ser Leu His Phe Asp Pro Glu Ala Cys Ser Phe Arg Glu Leu Leu Leu 115
120 125 Glu Asp Gly Tyr Asn Val Tyr Gln Ser Glu Ala His Gly Leu Pro
Leu 130 135 140 His Leu Pro Gly Asn Lys Ser Pro His Arg Asp Pro Ala
Pro Arg Gly 145 150 155 160 Pro Ala Arg Phe Leu Pro Leu Pro Gly Leu
Pro Pro Ala Leu Pro Glu 165 170 175 Pro Pro Gly Ile Leu Ala Pro Gln
Pro Pro Asp Val Gly Ser Ser Asp 180 185 190 Pro Leu Ser Met Val Gly
Pro Ser Gln Gly Arg Ser Pro Ser Tyr Ala 195 200 205 Ser
343018DNAHomo sapiens 34cggcaaaaag gagggaatcc agtctaggat cctcacacca
gctacttgca agggagaagg 60aaaaggccag taaggcctgg gccaggagag tcccgacagg
agtgtcaggt ttcaatctca 120gcaccagcca ctcagagcag ggcacgatgt
tgggggcccg cctcaggctc tgggtctgtg 180ccttgtgcag cgtctgcagc
atgagcgtcc tcagagccta tcccaatgcc tccccactgc 240tcggctccag
ctggggtggc ctgatccacc tgtacacagc cacagccagg aacagctacc
300acctgcagat ccacaagaat ggccatgtgg atggcgcacc ccatcagacc
atctacagtg 360ccctgatgat cagatcagag gatgctggct ttgtggtgat
tacaggtgtg atgagcagaa 420gatacctctg catggatttc agaggcaaca
tttttggatc acactatttc gacccggaga 480actgcaggtt ccaacaccag
acgctggaaa acgggtacga cgtctaccac tctcctcagt 540atcacttcct
ggtcagtctg ggccgggcga agagagcctt cctgccaggc atgaacccac
600ccccgtactc ccagttcctg tcccggagga acgagatccc cctaattcac
ttcaacaccc 660ccataccacg gcggcacacc cggagcgccg aggacgactc
ggagcgggac cccctgaacg 720tgctgaagcc ccgggcccgg atgaccccgg
ccccggcctc ctgttcacag gagctcccga 780gcgccgagga caacagcccg
atggccagtg acccattagg ggtggtcagg ggcggtcgag 840tgaacacgca
cgctggggga acgggcccgg aaggctgccg ccccttcgcc aagttcatct
900agggtcgctg gaagggcacc ctctttaacc catccctcag caaacgcagc
tcttcccaag 960gaccaggtcc cttgacgttc cgaggatggg aaaggtgaca
ggggcatgta tggaatttgc 1020tgcttctctg gggtcccttc cacaggaggt
cctgtgagaa ccaacctttg aggcccaagt 1080catggggttt caccgccttc
ctcactccat atagaacacc tttcccaata ggaaacccca 1140acaggtaaac
tagaaatttc cccttcatga aggtagagag aaggggtctc tcccaacata
1200tttctcttcc ttgtgcctct cctctttatc acttttaagc ataaaaaaaa
aaaaaaaaaa 1260aaaaaaaaaa aaaagcagtg ggttcctgag ctcaagactt
tgaaggtgta gggaagagga 1320aatcggagat cccagaagct tctccactgc
cctatgcatt tatgttagat gccccgatcc 1380cactggcatt tgagtgtgca
aaccttgaca ttaacagctg aatggggcaa gttgatgaaa 1440acactacttt
caagccttcg ttcttccttg agcatctctg gggaagagct gtcaaaagac
1500tggtggtagg ctggtgaaaa cttgacagct agacttgatg cttgctgaaa
tgaggcagga 1560atcataatag aaaactcagc ctccctacag ggtgagcacc
ttctgtctcg ctgtctccct 1620ctgtgcagcc acagccagag ggcccagaat
ggccccactc tgttcccaag cagttcatga 1680tacagcctca ccttttggcc
ccatctctgg tttttgaaaa tttggtctaa ggaataaata 1740gcttttacac
tggctcacga aaatctgccc tgctagaatt tgcttttcaa aatggaaata
1800aattccaact ctcctaagag gcatttaatt aaggctctac ttccaggttg
agtaggaatc 1860cattctgaac aaactacaaa aatgtgactg ggaagggggc
tttgagagac tgggactgct 1920ctgggttagg ttttctgtgg actgaaaaat
cgtgtccttt tctctaaatg aagtggcatc 1980aaggactcag ggggaaagaa
atcaggggac atgttataga agttatgaaa agacaaccac 2040atggtcaggc
tcttgtctgt ggtctctagg gctctgcagc agcagtggct cttcgattag
2100ttaaaactct cctaggctga cacatctggg tctcaatccc cttggaaatt
cttggtgcat 2160taaatgaagc cttaccccat tactgcggtt cttcctgtaa
gggggctcca ttttcctccc 2220tctctttaaa tgaccaccta aaggacagta
tattaacaag caaagtcgat tcaacaacag 2280cttcttccca gtcacttttt
tttttctcac tgccatcaca tactaacctt atactttgat 2340ctattctttt
tggttatgag agaaatgttg ggcaactgtt tttacctgat ggttttaagc
2400tgaacttgaa ggactggttc ctattctgaa acagtaaaac tatgtataat
agtatatagc 2460catgcatggc aaatatttta atatttctgt tttcatttcc
tgttggaaat attatcctgc 2520ataatagcta ttggaggctc ctcagtgaaa
gatcccaaaa ggattttggt ggaaaactag 2580ttgtaatctc acaaactcaa
cactaccatc aggggttttc tttatggcaa agccaaaata 2640gctcctacaa
tttcttatat ccctcgtcat gtggcagtat ttatttattt atttggaagt
2700ttgcctatcc ttctatattt atagatattt ataaaaatgt aacccctttt
tcctttcttc 2760tgtttaaaat aaaaataaaa tttatctcag cttctgttag
cttatcctct ttgtagtact 2820acttaaaagc atgtcggaat ataagaataa
aaaggattat gggaggggaa cattagggaa 2880atccagagaa ggcaaaattg
aaaaaaagat tttagaattt taaaattttc aaagatttct 2940tccattcata
aggagactca atgattttaa ttgatctaga cagaattatt taagttttat
3000caatattgga tttctggt 301835251PRTHomo sapiens 35Met Leu Gly Ala
Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser
Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30
Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35
40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly
Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser
Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg
Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser
His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr
Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His
Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro
Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160
Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165
170 175 His Thr Arg Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn
Val 180 185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser
Cys Ser Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met
Ala Ser Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn
Thr His Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Cys Arg Pro
Phe Ala Lys Phe Ile 245 250 36251PRTHomo sapiens 36Met Leu Gly Ala
Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser
Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30
Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35
40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly
Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser
Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg
Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser
His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr
Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His
Phe Leu Val Ser Leu Gly Arg Ala Lys
Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln
Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe
Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp
Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala
Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro
Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly
Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230
235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile 245 250
37431PRTHomo sapiens 37Tyr Asp Thr Phe Pro Lys Asn Phe Phe Trp Gly
Ile Gly Thr Gly Ala 1 5 10 15 Leu Gln Val Glu Gly Ser Trp Lys Lys
Asp Gly Lys Gly Pro Ser Ile 20 25 30 Trp Asp His Phe Ile His Thr
His Leu Lys Asn Val Ser Ser Thr Asn 35 40 45 Gly Ser Ser Asp Ser
Tyr Ile Phe Leu Glu Lys Asp Leu Ser Ala Leu 50 55 60 Asp Phe Ile
Gly Val Ser Phe Tyr Gln Phe Ser Ile Ser Trp Pro Arg 65 70 75 80 Leu
Phe Pro Asp Gly Ile Val Thr Val Ala Asn Ala Lys Gly Leu Gln 85 90
95 Tyr Tyr Ser Thr Leu Leu Asp Ala Leu Val Leu Arg Asn Ile Glu Pro
100 105 110 Ile Val Thr Leu Tyr His Trp Asp Leu Pro Leu Ala Leu Gln
Glu Lys 115 120 125 Tyr Gly Gly Trp Lys Asn Asp Thr Ile Ile Asp Ile
Phe Asn Asp Tyr 130 135 140 Ala Thr Tyr Cys Phe Gln Met Phe Gly Asp
Arg Val Lys Tyr Trp Ile 145 150 155 160 Thr Ile His Asn Pro Tyr Leu
Val Ala Trp His Gly Tyr Gly Thr Gly 165 170 175 Met His Ala Pro Gly
Glu Lys Gly Asn Leu Ala Ala Val Tyr Thr Val 180 185 190 Gly His Asn
Leu Ile Lys Ala His Ser Lys Val Trp His Asn Tyr Asn 195 200 205 Thr
His Phe Arg Pro His Gln Lys Gly Trp Leu Ser Ile Thr Leu Gly 210 215
220 Ser His Trp Ile Glu Pro Asn Arg Ser Glu Asn Thr Met Asp Ile Phe
225 230 235 240 Lys Cys Gln Gln Ser Met Val Ser Val Leu Gly Trp Phe
Ala Asn Pro 245 250 255 Ile His Gly Asp Gly Asp Tyr Pro Glu Gly Met
Arg Lys Lys Leu Phe 260 265 270 Ser Val Leu Pro Ile Phe Ser Glu Ala
Glu Lys His Glu Met Arg Gly 275 280 285 Thr Ala Asp Phe Phe Ala Phe
Ser Phe Gly Pro Asn Asn Phe Lys Pro 290 295 300 Leu Asn Thr Met Ala
Lys Met Gly Gln Asn Val Ser Leu Asn Leu Arg 305 310 315 320 Glu Ala
Leu Asn Trp Ile Lys Leu Glu Tyr Asn Asn Pro Arg Ile Leu 325 330 335
Ile Ala Glu Asn Gly Trp Phe Thr Asp Ser Arg Val Lys Thr Glu Asp 340
345 350 Thr Thr Ala Ile Tyr Met Met Lys Asn Phe Leu Ser Gln Val Leu
Gln 355 360 365 Ala Ile Arg Leu Asp Glu Ile Arg Val Phe Gly Tyr Thr
Ala Trp Ser 370 375 380 Leu Leu Asp Gly Phe Glu Trp Gln Asp Ala Tyr
Thr Ile Arg Arg Gly 385 390 395 400 Leu Phe Tyr Val Asp Phe Asn Ser
Lys Gln Lys Glu Arg Lys Pro Lys 405 410 415 Ser Ser Ala His Tyr Tyr
Lys Gln Ile Ile Arg Glu Asn Gly Phe 420 425 430 38397PRTHomo
sapiens 38Thr Arg Pro Ala Gln Cys Thr Asp Phe Val Asn Ile Lys Lys
Gln Leu 1 5 10 15 Glu Met Leu Ala Arg Met Lys Val Thr His Tyr Arg
Phe Ala Leu Asp 20 25 30 Trp Ala Ser Val Leu Pro Thr Gly Asn Leu
Ser Ala Val Asn Arg Gln 35 40 45 Ala Leu Arg Tyr Tyr Arg Cys Val
Val Ser Glu Gly Leu Lys Leu Gly 50 55 60 Ile Ser Ala Met Val Thr
Leu Tyr Tyr Pro Thr His Ala His Leu Gly 65 70 75 80 Leu Pro Glu Pro
Leu Leu His Ala Asp Gly Trp Leu Asn Pro Ser Thr 85 90 95 Ala Glu
Ala Phe Gln Ala Tyr Ala Gly Leu Cys Phe Gln Glu Leu Gly 100 105 110
Asp Leu Val Lys Leu Trp Ile Thr Ile Asn Glu Pro Asn Arg Leu Ser 115
120 125 Asp Ile Tyr Asn Arg Ser Gly Asn Asp Thr Tyr Gly Ala Ala His
Asn 130 135 140 Leu Leu Val Ala His Ala Leu Ala Trp Arg Leu Tyr Asp
Arg Gln Phe 145 150 155 160 Arg Pro Ser Gln Arg Gly Ala Val Ser Leu
Ser Leu His Ala Asp Trp 165 170 175 Ala Glu Pro Ala Asn Pro Tyr Ala
Asp Ser His Trp Arg Ala Ala Glu 180 185 190 Arg Phe Leu Gln Phe Glu
Ile Ala Trp Phe Ala Glu Pro Leu Phe Lys 195 200 205 Thr Gly Asp Tyr
Pro Ala Ala Met Arg Glu Tyr Ile Ala Ser Lys His 210 215 220 Arg Arg
Gly Leu Ser Ser Ser Ala Leu Pro Arg Leu Thr Glu Ala Glu 225 230 235
240 Arg Arg Leu Leu Lys Gly Thr Val Asp Phe Cys Ala Leu Asn His Phe
245 250 255 Thr Thr Arg Phe Val Met His Glu Gln Leu Ala Gly Ser Arg
Tyr Asp 260 265 270 Ser Asp Arg Asp Ile Gln Phe Leu Gln Asp Ile Thr
Arg Leu Ser Ser 275 280 285 Pro Thr Arg Leu Ala Val Ile Pro Trp Gly
Val Arg Lys Leu Leu Arg 290 295 300 Trp Val Arg Arg Asn Tyr Gly Asp
Met Asp Ile Tyr Ile Thr Ala Ser 305 310 315 320 Gly Ile Asp Asp Gln
Ala Leu Glu Asp Asp Arg Leu Arg Lys Tyr Tyr 325 330 335 Leu Gly Lys
Tyr Leu Gln Glu Val Leu Lys Ala Tyr Leu Ile Asp Lys 340 345 350 Val
Arg Ile Lys Gly Tyr Tyr Ala Phe Lys Leu Ala Glu Glu Lys Ser 355 360
365 Lys Pro Arg Phe Gly Phe Phe Thr Ser Asp Phe Lys Ala Lys Ser Ser
370 375 380 Ile Gln Phe Tyr Asn Lys Val Ile Ser Ser Arg Gly Phe 385
390 395 39946PRTHomo sapiens 39Gly Phe Ser Gly Asp Gly Arg Ala Ile
Trp Ser Lys Asn Pro Asn Phe 1 5 10 15 Thr Pro Val Asn Glu Ser Gln
Leu Phe Leu Tyr Asp Thr Phe Pro Lys 20 25 30 Asn Phe Phe Trp Gly
Ile Gly Thr Gly Ala Leu Gln Val Glu Gly Ser 35 40 45 Trp Lys Lys
Asp Gly Lys Gly Pro Ser Ile Trp Asp His Phe Ile His 50 55 60 Thr
His Leu Lys Asn Val Ser Ser Thr Asn Gly Ser Ser Asp Ser Tyr 65 70
75 80 Ile Phe Leu Glu Lys Asp Leu Ser Ala Leu Asp Phe Ile Gly Val
Ser 85 90 95 Phe Tyr Gln Phe Ser Ile Ser Trp Pro Arg Leu Phe Pro
Asp Gly Ile 100 105 110 Val Thr Val Ala Asn Ala Lys Gly Leu Gln Tyr
Tyr Ser Thr Leu Leu 115 120 125 Asp Ala Leu Val Leu Arg Asn Ile Glu
Pro Ile Val Thr Leu Tyr His 130 135 140 Trp Asp Leu Pro Leu Ala Leu
Gln Glu Lys Tyr Gly Gly Trp Lys Asn 145 150 155 160 Asp Thr Ile Ile
Asp Ile Phe Asn Asp Tyr Ala Thr Tyr Cys Phe Gln 165 170 175 Met Phe
Gly Asp Arg Val Lys Tyr Trp Ile Thr Ile His Asn Pro Tyr 180 185 190
Leu Val Ala Trp His Gly Tyr Gly Thr Gly Met His Ala Pro Gly Glu 195
200 205 Lys Gly Asn Leu Ala Ala Val Tyr Thr Val Gly His Asn Leu Ile
Lys 210 215 220 Ala His Ser Lys Val Trp His Asn Tyr Asn Thr His Phe
Arg Pro His 225 230 235 240 Gln Lys Gly Trp Leu Ser Ile Thr Leu Gly
Ser His Trp Ile Glu Pro 245 250 255 Asn Arg Ser Glu Asn Thr Met Asp
Ile Phe Lys Cys Gln Gln Ser Met 260 265 270 Val Ser Val Leu Gly Trp
Phe Ala Asn Pro Ile His Gly Asp Gly Asp 275 280 285 Tyr Pro Glu Gly
Met Arg Lys Lys Leu Phe Ser Val Leu Pro Ile Phe 290 295 300 Ser Glu
Ala Glu Lys His Glu Met Arg Gly Thr Ala Asp Phe Phe Ala 305 310 315
320 Phe Ser Phe Gly Pro Asn Asn Phe Lys Pro Leu Asn Thr Met Ala Lys
325 330 335 Met Gly Gln Asn Val Ser Leu Asn Leu Arg Glu Ala Leu Asn
Trp Ile 340 345 350 Lys Leu Glu Tyr Asn Asn Pro Arg Ile Leu Ile Ala
Glu Asn Gly Trp 355 360 365 Phe Thr Asp Ser Arg Val Lys Thr Glu Asp
Thr Thr Ala Ile Tyr Met 370 375 380 Met Lys Asn Phe Leu Ser Gln Val
Leu Gln Ala Ile Arg Leu Asp Glu 385 390 395 400 Ile Arg Val Phe Gly
Tyr Thr Ala Trp Ser Leu Leu Asp Gly Phe Glu 405 410 415 Trp Gln Asp
Ala Tyr Thr Ile Arg Arg Gly Leu Phe Tyr Val Asp Phe 420 425 430 Asn
Ser Lys Gln Lys Glu Arg Lys Pro Lys Ser Ser Ala His Tyr Tyr 435 440
445 Lys Gln Ile Ile Arg Glu Asn Gly Phe Ser Leu Lys Glu Ser Thr Pro
450 455 460 Asp Val Gln Gly Gln Phe Pro Cys Asp Phe Ser Trp Gly Val
Thr Glu 465 470 475 480 Ser Val Leu Lys Pro Glu Ser Val Ala Ser Ser
Pro Gln Phe Ser Asp 485 490 495 Pro His Leu Tyr Val Trp Asn Ala Thr
Gly Asn Arg Leu Leu His Arg 500 505 510 Val Glu Gly Val Arg Leu Lys
Thr Arg Pro Ala Gln Cys Thr Asp Phe 515 520 525 Val Asn Ile Lys Lys
Gln Leu Glu Met Leu Ala Arg Met Lys Val Thr 530 535 540 His Tyr Arg
Phe Ala Leu Asp Trp Ala Ser Val Leu Pro Thr Gly Asn 545 550 555 560
Leu Ser Ala Val Asn Arg Gln Ala Leu Arg Tyr Tyr Arg Cys Val Val 565
570 575 Ser Glu Gly Leu Lys Leu Gly Ile Ser Ala Met Val Thr Leu Tyr
Tyr 580 585 590 Pro Thr His Ala His Leu Gly Leu Pro Glu Pro Leu Leu
His Ala Asp 595 600 605 Gly Trp Leu Asn Pro Ser Thr Ala Glu Ala Phe
Gln Ala Tyr Ala Gly 610 615 620 Leu Cys Phe Gln Glu Leu Gly Asp Leu
Val Lys Leu Trp Ile Thr Ile 625 630 635 640 Asn Glu Pro Asn Arg Leu
Ser Asp Ile Tyr Asn Arg Ser Gly Asn Asp 645 650 655 Thr Tyr Gly Ala
Ala His Asn Leu Leu Val Ala His Ala Leu Ala Trp 660 665 670 Arg Leu
Tyr Asp Arg Gln Phe Arg Pro Ser Gln Arg Gly Ala Val Ser 675 680 685
Leu Ser Leu His Ala Asp Trp Ala Glu Pro Ala Asn Pro Tyr Ala Asp 690
695 700 Ser His Trp Arg Ala Ala Glu Arg Phe Leu Gln Phe Glu Ile Ala
Trp 705 710 715 720 Phe Ala Glu Pro Leu Phe Lys Thr Gly Asp Tyr Pro
Ala Ala Met Arg 725 730 735 Glu Tyr Ile Ala Ser Lys His Arg Arg Gly
Leu Ser Ser Ser Ala Leu 740 745 750 Pro Arg Leu Thr Glu Ala Glu Arg
Arg Leu Leu Lys Gly Thr Val Asp 755 760 765 Phe Cys Ala Leu Asn His
Phe Thr Thr Arg Phe Val Met His Glu Gln 770 775 780 Leu Ala Gly Ser
Arg Tyr Asp Ser Asp Arg Asp Ile Gln Phe Leu Gln 785 790 795 800 Asp
Ile Thr Arg Leu Ser Ser Pro Thr Arg Leu Ala Val Ile Pro Trp 805 810
815 Gly Val Arg Lys Leu Leu Arg Trp Val Arg Arg Asn Tyr Gly Asp Met
820 825 830 Asp Ile Tyr Ile Thr Ala Ser Gly Ile Asp Asp Gln Ala Leu
Glu Asp 835 840 845 Asp Arg Leu Arg Lys Tyr Tyr Leu Gly Lys Tyr Leu
Gln Glu Val Leu 850 855 860 Lys Ala Tyr Leu Ile Asp Lys Val Arg Ile
Lys Gly Tyr Tyr Ala Phe 865 870 875 880 Lys Leu Ala Glu Glu Lys Ser
Lys Pro Arg Phe Gly Phe Phe Thr Ser 885 890 895 Asp Phe Lys Ala Lys
Ser Ser Ile Gln Phe Tyr Asn Lys Val Ile Ser 900 905 910 Ser Arg Gly
Phe Pro Phe Glu Asn Ser Ser Ser Arg Cys Ser Gln Thr 915 920 925 Gln
Glu Asn Thr Glu Cys Thr Val Cys Leu Phe Leu Val Gln Lys Lys 930 935
940 Pro Leu 945 401195PRTHomo sapiens 40Glu Pro Gly Asp Gly Ala Gln
Thr Trp Ala Arg Phe Ser Arg Pro Pro 1 5 10 15 Ala Pro Glu Ala Ala
Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly Phe 20 25 30 Leu Trp Ala
Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp Gln 35 40 45 Gln
His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His His Pro 50 55
60 Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu Gly Ala
65 70 75 80 Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val Ala Ser Asp
Ser Tyr 85 90 95 Asn Asn Val Phe Arg Asp Thr Glu Ala Leu Arg Glu
Leu Gly Val Thr 100 105 110 His Tyr Arg Phe Ser Ile Ser Trp Ala Arg
Val Leu Pro Asn Gly Ser 115 120 125 Ala Gly Val Pro Asn Arg Glu Gly
Leu Arg Tyr Tyr Arg Arg Leu Leu 130 135 140 Glu Arg Leu Arg Glu Leu
Gly Val Gln Pro Val Val Thr Leu Tyr His 145 150 155 160 Trp Asp Leu
Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala Asn 165 170 175 Arg
Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe Arg 180 185
190 His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro Tyr
195 200 205 Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro
Gly Ile 210 215 220 Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val Ala His
Asn Leu Leu Leu 225 230 235 240 Ala His Ala Lys Val Trp His Leu Tyr
Asn Thr Ser Phe Arg Pro Thr 245 250 255 Gln Gly Gly Gln Val Ser Ile
Ala Leu Ser Ser His Trp Ile Asn Pro 260 265 270 Arg Arg Met Thr Asp
His Ser Ile Lys Glu Cys Gln Lys Ser Leu Asp 275 280 285 Phe Val Leu
Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp Tyr 290 295 300 Pro
Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe Thr 305 310
315 320 Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala
Leu 325 330 335 Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro
His Met Lys 340 345 350 Phe Arg Gln Leu Glu Ser Pro Asn Leu Arg Gln
Leu Leu Ser Trp Ile 355 360 365 Asp Leu Glu Phe Asn His Pro Gln Ile
Phe Ile Val Glu Asn Gly Trp 370 375 380 Phe Val Ser Gly Thr Thr Lys
Arg Asp Asp Ala Lys Tyr Met Tyr Tyr 385 390 395 400 Leu Lys Lys Phe
Ile Met Glu Thr Leu Lys
Ala Ile Lys Leu Asp Gly 405 410 415 Val Asp Val Ile Gly Tyr Thr Ala
Trp Ser Leu Met Asp Gly Phe Glu 420 425 430 Trp His Arg Gly Tyr Ser
Ile Arg Arg Gly Leu Phe Tyr Val Asp Phe 435 440 445 Leu Ser Gln Asp
Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe Tyr 450 455 460 Gln Lys
Leu Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn Gln 465 470 475
480 Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val Asp
485 490 495 Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp
Leu Asn 500 505 510 Val Tyr Leu Trp Asp Val His His Ser Lys Arg Leu
Ile Lys Val Asp 515 520 525 Gly Val Val Thr Lys Lys Arg Lys Ser Tyr
Cys Val Asp Phe Ala Ala 530 535 540 Ile Gln Pro Gln Ile Ala Leu Leu
Gln Glu Met His Val Thr His Phe 545 550 555 560 Arg Phe Ser Leu Asp
Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln Ser 565 570 575 Gln Val Asn
His Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser Glu 580 585 590 Leu
Val Arg Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro Met 595 600
605 Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala Trp
610 615 620 Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg
Leu Cys 625 630 635 640 Phe Gln Glu Leu Gly His His Val Lys Leu Trp
Ile Thr Met Asn Glu 645 650 655 Pro Tyr Thr Arg Asn Met Thr Tyr Ser
Ala Gly His Asn Leu Leu Lys 660 665 670 Ala His Ala Leu Ala Trp His
Val Tyr Asn Glu Lys Phe Arg His Ala 675 680 685 Gln Asn Gly Lys Ile
Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu Pro 690 695 700 Ala Cys Pro
Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val Leu 705 710 715 720
Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly Asp 725
730 735 Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe
Leu 740 745 750 Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln
Gly Thr Phe 755 760 765 Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile
Leu Val Asp Ser Glu 770 775 780 Lys Glu Asp Pro Ile Lys Tyr Asn Asp
Tyr Leu Glu Val Gln Glu Met 785 790 795 800 Thr Asp Ile Thr Trp Leu
Asn Ser Pro Ser Gln Val Ala Val Val Pro 805 810 815 Trp Gly Leu Arg
Lys Val Leu Asn Trp Leu Lys Phe Lys Tyr Gly Asp 820 825 830 Leu Pro
Met Tyr Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu His Ala 835 840 845
Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn Glu 850
855 860 Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr
Phe 865 870 875 880 Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe
Gly Leu Tyr Arg 885 890 895 Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala
Ser Met Lys His Tyr Arg 900 905 910 Lys Ile Ile Asp Ser Asn Gly Phe
Pro Gly Pro Glu Thr Leu Glu Arg 915 920 925 Phe Cys Pro Glu Glu Phe
Thr Val Cys Thr Glu Cys Ser Phe Phe His 930 935 940 Thr Arg Lys Ser
Leu Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 945 950 955 960 Ser
Gly Gly Gly Gly Ser Leu Lys Tyr Pro Asn Ala Ser Pro Leu Leu 965 970
975 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg
980 985 990 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp
Gly Ala 995 1000 1005 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile
Arg Ser Glu Asp 1010 1015 1020 Ala Gly Phe Val Val Ile Thr Gly Val
Met Ser Arg Arg Tyr Leu 1025 1030 1035 Cys Met Asp Phe Arg Gly Asn
Ile Phe Gly Ser His Tyr Phe Asp 1040 1045 1050 Pro Glu Asn Cys Arg
Phe Gln His Gln Thr Leu Glu Asn Gly Tyr 1055 1060 1065 Asp Val Tyr
His Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly 1070 1075 1080 Arg
Ala Lys Arg Ala Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr 1085 1090
1095 Ser Gln Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu Ile His Phe
1100 1105 1110 Asn Thr Pro Ile Pro Arg Arg His Thr Arg Ser Ala Glu
Asp Asp 1115 1120 1125 Ser Glu Arg Asp Pro Leu Asn Val Leu Lys Pro
Arg Ala Arg Met 1130 1135 1140 Thr Pro Ala Pro Ala Ser Cys Ser Gln
Glu Leu Pro Ser Ala Glu 1145 1150 1155 Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu Gly Val Val Arg Gly 1160 1165 1170 Gly Arg Val Asn Thr
His Ala Gly Gly Thr Gly Pro Glu Gly Cys 1175 1180 1185 Arg Pro Phe
Ala Lys Phe Ile 1190 1195 41 1195PRTHomo sapiens 41Glu Pro Gly Asp
Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro Pro 1 5 10 15 Ala Pro
Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly Phe 20 25 30
Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp Gln 35
40 45 Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr His His
Pro 50 55 60 Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser Leu Pro
Leu Gly Ala 65 70 75 80 Pro Ser Pro Leu Gln Pro Ala Thr Gly Asp Val
Ala Ser Asp Ser Tyr 85 90 95 Asn Asn Val Phe Arg Asp Thr Glu Ala
Leu Arg Glu Leu Gly Val Thr 100 105 110 His Tyr Arg Phe Ser Ile Ser
Trp Ala Arg Val Leu Pro Asn Gly Ser 115 120 125 Ala Gly Val Pro Asn
Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu Leu 130 135 140 Glu Arg Leu
Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr His 145 150 155 160
Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala Asn 165
170 175 Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu Cys Phe
Arg 180 185 190 His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr Ile Asp
Asn Pro Tyr 195 200 205 Val Val Ala Trp His Gly Tyr Ala Thr Gly Arg
Leu Ala Pro Gly Ile 210 215 220 Arg Gly Ser Pro Arg Leu Gly Tyr Leu
Val Ala His Asn Leu Leu Leu 225 230 235 240 Ala His Ala Lys Val Trp
His Leu Tyr Asn Thr Ser Phe Arg Pro Thr 245 250 255 Gln Gly Gly Gln
Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn Pro 260 265 270 Arg Arg
Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu Asp 275 280 285
Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly Asp Tyr 290
295 300 Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe
Thr 305 310 315 320 Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr Ala Asp
Phe Phe Ala Leu 325 330 335 Cys Phe Gly Pro Thr Leu Ser Phe Gln Leu
Leu Asp Pro His Met Lys 340 345 350 Phe Arg Gln Leu Glu Ser Pro Asn
Leu Arg Gln Leu Leu Ser Trp Ile 355 360 365 Asp Leu Glu Phe Asn His
Pro Gln Ile Phe Ile Val Glu Asn Gly Trp 370 375 380 Phe Val Ser Gly
Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr Tyr 385 390 395 400 Leu
Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp Gly 405 410
415 Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly Phe Glu
420 425 430 Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe Tyr Val
Asp Phe 435 440 445 Leu Ser Gln Asp Lys Met Leu Leu Pro Lys Ser Ser
Ala Leu Phe Tyr 450 455 460 Gln Lys Leu Ile Glu Lys Asn Gly Phe Pro
Pro Leu Pro Glu Asn Gln 465 470 475 480 Pro Leu Glu Gly Thr Phe Pro
Cys Asp Phe Ala Trp Gly Val Val Asp 485 490 495 Asn Tyr Ile Gln Val
Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu Asn 500 505 510 Val Tyr Leu
Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val Asp 515 520 525 Gly
Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala Ala 530 535
540 Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val Thr His Phe
545 550 555 560 Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro Leu Gly
Asn Gln Ser 565 570 575 Gln Val Asn His Thr Ile Leu Gln Tyr Tyr Arg
Cys Met Ala Ser Glu 580 585 590 Leu Val Arg Val Asn Ile Thr Pro Val
Val Ala Leu Trp Gln Pro Met 595 600 605 Ala Pro Asn Gln Gly Leu Pro
Arg Leu Leu Ala Arg Gln Gly Ala Trp 610 615 620 Glu Asn Pro Tyr Thr
Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu Cys 625 630 635 640 Phe Gln
Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn Glu 645 650 655
Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu Lys 660
665 670 Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe Arg His
Ala 675 680 685 Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp
Ile Glu Pro 690 695 700 Ala Cys Pro Phe Ser Gln Lys Asp Lys Glu Val
Ala Glu Arg Val Leu 705 710 715 720 Glu Phe Asp Ile Gly Trp Leu Ala
Glu Pro Ile Phe Gly Ser Gly Asp 725 730 735 Tyr Pro Trp Val Met Arg
Asp Trp Leu Asn Gln Arg Asn Asn Phe Leu 740 745 750 Leu Pro Tyr Phe
Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr Phe 755 760 765 Asp Phe
Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser Glu 770 775 780
Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln Glu Met 785
790 795 800 Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val Ala Val
Val Pro 805 810 815 Trp Gly Leu Arg Lys Val Leu Asn Trp Leu Lys Phe
Lys Tyr Gly Asp 820 825 830 Leu Pro Met Tyr Ile Ile Ser Asn Gly Ile
Asp Asp Gly Leu His Ala 835 840 845 Glu Asp Asp Gln Leu Arg Val Tyr
Tyr Met Gln Asn Tyr Ile Asn Glu 850 855 860 Ala Leu Lys Ala His Ile
Leu Asp Gly Ile Asn Leu Cys Gly Tyr Phe 865 870 875 880 Ala Tyr Ser
Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr Arg 885 890 895 Tyr
Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr Arg 900 905
910 Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu Glu Arg
915 920 925 Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu Cys Ser Phe
Phe His 930 935 940 Thr Arg Lys Ser Leu Gly Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly 945 950 955 960 Ser Gly Gly Gly Gly Ser Leu Lys Tyr
Pro Asn Ala Ser Pro Leu Leu 965 970 975 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 980 985 990 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 995 1000 1005 Pro His
Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp 1010 1015 1020
Ala Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu 1025
1030 1035 Cys Met Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe
Asp 1040 1045 1050 Pro Glu Asn Cys Arg Phe Gln His Gln Thr Leu Glu
Asn Gly Tyr 1055 1060 1065 Asp Val Tyr His Ser Pro Gln Tyr His Phe
Leu Val Ser Leu Gly 1070 1075 1080 Arg Ala Lys Arg Ala Phe Leu Pro
Gly Met Asn Pro Pro Pro Tyr 1085 1090 1095 Ser Gln Phe Leu Ser Arg
Arg Asn Glu Ile Pro Leu Ile His Phe 1100 1105 1110 Asn Thr Pro Ile
Pro Arg Arg His Thr Gln Ser Ala Glu Asp Asp 1115 1120 1125 Ser Glu
Arg Asp Pro Leu Asn Val Leu Lys Pro Arg Ala Arg Met 1130 1135 1140
Thr Pro Ala Pro Ala Ser Cys Ser Gln Glu Leu Pro Ser Ala Glu 1145
1150 1155 Asp Asn Ser Pro Met Ala Ser Asp Pro Leu Gly Val Val Arg
Gly 1160 1165 1170 Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly Pro
Glu Gly Cys 1175 1180 1185 Arg Pro Phe Ala Lys Phe Ile 1190 1195
42227PRTHomo sapiens 42Tyr Pro Asn Ala Ser Pro Leu Leu Gly Ser Ser
Trp Gly Gly Leu Ile 1 5 10 15 His Leu Tyr Thr Ala Thr Ala Arg Asn
Ser Tyr His Leu Gln Ile His 20 25 30 Lys Asn Gly His Val Asp Gly
Ala Pro His Gln Thr Ile Tyr Ser Ala 35 40 45 Leu Met Ile Arg Ser
Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val 50 55 60 Met Ser Arg
Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly 65 70 75 80 Ser
His Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His Gln Thr Leu 85 90
95 Glu Asn Gly Tyr Asp Val Tyr His Ser Pro Gln Tyr His Phe Leu Val
100 105 110 Ser Leu Gly Arg Ala Lys Arg Ala Phe Leu Pro Gly Met Asn
Pro Pro 115 120 125 Pro Tyr Ser Gln Phe Leu Ser Arg Arg Asn Glu Ile
Pro Leu Ile His 130 135 140 Phe Asn Thr Pro Ile Pro Arg Arg His Thr
Arg Ser Ala Glu Asp Asp 145 150 155 160 Ser Glu Arg Asp Pro Leu Asn
Val Leu Lys Pro Arg Ala Arg Met Thr 165 170 175 Pro Ala Pro Ala Ser
Cys Ser Gln Glu Leu Pro Ser Ala Glu Asp Asn 180 185 190 Ser Pro Met
Ala Ser Asp Pro Leu Gly Val Val Arg Gly Gly Arg Val 195 200 205 Asn
Thr His Ala Gly Gly Thr Gly Pro Glu Gly Cys Arg Pro Phe Ala 210 215
220 Lys Phe Ile 225 43227PRTHomo sapiens 43Tyr Pro Asn Ala Ser Pro
Leu Leu Gly Ser Ser Trp Gly Gly Leu Ile 1 5 10 15 His Leu Tyr Thr
Ala Thr Ala Arg Asn Ser Tyr His Leu Gln Ile His 20 25 30 Lys Asn
Gly His Val Asp Gly Ala Pro His Gln Thr Ile Tyr Ser Ala 35 40 45
Leu Met Ile Arg Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val 50
55
60 Met Ser Arg Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly
65 70 75 80 Ser His Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His Gln
Thr Leu 85 90 95 Glu Asn Gly Tyr Asp Val Tyr His Ser Pro Gln Tyr
His Phe Leu Val 100 105 110 Ser Leu Gly Arg Ala Lys Arg Ala Phe Leu
Pro Gly Met Asn Pro Pro 115 120 125 Pro Tyr Ser Gln Phe Leu Ser Arg
Arg Asn Glu Ile Pro Leu Ile His 130 135 140 Phe Asn Thr Pro Ile Pro
Arg Arg His Thr Gln Ser Ala Glu Asp Asp 145 150 155 160 Ser Glu Arg
Asp Pro Leu Asn Val Leu Lys Pro Arg Ala Arg Met Thr 165 170 175 Pro
Ala Pro Ala Ser Cys Ser Gln Glu Leu Pro Ser Ala Glu Asp Asn 180 185
190 Ser Pro Met Ala Ser Asp Pro Leu Gly Val Val Arg Gly Gly Arg Val
195 200 205 Asn Thr His Ala Gly Gly Thr Gly Pro Glu Gly Cys Arg Pro
Phe Ala 210 215 220 Lys Phe Ile 225 44982PRTHomo sapiens 44Met Pro
Ala Ser Ala Pro Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15
Leu Ser Leu Leu Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20
25 30 Ala Glu Pro Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg
Pro 35 40 45 Pro Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe
Pro Asp Gly 50 55 60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln
Thr Glu Gly Gly Trp 65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile
Trp Asp Thr Phe Thr His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp
Ser Arg Asn Ala Ser Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu
Gln Pro Ala Thr Gly Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn
Val Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr
His Tyr Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150
155 160 Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg
Leu 165 170 175 Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val
Thr Leu Tyr 180 185 190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala
Tyr Gly Gly Trp Ala 195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg
Asp Tyr Ala Glu Leu Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val
Lys Tyr Trp Ile Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala
Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg
Gly Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270
Leu Ala His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275
280 285 Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile
Asn 290 295 300 Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln
Lys Ser Leu 305 310 315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro
Val Phe Ile Asp Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn
Leu Ser Ser Ile Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys
Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly
Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe
Arg Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395
400 Ile Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly
405 410 415 Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr
Met Tyr 420 425 430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala
Ile Lys Leu Asp 435 440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp
Ser Leu Met Asp Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile
Arg Arg Gly Leu Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp
Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys
Leu Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln
Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520
525 Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu
530 535 540 Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile
Lys Val 545 550 555 560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr
Cys Val Asp Phe Ala 565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu
Gln Glu Met His Val Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp
Ala Leu Ile Leu Pro Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His
Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val
Arg Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640
Met Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645
650 655 Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg
Leu 660 665 670 Cys Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile
Thr Met Asn 675 680 685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala
Gly His Asn Leu Leu 690 695 700 Lys Ala His Ala Leu Ala Trp His Val
Tyr Asn Glu Lys Phe Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile
Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro
Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu
Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765
Asp Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770
775 780 Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly
Thr 785 790 795 800 Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile
Leu Val Asp Ser 805 810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp
Tyr Leu Glu Val Gln Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn
Ser Pro Ser Gln Val Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys
Val Leu Asn Trp Leu Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met
Tyr Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala
Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890
895 Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr
900 905 910 Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly
Leu Tyr 915 920 925 Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser
Met Lys His Tyr 930 935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro
Gly Pro Glu Thr Leu Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe
Thr Val Cys Thr Glu Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser
Leu 980 45974PRTHomo sapiens 45Met Ser Val Leu Thr Gln Val Leu Ala
Leu Leu Leu Leu Trp Leu Thr 1 5 10 15 Gly Leu Gly Gly Arg Arg Leu
Arg Ala Glu Pro Gly Asp Gly Ala Gln 20 25 30 Thr Trp Ala Arg Phe
Ser Arg Pro Pro Ala Pro Glu Ala Ala Gly Leu 35 40 45 Phe Gln Gly
Thr Phe Pro Asp Gly Phe Leu Trp Ala Val Gly Ser Ala 50 55 60 Ala
Tyr Gln Thr Glu Gly Gly Trp Gln Gln His Gly Lys Gly Ala Ser 65 70
75 80 Ile Trp Asp Thr Phe Thr His His Pro Leu Ala Pro Pro Gly Asp
Ser 85 90 95 Arg Asn Ala Ser Leu Pro Leu Gly Ala Pro Ser Pro Leu
Gln Pro Ala 100 105 110 Thr Gly Asp Val Ala Ser Asp Ser Tyr Asn Asn
Val Phe Arg Asp Thr 115 120 125 Glu Ala Leu Arg Glu Leu Gly Val Thr
His Tyr Arg Phe Ser Ile Ser 130 135 140 Trp Ala Arg Val Leu Pro Asn
Gly Ser Ala Gly Val Pro Asn Arg Glu 145 150 155 160 Gly Leu Arg Tyr
Tyr Arg Arg Leu Leu Glu Arg Leu Arg Glu Leu Gly 165 170 175 Val Gln
Pro Val Val Thr Leu Tyr His Trp Asp Leu Pro Gln Arg Leu 180 185 190
Gln Asp Ala Tyr Gly Gly Trp Ala Asn Arg Ala Leu Ala Asp His Phe 195
200 205 Arg Asp Tyr Ala Glu Leu Cys Phe Arg His Phe Gly Gly Gln Val
Lys 210 215 220 Tyr Trp Ile Thr Ile Asp Asn Pro Tyr Val Val Ala Trp
His Gly Tyr 225 230 235 240 Ala Thr Gly Arg Leu Ala Pro Gly Ile Arg
Gly Ser Pro Arg Leu Gly 245 250 255 Tyr Leu Val Ala His Asn Leu Leu
Leu Ala His Ala Lys Val Trp His 260 265 270 Leu Tyr Asn Thr Ser Phe
Arg Pro Thr Gln Gly Gly Gln Val Ser Ile 275 280 285 Ala Leu Ser Ser
His Trp Ile Asn Pro Arg Arg Met Thr Asp His Ser 290 295 300 Ile Lys
Glu Cys Gln Lys Ser Leu Asp Phe Val Leu Gly Trp Phe Ala 305 310 315
320 Lys Pro Val Phe Ile Asp Gly Asp Tyr Pro Glu Ser Met Lys Asn Asn
325 330 335 Leu Ser Ser Ile Leu Pro Asp Phe Thr Glu Ser Glu Lys Lys
Phe Ile 340 345 350 Lys Gly Thr Ala Asp Phe Phe Ala Leu Cys Phe Gly
Pro Thr Leu Ser 355 360 365 Phe Gln Leu Leu Asp Pro His Met Lys Phe
Arg Gln Leu Glu Ser Pro 370 375 380 Asn Leu Arg Gln Leu Leu Ser Trp
Ile Asp Leu Glu Phe Asn His Pro 385 390 395 400 Gln Ile Phe Ile Val
Glu Asn Gly Trp Phe Val Ser Gly Thr Thr Lys 405 410 415 Arg Asp Asp
Ala Lys Tyr Met Tyr Tyr Leu Lys Lys Phe Ile Met Glu 420 425 430 Thr
Leu Lys Ala Ile Lys Leu Asp Gly Val Asp Val Ile Gly Tyr Thr 435 440
445 Ala Trp Ser Leu Met Asp Gly Phe Glu Trp His Arg Gly Tyr Ser Ile
450 455 460 Arg Arg Gly Leu Phe Tyr Val Asp Phe Leu Ser Gln Asp Lys
Met Leu 465 470 475 480 Leu Pro Lys Ser Ser Ala Leu Phe Tyr Gln Lys
Leu Ile Glu Lys Asn 485 490 495 Gly Phe Pro Pro Leu Pro Glu Asn Gln
Pro Leu Glu Gly Thr Phe Pro 500 505 510 Cys Asp Phe Ala Trp Gly Val
Val Asp Asn Tyr Ile Gln Val Asp Thr 515 520 525 Thr Leu Ser Gln Phe
Thr Asp Leu Asn Val Tyr Leu Trp Asp Val His 530 535 540 His Ser Lys
Arg Leu Ile Lys Val Asp Gly Val Val Thr Lys Lys Arg 545 550 555 560
Lys Ser Tyr Cys Val Asp Phe Ala Ala Ile Gln Pro Gln Ile Ala Leu 565
570 575 Leu Gln Glu Met His Val Thr His Phe Arg Phe Ser Leu Asp Trp
Ala 580 585 590 Leu Ile Leu Pro Leu Gly Asn Gln Ser Gln Val Asn His
Thr Ile Leu 595 600 605 Gln Tyr Tyr Arg Cys Met Ala Ser Glu Leu Val
Arg Val Asn Ile Thr 610 615 620 Pro Val Val Ala Leu Trp Gln Pro Met
Ala Pro Asn Gln Gly Leu Pro 625 630 635 640 Arg Leu Leu Ala Arg Gln
Gly Ala Trp Glu Asn Pro Tyr Thr Ala Leu 645 650 655 Ala Phe Ala Glu
Tyr Ala Arg Leu Cys Phe Gln Glu Leu Gly His His 660 665 670 Val Lys
Leu Trp Ile Thr Met Asn Glu Pro Tyr Thr Arg Asn Met Thr 675 680 685
Tyr Ser Ala Gly His Asn Leu Leu Lys Ala His Ala Leu Ala Trp His 690
695 700 Val Tyr Asn Glu Lys Phe Arg His Ala Gln Asn Gly Lys Ile Ser
Ile 705 710 715 720 Ala Leu Gln Ala Asp Trp Ile Glu Pro Ala Cys Pro
Phe Ser Gln Lys 725 730 735 Asp Lys Glu Val Ala Glu Arg Val Leu Glu
Phe Asp Ile Gly Trp Leu 740 745 750 Ala Glu Pro Ile Phe Gly Ser Gly
Asp Tyr Pro Trp Val Met Arg Asp 755 760 765 Trp Leu Asn Gln Arg Asn
Asn Phe Leu Leu Pro Tyr Phe Thr Glu Asp 770 775 780 Glu Lys Lys Leu
Ile Gln Gly Thr Phe Asp Phe Leu Ala Leu Ser His 785 790 795 800 Tyr
Thr Thr Ile Leu Val Asp Ser Glu Lys Glu Asp Pro Ile Lys Tyr 805 810
815 Asn Asp Tyr Leu Glu Val Gln Glu Met Thr Asp Ile Thr Trp Leu Asn
820 825 830 Ser Pro Ser Gln Val Ala Val Val Pro Trp Gly Leu Arg Lys
Val Leu 835 840 845 Asn Trp Leu Lys Phe Lys Tyr Gly Asp Leu Pro Met
Tyr Ile Ile Ser 850 855 860 Asn Gly Ile Asp Asp Gly Leu His Ala Glu
Asp Asp Gln Leu Arg Val 865 870 875 880 Tyr Tyr Met Gln Asn Tyr Ile
Asn Glu Ala Leu Lys Ala His Ile Leu 885 890 895 Asp Gly Ile Asn Leu
Cys Gly Tyr Phe Ala Tyr Ser Phe Asn Asp Arg 900 905 910 Thr Ala Pro
Arg Phe Gly Leu Tyr Arg Tyr Ala Ala Asp Gln Phe Glu 915 920 925 Pro
Lys Ala Ser Met Lys His Tyr Arg Lys Ile Ile Asp Ser Asn Gly 930 935
940 Phe Pro Gly Pro Glu Thr Leu Glu Arg Phe Cys Pro Glu Glu Phe Thr
945 950 955 960 Val Cys Thr Glu Cys Ser Phe Phe His Thr Arg Lys Ser
Leu 965 970 464380DNAHomo sapiens 46atgcccgcca gcgccccgcc
gcgccgcccg cggccgccgc cgccgtcgct gtcgctgctg 60ctggtgctgc tgggcctggg
cggccgccgc ctgcgtgcgg agccgggcga cggcgcgcag 120acctgggccc
gtttctcgcg gcctcctgcc cccgaggccg cgggcctctt ccagggcacc
180ttccccgacg gcttcctctg ggccgtgggc agcgccgcct accagaccga
gggcggctgg 240cagcagcacg gcaagggtgc gtccatctgg gatacgttca
cccaccaccc cctggcaccc 300ccgggagact cccggaacgc cagtctgccg
ttgggcgccc cgtcgccgct gcagcccgcc 360accggggacg tagccagcga
cagctacaac aacgtcttcc gcgacacgga ggcgctgcgc 420gagctcgggg
tcactcacta ccgcttctcc atctcgtggg cgcgagtgct ccccaatggc
480agcgcgggcg tccccaaccg cgaggggctg cgctactacc ggcgcctgct
ggagcggctg 540cgggagctgg gcgtgcagcc cgtggtcacc ctgtaccact
gggacctgcc ccagcgcctg 600caggacgcct acggcggctg ggccaaccgc
gccctggccg accacttcag ggattacgcg 660gagctctgct tccgccactt
cggcggtcag gtcaagtact ggatcaccat cgacaacccc 720tacgtggtgg
cctggcacgg ctacgccacc gggcgcctgg cccccggcat ccggggcagc
780ccgcggctcg ggtacctggt ggcgcacaac ctcctcctgg ctcatgccaa
agtctggcat 840ctctacaata cttctttccg tcccactcag ggaggtcagg
tgtccattgc cctaagctct 900cactggatca atcctcgaag aatgaccgac
cacagcatca aagaatgtca aaaatctctg 960gactttgtac taggttggtt
tgccaaaccc gtatttattg atggtgacta tcccgagagc 1020atgaagaata
acctttcatc tattctgcct gattttactg aatctgagaa aaagttcatc
1080aaaggaactg ctgacttttt tgctctttgc tttggaccca ccttgagttt
tcaacttttg 1140gaccctcaca tgaagttccg
ccaattggaa tctcccaacc tgaggcaact gctttcctgg 1200attgaccttg
aatttaacca tcctcaaata tttattgtgg aaaatggctg gtttgtctca
1260gggaccacca agagagatga tgccaaatat atgtattacc tcaaaaagtt
catcatggaa 1320accttaaaag ccatcaagct ggatggggtg gatgtcatcg
ggtataccgc atggtccctc 1380atggatggtt tcgagtggca cagaggttac
agcatcaggc gtggactctt ctatgttgac 1440tttctaagcc aggacaagat
gttgttgcca aagtcttcag ccttgttcta ccaaaagctg 1500atagagaaaa
atggcttccc tcctttacct gaaaatcagc ccctagaagg gacatttccc
1560tgtgactttg cttggggagt tgttgacaac tacattcaag tagataccac
tctgtctcag 1620tttaccgacc tgaatgttta cctgtgggat gtccaccaca
gtaaaaggct tattaaagtg 1680gatggggttg tgaccaagaa gaggaaatcc
tactgtgttg actttgctgc catccagccc 1740cagatcgctt tactccagga
aatgcacgtt acacattttc gcttctccct ggactgggcc 1800ctgattctcc
ctctgggtaa ccagtcccag gtgaaccaca ccatcctgca gtactatcgc
1860tgcatggcca gcgagcttgt ccgtgtcaac atcaccccag tggtggccct
gtggcagcct 1920atggccccga accaaggact gccgcgcctc ctggccaggc
agggcgcctg ggagaacccc 1980tacactgccc tggcctttgc agagtatgcc
cgactgtgct ttcaagagct cggccatcac 2040gtcaagcttt ggataacgat
gaatgagccg tatacaagga atatgacata cagtgctggc 2100cacaaccttc
tgaaggccca tgccctggct tggcatgtgt acaatgaaaa gtttaggcat
2160gctcagaatg ggaaaatatc catagccttg caggctgatt ggatagaacc
tgcctgccct 2220ttctcccaaa aggacaaaga ggtggccgag agagttttgg
aatttgacat tggctggctg 2280gctgagccca ttttcggctc tggagattat
ccatgggtga tgagggactg gctgaaccaa 2340agaaacaatt ttcttcttcc
ttatttcact gaagatgaaa aaaagctaat ccagggtacc 2400tttgactttt
tggctttaag ccattatacc accatccttg tagactcaga aaaagaagat
2460ccaataaaat acaatgatta cctagaagtg caagaaatga ccgacatcac
gtggctcaac 2520tcccccagtc aggtggcggt agtgccctgg gggttgcgca
aagtgctgaa ctggctgaag 2580ttcaagtacg gagacctccc catgtacata
atatccaacg gaatcgatga cgggctgcat 2640gctgaggacg accagctgag
ggtgtattat atgcagaatt acataaacga agctctcaaa 2700gcccacatac
tggatggtat caatctttgc ggatactttg cttattcgtt taacgaccgc
2760acagctccga ggtttggcct ctatcgttat gctgcagatc agtttgagcc
caaggcatcc 2820atgaaacatt acaggaaaat tattgacagc aatggtttcc
cgggcccaga aactctggaa 2880agattttgtc cagaagaatt caccgtgtgt
actgagtgca gtttttttca cacccgaaag 2940tctttaggat ccggaggtgg
aggttcagga ggtggaggtt caggaggtgg aggttcactt 3000aagtatccca
atgcctcccc actgctcggc tccagctggg gtggcctgat ccacctgtac
3060acagccacag ccaggaacag ctaccacctg cagatccaca agaatggcca
tgtggatggc 3120gcaccccatc agaccatcta cagtgccctg atgatcagat
cagaggatgc tggctttgtg 3180gtgattacag gtgtgatgag cagaagatac
ctctgcatgg atttcagagg caacattttt 3240ggatcacact atttcgaccc
ggagaactgc aggttccaac accagacgct ggaaaacggg 3300tacgacgtct
accactctcc tcagtatcac ttcctggtca gtctgggccg ggcgaagaga
3360gccttcctgc caggcatgaa cccacccccg tactcccagt tcctgtcccg
gaggaacgag 3420atccccctaa ttcacttcaa cacccccata ccacggcggc
acacccagag cgccgaggac 3480gactcggagc gggaccccct gaacgtgctg
aagccccggg cccggatgac cccggccccg 3540gcctcctgtt cacaggagct
cccgagcgcc gaggacaaca gcccgatggc cagtgaccca 3600ttaggggtgg
tcaggggcgg tcgagtgaac acgcacgctg ggggaacggg cccggaaggc
3660tgccgcccct tcgccaagtt catcggaggt ggaggttcaa aaacccacac
gtgtcctcct 3720tgtcctgccc cagaagcagc aggtggtcca tcagtttttc
ttttccctcc caaacccaag 3780gatacgctga tgatctctcg cacgcctgag
gtgacatgcg tcgtagtaga cgtgagccac 3840gaagatcccg aggtgaagtt
caattggtat gtggacggag tagaagtgca taacgcgaaa 3900actaagccgc
gcgaggaaca atataacagt acttacaggg tggtatccgt gctcacagtc
3960ctgcaccagg actggctgaa cggtaaggaa tacaagtgca aagtaagcaa
caaggcactt 4020cccgcgccta ttgagaaaac aatctccaag gcgaagggac
aaccaagaga acctcaggtt 4080tacactctcc cgccttccag ggaagagatg
accaaaaatc aagtttccct gacttgcctc 4140gtcaaaggat tctacccttc
cgacattgct gttgaatggg aaagcaatgg acaaccagag 4200aacaactaca
agacaacacc cccggtgctg gatagtgacg gatctttctt tctctactca
4260aagctgaccg tggataagtc caggtggcag cagggaaacg tgttttcctg
ctctgtcatg 4320catgaagcgc tgcataatca ctatacccag aagtctctga
gcttgagccc aggcaagtaa 4380471459PRTHomo sapiens 47Met Pro Ala Ser
Ala Pro Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser
Leu Leu Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30
Ala Glu Pro Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35
40 45 Pro Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp
Gly 50 55 60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu
Gly Gly Trp 65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp
Thr Phe Thr His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg
Asn Ala Ser Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro
Ala Thr Gly Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe
Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr
Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160
Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165
170 175 Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu
Tyr 180 185 190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly
Gly Trp Ala 195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr
Ala Glu Leu Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr
Trp Ile Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His
Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser
Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala
His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285
Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290
295 300 Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser
Leu 305 310 315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe
Ile Asp Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser
Ser Ile Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile
Lys Gly Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr
Leu Ser Phe Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln
Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile
Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410
415 Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr
420 425 430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys
Leu Asp 435 440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu
Met Asp Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg
Gly Leu Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met
Leu Leu Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile
Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu
Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520 525 Asp
Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu 530 535
540 Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val
545 550 555 560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val
Asp Phe Ala 565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu
Met His Val Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp Ala Leu
Ile Leu Pro Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His Thr Ile
Leu Gln Tyr Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val Arg Val
Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640 Met Ala
Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645 650 655
Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660
665 670 Cys Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met
Asn 675 680 685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His
Asn Leu Leu 690 695 700 Lys Ala His Ala Leu Ala Trp His Val Tyr Asn
Glu Lys Phe Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile Ser Ile
Ala Leu Gln Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro Phe Ser
Gln Lys Asp Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu Phe Asp
Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765 Asp Tyr
Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770 775 780
Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr 785
790 795 800 Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val
Asp Ser 805 810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu
Glu Val Gln Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn Ser Pro
Ser Gln Val Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys Val Leu
Asn Trp Leu Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met Tyr Ile
Ile Ser Asn Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala Glu Asp
Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890 895 Glu
Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900 905
910 Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr
915 920 925 Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys
His Tyr 930 935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro
Glu Thr Leu Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe Thr Val
Cys Thr Glu Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser Leu Gly
Ser Gly Gly Gly Gly Ser Gly Gly Gly 980 985 990 Gly Ser Gly Gly Gly
Gly Ser Leu Lys Tyr Pro Asn Ala Ser Pro Leu 995 1000 1005 Leu Gly
Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr 1010 1015 1020
Ala Arg Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val 1025
1030 1035 Asp Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile
Arg 1040 1045 1050 Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val
Met Ser Arg 1055 1060 1065 Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn
Ile Phe Gly Ser His 1070 1075 1080 Tyr Phe Asp Pro Glu Asn Cys Arg
Phe Gln His Gln Thr Leu Glu 1085 1090 1095 Asn Gly Tyr Asp Val Tyr
His Ser Pro Gln Tyr His Phe Leu Val 1100 1105 1110 Ser Leu Gly Arg
Ala Lys Arg Ala Phe Leu Pro Gly Met Asn Pro 1115 1120 1125 Pro Pro
Tyr Ser Gln Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu 1130 1135 1140
Ile His Phe Asn Thr Pro Ile Pro Arg Arg His Thr Gln Ser Ala 1145
1150 1155 Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys Pro
Arg 1160 1165 1170 Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln
Glu Leu Pro 1175 1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu Gly Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn Thr
His Ala Gly Gly Thr Gly Pro 1205 1210 1215 Glu Gly Cys Arg Pro Phe
Ala Lys Phe Ile Gly Gly Gly Gly Ser 1220 1225 1230 Lys Thr His Thr
Cys Pro Pro Cys Pro Ala Pro Glu Ala Ala Gly 1235 1240 1245 Gly Pro
Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 1250 1255 1260
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val 1265
1270 1275 Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp
Gly 1280 1285 1290 Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu
Glu Gln Tyr 1295 1300 1305 Asn Ser Thr Tyr Arg Val Val Ser Val Leu
Thr Val Leu His Gln 1310 1315 1320 Asp Trp Leu Asn Gly Lys Glu Tyr
Lys Cys Lys Val Ser Asn Lys 1325 1330 1335 Ala Leu Pro Ala Pro Ile
Glu Lys Thr Ile Ser Lys Ala Lys Gly 1340 1345 1350 Gln Pro Arg Glu
Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu 1355 1360 1365 Glu Met
Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly 1370 1375 1380
Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln 1385
1390 1395 Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser
Asp 1400 1405 1410 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp
Lys Ser Arg 1415 1420 1425 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser
Val Met His Glu Ala 1430 1435 1440 Leu His Asn His Tyr Thr Gln Lys
Ser Leu Ser Leu Ser Pro Gly 1445 1450 1455 Lys 484353DNAHomo
sapiens 48atgcccgcca gcgccccgcc gcgccgcccg cggccgccgc cgccgtcgct
gtcgctgctg 60ctggtgctgc tgggcctggg cggccgccgc ctgcgtgcgg agccgggcga
cggcgcgcag 120acctgggccc gtttctcgcg gcctcctgcc cccgaggccg
cgggcctctt ccagggcacc 180ttccccgacg gcttcctctg ggccgtgggc
agcgccgcct accagaccga gggcggctgg 240cagcagcacg gcaagggtgc
gtccatctgg gatacgttca cccaccaccc cctggcaccc 300ccgggagact
cccggaacgc cagtctgccg ttgggcgccc cgtcgccgct gcagcccgcc
360accggggacg tagccagcga cagctacaac aacgtcttcc gcgacacgga
ggcgctgcgc 420gagctcgggg tcactcacta ccgcttctcc atctcgtggg
cgcgagtgct ccccaatggc 480agcgcgggcg tccccaaccg cgaggggctg
cgctactacc ggcgcctgct ggagcggctg 540cgggagctgg gcgtgcagcc
cgtggtcacc ctgtaccact gggacctgcc ccagcgcctg 600caggacgcct
acggcggctg ggccaaccgc gccctggccg accacttcag ggattacgcg
660gagctctgct tccgccactt cggcggtcag gtcaagtact ggatcaccat
cgacaacccc 720tacgtggtgg cctggcacgg ctacgccacc gggcgcctgg
cccccggcat ccggggcagc 780ccgcggctcg ggtacctggt ggcgcacaac
ctcctcctgg ctcatgccaa agtctggcat 840ctctacaata cttctttccg
tcccactcag ggaggtcagg tgtccattgc cctaagctct 900cactggatca
atcctcgaag aatgaccgac cacagcatca aagaatgtca aaaatctctg
960gactttgtac taggttggtt tgccaaaccc gtatttattg atggtgacta
tcccgagagc 1020atgaagaata acctttcatc tattctgcct gattttactg
aatctgagaa aaagttcatc 1080aaaggaactg ctgacttttt tgctctttgc
tttggaccca ccttgagttt tcaacttttg 1140gaccctcaca tgaagttccg
ccaattggaa tctcccaacc tgaggcaact gctttcctgg 1200attgaccttg
aatttaacca tcctcaaata tttattgtgg aaaatggctg gtttgtctca
1260gggaccacca agagagatga tgccaaatat atgtattacc tcaaaaagtt
catcatggaa 1320accttaaaag ccatcaagct ggatggggtg gatgtcatcg
ggtataccgc atggtccctc 1380atggatggtt tcgagtggca cagaggttac
agcatcaggc gtggactctt ctatgttgac 1440tttctaagcc aggacaagat
gttgttgcca aagtcttcag ccttgttcta ccaaaagctg 1500atagagaaaa
atggcttccc tcctttacct gaaaatcagc ccctagaagg gacatttccc
1560tgtgactttg cttggggagt tgttgacaac tacattcaag tagataccac
tctgtctcag 1620tttaccgacc tgaatgttta cctgtgggat gtccaccaca
gtaaaaggct tattaaagtg 1680gatggggttg tgaccaagaa gaggaaatcc
tactgtgttg actttgctgc catccagccc 1740cagatcgctt tactccagga
aatgcacgtt acacattttc gcttctccct ggactgggcc 1800ctgattctcc
ctctgggtaa ccagtcccag gtgaaccaca ccatcctgca gtactatcgc
1860tgcatggcca gcgagcttgt ccgtgtcaac atcaccccag tggtggccct
gtggcagcct 1920atggccccga accaaggact gccgcgcctc ctggccaggc
agggcgcctg ggagaacccc 1980tacactgccc tggcctttgc agagtatgcc
cgactgtgct ttcaagagct cggccatcac 2040gtcaagcttt ggataacgat
gaatgagccg tatacaagga atatgacata cagtgctggc 2100cacaaccttc
tgaaggccca tgccctggct
tggcatgtgt acaatgaaaa gtttaggcat 2160gctcagaatg ggaaaatatc
catagccttg caggctgatt ggatagaacc tgcctgccct 2220ttctcccaaa
aggacaaaga ggtggccgag agagttttgg aatttgacat tggctggctg
2280gctgagccca ttttcggctc tggagattat ccatgggtga tgagggactg
gctgaaccaa 2340agaaacaatt ttcttcttcc ttatttcact gaagatgaaa
aaaagctaat ccagggtacc 2400tttgactttt tggctttaag ccattatacc
accatccttg tagactcaga aaaagaagat 2460ccaataaaat acaatgatta
cctagaagtg caagaaatga ccgacatcac gtggctcaac 2520tcccccagtc
aggtggcggt agtgccctgg gggttgcgca aagtgctgaa ctggctgaag
2580ttcaagtacg gagacctccc catgtacata atatccaacg gaatcgatga
cgggctgcat 2640gctgaggacg accagctgag ggtgtattat atgcagaatt
acataaacga agctctcaaa 2700gcccacatac tggatggtat caatctttgc
ggatactttg cttattcgtt taacgaccgc 2760acagctccga ggtttggcct
ctatcgttat gctgcagatc agtttgagcc caaggcatcc 2820atgaaacatt
acaggaaaat tattgacagc aatggtttcc cgggcccaga aactctggaa
2880agattttgtc cagaagaatt caccgtgtgt actgagtgca gtttttttca
cacccgaaag 2940tctttaggat ccggaggtgg aggttcagga ggtggaggtt
caggaggtgg aggttcactt 3000aagtatccca atgcctcccc actgctcggc
tccagctggg gtggcctgat ccacctgtac 3060acagccacag ccaggaacag
ctaccacctg cagatccaca agaatggcca tgtggatggc 3120gcaccccatc
agaccatcta cagtgccctg atgatcagat cagaggatgc tggctttgtg
3180gtgattacag gtgtgatgag cagaagatac ctctgcatgg atttcagagg
caacattttt 3240ggatcacact atttcgaccc ggagaactgc aggttccaac
accagacgct ggaaaacggg 3300tacgacgtct accactctcc tcagtatcac
ttcctggtca gtctgggccg ggcgaagaga 3360gccttcctgc caggcatgaa
cccacccccg tactcccagt tcctgtcccg gaggaacgag 3420atccccctaa
ttcacttcaa cacccccata ccacggcggc acacccagag cgccgaggac
3480gactcggagc gggaccccct gaacgtgctg aagccccggg cccggatgac
cccggccccg 3540gcctcctgtt cacaggagct cccgagcgcc gaggacaaca
gcccgatggc cagtgaccca 3600ttaggggtgg tcaggggcgg tcgagtgaac
acgcacgctg ggggaacggg cccggaaggc 3660tgccgcccct tcgccaagtt
catcggaggt ggaggttcag ccccagaagc agcaggtggt 3720ccatcagttt
ttcttttccc tcccaaaccc aaggatacgc tgatgatctc tcgcacgcct
3780gaggtgacat gcgtcgtagt agacgtgagc cacgaagatc ccgaggtgaa
gttcaattgg 3840tatgtggacg gagtagaagt gcataacgcg aaaactaagc
cgcgcgagga acaatataac 3900agtacttaca gggtggtatc cgtgctcaca
gtcctgcacc aggactggct gaacggtaag 3960gaatacaagt gcaaagtaag
caacaaggca cttcccgcgc ctattgagaa aacaatctcc 4020aaggcgaagg
gacaaccaag agaacctcag gtttacactc tcccgccttc cagggaagag
4080atgaccaaaa atcaagtttc cctgacttgc ctcgtcaaag gattctaccc
ttccgacatt 4140gctgttgaat gggaaagcaa tggacaacca gagaacaact
acaagacaac acccccggtg 4200ctggatagtg acggatcttt ctttctctac
tcaaagctga ccgtggataa gtccaggtgg 4260cagcagggaa acgtgttttc
ctgctctgtc atgcatgaag cgctgcataa tcactatacc 4320cagaagtctc
tgagcttgag cccaggcaag taa 4353491450PRTHomo sapiens 49Met Pro Ala
Ser Ala Pro Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu
Ser Leu Leu Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25
30 Ala Glu Pro Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro
35 40 45 Pro Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro
Asp Gly 50 55 60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr
Glu Gly Gly Trp 65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp
Asp Thr Phe Thr His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser
Arg Asn Ala Ser Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln
Pro Ala Thr Gly Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val
Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His
Tyr Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155
160 Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu
165 170 175 Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr
Leu Tyr 180 185 190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr
Gly Gly Trp Ala 195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp
Tyr Ala Glu Leu Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys
Tyr Trp Ile Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp
His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly
Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu
Ala His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280
285 Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn
290 295 300 Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys
Ser Leu 305 310 315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val
Phe Ile Asp Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu
Ser Ser Ile Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe
Ile Lys Gly Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro
Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg
Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400
Ile Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405
410 415 Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met
Tyr 420 425 430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile
Lys Leu Asp 435 440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser
Leu Met Asp Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg
Arg Gly Leu Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys
Met Leu Leu Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu
Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro
Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520 525
Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu 530
535 540 Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys
Val 545 550 555 560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys
Val Asp Phe Ala 565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln
Glu Met His Val Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp Ala
Leu Ile Leu Pro Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His Thr
Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val Arg
Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640 Met
Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645 650
655 Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu
660 665 670 Cys Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr
Met Asn 675 680 685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly
His Asn Leu Leu 690 695 700 Lys Ala His Ala Leu Ala Trp His Val Tyr
Asn Glu Lys Phe Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile Ser
Ile Ala Leu Gln Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro Phe
Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu Phe
Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765 Asp
Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770 775
780 Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr
785 790 795 800 Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu
Val Asp Ser 805 810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr
Leu Glu Val Gln Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn Ser
Pro Ser Gln Val Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys Val
Leu Asn Trp Leu Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met Tyr
Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala Glu
Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890 895
Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900
905 910 Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu
Tyr 915 920 925 Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met
Lys His Tyr 930 935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly
Pro Glu Thr Leu Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe Thr
Val Cys Thr Glu Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser Leu
Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly 980 985 990 Gly Ser Gly Gly
Gly Gly Ser Leu Lys Tyr Pro Asn Ala Ser Pro Leu 995 1000 1005 Leu
Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr 1010 1015
1020 Ala Arg Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val
1025 1030 1035 Asp Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met
Ile Arg 1040 1045 1050 Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly
Val Met Ser Arg 1055 1060 1065 Arg Tyr Leu Cys Met Asp Phe Arg Gly
Asn Ile Phe Gly Ser His 1070 1075 1080 Tyr Phe Asp Pro Glu Asn Cys
Arg Phe Gln His Gln Thr Leu Glu 1085 1090 1095 Asn Gly Tyr Asp Val
Tyr His Ser Pro Gln Tyr His Phe Leu Val 1100 1105 1110 Ser Leu Gly
Arg Ala Lys Arg Ala Phe Leu Pro Gly Met Asn Pro 1115 1120 1125 Pro
Pro Tyr Ser Gln Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu 1130 1135
1140 Ile His Phe Asn Thr Pro Ile Pro Arg Arg His Thr Gln Ser Ala
1145 1150 1155 Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys
Pro Arg 1160 1165 1170 Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser
Gln Glu Leu Pro 1175 1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala
Ser Asp Pro Leu Gly Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn
Thr His Ala Gly Gly Thr Gly Pro 1205 1210 1215 Glu Gly Cys Arg Pro
Phe Ala Lys Phe Ile Gly Gly Gly Gly Ser 1220 1225 1230 Ala Pro Glu
Ala Ala Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 1235 1240 1245 Lys
Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr 1250 1255
1260 Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe
1265 1270 1275 Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys
Thr Lys 1280 1285 1290 Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg
Val Val Ser Val 1295 1300 1305 Leu Thr Val Leu His Gln Asp Trp Leu
Asn Gly Lys Glu Tyr Lys 1310 1315 1320 Cys Lys Val Ser Asn Lys Ala
Leu Pro Ala Pro Ile Glu Lys Thr 1325 1330 1335 Ile Ser Lys Ala Lys
Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 1340 1345 1350 Leu Pro Pro
Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu 1355 1360 1365 Thr
Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu 1370 1375
1380 Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
1385 1390 1395 Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser
Lys Leu 1400 1405 1410 Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn
Val Phe Ser Cys 1415 1420 1425 Ser Val Met His Glu Ala Leu His Asn
His Tyr Thr Gln Lys Ser 1430 1435 1440 Leu Ser Leu Ser Pro Gly Lys
1445 1450 50 1449DNAHomo sapiens 50atgttggggg cccgcctcag gctctgggtc
tgtgccttgt gcagcgtctg cagcatgagc 60gtcctcagag cctatcccaa tgcctcccca
ctgctcggct ccagctgggg tggcctgatc 120cacctgtaca cagccacagc
caggaacagc taccacctgc agatccacaa gaatggccat 180gtggatggcg
caccccatca gaccatctac agtgccctga tgatcagatc agaggatgct
240ggctttgtgg tgattacagg tgtgatgagc agaagatacc tctgcatgga
tttcagaggc 300aacatttttg gatcacacta tttcgacccg gagaactgca
ggttccaaca ccagacgctg 360gaaaacgggt acgacgtcta ccactctcct
cagtatcact tcctggtcag tctgggccgg 420gcgaagagag ccttcctgcc
aggcatgaac ccacccccgt actcccagtt cctgtcccgg 480aggaacgaga
tccccctaat tcacttcaac acccccatac cacggcggca cacccagagc
540gccgaggacg actcggagcg ggaccccctg aacgtgctga agccccgggc
ccggatgacc 600ccggccccgg cctcctgttc acaggagctc ccgagcgccg
aggacaacag cccgatggcc 660agtgacccat taggggtggt caggggcggt
cgagtgaaca cgcacgctgg gggaacgggc 720ccggaaggct gccgcccctt
cgccaagttc atcggaggtg gaggttcaaa aacccacacg 780tgtcctcctt
gtcctgcccc agaagcagca ggtggtccat cagtttttct tttccctccc
840aaacccaagg atacgctgat gatctctcgc acgcctgagg tgacatgcgt
cgtagtagac 900gtgagccacg aagatcccga ggtgaagttc aattggtatg
tggacggagt agaagtgcat 960aacgcgaaaa ctaagccgcg cgaggaacaa
tataacagta cttacagggt ggtatccgtg 1020ctcacagtcc tgcaccagga
ctggctgaac ggtaaggaat acaagtgcaa agtaagcaac 1080aaggcacttc
ccgcgcctat tgagaaaaca atctccaagg cgaagggaca accaagagaa
1140cctcaggttt acactctccc gccttccagg gaagagatga ccaaaaatca
agtttccctg 1200acttgcctcg tcaaaggatt ctacccttcc gacattgctg
ttgaatggga aagcaatgga 1260caaccagaga acaactacaa gacaacaccc
ccggtgctgg atagtgacgg atctttcttt 1320ctctactcaa agctgaccgt
ggataagtcc aggtggcagc agggaaacgt gttttcctgc 1380tctgtcatgc
atgaagcgct gcataatcac tatacccaga agtctctgag cttgagccca
1440ggcaagtaa 144951482PRTHomo sapiens 51Met Leu Gly Ala Arg Leu
Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser
Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser
Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45
Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50
55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp
Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr
Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr
Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu
Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu
Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met
Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn
Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175
His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180
185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser
Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His
Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala
Lys Phe Ile Gly Gly Gly Gly Ser
245 250 255 Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Ala Ala
Gly Gly 260 265 270 Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp
Thr Leu Met Ile 275 280 285 Ser Arg Thr Pro Glu Val Thr Cys Val Val
Val Asp Val Ser His Glu 290 295 300 Asp Pro Glu Val Lys Phe Asn Trp
Tyr Val Asp Gly Val Glu Val His 305 310 315 320 Asn Ala Lys Thr Lys
Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg 325 330 335 Val Val Ser
Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys 340 345 350 Glu
Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu 355 360
365 Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr
370 375 380 Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
Ser Leu 385 390 395 400 Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp
Ile Ala Val Glu Trp 405 410 415 Glu Ser Asn Gly Gln Pro Glu Asn Asn
Tyr Lys Thr Thr Pro Pro Val 420 425 430 Leu Asp Ser Asp Gly Ser Phe
Phe Leu Tyr Ser Lys Leu Thr Val Asp 435 440 445 Lys Ser Arg Trp Gln
Gln Gly Asn Val Phe Ser Cys Ser Val Met His 450 455 460 Glu Ala Leu
His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro 465 470 475 480
Gly Lys 521422DNAHomo sapiens 52atgttggggg cccgcctcag gctctgggtc
tgtgccttgt gcagcgtctg cagcatgagc 60gtcctcagag cctatcccaa tgcctcccca
ctgctcggct ccagctgggg tggcctgatc 120cacctgtaca cagccacagc
caggaacagc taccacctgc agatccacaa gaatggccat 180gtggatggcg
caccccatca gaccatctac agtgccctga tgatcagatc agaggatgct
240ggctttgtgg tgattacagg tgtgatgagc agaagatacc tctgcatgga
tttcagaggc 300aacatttttg gatcacacta tttcgacccg gagaactgca
ggttccaaca ccagacgctg 360gaaaacgggt acgacgtcta ccactctcct
cagtatcact tcctggtcag tctgggccgg 420gcgaagagag ccttcctgcc
aggcatgaac ccacccccgt actcccagtt cctgtcccgg 480aggaacgaga
tccccctaat tcacttcaac acccccatac cacggcggca cacccagagc
540gccgaggacg actcggagcg ggaccccctg aacgtgctga agccccgggc
ccggatgacc 600ccggccccgg cctcctgttc acaggagctc ccgagcgccg
aggacaacag cccgatggcc 660agtgacccat taggggtggt caggggcggt
cgagtgaaca cgcacgctgg gggaacgggc 720ccggaaggct gccgcccctt
cgccaagttc atcggaggtg gaggttcagc cccagaagca 780gcaggtggtc
catcagtttt tcttttccct cccaaaccca aggatacgct gatgatctct
840cgcacgcctg aggtgacatg cgtcgtagta gacgtgagcc acgaagatcc
cgaggtgaag 900ttcaattggt atgtggacgg agtagaagtg cataacgcga
aaactaagcc gcgcgaggaa 960caatataaca gtacttacag ggtggtatcc
gtgctcacag tcctgcacca ggactggctg 1020aacggtaagg aatacaagtg
caaagtaagc aacaaggcac ttcccgcgcc tattgagaaa 1080acaatctcca
aggcgaaggg acaaccaaga gaacctcagg tttacactct cccgccttcc
1140agggaagaga tgaccaaaaa tcaagtttcc ctgacttgcc tcgtcaaagg
attctaccct 1200tccgacattg ctgttgaatg ggaaagcaat ggacaaccag
agaacaacta caagacaaca 1260cccccggtgc tggatagtga cggatctttc
tttctctact caaagctgac cgtggataag 1320tccaggtggc agcagggaaa
cgtgttttcc tgctctgtca tgcatgaagc gctgcataat 1380cactataccc
agaagtctct gagcttgagc ccaggcaagt aa 142253473PRTHomo sapiens 53Met
Leu Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10
15 Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu
20 25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr
Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His
Val Asp Gly Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met
Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val
Met Ser Arg Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile
Phe Gly Ser His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln
His Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro
Gln Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140
Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145
150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro
Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp
Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala
Pro Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn
Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly
Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly
Cys Arg Pro Phe Ala Lys Phe Ile Gly Gly Gly Gly Ser 245 250 255 Ala
Pro Glu Ala Ala Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 260 265
270 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val
275 280 285 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn
Trp Tyr 290 295 300 Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys
Pro Arg Glu Glu 305 310 315 320 Gln Tyr Asn Ser Thr Tyr Arg Val Val
Ser Val Leu Thr Val Leu His 325 330 335 Gln Asp Trp Leu Asn Gly Lys
Glu Tyr Lys Cys Lys Val Ser Asn Lys 340 345 350 Ala Leu Pro Ala Pro
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 355 360 365 Pro Arg Glu
Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met 370 375 380 Thr
Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 385 390
395 400 Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn
Asn 405 410 415 Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser
Phe Phe Leu 420 425 430 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp
Gln Gln Gly Asn Val 435 440 445 Phe Ser Cys Ser Val Met His Glu Ala
Leu His Asn His Tyr Thr Gln 450 455 460 Lys Ser Leu Ser Leu Ser Pro
Gly Lys 465 470 541228PRTHomo sapiens 54Met Pro Ala Ser Ala Pro Pro
Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu
Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro
Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro
Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55
60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp
65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr
His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser
Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly
Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr
Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser
Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly
Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu
Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185
190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala
195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu
Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr
Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala
Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu
Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys
Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly
Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro
Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310
315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly
Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu
Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr
Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe
Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser
Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu
Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe
Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430
Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435
440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly
Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe
Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro
Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn
Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Thr
Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520 525 Asp Asn Tyr Ile
Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu 530 535 540 Asn Val
Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val 545 550 555
560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala
565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val
Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro
Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His Thr Ile Leu Gln Tyr
Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val Arg Val Asn Ile Thr
Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640 Met Ala Pro Asn Gln
Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645 650 655 Trp Glu Asn
Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660 665 670 Cys
Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn 675 680
685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu
690 695 700 Lys Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe
Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln
Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro Phe Ser Gln Lys Asp
Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu Phe Asp Ile Gly Trp
Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765 Asp Tyr Pro Trp Val
Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770 775 780 Leu Leu Pro
Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr 785 790 795 800
Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser 805
810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln
Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val
Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu
Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met Tyr Ile Ile Ser Asn
Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala Glu Asp Asp Gln Leu
Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890 895 Glu Ala Leu Lys
Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900 905 910 Phe Ala
Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr 915 920 925
Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr 930
935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu
Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu
Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser Leu Gly Ser Gly Gly
Gly Gly Ser Gly Gly Gly 980 985 990 Gly Ser Gly Gly Gly Gly Ser Leu
Lys Tyr Pro Asn Ala Ser Pro Leu 995 1000 1005 Leu Gly Ser Ser Trp
Gly Gly Leu Ile His Leu Tyr Thr Ala Thr 1010 1015 1020 Ala Arg Asn
Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val 1025 1030 1035 Asp
Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg 1040 1045
1050 Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val Met Ser Arg
1055 1060 1065 Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly
Ser His 1070 1075 1080 Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His
Gln Thr Leu Glu 1085 1090 1095 Asn Gly Tyr Asp Val Tyr His Ser Pro
Gln Tyr His Phe Leu Val 1100 1105 1110 Ser Leu Gly Arg Ala Lys Arg
Ala Phe Leu Pro Gly Met Asn Pro 1115 1120 1125 Pro Pro Tyr Ser Gln
Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu 1130 1135 1140 Ile His Phe
Asn Thr Pro Ile Pro Arg Arg His Thr Gln Ser Ala 1145 1150 1155 Glu
Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys Pro Arg 1160 1165
1170 Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln Glu Leu Pro
1175 1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu
Gly Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn Thr His Ala Gly
Gly Thr Gly Pro 1205 1210 1215 Glu Gly Cys Arg Pro Phe Ala Lys Phe
Ile 1220 1225 55 1228PRTHomo sapiens 55Met Pro Ala Ser Ala Pro Pro
Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu
Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro
Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro
Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55
60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp
65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr
His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser
Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly
Asp Val Ala Ser Asp Ser 115 120 125 Tyr
Asn Asn Val Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135
140 Thr His Tyr Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly
145 150 155 160 Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr
Arg Arg Leu 165 170 175 Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro
Val Val Thr Leu Tyr 180 185 190 His Trp Asp Leu Pro Gln Arg Leu Gln
Asp Ala Tyr Gly Gly Trp Ala 195 200 205 Asn Arg Ala Leu Ala Asp His
Phe Arg Asp Tyr Ala Glu Leu Cys Phe 210 215 220 Arg His Phe Gly Gly
Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val
Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255
Ile Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260
265 270 Leu Ala His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg
Pro 275 280 285 Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His
Trp Ile Asn 290 295 300 Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu
Cys Gln Lys Ser Leu 305 310 315 320 Asp Phe Val Leu Gly Trp Phe Ala
Lys Pro Val Phe Ile Asp Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys
Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu
Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys
Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met 370 375 380
Lys Phe Arg Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385
390 395 400 Ile Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu
Asn Gly 405 410 415 Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala
Lys Tyr Met Tyr 420 425 430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu
Lys Ala Ile Lys Leu Asp 435 440 445 Gly Val Asp Val Ile Gly Tyr Thr
Ala Trp Ser Leu Met Asp Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr
Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser
Gln Asp Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr
Gln Lys Leu Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505
510 Gln Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val
515 520 525 Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr
Asp Leu 530 535 540 Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg
Leu Ile Lys Val 545 550 555 560 Asp Gly Val Val Thr Lys Lys Arg Lys
Ser Tyr Cys Val Asp Phe Ala 565 570 575 Ala Ile Gln Pro Gln Ile Ala
Leu Leu Gln Glu Met His Val Thr His 580 585 590 Phe Arg Phe Ser Leu
Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln 595 600 605 Ser Gln Val
Asn His Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser 610 615 620 Glu
Leu Val Arg Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro 625 630
635 640 Met Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly
Ala 645 650 655 Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr
Ala Arg Leu 660 665 670 Cys Phe Gln Glu Leu Gly His His Val Lys Leu
Trp Ile Thr Met Asn 675 680 685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr
Ser Ala Gly His Asn Leu Leu 690 695 700 Lys Ala His Ala Leu Ala Trp
His Val Tyr Asn Glu Lys Phe Arg His 705 710 715 720 Ala Gln Asn Gly
Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu 725 730 735 Pro Ala
Cys Pro Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val 740 745 750
Leu Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly 755
760 765 Asp Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn
Phe 770 775 780 Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile
Gln Gly Thr 785 790 795 800 Phe Asp Phe Leu Ala Leu Ser His Tyr Thr
Thr Ile Leu Val Asp Ser 805 810 815 Glu Lys Glu Asp Pro Ile Lys Tyr
Asn Asp Tyr Leu Glu Val Gln Glu 820 825 830 Met Thr Asp Ile Thr Trp
Leu Asn Ser Pro Ser Gln Val Ala Val Val 835 840 845 Pro Trp Gly Leu
Arg Lys Val Leu Asn Trp Leu Lys Phe Lys Tyr Gly 850 855 860 Asp Leu
Pro Met Tyr Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu His 865 870 875
880 Ala Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn
885 890 895 Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys
Gly Tyr 900 905 910 Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg
Phe Gly Leu Tyr 915 920 925 Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys
Ala Ser Met Lys His Tyr 930 935 940 Arg Lys Ile Ile Asp Ser Asn Gly
Phe Pro Gly Pro Glu Thr Leu Glu 945 950 955 960 Arg Phe Cys Pro Glu
Glu Phe Thr Val Cys Thr Glu Cys Ser Phe Phe 965 970 975 His Thr Arg
Lys Ser Leu Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly 980 985 990 Gly
Ser Gly Gly Gly Gly Ser Leu Lys Tyr Pro Asn Ala Ser Pro Leu 995
1000 1005 Leu Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala
Thr 1010 1015 1020 Ala Arg Asn Ser Tyr His Leu Gln Ile His Lys Asn
Gly His Val 1025 1030 1035 Asp Gly Ala Pro His Gln Thr Ile Tyr Ser
Ala Leu Met Ile Arg 1040 1045 1050 Ser Glu Asp Ala Gly Phe Val Val
Ile Thr Gly Val Met Ser Arg 1055 1060 1065 Arg Tyr Leu Cys Met Asp
Phe Arg Gly Asn Ile Phe Gly Ser His 1070 1075 1080 Tyr Phe Asp Pro
Glu Asn Cys Arg Phe Gln His Gln Thr Leu Glu 1085 1090 1095 Asn Gly
Tyr Asp Val Tyr His Ser Pro Gln Tyr His Phe Leu Val 1100 1105 1110
Ser Leu Gly Arg Ala Lys Arg Ala Phe Leu Pro Gly Met Asn Pro 1115
1120 1125 Pro Pro Tyr Ser Gln Phe Leu Ser Arg Arg Asn Glu Ile Pro
Leu 1130 1135 1140 Ile His Phe Asn Thr Pro Ile Pro Arg Arg His Thr
Gln Ser Ala 1145 1150 1155 Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn
Val Leu Lys Pro Arg 1160 1165 1170 Ala Arg Met Thr Pro Ala Pro Ala
Ser Cys Ser Gln Glu Leu Pro 1175 1180 1185 Ser Ala Glu Asp Asn Ser
Pro Met Ala Ser Asp Pro Leu Gly Val 1190 1195 1200 Val Arg Gly Gly
Arg Val Asn Thr His Ala Gly Gly Thr Gly Pro 1205 1210 1215 Glu Gly
Ser Arg Pro Phe Ala Lys Phe Ile 1220 1225 561228PRTHomo sapiens
56Met Pro Ala Ser Ala Pro Pro Arg Arg Pro Arg Pro Pro Pro Pro Ser 1
5 10 15 Leu Ser Leu Leu Leu Val Leu Leu Gly Leu Gly Gly Arg Arg Leu
Arg 20 25 30 Ala Glu Pro Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe
Ser Arg Pro 35 40 45 Pro Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly
Thr Phe Pro Asp Gly 50 55 60 Phe Leu Trp Ala Val Gly Ser Ala Ala
Tyr Gln Thr Glu Gly Gly Trp 65 70 75 80 Gln Gln His Gly Lys Gly Ala
Ser Ile Trp Asp Thr Phe Thr His His 85 90 95 Pro Leu Ala Pro Pro
Gly Asp Ser Arg Asn Ala Ser Leu Pro Leu Gly 100 105 110 Ala Pro Ser
Pro Leu Gln Pro Ala Thr Gly Asp Val Ala Ser Asp Ser 115 120 125 Tyr
Asn Asn Val Phe Arg Asp Thr Glu Ala Leu Arg Glu Leu Gly Val 130 135
140 Thr His Tyr Arg Phe Ser Ile Ser Trp Ala Arg Val Leu Pro Asn Gly
145 150 155 160 Ser Ala Gly Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr
Arg Arg Leu 165 170 175 Leu Glu Arg Leu Arg Glu Leu Gly Val Gln Pro
Val Val Thr Leu Tyr 180 185 190 His Trp Asp Leu Pro Gln Arg Leu Gln
Asp Ala Tyr Gly Gly Trp Ala 195 200 205 Asn Arg Ala Leu Ala Asp His
Phe Arg Asp Tyr Ala Glu Leu Cys Phe 210 215 220 Arg His Phe Gly Gly
Gln Val Lys Tyr Trp Ile Thr Ile Asp Asn Pro 225 230 235 240 Tyr Val
Val Ala Trp His Gly Tyr Ala Thr Gly Arg Leu Ala Pro Gly 245 250 255
Ile Arg Gly Ser Pro Arg Leu Gly Tyr Leu Val Ala His Asn Leu Leu 260
265 270 Leu Ala His Ala Lys Val Trp His Leu Tyr Asn Thr Ser Phe Arg
Pro 275 280 285 Thr Gln Gly Gly Gln Val Ser Ile Ala Leu Ser Ser His
Trp Ile Asn 290 295 300 Pro Arg Arg Met Thr Asp His Ser Ile Lys Glu
Cys Gln Lys Ser Leu 305 310 315 320 Asp Phe Val Leu Gly Trp Phe Ala
Lys Pro Val Phe Ile Asp Gly Asp 325 330 335 Tyr Pro Glu Ser Met Lys
Asn Asn Leu Ser Ser Ile Leu Pro Asp Phe 340 345 350 Thr Glu Ser Glu
Lys Lys Phe Ile Lys Gly Thr Ala Asp Phe Phe Ala 355 360 365 Leu Cys
Phe Gly Pro Thr Leu Ser Phe Gln Leu Leu Asp Pro His Met 370 375 380
Lys Phe Arg Gln Leu Glu Ser Pro Asn Leu Arg Gln Leu Leu Ser Trp 385
390 395 400 Ile Asp Leu Glu Phe Asn His Pro Gln Ile Phe Ile Val Glu
Asn Gly 405 410 415 Trp Phe Val Ser Gly Thr Thr Lys Arg Asp Asp Ala
Lys Tyr Met Tyr 420 425 430 Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu
Lys Ala Ile Lys Leu Asp 435 440 445 Gly Val Asp Val Ile Gly Tyr Thr
Ala Trp Ser Leu Met Asp Gly Phe 450 455 460 Glu Trp His Arg Gly Tyr
Ser Ile Arg Arg Gly Leu Phe Tyr Val Asp 465 470 475 480 Phe Leu Ser
Gln Asp Lys Met Leu Leu Pro Lys Ser Ser Ala Leu Phe 485 490 495 Tyr
Gln Lys Leu Ile Glu Lys Asn Gly Phe Pro Pro Leu Pro Glu Asn 500 505
510 Gln Pro Leu Glu Gly Thr Phe Pro Cys Asp Phe Ala Trp Gly Val Val
515 520 525 Asp Asn Tyr Ile Gln Val Asp Thr Thr Leu Ser Gln Phe Thr
Asp Leu 530 535 540 Asn Val Tyr Leu Trp Asp Val His His Ser Lys Arg
Leu Ile Lys Val 545 550 555 560 Asp Gly Val Val Thr Lys Lys Arg Lys
Ser Tyr Cys Val Asp Phe Ala 565 570 575 Ala Ile Gln Pro Gln Ile Ala
Leu Leu Gln Glu Met His Val Thr His 580 585 590 Phe Arg Phe Ser Leu
Asp Trp Ala Leu Ile Leu Pro Leu Gly Asn Gln 595 600 605 Ser Gln Val
Asn His Thr Ile Leu Gln Tyr Tyr Arg Cys Met Ala Ser 610 615 620 Glu
Leu Val Arg Val Asn Ile Thr Pro Val Val Ala Leu Trp Gln Pro 625 630
635 640 Met Ala Pro Asn Gln Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly
Ala 645 650 655 Trp Glu Asn Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr
Ala Arg Leu 660 665 670 Cys Phe Gln Glu Leu Gly His His Val Lys Leu
Trp Ile Thr Met Asn 675 680 685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr
Ser Ala Gly His Asn Leu Leu 690 695 700 Lys Ala His Ala Leu Ala Trp
His Val Tyr Asn Glu Lys Phe Arg His 705 710 715 720 Ala Gln Asn Gly
Lys Ile Ser Ile Ala Leu Gln Ala Asp Trp Ile Glu 725 730 735 Pro Ala
Cys Pro Phe Ser Gln Lys Asp Lys Glu Val Ala Glu Arg Val 740 745 750
Leu Glu Phe Asp Ile Gly Trp Leu Ala Glu Pro Ile Phe Gly Ser Gly 755
760 765 Asp Tyr Pro Trp Val Met Arg Asp Trp Leu Asn Gln Arg Asn Asn
Phe 770 775 780 Leu Leu Pro Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile
Gln Gly Thr 785 790 795 800 Phe Asp Phe Leu Ala Leu Ser His Tyr Thr
Thr Ile Leu Val Asp Ser 805 810 815 Glu Lys Glu Asp Pro Ile Lys Tyr
Asn Asp Tyr Leu Glu Val Gln Glu 820 825 830 Met Thr Asp Ile Thr Trp
Leu Asn Ser Pro Ser Gln Val Ala Val Val 835 840 845 Pro Trp Gly Leu
Arg Lys Val Leu Asn Trp Leu Lys Phe Lys Tyr Gly 850 855 860 Asp Leu
Pro Met Tyr Ile Ile Ser Asn Gly Ile Asp Asp Gly Leu His 865 870 875
880 Ala Glu Asp Asp Gln Leu Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn
885 890 895 Glu Ala Leu Lys Ala His Ile Leu Asp Gly Ile Asn Leu Cys
Gly Tyr 900 905 910 Phe Ala Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg
Phe Gly Leu Tyr 915 920 925 Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys
Ala Ser Met Lys His Tyr 930 935 940 Arg Lys Ile Ile Asp Ser Asn Gly
Phe Pro Gly Pro Glu Thr Leu Glu 945 950 955 960 Arg Phe Cys Pro Glu
Glu Phe Thr Val Cys Thr Glu Cys Ser Phe Phe 965 970 975 His Thr Arg
Lys Ser Leu Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly 980 985 990 Gly
Ser Gly Gly Gly Gly Ser Leu Lys Tyr Pro Asn Ala Ser Pro Leu 995
1000 1005 Leu Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala
Thr 1010 1015 1020 Ala Arg Asn Ser Tyr His Leu Gln Ile His Lys Asn
Gly His Val 1025 1030 1035 Asp Gly Ala Pro His Gln Thr Ile Tyr Ser
Ala Leu Met Ile Arg 1040 1045 1050 Ser Glu Asp Ala Gly Phe Val Val
Ile Thr Gly Val Met Ser Arg 1055 1060 1065 Arg Tyr Leu Cys Met Asp
Phe Arg Gly Asn Ile Phe Gly Ser His 1070 1075 1080 Tyr Phe Asp Pro
Glu Asn Cys Arg Phe Gln His Gln Thr Leu Glu 1085 1090 1095 Asn Gly
Tyr Asp Val Tyr His Ser Pro Gln Tyr His Phe Leu Val 1100 1105 1110
Ser Leu Gly Arg Ala Lys Arg Ala Phe Leu Pro Gly Met Asn Pro 1115
1120 1125 Pro Pro Tyr Ser Ala Phe Leu Ser Arg Arg Asn Glu Ile Pro
Leu 1130 1135 1140 Ile His Phe Asn Thr Pro Ile Pro Arg Arg His Thr
Gln Ser Ala 1145 1150 1155 Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn
Val Leu Lys Pro Arg 1160 1165 1170 Ala Arg Met Thr Pro Ala Pro Ala
Ser Cys Ser Gln Glu Leu Pro 1175
1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu Gly
Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly
Thr Gly Pro 1205 1210 1215 Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile
1220 1225 57 1228PRTHomo sapiens 57Met Pro Ala Ser Ala Pro Pro Arg
Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu Val
Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro Gly
Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro Ala
Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55 60
Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp 65
70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr
His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser
Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly
Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr
Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser
Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly
Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu
Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185
190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala
195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu
Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr
Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala
Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu
Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys
Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly
Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro
Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310
315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly
Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu
Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr
Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe
Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser
Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu
Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe
Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430
Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435
440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly
Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe
Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro
Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn
Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Thr
Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520 525 Asp Asn Tyr Ile
Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu 530 535 540 Asn Val
Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val 545 550 555
560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala
565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val
Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro
Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His Thr Ile Leu Gln Tyr
Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val Arg Val Asn Ile Thr
Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640 Met Ala Pro Asn Gln
Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645 650 655 Trp Glu Asn
Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660 665 670 Cys
Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn 675 680
685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu
690 695 700 Lys Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe
Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln
Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro Phe Ser Gln Lys Asp
Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu Phe Asp Ile Gly Trp
Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765 Asp Tyr Pro Trp Val
Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770 775 780 Leu Leu Pro
Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr 785 790 795 800
Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser 805
810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln
Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val
Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu
Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met Tyr Ile Ile Ser Asn
Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala Glu Asp Asp Gln Leu
Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890 895 Glu Ala Leu Lys
Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900 905 910 Phe Ala
Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr 915 920 925
Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr 930
935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu
Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu
Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser Leu Gly Ser Gly Gly
Gly Gly Ser Gly Gly Gly 980 985 990 Gly Ser Gly Gly Gly Gly Ser Leu
Lys Tyr Pro Asn Ala Ser Pro Leu 995 1000 1005 Leu Gly Ser Ser Trp
Gly Gly Leu Ile His Leu Tyr Thr Ala Thr 1010 1015 1020 Ala Arg Asn
Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val 1025 1030 1035 Asp
Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg 1040 1045
1050 Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val Met Ser Arg
1055 1060 1065 Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly
Ser His 1070 1075 1080 Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His
Gln Thr Leu Glu 1085 1090 1095 Asn Gly Tyr Asp Val Tyr His Ser Pro
Gln Tyr His Phe Leu Val 1100 1105 1110 Ser Leu Gly Arg Ala Lys Arg
Ala Phe Leu Pro Gly Met Asn Pro 1115 1120 1125 Pro Pro Tyr Ser Gln
Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu 1130 1135 1140 Ile His Phe
Asn Thr Pro Ile Pro Arg Arg His Thr Gln Ser Ala 1145 1150 1155 Glu
Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys Pro Arg 1160 1165
1170 Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln Glu Leu Pro
1175 1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu
Gly Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn Thr His Ala Gly
Gly Thr Gly Pro 1205 1210 1215 Glu Gly Ser Arg Pro Phe Ala Lys Phe
Ile 1220 1225 58 1228PRTHomo sapiens 58Met Pro Ala Ser Ala Pro Pro
Arg Arg Pro Arg Pro Pro Pro Pro Ser 1 5 10 15 Leu Ser Leu Leu Leu
Val Leu Leu Gly Leu Gly Gly Arg Arg Leu Arg 20 25 30 Ala Glu Pro
Gly Asp Gly Ala Gln Thr Trp Ala Arg Phe Ser Arg Pro 35 40 45 Pro
Ala Pro Glu Ala Ala Gly Leu Phe Gln Gly Thr Phe Pro Asp Gly 50 55
60 Phe Leu Trp Ala Val Gly Ser Ala Ala Tyr Gln Thr Glu Gly Gly Trp
65 70 75 80 Gln Gln His Gly Lys Gly Ala Ser Ile Trp Asp Thr Phe Thr
His His 85 90 95 Pro Leu Ala Pro Pro Gly Asp Ser Arg Asn Ala Ser
Leu Pro Leu Gly 100 105 110 Ala Pro Ser Pro Leu Gln Pro Ala Thr Gly
Asp Val Ala Ser Asp Ser 115 120 125 Tyr Asn Asn Val Phe Arg Asp Thr
Glu Ala Leu Arg Glu Leu Gly Val 130 135 140 Thr His Tyr Arg Phe Ser
Ile Ser Trp Ala Arg Val Leu Pro Asn Gly 145 150 155 160 Ser Ala Gly
Val Pro Asn Arg Glu Gly Leu Arg Tyr Tyr Arg Arg Leu 165 170 175 Leu
Glu Arg Leu Arg Glu Leu Gly Val Gln Pro Val Val Thr Leu Tyr 180 185
190 His Trp Asp Leu Pro Gln Arg Leu Gln Asp Ala Tyr Gly Gly Trp Ala
195 200 205 Asn Arg Ala Leu Ala Asp His Phe Arg Asp Tyr Ala Glu Leu
Cys Phe 210 215 220 Arg His Phe Gly Gly Gln Val Lys Tyr Trp Ile Thr
Ile Asp Asn Pro 225 230 235 240 Tyr Val Val Ala Trp His Gly Tyr Ala
Thr Gly Arg Leu Ala Pro Gly 245 250 255 Ile Arg Gly Ser Pro Arg Leu
Gly Tyr Leu Val Ala His Asn Leu Leu 260 265 270 Leu Ala His Ala Lys
Val Trp His Leu Tyr Asn Thr Ser Phe Arg Pro 275 280 285 Thr Gln Gly
Gly Gln Val Ser Ile Ala Leu Ser Ser His Trp Ile Asn 290 295 300 Pro
Arg Arg Met Thr Asp His Ser Ile Lys Glu Cys Gln Lys Ser Leu 305 310
315 320 Asp Phe Val Leu Gly Trp Phe Ala Lys Pro Val Phe Ile Asp Gly
Asp 325 330 335 Tyr Pro Glu Ser Met Lys Asn Asn Leu Ser Ser Ile Leu
Pro Asp Phe 340 345 350 Thr Glu Ser Glu Lys Lys Phe Ile Lys Gly Thr
Ala Asp Phe Phe Ala 355 360 365 Leu Cys Phe Gly Pro Thr Leu Ser Phe
Gln Leu Leu Asp Pro His Met 370 375 380 Lys Phe Arg Gln Leu Glu Ser
Pro Asn Leu Arg Gln Leu Leu Ser Trp 385 390 395 400 Ile Asp Leu Glu
Phe Asn His Pro Gln Ile Phe Ile Val Glu Asn Gly 405 410 415 Trp Phe
Val Ser Gly Thr Thr Lys Arg Asp Asp Ala Lys Tyr Met Tyr 420 425 430
Tyr Leu Lys Lys Phe Ile Met Glu Thr Leu Lys Ala Ile Lys Leu Asp 435
440 445 Gly Val Asp Val Ile Gly Tyr Thr Ala Trp Ser Leu Met Asp Gly
Phe 450 455 460 Glu Trp His Arg Gly Tyr Ser Ile Arg Arg Gly Leu Phe
Tyr Val Asp 465 470 475 480 Phe Leu Ser Gln Asp Lys Met Leu Leu Pro
Lys Ser Ser Ala Leu Phe 485 490 495 Tyr Gln Lys Leu Ile Glu Lys Asn
Gly Phe Pro Pro Leu Pro Glu Asn 500 505 510 Gln Pro Leu Glu Gly Thr
Phe Pro Cys Asp Phe Ala Trp Gly Val Val 515 520 525 Asp Asn Tyr Ile
Gln Val Asp Thr Thr Leu Ser Gln Phe Thr Asp Leu 530 535 540 Asn Val
Tyr Leu Trp Asp Val His His Ser Lys Arg Leu Ile Lys Val 545 550 555
560 Asp Gly Val Val Thr Lys Lys Arg Lys Ser Tyr Cys Val Asp Phe Ala
565 570 575 Ala Ile Gln Pro Gln Ile Ala Leu Leu Gln Glu Met His Val
Thr His 580 585 590 Phe Arg Phe Ser Leu Asp Trp Ala Leu Ile Leu Pro
Leu Gly Asn Gln 595 600 605 Ser Gln Val Asn His Thr Ile Leu Gln Tyr
Tyr Arg Cys Met Ala Ser 610 615 620 Glu Leu Val Arg Val Asn Ile Thr
Pro Val Val Ala Leu Trp Gln Pro 625 630 635 640 Met Ala Pro Asn Gln
Gly Leu Pro Arg Leu Leu Ala Arg Gln Gly Ala 645 650 655 Trp Glu Asn
Pro Tyr Thr Ala Leu Ala Phe Ala Glu Tyr Ala Arg Leu 660 665 670 Cys
Phe Gln Glu Leu Gly His His Val Lys Leu Trp Ile Thr Met Asn 675 680
685 Glu Pro Tyr Thr Arg Asn Met Thr Tyr Ser Ala Gly His Asn Leu Leu
690 695 700 Lys Ala His Ala Leu Ala Trp His Val Tyr Asn Glu Lys Phe
Arg His 705 710 715 720 Ala Gln Asn Gly Lys Ile Ser Ile Ala Leu Gln
Ala Asp Trp Ile Glu 725 730 735 Pro Ala Cys Pro Phe Ser Gln Lys Asp
Lys Glu Val Ala Glu Arg Val 740 745 750 Leu Glu Phe Asp Ile Gly Trp
Leu Ala Glu Pro Ile Phe Gly Ser Gly 755 760 765 Asp Tyr Pro Trp Val
Met Arg Asp Trp Leu Asn Gln Arg Asn Asn Phe 770 775 780 Leu Leu Pro
Tyr Phe Thr Glu Asp Glu Lys Lys Leu Ile Gln Gly Thr 785 790 795 800
Phe Asp Phe Leu Ala Leu Ser His Tyr Thr Thr Ile Leu Val Asp Ser 805
810 815 Glu Lys Glu Asp Pro Ile Lys Tyr Asn Asp Tyr Leu Glu Val Gln
Glu 820 825 830 Met Thr Asp Ile Thr Trp Leu Asn Ser Pro Ser Gln Val
Ala Val Val 835 840 845 Pro Trp Gly Leu Arg Lys Val Leu Asn Trp Leu
Lys Phe Lys Tyr Gly 850 855 860 Asp Leu Pro Met Tyr Ile Ile Ser Asn
Gly Ile Asp Asp Gly Leu His 865 870 875 880 Ala Glu Asp Asp Gln Leu
Arg Val Tyr Tyr Met Gln Asn Tyr Ile Asn 885 890 895 Glu Ala Leu Lys
Ala His Ile Leu Asp Gly Ile Asn Leu Cys Gly Tyr 900 905 910 Phe Ala
Tyr Ser Phe Asn Asp Arg Thr Ala Pro Arg Phe Gly Leu Tyr 915 920 925
Arg Tyr Ala Ala Asp Gln Phe Glu Pro Lys Ala Ser Met Lys His Tyr 930
935 940 Arg Lys Ile Ile Asp Ser Asn Gly Phe Pro Gly Pro Glu Thr Leu
Glu 945 950 955 960 Arg Phe Cys Pro Glu Glu Phe Thr Val Cys Thr Glu
Cys Ser Phe Phe 965 970 975 His Thr Arg Lys Ser Leu Gly Ser Gly Gly
Gly Gly Ser Gly Gly Gly 980 985 990 Gly Ser Gly Gly Gly Gly Ser Leu
Lys Tyr Pro Asn Ala Ser Pro Leu 995 1000 1005 Leu Gly Ser Ser
Trp
Gly Gly Leu Ile His Leu Tyr Thr Ala Thr 1010 1015 1020 Ala Arg Asn
Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val 1025 1030 1035 Asp
Gly Ala Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg 1040 1045
1050 Ser Glu Asp Ala Gly Phe Val Val Ile Thr Gly Val Met Ser Arg
1055 1060 1065 Arg Tyr Leu Cys Met Asp Phe Arg Gly Asn Ile Phe Gly
Ser His 1070 1075 1080 Tyr Phe Asp Pro Glu Asn Cys Arg Phe Gln His
Gln Thr Leu Glu 1085 1090 1095 Asn Gly Tyr Asp Val Tyr His Ser Pro
Gln Tyr His Phe Leu Val 1100 1105 1110 Ser Leu Gly Arg Ala Lys Arg
Ala Phe Leu Pro Gly Met Asn Pro 1115 1120 1125 Pro Pro Tyr Ser Ala
Phe Leu Ser Arg Arg Asn Glu Ile Pro Leu 1130 1135 1140 Ile His Phe
Asn Thr Pro Ile Pro Arg Arg His Thr Gln Ser Ala 1145 1150 1155 Glu
Asp Asp Ser Glu Arg Asp Pro Leu Asn Val Leu Lys Pro Arg 1160 1165
1170 Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln Glu Leu Pro
1175 1180 1185 Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu
Gly Val 1190 1195 1200 Val Arg Gly Gly Arg Val Asn Thr His Ala Gly
Gly Thr Gly Pro 1205 1210 1215 Glu Gly Ser Arg Pro Phe Ala Lys Phe
Ile 1220 1225 59482PRTHomo sapiens 59Met Leu Gly Ala Arg Leu Arg
Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val
Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser
Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn
Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55
60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala
65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu
Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe
Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn
Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val
Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn
Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu
Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His
Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185
190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln
195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp
Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala
Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys
Phe Ile Gly Gly Gly Gly Ser 245 250 255 Lys Thr His Thr Cys Pro Pro
Cys Pro Ala Pro Glu Ala Ala Gly Gly 260 265 270 Pro Ser Val Phe Leu
Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile 275 280 285 Ser Arg Thr
Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu 290 295 300 Asp
Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His 305 310
315 320 Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr
Arg 325 330 335 Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu
Asn Gly Lys 340 345 350 Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu
Pro Ala Pro Ile Glu 355 360 365 Lys Thr Ile Ser Lys Ala Lys Gly Gln
Pro Arg Glu Pro Gln Val Tyr 370 375 380 Thr Leu Pro Pro Ser Arg Glu
Glu Met Thr Lys Asn Gln Val Ser Leu 385 390 395 400 Thr Cys Leu Val
Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp 405 410 415 Glu Ser
Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val 420 425 430
Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp 435
440 445 Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met
His 450 455 460 Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser
Leu Ser Pro 465 470 475 480 Gly Lys 60482PRTHomo sapiens 60Met Leu
Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15
Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20
25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala
Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val
Asp Gly Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile
Arg Ser Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met
Ser Arg Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe
Gly Ser His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His
Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln
Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe
Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150
155 160 Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg
Arg 165 170 175 His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro
Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro
Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser
Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg
Val Asn Thr His Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Ser
Arg Pro Phe Ala Lys Phe Ile Gly Gly Gly Gly Ser 245 250 255 Lys Thr
His Thr Cys Pro Pro Cys Pro Ala Pro Glu Ala Ala Gly Gly 260 265 270
Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile 275
280 285 Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His
Glu 290 295 300 Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val
Glu Val His 305 310 315 320 Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln
Tyr Asn Ser Thr Tyr Arg 325 330 335 Val Val Ser Val Leu Thr Val Leu
His Gln Asp Trp Leu Asn Gly Lys 340 345 350 Glu Tyr Lys Cys Lys Val
Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu 355 360 365 Lys Thr Ile Ser
Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr 370 375 380 Thr Leu
Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu 385 390 395
400 Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp
405 410 415 Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
Pro Val 420 425 430 Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys
Leu Thr Val Asp 435 440 445 Lys Ser Arg Trp Gln Gln Gly Asn Val Phe
Ser Cys Ser Val Met His 450 455 460 Glu Ala Leu His Asn His Tyr Thr
Gln Lys Ser Leu Ser Leu Ser Pro 465 470 475 480 Gly Lys
61482PRTHomo sapiens 61Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Ala Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gly Gly
Gly Gly Ser 245 250 255 Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro
Glu Ala Ala Gly Gly 260 265 270 Pro Ser Val Phe Leu Phe Pro Pro Lys
Pro Lys Asp Thr Leu Met Ile 275 280 285 Ser Arg Thr Pro Glu Val Thr
Cys Val Val Val Asp Val Ser His Glu 290 295 300 Asp Pro Glu Val Lys
Phe Asn Trp Tyr Val Asp Gly Val Glu Val His 305 310 315 320 Asn Ala
Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg 325 330 335
Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys 340
345 350 Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile
Glu 355 360 365 Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro
Gln Val Tyr 370 375 380 Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys
Asn Gln Val Ser Leu 385 390 395 400 Thr Cys Leu Val Lys Gly Phe Tyr
Pro Ser Asp Ile Ala Val Glu Trp 405 410 415 Glu Ser Asn Gly Gln Pro
Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val 420 425 430 Leu Asp Ser Asp
Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp 435 440 445 Lys Ser
Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His 450 455 460
Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro 465
470 475 480 Gly Lys 62482PRTHomo sapiens 62Met Leu Gly Ala Arg Leu
Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser
Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser
Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45
Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50
55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp
Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr
Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr
Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu
Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu
Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met
Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn
Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175
His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180
185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser
Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His
Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Ser Arg Pro Phe Ala
Lys Phe Ile Gly Gly Gly Gly Ser 245 250 255 Lys Thr His Thr Cys Pro
Pro Cys Pro Ala Pro Glu Ala Ala Gly Gly 260 265 270 Pro Ser Val Phe
Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile 275 280 285 Ser Arg
Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu 290 295 300
Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His 305
310 315 320 Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
Tyr Arg 325 330 335 Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp
Leu Asn Gly Lys 340 345 350 Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala
Leu Pro Ala Pro Ile Glu 355 360 365 Lys Thr Ile Ser Lys Ala Lys Gly
Gln Pro Arg Glu Pro Gln Val Tyr 370 375 380 Thr Leu Pro Pro Ser Arg
Glu Glu Met Thr Lys Asn Gln Val Ser Leu 385 390 395 400 Thr Cys Leu
Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp 405 410 415 Glu
Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val 420 425
430 Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp
435 440 445 Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
Met His 450 455 460 Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu
Ser Leu Ser Pro 465 470 475 480 Gly Lys 63482PRTHomo sapiens 63Met
Leu Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10
15 Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu
20 25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr
Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His
Val Asp Gly Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met
Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val
Met Ser Arg Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile
Phe Gly Ser His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln
His Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro
Gln Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 130
135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Ala Phe Leu Ser
Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro
Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp Asp Ser Glu
Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala Arg Met Thr
Pro Ala Pro Ala Ser Ser Ser Gln 195 200 205 Glu Leu Pro Ser Ala Glu
Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly Val Val Arg
Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230 235 240 Pro
Glu Gly Ser Arg Pro Phe Ala Lys Phe Ile Gly Gly Gly Gly Ser 245 250
255 Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Ala Ala Gly Gly
260 265 270 Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
Met Ile 275 280 285 Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp
Val Ser His Glu 290 295 300 Asp Pro Glu Val Lys Phe Asn Trp Tyr Val
Asp Gly Val Glu Val His 305 310 315 320 Asn Ala Lys Thr Lys Pro Arg
Glu Glu Gln Tyr Asn Ser Thr Tyr Arg 325 330 335 Val Val Ser Val Leu
Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys 340 345 350 Glu Tyr Lys
Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu 355 360 365 Lys
Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr 370 375
380 Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu
385 390 395 400 Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala
Val Glu Trp 405 410 415 Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys
Thr Thr Pro Pro Val 420 425 430 Leu Asp Ser Asp Gly Ser Phe Phe Leu
Tyr Ser Lys Leu Thr Val Asp 435 440 445 Lys Ser Arg Trp Gln Gln Gly
Asn Val Phe Ser Cys Ser Val Met His 450 455 460 Glu Ala Leu His Asn
His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro 465 470 475 480 Gly Lys
64473PRTHomo sapiens 64Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile Gly Gly
Gly Gly Ser 245 250 255 Ala Pro Glu Ala Ala Gly Gly Pro Ser Val Phe
Leu Phe Pro Pro Lys 260 265 270 Pro Lys Asp Thr Leu Met Ile Ser Arg
Thr Pro Glu Val Thr Cys Val 275 280 285 Val Val Asp Val Ser His Glu
Asp Pro Glu Val Lys Phe Asn Trp Tyr 290 295 300 Val Asp Gly Val Glu
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 305 310 315 320 Gln Tyr
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 325 330 335
Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 340
345 350 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly
Gln 355 360 365 Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg
Glu Glu Met 370 375 380 Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val
Lys Gly Phe Tyr Pro 385 390 395 400 Ser Asp Ile Ala Val Glu Trp Glu
Ser Asn Gly Gln Pro Glu Asn Asn 405 410 415 Tyr Lys Thr Thr Pro Pro
Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 420 425 430 Tyr Ser Lys Leu
Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 435 440 445 Phe Ser
Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 450 455 460
Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 65473PRTHomo sapiens
65Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1
5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu
Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala
Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly
His Val Asp Gly Ala 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu
Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly
Val Met Ser Arg Arg Tyr Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn
Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe
Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser
Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135
140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr Ser Gln Phe Leu Ser Arg
145 150 155 160 Arg Asn Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile
Pro Arg Arg 165 170 175 His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg
Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro
Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp
Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215 220 Gly Val Val Arg Gly
Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu
Gly Ser Arg Pro Phe Ala Lys Phe Ile Gly Gly Gly Gly Ser 245 250 255
Ala Pro Glu Ala Ala Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 260
265 270 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys
Val 275 280 285 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe
Asn Trp Tyr 290 295 300 Val Asp Gly Val Glu Val His Asn Ala Lys Thr
Lys Pro Arg Glu Glu 305 310 315 320 Gln Tyr Asn Ser Thr Tyr Arg Val
Val Ser Val Leu Thr Val Leu His 325 330 335 Gln Asp Trp Leu Asn Gly
Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 340 345 350 Ala Leu Pro Ala
Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 355 360 365 Pro Arg
Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met 370 375 380
Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 385
390 395 400 Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu
Asn Asn 405 410 415 Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly
Ser Phe Phe Leu 420 425 430 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg
Trp Gln Gln Gly Asn Val 435 440 445 Phe Ser Cys Ser Val Met His Glu
Ala Leu His Asn His Tyr Thr Gln 450 455 460 Lys Ser Leu Ser Leu Ser
Pro Gly Lys 465 470 66473PRTHomo sapiens 66Met Leu Gly Ala Arg Leu
Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser
Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser
Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45
Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50
55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp
Ala 65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr
Leu Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr
Phe Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu
Asn Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu
Val Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met
Asn Pro Pro Pro Tyr Ser Ala Phe Leu Ser Arg 145 150 155 160 Arg Asn
Glu Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175
His Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180
185 190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser
Gln 195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser
Asp Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His
Ala Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala
Lys Phe Ile Gly Gly Gly Gly Ser 245 250 255 Ala Pro Glu Ala Ala Gly
Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 260 265 270 Pro Lys Asp Thr
Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 275 280 285 Val Val
Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 290 295 300
Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 305
310 315 320 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val
Leu His 325 330 335 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys
Val Ser Asn Lys 340 345 350 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile
Ser Lys Ala Lys Gly Gln 355 360 365 Pro Arg Glu Pro Gln Val Tyr Thr
Leu Pro Pro Ser Arg Glu Glu Met 370 375 380 Thr Lys Asn Gln Val Ser
Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 385 390 395 400 Ser Asp Ile
Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 405 410 415 Tyr
Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 420 425
430 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val
435 440 445 Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr
Thr Gln 450 455 460 Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470
67473PRTHomo sapiens 67Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Gln Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Ser Arg Pro Phe Ala Lys Phe Ile Gly Gly
Gly Gly Ser 245 250 255 Ala Pro Glu Ala Ala Gly Gly Pro Ser Val Phe
Leu Phe Pro Pro Lys 260 265 270 Pro Lys Asp Thr Leu Met Ile Ser Arg
Thr Pro Glu Val Thr Cys Val 275 280 285 Val Val Asp Val Ser His Glu
Asp Pro Glu Val Lys Phe Asn Trp Tyr 290 295 300 Val Asp Gly Val Glu
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 305 310 315 320 Gln Tyr
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 325 330 335
Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 340
345 350 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly
Gln 355 360 365 Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg
Glu Glu Met 370 375 380 Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val
Lys Gly Phe Tyr Pro 385 390 395 400 Ser Asp Ile Ala Val Glu Trp Glu
Ser Asn Gly Gln Pro Glu Asn Asn 405 410 415 Tyr Lys Thr Thr Pro Pro
Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 420 425 430 Tyr Ser Lys Leu
Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 435 440 445 Phe Ser
Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 450 455 460
Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470 68473PRTHomo sapiens
68Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys Ala Leu Cys Ser Val 1
5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr Pro Asn Ala Ser Pro Leu
Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu Ile His Leu Tyr Thr Ala
Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile His Lys Asn Gly
His Val Asp Gly Ala 50 55
60 Pro His Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala
65 70 75 80 Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu
Cys Met 85 90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe
Asp Pro Glu Asn 100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn
Gly Tyr Asp Val Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val
Ser Leu Gly Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn
Pro Pro Pro Tyr Ser Ala Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu
Ile Pro Leu Ile His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His
Thr Gln Ser Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185
190 Leu Lys Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Ser Ser Gln
195 200 205 Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp
Pro Leu 210 215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala
Gly Gly Thr Gly 225 230 235 240 Pro Glu Gly Ser Arg Pro Phe Ala Lys
Phe Ile Gly Gly Gly Gly Ser 245 250 255 Ala Pro Glu Ala Ala Gly Gly
Pro Ser Val Phe Leu Phe Pro Pro Lys 260 265 270 Pro Lys Asp Thr Leu
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 275 280 285 Val Val Asp
Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 290 295 300 Val
Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 305 310
315 320 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu
His 325 330 335 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val
Ser Asn Lys 340 345 350 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser
Lys Ala Lys Gly Gln 355 360 365 Pro Arg Glu Pro Gln Val Tyr Thr Leu
Pro Pro Ser Arg Glu Glu Met 370 375 380 Thr Lys Asn Gln Val Ser Leu
Thr Cys Leu Val Lys Gly Phe Tyr Pro 385 390 395 400 Ser Asp Ile Ala
Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 405 410 415 Tyr Lys
Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 420 425 430
Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 435
440 445 Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr
Gln 450 455 460 Lys Ser Leu Ser Leu Ser Pro Gly Lys 465 470
69251PRTHomo sapiens 69Met Leu Gly Ala Arg Leu Arg Leu Trp Val Cys
Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Leu Arg Ala Tyr
Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly Leu
Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu
Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His Gln
Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80 Gly
Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85 90
95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asp Pro Glu Asn
100 105 110 Cys Arg Phe Gln His Gln Thr Leu Glu Asn Gly Tyr Asp Val
Tyr His 115 120 125 Ser Pro Gln Tyr His Phe Leu Val Ser Leu Gly Arg
Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro Tyr
Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu Ile
His Phe Asn Thr Pro Ile Pro Arg Arg 165 170 175 His Thr Arg Ser Ala
Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro
Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205 Glu
Leu Pro Ser Ala Glu Asp Asn Ser Pro Met Ala Ser Asp Pro Leu 210 215
220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr Gly
225 230 235 240 Pro Glu Gly Cys Arg Pro Phe Ala Lys Phe Ile 245 250
70251PRTMacaca mulatta 70Met Leu Gly Ala Arg Leu Arg Leu Trp Val
Cys Ala Leu Cys Ser Val 1 5 10 15 Cys Ser Met Ser Val Ile Arg Ala
Tyr Pro Asn Ala Ser Pro Leu Leu 20 25 30 Gly Ser Ser Trp Gly Gly
Leu Ile His Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His
Leu Gln Ile His Lys Asn Gly His Val Asp Gly Ala 50 55 60 Pro His
Gln Thr Ile Tyr Ser Ala Leu Met Ile Arg Ser Glu Asp Ala 65 70 75 80
Gly Phe Val Val Ile Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met 85
90 95 Asp Phe Arg Gly Asn Ile Phe Gly Ser His Tyr Phe Asn Pro Glu
Asn 100 105 110 Cys Arg Phe Arg His Trp Thr Leu Glu Asn Gly Tyr Asp
Val Tyr His 115 120 125 Ser Pro Gln His His Phe Leu Val Ser Leu Gly
Arg Ala Lys Arg Ala 130 135 140 Phe Leu Pro Gly Met Asn Pro Pro Pro
Tyr Ser Gln Phe Leu Ser Arg 145 150 155 160 Arg Asn Glu Ile Pro Leu
Ile His Phe Asn Thr Pro Arg Pro Arg Arg 165 170 175 His Thr Arg Ser
Ala Glu Asp Asp Ser Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys
Pro Arg Ala Arg Met Thr Pro Ala Pro Ala Ser Cys Ser Gln 195 200 205
Glu Leu Pro Ser Ala Glu Asp Asn Ser Pro Val Ala Ser Asp Pro Leu 210
215 220 Gly Val Val Arg Gly Gly Arg Val Asn Thr His Ala Gly Gly Thr
Gly 225 230 235 240 Pro Glu Ala Cys Arg Pro Phe Pro Lys Phe Ile 245
250 71245PRTBos taurus 71Met Leu Gly Ala Arg Leu Gly Leu Trp Val
Cys Thr Leu Ser Cys Val 1 5 10 15 Val Gln Ala Tyr Pro Asn Ser Ser
Pro Leu Leu Gly Ser Ser Trp Gly 20 25 30 Gly Leu Thr His Leu Tyr
Thr Ala Thr Ala Arg Asn Ser Tyr His Leu 35 40 45 Gln Ile His Gly
Asp Gly His Val Asp Gly Ser Pro Gln Gln Thr Val 50 55 60 Tyr Ser
Ala Leu Met Ile Arg Ser Glu Asp Ala Gly Phe Val Val Ile 65 70 75 80
Thr Gly Val Met Ser Arg Arg Tyr Leu Cys Met Asp Phe Thr Gly Asn 85
90 95 Ile Phe Gly Ser His His Phe Ser Pro Glu Ser Cys Arg Phe Arg
Gln 100 105 110 Arg Thr Leu Glu Asn Gly Tyr Asp Val Tyr His Ser Pro
Gln His Arg 115 120 125 Phe Leu Val Ser Leu Gly Arg Ala Lys Arg Ala
Phe Leu Pro Gly Thr 130 135 140 Asn Pro Pro Pro Tyr Ala Gln Phe Leu
Ser Arg Arg Asn Glu Ile Pro 145 150 155 160 Leu Pro His Phe Ala Ala
Thr Ala Arg Pro Arg Arg His Thr Arg Ser 165 170 175 Ala His Asp Ser
Gly Asp Pro Leu Ser Val Leu Lys Pro Arg Ala Arg 180 185 190 Ala Thr
Pro Val Pro Ala Ala Cys Ser Gln Glu Leu Pro Ser Ala Glu 195 200 205
Asp Ser Gly Pro Ala Ala Ser Asp Pro Leu Gly Val Leu Arg Gly His 210
215 220 Arg Leu Asp Val Arg Ala Gly Ser Ala Gly Ala Glu Arg Cys Arg
Pro 225 230 235 240 Phe Pro Gly Phe Ala 245 72251PRTMus musculus
72Met Leu Gly Thr Cys Leu Arg Leu Leu Val Gly Val Leu Cys Thr Val 1
5 10 15 Cys Ser Leu Gly Thr Ala Arg Ala Tyr Pro Asp Thr Ser Pro Leu
Leu 20 25 30 Gly Ser Asn Trp Gly Ser Leu Thr His Leu Tyr Thr Ala
Thr Ala Arg 35 40 45 Thr Ser Tyr His Leu Gln Ile His Arg Asp Gly
His Val Asp Gly Thr 50 55 60 Pro His Gln Thr Ile Tyr Ser Ala Leu
Met Ile Thr Ser Glu Asp Ala 65 70 75 80 Gly Ser Val Val Ile Thr Gly
Ala Met Thr Arg Arg Phe Leu Cys Met 85 90 95 Asp Leu His Gly Asn
Ile Phe Gly Ser Leu His Phe Ser Pro Glu Asn 100 105 110 Cys Lys Phe
Arg Gln Trp Thr Leu Glu Asn Gly Tyr Asp Val Tyr Leu 115 120 125 Ser
Gln Lys His His Tyr Leu Val Ser Leu Gly Arg Ala Lys Arg Ile 130 135
140 Phe Gln Pro Gly Thr Asn Pro Pro Pro Phe Ser Gln Phe Leu Ala Arg
145 150 155 160 Arg Asn Glu Val Pro Leu Leu His Phe Tyr Thr Val Arg
Pro Arg Arg 165 170 175 His Thr Arg Ser Ala Glu Asp Pro Pro Glu Arg
Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Pro Arg Ala Thr Pro
Val Pro Val Ser Cys Ser Arg 195 200 205 Glu Leu Pro Ser Ala Glu Glu
Gly Gly Pro Ala Ala Ser Asp Pro Leu 210 215 220 Gly Val Leu Arg Arg
Gly Arg Gly Asp Ala Arg Gly Gly Ala Gly Gly 225 230 235 240 Ala Asp
Arg Cys Arg Pro Phe Pro Arg Phe Val 245 250 73251PRTRattus
norvegicus 73Met Leu Gly Ala Cys Leu Arg Leu Leu Val Gly Ala Leu
Cys Thr Val 1 5 10 15 Cys Ser Leu Gly Thr Ala Arg Ala Tyr Ser Asp
Thr Ser Pro Leu Leu 20 25 30 Gly Ser Asn Trp Gly Ser Leu Thr His
Leu Tyr Thr Ala Thr Ala Arg 35 40 45 Asn Ser Tyr His Leu Gln Ile
His Arg Asp Gly His Val Asp Gly Thr 50 55 60 Pro His Gln Thr Ile
Tyr Ser Ala Leu Met Ile Thr Ser Glu Asp Ala 65 70 75 80 Gly Ser Val
Val Ile Ile Gly Ala Met Thr Arg Arg Phe Leu Cys Met 85 90 95 Asp
Leu Arg Gly Asn Ile Phe Gly Ser Tyr His Phe Ser Pro Glu Asn 100 105
110 Cys Arg Phe Arg Gln Trp Thr Leu Glu Asn Gly Tyr Asp Val Tyr Leu
115 120 125 Ser Pro Lys His His Tyr Leu Val Ser Leu Gly Arg Ser Lys
Arg Ile 130 135 140 Phe Gln Pro Gly Thr Asn Pro Pro Pro Phe Ser Gln
Phe Leu Ala Arg 145 150 155 160 Arg Asn Glu Val Pro Leu Leu His Phe
Tyr Thr Ala Arg Pro Arg Arg 165 170 175 His Thr Arg Ser Ala Glu Asp
Pro Pro Glu Arg Asp Pro Leu Asn Val 180 185 190 Leu Lys Pro Arg Pro
Arg Ala Thr Pro Ile Pro Val Ser Cys Ser Arg 195 200 205 Glu Leu Pro
Ser Ala Glu Glu Gly Gly Pro Ala Ala Ser Asp Pro Leu 210 215 220 Gly
Val Leu Arg Arg Gly Arg Gly Asp Ala Arg Arg Gly Ala Gly Gly 225 230
235 240 Thr Asp Arg Cys Arg Pro Phe Pro Arg Phe Val 245 250
746PRTArtificial SequenceDescription of Artificial Sequence
Synthetic 6xHis tag 74His His His His His His 1 5
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