U.S. patent application number 14/885732 was filed with the patent office on 2016-02-04 for t7 rna polymerase variants with enhanced thermostability.
The applicant listed for this patent is ROCHE DIAGNOSTICS OPERATIONS, INC.. Invention is credited to Michael Greif, Rainer Mueller, Christian Rudolph, Armin Ruf, Manfred Schmidt, Harald Sobek, Johann-Peter Thalhofer.
Application Number | 20160032261 14/885732 |
Document ID | / |
Family ID | 42460013 |
Filed Date | 2016-02-04 |
United States Patent
Application |
20160032261 |
Kind Code |
A1 |
Sobek; Harald ; et
al. |
February 4, 2016 |
T7 RNA POLYMERASE VARIANTS WITH ENHANCED THERMOSTABILITY
Abstract
The present invention provides improved variants of T7 RNA
polymerase by introducing novel mutations which lead to improved
thermostability of the enzyme. According to the invention, amino
acid substitutions at the positions Val426, Ser633, Val650, Thr654,
Ala702, Val795, and combinations thereof are advantageous.
Inventors: |
Sobek; Harald; (Penzberg,
DE) ; Thalhofer; Johann-Peter; (Weilheim, DE)
; Mueller; Rainer; (Penzberg, DE) ; Schmidt;
Manfred; (Penzberg, DE) ; Greif; Michael;
(Penzberg, DE) ; Ruf; Armin; (Freiburg, DE)
; Rudolph; Christian; (Penzberg, DE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
ROCHE DIAGNOSTICS OPERATIONS, INC. |
Indianapolis |
IN |
US |
|
|
Family ID: |
42460013 |
Appl. No.: |
14/885732 |
Filed: |
October 16, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13069514 |
Mar 23, 2011 |
9193959 |
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14885732 |
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Current U.S.
Class: |
435/91.21 ;
435/194 |
Current CPC
Class: |
C12N 9/1247 20130101;
C12Y 207/07006 20130101; C12P 19/34 20130101 |
International
Class: |
C12N 9/12 20060101
C12N009/12; C12P 19/34 20060101 C12P019/34 |
Foreign Application Data
Date |
Code |
Application Number |
Apr 16, 2010 |
EP |
10004059.1 |
Claims
1. A variant polypeptide of T7 RNA polymerase (T7 variant), the T7
variant having (i) a DNA-dependent RNA polymerase activity, (ii) an
enhanced thermostability compared to SEQ ID NO: 2 (wild-type
reference), and (iii) a different composition of amino acids
compared to the wild-type reference, wherein the T7 variant
comprises a polypeptide of the wild-type reference in which at
least one amino acid and up to four amino acids at different
positions are substituted, wherein a different amino acid
substitutes for an amino acid selected from the group consisting of
Val426, Ser633, Val650, Thr654, Ala702, and Val795, numbered from
the N-terminus of the wild-type reference, and wherein the
different amino acid is selected from the group consisting of Leu,
Ile and Phe when the different amino acid substitutes for Val426
(Val426Leu, Val426Ile, Val426Phe), Val and Met when the different
amino acid substitutes for Ser633 (Ser633Val, Ser633Met), Leu when
the different amino acid substitutes for Val650 (Val650Leu), Leu
when the different amino acid substitutes for Thr654 (Thr654Leu),
Val when the different amino acid substitutes for Ala702
(Ala702Val), and Ile when the different amino acid substitutes for
Val795 (Val795Ile).
2-12. (canceled)
13. A method for synthesizing an RNA molecule comprising the steps
of (a) providing a template DNA molecule comprising a T7 promoter,
the T7 promoter being functionally linked to a target nucleotide
sequence to be transcribed; (b) contacting the template DNA of step
(a) with a variant polypeptide of T7 RNA polymerase (T7 variant)
according to claim 1; followed by (c) incubating the template DNA
and the T7 variant in the presence of ribonucleoside triphosphates,
thereby synthesizing the RNA molecule.
14-15. (canceled)
Description
RELATED APPLICATIONS
[0001] This application claims priority to European application EP
10004059.1 filed Apr. 16, 2010.
SEQUENCE LISTING
[0002] The instant application contains a Sequence Listing which
has been submitted in ASCII format via EFS-Web and is hereby
incorporated by reference in its entirety. Said ASCII copy, created
on Mar. 17, 2011, is named 26576US.txt, and is 297,574 bytes in
size.
FIELD OF THE INVENTION
[0003] The invention pertains to the field of biochemistry,
particularly to the field of enzyme engineering.
[0004] The present invention provides improved variants of T7 RNA
polymerase by introducing novel mutations which lead to improved
thermostability of the enzyme. According to the invention, amino
acid substitutions at the positions Val426, Ser633, Val650, Thr654,
Ala702, Val795, and combinations thereof are advantageous.
BACKGROUND OF THE INVENTION
[0005] T7 RNA polymerase (E.C. 2.7.7.6.; herein also referred to as
"T7 polymerase" or "T7") is a monomeric bacteriophage encoded DNA
directed RNA polymerase which catalyzes the formation of RNA in the
5'.fwdarw.3' direction. In the process of initiation of
transcription T7 recognizes a specific promoter sequence, the T7
promoter. T7 consists of 883 amino acids and has a molecular weight
of 99 kDa. On the level of amino acid sequence T7 is highly
homologous to T3 RNA polymerase and, to a lesser extent, SP6 RNA
polymerase. The three-dimensional structure of T7 is very similar
to other polymerases with different template and substrate
specificities, despite low sequence similarity. T7 consists of
different domains, the N-terminal domain, the "thumb", the "palm"
and the "fingers" (Sousa, R., and Mukherjee, S., Prog. Nucl. Acid
Res. Mol. Biol. 73 (2003) 1-41).
[0006] Detailed studies of the transcription reaction showed that
the enzyme acts like a molecular machine showing well concerted
movements of flexible parts of the enzyme (Steitz, T. A., EMBO J.
25 (2006) 3458-3468; Steitz, T. A., Curr. Opin. Struct. Biol. 14
(2004) 4-9; Yin, Y. W., and Steitz, T. A., Cell 116 (2004)
393-404).
[0007] Several structures of T7 in complex with promoter DNA were
solved and are available in the Protein Data Bank (pdb). The
structure of the initiation complex of T7 RNA polymerase was solved
at high resolution (Cheetham, G. M. T., et al., Nature 399 (1999)
80-83; Cheetham, G. M. T., and Steitz, T. A., Science 286 (1999)
2305-2309). The structure of the elongation complex solved at 2.9 A
resolution showed the rearrangement of the N-terminal region
(Tahirov, T. H., et al., Nature 420 (2002) 43-50). The structural
studies showed that the conformation of the N-terminal domain
changes between the initiation and elongation phases. Recently, the
structure of transcribing T7 in transition from initiation to
elongation phase was described (Durniak, K. J., et al., Science 322
(2008) 553-557).
[0008] The cloning and the expression of the gene encoding T7 has
been described (Studier et al., U.S. Pat. No. 4,952,496). T7 has
been studied intensively by mutagenesis to explore the
conformational changes during transcription (Ma, K., et. al., Proc.
Nat. Acad. Sci. 102 (2005) 17612-17617), to facilitate promoter
clearance (Guillerez, J., et al., Proc. Natl. Acad. Sci. 102 (2005)
5958-5963) or to study the abortive cycling phenomenon (He, B., et
al., J. Mol. Biol. 265 (1997) 275-288). Bonner, G., et al., J.
Biol. Chem. 269 (1994) 25120-25128 described a set of active site
mutants with altered elongation rates.
[0009] Due to the promoter specificity and high RNA polymerase
enzymatic activity, T7 is useful for a variety of applications in
molecular biology. In the field of recombinant protein expression
T7 is used for the high-level expression of recombinant genes in E.
coli (Studier, F. W., and Moffat, B. A., J. Mol. Biol. 189 (1986)
113-130). The synthesis of defined oligoribonucleotides was
described by Milligan, J. F., et al., Nucl. Aids Res. 15 (1987)
8783-8798.
[0010] In addition, T7 is used in nucleic acid amplification
methods for diagnostic purposes. A first example for such use is a
technique known as "Nucleic Acid Sequence Based Amplification"
(NASBA). This process comprises the steps of (a) adding a RNA
template to a reaction mixture, wherein a first primer anneals to a
complementary site at the 3' end of the template; (b) reverse
transcribing a DNA strand complementary to the RNA template,
wherein a RNA/DNA heteroduplex is formed; (c) degrading the RNA
strand of the heteroduplex by way of RNaseH activity; (d) annealing
a second primer to the 5' end of the DNA strand; (e) repeatedly
synthesizing a complementary RNA strand with T7 RNA polymerase,
wherein the synthesized RNA strand can serve again as a template in
step (a). The NASBA technique has been used to develop rapid
diagnostic tests for several pathogenic viruses, particularly those
with single-stranded RNA genomes.
[0011] A further example for a diagnostic isothermal amplification
method is "Transcription mediated amplification" (TMA) known to be
one of the most sensitive detection assays for hepatitis C virus
RNA in patient serum. For amplification of target RNA, two enzymes
are used which are reverse transcriptase (RT) and T7 RNA
polymerase. Complementary DNA (cDNA) of sample RNA is generated by
RT with RNAse H activity and a primer containing a T7-promoter at
the 5'-end. The RNA resulting of the RNA-DNA duplex is degradated
by RNAse H activity of the RT. Another primer then binds to the
cDNA already containing the T7-promotor sequence from the first
primer and a double-stranded DNA is synthesized by the DNA
polymerase activity of the RT. The T7 RNA polymerase recognizes the
T7-promoter sequence within the double-stranded DNA molecule and
synthesizes numerous RNA antisense transcripts. Each of the newly
produced RNA amplicons re-enters the TMA process and serves as a
template for a new round of RT to double-stranded DNA including the
T7-promotor and transcription of antisense amplicons. The
circulation of antisense transcripts into the amplification process
results in exponential amplification of target RNA.
[0012] For NASBA, TMA and related methods as well as for other
applications it would be advantageous if the reaction temperature
could be elevated to improve the reaction kinetics. E.g., higher
reaction temperatures of isothermal amplification could allow the
amplification of RNA having secondary structures. It has also been
shown with the polymerase chain reaction (PCR) technology that high
annealing temperatures allow the specific hybridization of a primer
to its target resulting in a highly specific amplification. With
the same advantage, more thermostable enzymes could in principle
also be applied isothermal amplifications.
[0013] Therefore, there is a need of a T7 RNA polymerase with
improved stability and activity at higher reaction
temperatures.
[0014] The stability of T7 RNA polymerase has been studied
extensively. Thermal and urea-induced unfolding of was studied by
Protasevich et al. using calorimetry, circular dichronism and
fluorescence (Protasevich, I. I., et al., FEBS Lett 349 (1994)
429-432). Under the conditions used the enzyme unfolded at
48.3.degree. C. Thermal unfolding was also studied by Griko et al.
using calorimetric methods (Griko, Y., et al., Prot. Sci. 10 (2001)
845-853). A smaller 22 kDa N-terminal part of the enzyme was shown
to increase the thermostability of the C-terminal 77 kDa
domain.
[0015] By introducing point mutations in the sequence of the
wild-type enzyme T7 variants were generated in which the stability
of T7 RNA polymerase was enhanced. The patents U.S. Pat. No.
6,524,828 and EP 1 261 696 describe four distinct amino acid
exchanges in the T7 RNA polymerase polypeptide (Ser430Pro,
Ser633Pro, Phe849Ile and Phe880Tyr) which stabilize the enzyme.
Combinations of two or more of these mutations in a modified T7
polypeptide result in even more stable enzyme variants.
[0016] The aim of the present invention was to extend the
collection of stabilizing mutations by creating novel mutations in
T7 RNA polymerase which lead to improved stability. It is further
desired to combine several of these mutations in a single T7
variant (double-, triple-, quadruple-, multiple-mutant), provided
the combined mutations lead to an even increased stability, that is
to say thermostability. According to the invention, new mutations
are found giving rise to T7 variants which exhibit high stability
in thermal unfolding assays.
SUMMARY OF THE INVENTION
[0017] A first aspect of the invention is a variant polypeptide of
T7 RNA polymerase (T7 variant), said T7 variant being characterized
by (i) a DNA-dependent RNA polymerase activity, (ii) an enhanced
thermostability compared to the 883-amino acid T7 RNA polymerase
polypeptide of SEQ ID NO: 2 (wild-type reference), and (iii) a
different composition of amino acids compared to the wild-type
reference, wherein the variant comprises a polypeptide of the
wild-type reference in which at least one amino acid and up to four
amino acids at different positions is/are substituted, wherein a
different amino acid substitutes for an amino acid selected from
the group consisting of Val426, Ser633, Val650, Thr654, Ala702, and
Val795, numbered from the N-terminus of the wild-type reference,
and wherein the different amino acid is selected from the group
consisting of Leu, Ile and Phe when the different amino acid
substitutes for Val426 (Val426Leu, Val426Ile, Val426Phe), Val and
Met when the different amino acid substitutes for Ser633
(Ser633Val, Ser633Met), Leu when the different amino acid
substitutes for Val650 (Val650Leu), Leu when the different amino
acid substitutes for Thr654 (Thr654Leu), Val when the different
amino acid substitutes for Ala702 (Ala702Val), Ile when the
different amino acid substitutes for Val795 (Val795Ile).
[0018] In a first embodiment, a different amino acid substitutes
for an amino acid selected from the group consisting of Val426,
Val650, Ala702, and Val795. In a preferred embodiment, four amino
acids at different positions are substituted, and the different
amino acids are Val426Leu, Val650Leu, Ala702Val, and Val795Ile. In
yet a further preferred embodiment, a different amino acid
substitutes for an amino acid selected from the group consisting of
Val426, Ala702, and Val795, and wherein up to three amino acids at
different positions is/are substituted. In yet a further preferred
embodiment, two or three amino acids at different positions are
substituted, and the different amino acids are selected from the
group consisting of Val426Leu, Ala702Val, and Val795Ile. In yet a
further preferred embodiment, three amino acids at different
positions are substituted and the different amino acids are
Val426Leu, Ala702Val, and Val795Ile.
[0019] A second aspect of the invention is a method to produce a
variant polypeptide of T7 RNA polymerase (T7 variant), said T7
variant being characterized by (i) a DNA-dependent RNA polymerase
activity, (ii) an enhanced thermostability compared to the
883-amino acid T7 RNA polymerase polypeptide of SEQ ID NO: 2
(wild-type reference), and (iii) a different composition of amino
acids compared to the wild-type reference, said method comprising
the steps of (a) selecting an amino acid from the group consisting
of Val426, Ser633, Val650, Thr654, Ala702, and Val795, numbered
from the N-terminus of the wild-type reference; (b) substituting a
selected amino acid with a different amino acid to form a T7
variant, wherein the different amino acid is selected from the
group consisting of Leu, Ile and Phe when the different amino acid
substitutes for Val426 (Val426Leu, Val426Ile, Val426Phe), Val and
Met when the different amino acid substitutes for Ser633
(Ser633Val, Ser633Met), Leu when the different amino acid
substitutes for Val650 (Val650Leu), Leu when the different amino
acid substitutes for Thr654 (Thr654Leu), Val when the different
amino acid substitutes for Ala702 (Ala702Val), Ile when the
different amino acid substitutes for Val795 (Val795Ile), wherein of
the wild-type reference at least one amino acid and up to four
amino acids at different positions is/are substituted; (c)
expressing a nucleic acid molecule encoding the T7 variant with the
substituted amino acids of step (b) in a transformed host organism,
and isolating the expressed T7 variant from the host organism;
thereby producing the T7 variant.
[0020] A third aspect of the invention is a method to produce a
nucleic acid molecule encoding a variant polypeptide of T7 RNA
polymerase (T7 variant), said T7 variant being characterized by (i)
a DNA-dependent RNA polymerase activity, (ii) an enhanced
thermostability compared to the 883-amino acid T7 RNA polymerase
polypeptide of SEQ ID NO: 2 (wild-type reference), and (iii) a
different composition of amino acids compared to the wild-type
reference, said method comprising the steps of (a) reverse
transcribing an amino acid sequence of a polypeptide according to
the invention or an amino acid sequence of a polypeptide obtainable
by the method according to the invention, thereby obtaining a
nucleic acid sequence; followed by (b) synthesizing a nucleic acid
molecule with the nucleic acid sequence obtained after performing
step (a); thereby producing the nucleic acid molecule encoding the
T7 variant.
[0021] A preferred embodiment of the invention is a nucleic acid
molecule with a sequence encoding a variant polypeptide of T7 RNA
polymerase (T7 variant), said T7 variant being characterized by (i)
a DNA-dependent RNA polymerase activity, (ii) an enhanced
thermostability compared to the 883-amino acid T7 RNA polymerase
polypeptide of SEQ ID NO: 2 (wild-type reference), and (iii) a
different composition of amino acids compared to the wild-type
reference, said nucleic acid being obtainable by the method of the
invention. Yet, a further preferred embodiment of the invention is
a nucleic acid molecule, wherein said nucleic acid is selected from
the group consisting of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 19,
SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 29, SEQ ID
NO: 37, SEQ ID NO: 39, SEQ ID NO: 41, SEQ ID NO: 43, and SEQ ID NO:
47.
[0022] A fourth aspect of the invention is an expression vector
comprising a nucleic acid molecule according to the invention,
wherein said nucleic acid molecule is functionally linked to one or
more sequences capable of controlling transcription and/or
translation.
[0023] A fifth aspect of the invention is a host organism capable
of recombinant expression of a polypeptide, wherein said host
organism is transformed with an expression vector according to the
invention.
[0024] A sixth aspect of the invention is a method to synthesize a
RNA molecule, comprising the steps of (a) providing a template DNA
molecule comprising a T7 promoter, said T7 promoter being
functionally linked to a target nucleotide sequence to be
transcribed; (b) contacting the template DNA of step (a) with a
variant polypeptide of T7 RNA polymerase (T7 variant) according to
the invention or a T7 variant obtainable by the method of the
inevtion; followed by (c) incubating the template DNA and the T7
variant in the presence of ribonucleoside triphosphates; thereby
synthesizing the RNA molecule.
[0025] A seventh aspect of the invention is a composition
comprising a template DNA molecule with a T7 promoter functionally
coupled to a target nucleotide sequence to be transcribed,
ribonucleoside triphosphates, and a variant polypeptide of T7 RNA
polymerase (T7 variant) according to the invention or a T7 variant
obtainable by the method according to the invention.
[0026] An eighth aspect of the invention is a kit comprising, in
separate containers, a variant polypeptide of T7 RNA polymerase (T7
variant) according to the invention or a T7 variant obtainable by
the method according to the invention and a buffer with one or more
ribonucleoside triphosphates.
DETAILED DESCRIPTION OF THE INVENTION
[0027] The present invention aims at providing the skilled person
with novel variants of T7 polymerase which are characterized by (i)
a DNA-dependent RNA polymerase enzymatic activity, (ii) an enhanced
thermostability compared to the wild-type T7 RNA polymerase
polypeptide (wild-type reference), and (iii) a different
composition of amino acids compared to the wild-type reference.
Such a "variant" is an allelic form of the wild-type T7 protein,
wherein said variant is generated by way of amino acid
substitution.
[0028] Certain terms are used with particular meaning or are
defined for the first time in this description of the present
invention. For the purposes of the invention, the terms used are
defined by their art-accepted definitions, when such exist, except
that when those definitions conflict or partially conflict with the
definitions set forth below. In the event of a conflict in
definition, the meaning of a term is first defined by any of the
definitions set forth in this document.
[0029] The term "comprising" is used in the description of the
invention and in the claims to mean "including, but not necessarily
limited to".
[0030] The articles "a" and "an" are used herein to refer to one or
to more than one (i.e., to at least one) of the grammatical object
of the article. By way of example, "a compound" means one compound
or more than one compound.
[0031] If not stated otherwise, it is understood that the term
"about" in combination with a numerical value n ("about n")
indicates a value x in the interval given by the numerical value
.+-.5% of the value, i.e., n-0.05*n.times.n+0.05*n. In case the
term "about" in combination with a numerical value n describes a
preferred embodiment of the invention, the value of n is most
preferred, if not indicated otherwise.
[0032] A nucleotide sequence "encodes" a peptide or polypeptide
when at least a portion of the nucleic acid, or its complement, can
be directly translated to provide the amino acid sequence of the
peptide or protein, or when the isolated nucleic acid can be used,
alone or as part of an expression vector, to express the peptide or
protein in vitro, in a prokaryotic host cell, or in a eukaryotic
host cell.
[0033] Where a nucleotide sequence is single-stranded, it is to be
understood that the complementary sequence of that nucleotide
sequence is also included within the scope of the present
invention.
[0034] The coding portion of a nucleotide sequence encoding a
peptide or a polypeptide begins with a start codon encoding
Methionine which thus becomes the N-terminal amino acid of the
primary translation product. As part of post-translational
processes, the N-terminal Methionine is frequently cleaved off,
e.g., by a Methionine aminopeptidase which is a ubiquituous enzyme.
In such a case, the primary translation product may give rise to a
mixture comprising members without N-terminal Methionine and
members retaining this amino acid as N-terminus. It is also
possible that the form of the enzyme without N-terminal Methionine
is the only one which can be isolated. However, the amino acid
sequences of the wild-type T7 polymerase and the T7 variants
according to the invention are described in the sequence listing
including N-terminal Methionine. But the present invention also
encompasses the said T7 variants which do not include N-terminal
Methionine.
[0035] For purposes of shorthand designation of T7 polymerase
variants described herein, it is noted that for each mutation a
number refers to the amino acid residue/position along the
reference amino acid sequence of the wild-type T7 polymerase
protein given in SEQ ID NO: 2. Amino acid identification uses the
three-letter abbreviations as well as the single-letter alphabet of
amino acids, i.e., Asp D Aspartic acid, Ile I Isoleucine, Thr T
Threonine, Leu L Leucine, Ser S Serine, Tyr Y Tyrosine, Glu E
Glutamic acid, Phe F Phenylalanine, Pro P Proline, His H Histidine,
Gly G Glycine, Lys K Lysine, Ala A Alanine, Arg R Arginine, Cys C
Cysteine, Trp W Tryptophan, Val V Valine, Gln Q Glutamine, Met M
Methionine, Asn N Asparagine. An amino acid at a particular
position in an amino acid sequence is given by its three-letter
abbreviation and a number. Accordingly, any of "Leu705" and "L705"
denote the leucine residue at amino acid position 705 in SEQ ID NO:
2. In any T7 mutant and/or T7 variant disclosed herein, a
substitution by a different amino acid is given as the three-letter
abbreviation added after the number indicating the position. E.g.,
"Leu705Ile" (=[Leu705Ile]) or "L705I" (=[L705I]) denote the
substitution of Leu at position 705 in SEQ ID NO: 2 by Ile (see #16
of Table 3). A Leu705Ile (=L705I) substitution results in an amino
acid sequence as given in SEQ ID NO: 28, encoded by the nucleotide
sequence of SEQ ID NO: 27. Preferred amino acid substitutions are
disclosed in Table 1 further below (see Example 1). Particularly
preferred variants according to the invention are characterized by
a plurality (preferred 2 to 4) of amino acid substitutions.
Examples therefor can be found in Table 4, for example--but not
limited to--#24 [Val426Leu, Val795Ile] or #25 [Val426Leu,
Ala702Val, Val7951Ile].
[0036] The term "polypeptide" or "protein" denotes a polymer
composed of a plurality of amino acid monomers joined by peptide
bonds. Preferably, the polymer comprises 50 or more monomers. A
preferred polypeptide or protein according to the invention is a T7
variant. A "peptide bond" is a covalent bond between a first amino
acid and a second amino acid in which the .alpha.-amino group of
the first amino acid is bonded to the .alpha.-carboxyl group of the
second amino acid.
[0037] The T7 variants of the invention also comprise fusion
proteins with an affinity tag such as, but not limited to, a
Histidine tag (His-tag). Well known to the skilled artisan, a
His-tag is an amino acid sequence containing several, preferably 3
to 7, more preferred 6 consecutive Histidines (SEQ ID NO: 53). In a
His-tag sequence the Histidines represent the essential portion.
But facultatively there are few additional amino acids comprised in
the His-tag. For example, a N-terminal T7 sequence including a
His-tag can comprise the sequence N-Met His His His His His His Gly
Ser- (residues 1-9 of SEQ ID NO: 52). To this end see SEQ ID NO: 52
comprising the foregoing amino acid sequence. In the present
exemplary His-tag the amino acids Gly and Ser form a linker to the
N-terminus of the T7 variant. The linker amino acids are part of
the His-tag and typically arise as a cloning artifact of the
nucleotide sequence encoding the His-tag (e.g., SEQ ID NO: 51).
Preferably, the linker sequence in the His-tag comprises 1 to 5
amino acids.
[0038] According to the invention, the N-terminal Methionine of a
T7 variant can be replaced by a His-tag. Alternatively, the
N-terminal sequence of the T7 variant according to the invention
can be extended by the His-tag. In such a case, the N-terminus of
the primary translation product of the T7 variant comprises a
N-terminal Methionine followed by the His-tag, followed by the
Methionine encoded by the start codon of the original T7 encoding
nucleotide sequence.
[0039] Purification of a His-tagged T7 wild-type or variant
polypeptide is efficiently performed by immobilized metal affinity
chromatography. This method is a widely employed method to purify
recombinant proteins containing a short affinity-tag consisting of
Histidine residues (His-tag). Immobilized metal-affinity
chromatography (described by Porath, J., et al., Nature 258 (1975)
598-599) is based on the interaction between a transition metal ion
(Co.sup.2+, Ni.sup.2+, Cu.sup.2+, Zn.sup.2+) immobilized on a
particulate metal chelating affinity matrix and specific amino acid
side chains. Histidine is the amino acid that exhibits the
strongest interaction with immobilized metal ion matrices, as
electron donor groups on the Histidine imidazole ring readily form
coordination bonds with the immobilized transition metal.
[0040] A "vector" is defined as a DNA which can comprise, i.e.,
carry, and maintain a DNA fragment according to the invention,
including, for example, phages and plasmids. These terms are
understood by those of skill in the art of genetic engineering. The
term "expression cassette" denotes a nucleotide sequence encoding a
pre-protein, operably linked to a promoter and a terminator. As for
vectors containing an expression cassette, the terms "vector" and
"expression vector" are used as synonyms.
[0041] The term "oligonucleotide" is used for a nucleic acid
molecule, DNA (or RNA), with less than 100 nucleotides in length.
Preferably, an oligonucleotide is about 75, about 50 or less
nucleotides in length
[0042] "Transformation" means introducing DNA into an organism,
i.e., a host organism, so that the DNA is replicable, either as an
extrachromosomal element or by chromosomal integration.
[0043] The term "expression" and the verb "to express" denote
transcription of DNA sequences and/or the translation of the
transcribed mRNA in a host organism resulting in a pre-protein,
i.e., not including post-translational processes.
[0044] A "promoter" is a regulatory nucleotide sequence that
stimulates transcription. These terms are understood by those of
skill in the art of genetic engineering. Like a promoter, a
"promoter element" stimulates transcription but constitutes a
sub-fragment of a larger promoter sequence.
[0045] The term "operably linked" refers to the association of two
or more nucleic acid fragments on a single vector so that the
function of one is affected by the other. For example, a promoter
is operably linked with a coding sequence, i.e., a nucleotide
sequence encoding a protein or a pre-protein, when it is capable of
affecting the expression of that coding sequence, i.e., that the
coding sequence is under the transcriptional control of the
promoter.
[0046] According to the invention, a first embodiment is a variant
polypeptide of T7 RNA polymerase (T7 variant), said T7 variant
being characterized by (i) a DNA-dependent RNA polymerase activity,
(ii) an enhanced thermostability compared to the 883-amino acid T7
RNA polymerase polypeptide of SEQ ID NO: 2 (wild-type reference),
and (iii) a different composition of amino acids compared to the
wild-type reference, wherein the variant comprises a polypeptide of
the wild-type reference in which at least one amino acid and up to
four amino acids at different positions is/are substituted, wherein
a different amino acid substitutes for an amino acid selected from
the group consisting of Val426, Ser633, Val650, Thr654, Ala702, and
Val795, numbered from the N-terminus of the wild-type reference,
and wherein the different amino acid is selected from the group
consisting of [0047] Leu, Ile and Phe when the different amino acid
substitutes for Val426 (Val426Leu, Val426Ile, Val426Phe), [0048]
Val and Met when the different amino acid substitutes for Ser633
(Ser633Val, Ser633Met), [0049] Leu when the different amino acid
substitutes for Val650 (Val650Leu), [0050] Leu when the different
amino acid substitutes for Thr654 (Thr654Leu), [0051] Val when the
different amino acid substitutes for Ala702 (Ala702Val), [0052] Ile
when the different amino acid substitutes for Val795
(Val795Ile).
[0053] The experimental work underlying the above selections
basically followed a rational approach to introduce at selected
positions predetermined amino acid substitutions in the T7
polypeptide. Among a large number of mutations tested, several
unexpectedly inhibited T7 polymerase activity, others--against the
prediction--did not noticeably enhance thermostability; only a few
mutations in fact showed the desired effects.
[0054] Since the development of site-directed mutagenesis
protocols, modification of enzyme-encoding nucleotide sequences has
become a powerful method in the field of protein engineering
(Winter, G., et al., Nature 299 (1982) 756-758). The knowledge of
the structure of an enzyme--combined with detailed biochemical data
concerning the principles underlying its function and
stability--offers the opportunity to rationally design enzymes with
improved properties. Examples for improvements are, e.g., enhanced
specificity, altered substrate spectrum, and thermostability. The
latter kind of improvements is pursued in the present case.
[0055] Fersht & Serrano discussed general principles of protein
stability derived from protein engineering experiments (Fersht, A.
R., and Serrano, L., Curr. Opin. Struct. Biol. 3 (1993) 75-83). The
specific interactions between amino acids in a protein and the
effect on stability were described. With regards to the
stabilization of a protein an improvement was exemplified by
"filling" hydrophobic cavities in the inner core of a protein with
hydrophobic amino acid residues. It was shown that by these means
the overall hydrophobic interaction in the protein core was
increased and that an increase of thermostability of the target
protein was achieved. Further developments in the field of
stabilization of protein structures were reviewed by Lee, B., and
Vasmatzis, G., Current Opinion Biotechn. 8 (1997) 423-428.
[0056] With the aim of applying this knowledge to the development
of the desired T7 variants, high resolution x-ray structures of T7
RNA polymerase deposited in data banks were carefully inspected.
Candidate sites in the protein structures were identified and sites
were selected where certain mutations could improve the stability
of the enzyme. The designed variants were synthesized, cloned,
expressed and purified. The stability of the enzyme variants was
examined and compared with the stability of the wild-type enzyme.
Table 3 in Example 1 summarizes the mutations considered in the
experimental work underlying the present invention.
[0057] To determine of the stability of wild-type T7 polymerase and
T7 variants two parameters were examined: (i) the half-life time
under a predetermined temperature regime, and (ii) the melting
temperature (T.sub.m).
[0058] In yet more detail, the present invention embodies the
following items.
[0059] 1. A variant polypeptide of T7 RNA polymerase (T7 variant),
said T7 variant being characterized by (i) a DNA-dependent RNA
polymerase activity, (ii) an enhanced thermostability compared to
the 883-amino acid T7 RNA polymerase polypeptide of SEQ ID NO: 2
(wild-type reference), and (iii) a different composition of amino
acids compared to the wild-type reference, [0060] wherein the
variant comprises a polypeptide of the wild-type reference in which
at least one amino acid and up to four amino acids at different
positions is/are substituted, [0061] wherein a different amino acid
substitutes for an amino acid selected from the group consisting of
Val426, Ser633, Val650, Thr654, Ala702, and Val795, numbered from
the N-terminus of the wild-type reference, and [0062] wherein the
different amino acid is selected from the group consisting of
[0063] Leu, Ile and Phe when the different amino acid substitutes
for Val426 (Val426Leu, Val426Ile, Val426Phe), [0064] Val and Met
when the different amino acid substitutes for Ser633 (Ser633Val,
Ser633Met), [0065] Leu when the different amino acid substitutes
for Val650 (Val650Leu), [0066] Leu when the different amino acid
substitutes for Thr654 (Thr654Leu), [0067] Val when the different
amino acid substitutes for Ala702 (Ala702Val), [0068] Ile when the
different amino acid substitutes for Val795 (Val795Ile).
[0069] 2. The T7 variant according to item 1, wherein a different
amino acid substitutes for an amino acid selected from the group
consisting of Val426, Val650, Ala702, and Val795.
[0070] 3. The T7 variant according to item 2, wherein four amino
acids at different positions are substituted, and the different
amino acids are Val426Leu, Val650Leu, Ala702Val, and Val795Ile.
[0071] 4. The T7 variant according to item 1, wherein a different
amino acid substitutes for an amino acid selected from the group
consisting of Val426, Ala702, and Val795, and wherein up to three
amino acids at different positions is/are substituted.
[0072] 5. The T7 variant according to item 4, wherein two or three
amino acids at different positions are substituted, and the
different amino acids are selected from the group consisting of
Val426Leu, Ala702Val, and Val795Ile.
[0073] 6. The T7 variant according to item 4, wherein three amino
acids at different positions are substituted and the different
amino acids are Val426Leu, Ala702Val, and Val1795Ile.
[0074] 7. The T7 variant according to any of the items 1 to 6,
additionally comprising a tag (affinity tag) capable of
specifically binding to an affinity chromatography matrix.
[0075] 8. The T7 variant according to any of the items 1 to 7,
wherein the affinity tag is a Histidine tag.
[0076] 9. The T7 variant according to item 8, wherein the Histidine
tag is a C-terminal tag.
[0077] 10. The T7 variant according to item 8, wherein the
Histidine tag is a N-terminal tag.
[0078] 11. The T7 variant according to item 10, additionally
comprising a linker sequence linking the Histidine tag with the
amino acid corresponding to Met1 or Asn2 of SEQ ID NO: 2.
[0079] 12. The T7 variant according to item 11, wherein the linker
sequence consists of between 1 and 4 amino acids.
[0080] 13. The T7 variant according to item 12, wherein the linker
sequence consists of between 1 and 2 amino acids.
[0081] 14. The T7 variant according to any of the items 1 to 13,
wherein at 50.degree. C. the half-life time of the variant is
between 12 min and about 320 min, and particularly between 12 min
and about 312 min; preferably, the T7 variant comprises (i) a
single amino acid substitution selected from the group consisting
of [Val426Leu], [Val426Ile], [Val426Phe], [Ser633Met], [Val650Leu],
[Thr654Leu], [Ala702Val], and [Val795Ile], or (ii) a double amino
acid substitution selected from the group consisting of [Ala702Val,
Val1795Ile], [Val426Leu, Ala702Val], and [Val426Leu, Val795Ile], or
(iii) the triple amino acid substitution [Val426Leu, Ala702Val,
Val795Ile], or (iv) the quadruple amino acid substitution
[Val426Leu, Val650Leu, Ala702Val, Val795Ile].
[0082] 15. The T7 variant according to item 14, wherein the
half-life time is between about 20 min and about 320 min, and
particularly between 20 min and about 312 min.
[0083] 16. The T7 variant according to item 14, wherein the
half-life time is between about 30 min and about 320 min, and
particularly between about 30 min and about 312 min.
[0084] 17. The T7 variant according to item 14, wherein the
half-life time is between about 40 min and about 320 min, and
particularly between about 40 min and about 312 min.
[0085] 18. The T7 variant according to item 14, wherein the
half-life time is between about 60 min and about 320 min, and
particularly between about 60 min and about 312 min.
[0086] 19. The T7 variant according to item 14, wherein the
half-life time is about 320 min.
[0087] 20. The T7 variant according to item 14, wherein the
half-life time is about 312 min.
[0088] 21. A method to produce a variant polypeptide of T7 RNA
polymerase (T7 variant), said T7 variant being characterized by (i)
a DNA-dependent RNA polymerase activity, (ii) an enhanced
thermostability compared to the 883-amino acid T7 RNA polymerase
polypeptide of SEQ ID NO: 2 (wild-type reference), and (iii) a
different composition of amino acids compared to the wild-type
reference, said method comprising the steps of [0089] (a) selecting
an amino acid from the group consisting of Val426, Ser633, Val650,
Thr654, Ala702, and Val795, numbered from the N-terminus of the
wild-type reference; [0090] (b) substituting a selected amino acid
with a different amino acid to form a T7 variant, wherein the
different amino acid is selected from the group consisting of
[0091] Leu, Ile and Phe when the different amino acid substitutes
for Val426 (Val126Leu, Val426Ile ,Val426Phe), [0092] Val and Met
when the different amino acid substitutes for Ser633 (Ser633Val,
Ser633Met), [0093] Leu when the different amino acid substitutes
for Val650 (Val650Leu), [0094] Leu when the different amino acid
substitutes for Thr654 (Thr654Leu), [0095] Val when the different
amino acid substitutes for Ala702 (Ala702Val), [0096] Ile when the
different amino acid substitutes for Val795 (Val795Ile), wherein at
least one amino acid of the wild-type reference and up to four
amino acids at different positions is/are substituted; [0097] (c)
expressing a nucleic acid molecule with a nucleotide sequence
encoding the T7 variant with the substituted amino acids of step
(b) in an expression system transformed host organism, and
isolating the expressed T7 variant from the expression system host
organism; thereby producing the T7 variant.
[0098] 22. The method of item 21, wherein the expression system is
selected from the group consisting of a transformed host organism
and a cell-free expression system.
[0099] 23. The method according to any of the items 21 and 22,
wherein in step (c) the nucleic acid molecule encodes a T7 variant
according to any of the items 1 to 20.
[0100] 24. A method to produce a nucleic acid molecule with a
nucleotide sequence encoding a variant polypeptide of T7 RNA
polymerase (T7 variant), said T7 variant being characterized by (i)
a DNA-dependent RNA polymerase activity, (ii) an enhanced
thermostability compared to the 883-amino acid T7 RNA polymerase
polypeptide of SEQ ID NO: 2 (wild-type reference), and (iii) a
different composition of amino acids compared to the wild-type
reference, said method comprising the steps of [0101] (a) reverse
transcribing an amino acid sequence of a polypeptide according to
any of the items 1 to 20 or an amino acid sequence of a polypeptide
obtainable by the method according to any of the items 21 to 23,
thereby obtaining a nucleic acid sequence; followed by [0102] (b)
synthesizing a nucleic acid molecule with the nucleic acid sequence
obtained after performing step (a); [0103] thereby producing the
nucleic acid molecule encoding the T7 variant.
[0104] 25. A nucleic acid molecule with a nucleotide sequence
encoding a variant polypeptide of T7 RNA polymerase (T7 variant),
said T7 variant being characterized by (i) a DNA-dependent RNA
polymerase activity, (ii) an enhanced thermostability compared to
the 883-amino acid T7 RNA polymerase polypeptide of SEQ ID NO: 2
(wild-type reference), and (iii) a different composition of amino
acids compared to the wild-type reference, said nucleic acid being
obtainable by the method of item 24.
[0105] 26. The nucleic acid molecule according to item 25, wherein
the nucleotide sequence of said nucleic acid comprises a sequence
selected from the group consisting of SEQ ID NO: 9, SEQ ID NO: 11,
SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID
NO: 29, SEQ ID NO: 37, SEQ ID NO: 39, SEQ ID NO: 41, SEQ ID NO: 43,
and SEQ ID NO: 47.
[0106] 27. An expression vector comprising a nucleic acid molecule
according to any of the items 25 and 26, wherein said nucleic acid
molecule is functionally linked to one or more nucleotide sequences
capable of controlling transcription and/or translation.
[0107] 28. A host organism capable of recombinant expression of a
polypeptide, wherein said host organism is transformed with an
expression vector according to item 27.
[0108] 29. A method to synthesize a RNA molecule, comprising the
steps of [0109] (a) providing a template DNA molecule comprising a
T7 promoter, said T7 promoter being functionally linked to a target
nucleotide sequence to be transcribed; [0110] (b) contacting the
template DNA of step (a) with a variant polypeptide of T7 RNA
polymerase (T7 variant) according to any of the items 1 to 20 or a
T7 variant obtainable by the method according to any of the items
21 to 23; followed by [0111] (c) incubating the template DNA and
the T7 variant in the presence of ribonucleoside triphosphates;
[0112] thereby synthesizing the RNA molecule.
[0113] 30. The method according to item 29, wherein step (c) is
performed at a temperature from 4.degree. C. to 55.degree. C., more
preferred at a temperature from 18.degree. C. to 50.degree. C., and
even more preferred at a temperature from 37.degree. C. to
50.degree. C.
[0114] 31. A composition comprising (i) a template DNA molecule,
the template DNA molecule comprising a
[0115] T7 promoter which is functionally coupled to a target
nucleotide sequence to be transcribed, (ii) ribonucleoside
triphosphates, (iii) an aqueous buffer, and (iv) a variant
polypeptide of T7 RNA polymerase (T7 variant) according to any of
the items 1 to 20 or a T7 variant obtainable by the method
according to any of the items 21 to 23.
[0116] 32. Use of a composition according to item 31 in a method
according to any of the items 29 and 30.
[0117] 33. A kit comprising, in separate containers, a variant
polypeptide of T7 RNA polymerase (T7 variant) according to any of
the items 1 to 20 or a T7 variant obtainable by the method
according to any of the items 21 to 23, and an aqueous buffer with
one or more ribonucleoside triphosphates.
[0118] 34. Use of a kit according to item 33 in a method according
to any of the items 29 and 30.
[0119] The following examples and sequence listing are provided to
aid the understanding of the present invention, the true scope of
which is set forth in the appended claims. It is understood that
modifications can be made in the procedures set forth without
departing from the spirit of the invention.
DESCRIPTION OF THE SEQUENCE LISTING
[0120] SEQ ID NO: 1 DNA (=nucleic acid; here and in the following
SEQ ID NO items to be read as DNA, if not indicated otherwise)
sequence encoding wild-type T7 DNA-dependent RNA polymerase,
including start codon encoding N-terminal Methionine; corresponding
to #1 in Table 3.
[0121] SEQ ID NO: 2 Wild-type T7 DNA-dependent RNA polymerase,
amino acid sequence including N-terminal Methionine; corresponding
to #1 in Table 3.
[0122] SEQ ID NO: 3 DNA sequence encoding the A319S variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #2 in Table 3.
[0123] SEQ ID NO: 4 A319S variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #2 in Table 3.
[0124] SEQ ID NO: 5 DNA sequence encoding the A319V variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #3 in Table 3.
[0125] SEQ ID NO: 6 A319V variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #3 in Table 3.
[0126] SEQ ID NO: 7 DNA sequence encoding the A319P variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #4 in Table 3.
[0127] SEQ ID NO: 8 A319P variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #4 in Table 3.
[0128] SEQ ID NO: 9 DNA sequence encoding the V426L variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #5 in Table 3.
[0129] SEQ ID NO: 10 V426L variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #5 in Table 3.
[0130] SEQ ID NO: 11 DNA sequence encoding the V4261 variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #6 in Table 3.
[0131] SEQ ID NO: 12 V4261 variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #6 in Table 3.
[0132] SEQ ID NO: 13 DNA sequence encoding the V426F variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #7 in Table 3.
[0133] SEQ ID NO: 14 V426F variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #7 in Table 3.
[0134] SEQ ID NO: 15 DNA sequence encoding the S633V variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #10 in Table 3.
[0135] SEQ ID NO: 16 S633V variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #10 in Table 3.
[0136] SEQ ID NO: 17 DNA sequence encoding the S633L variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #11 in Table 3.
[0137] SEQ ID NO: 18 S633L variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #11 in Table 3.
[0138] SEQ ID NO: 19 DNA sequence encoding the S633M variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #12 in Table 3.
[0139] SEQ ID NO: 20 S633M variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #12 in Table 3.
[0140] SEQ ID NO: 21 DNA sequence encoding the V650L variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #13 in Table 3.
[0141] SEQ ID NO: 22 V650L variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #13 in Table 3.
[0142] SEQ ID NO: 23 DNA sequence encoding the T654L variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #14 in Table 3.
[0143] SEQ ID NO: 24 T654L variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #14 in Table 3.
[0144] SEQ ID NO: 25 DNA sequence encoding the A702V variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #15 in Table 3.
[0145] SEQ ID NO: 26 A702V variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #15 in Table 3.
[0146] SEQ ID NO: 27 DNA sequence encoding the L7051 variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #16 in Table 3.
[0147] SEQ ID NO: 28 L7051 variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #16 in Table 3.
[0148] SEQ ID NO: 29 DNA sequence encoding the V7951 variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #18 in Table 3.
[0149] SEQ ID NO: 30 V7951 variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #18 in Table 3.
[0150] SEQ ID NO: 31 DNA sequence encoding the L809F variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #19 in Table 3.
[0151] SEQ ID NO: 32 L809F variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #19 in Table 3.
[0152] SEQ ID NO: 33 DNA sequence encoding the F814W variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #20 in Table 3.
[0153] SEQ ID NO: 34 F814W variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #20 in Table 3.
[0154] SEQ ID NO: 35 DNA sequence encoding the M861W variant of T7
DNA-dependent RNA polymerase, including start codon encoding
N-terminal Methionine; corresponding to #21 in Table 3.
[0155] SEQ ID NO: 36 M861W variant of T7 DNA-dependent RNA
polymerase, amino acid sequence including N-terminal Methionine;
corresponding to #21 in Table 3.
[0156] SEQ ID NO: 37 DNA sequence encoding the A702V, V795I variant
(double mutant) of T7 DNA-dependent RNA polymerase, including start
codon encoding N-terminal Methionine; corresponding to #22 in Table
3.
[0157] SEQ ID NO: 38 A702V, V795I variant (double mutant) of T7
DNA-dependent RNA polymerase, amino acid sequence including
N-terminal Methionine; corresponding to #22 in Table 3.
[0158] SEQ ID NO: 39 DNA sequence encoding the V426L, A702V variant
(double mutant) of T7 DNA-dependent RNA polymerase, including start
codon encoding N-terminal Methionine; corresponding to #23 in Table
3.
[0159] SEQ ID NO: 40 V426L, A702V variant (double mutant) of T7
DNA-dependent RNA polymerase, amino acid sequence including
N-terminal Methionine; corresponding to #23 in Table 3.
[0160] SEQ ID NO: 41 DNA sequence encoding the V426L, V795I variant
(double mutant) of T7 DNA-dependent RNA polymerase, including start
codon encoding N-terminal Methionine; corresponding to #24 in Table
3.
[0161] SEQ ID NO: 42 V426L, V795I variant (double mutant) of T7
DNA-dependent RNA polymerase, amino acid sequence including
N-terminal Methionine; corresponding to #24 in Table 3.
[0162] SEQ ID NO: 43 DNA sequence encoding the V426L, A702V, V795I
variant (triple mutant) of T7 DNA-dependent RNA polymerase,
including start codon encoding N-terminal Methionine; corresponding
to #25 in Table 3.
[0163] SEQ ID NO: 44 V426L, A702V, V795I variant (triple mutant) of
T7 DNA-dependent RNA polymerase, amino acid sequence including
N-terminal Methionine; corresponding to #25 in Table 3.
[0164] SEQ ID NO: 45 DNA sequence encoding the V426L, S633M, A702V,
V795I variant (quadruple mutant) of T7 DNA-dependent RNA
polymerase, including start codon encoding N-terminal Methionine;
corresponding to #26 in Table 3.
[0165] SEQ ID NO: 46 V426L, S633M, A702V, V795I variant (quadruple
mutant) of T7 DNA-dependent RNA polymerase, amino acid sequence
including N-terminal Methionine; corresponding to #26 in Table
3.
[0166] SEQ ID NO: 47 DNA sequence encoding the V426L, V650L, A702V,
V795I variant (quadruple mutant) of T7 DNA-dependent RNA
polymerase, including start codon encoding N-terminal Methionine;
corresponding to #27 in Table 3.
[0167] SEQ ID NO: 48 V426L, V650L, A702V, V795I variant (quadruple
mutant) of T7 DNA-dependent RNA polymerase, amino acid sequence
including N-terminal Methionine; corresponding to #27 in Table
3.
[0168] SEQ ID NO: 49 DNA sequence encoding the V426L, S633M, V650L,
A702V, V795I variant (quintuple mutant) of T7 DNA-dependent RNA
polymerase, including start codon encoding N-terminal Methionine;
corresponding to #28 in Table 3.
[0169] SEQ ID NO: 50 V426L, S633M, V650L, A702V, V795I variant
(quintuple mutant) of T7 DNA-dependent RNA polymerase, amino acid
sequence including N-terminal Methionine; corresponding to #28 in
Table 3.
[0170] SEQ ID NO: 51 DNA sequence encoding N-terminal Histidine tag
(His6) (SEQ ID NO: 54) with linker sequence, fused to the first two
N-terminal amino acids of T7 (Met and Asn).
[0171] SEQ ID NO: 52 Amino acid sequence of N-terminal Histidine
(His6) tag (SEQ ID NO: 54) with linker sequence, fused to the first
two N-terminal amino acids of T7 (Met and Asn).
Example 1: Design of Amino Acid Exchange Mutations in the T7
Polypeptide
[0172] X-ray structures of T7 RNA polymerase deposited in the
Protein Data Bank (codes: lcez [referring to Cheetham, G. M. T., et
al., Nature 399 (1999) 80-83], and 1s77 [referring to Yin, Y. W.,
and Steitz, T. A., Cell 116 (2004) 393-404]) were inspected to
identify candidate sites for the introduction of mutations to
increase the stability of the protein.
[0173] Selected positions of the T7 wild-type amino acid sequence
(according to SEQ ID NO: 2) are shown in Table 1 which also
provides amino acid substitution mutations expected to increase the
stability of the T7 polymerase protein. The underlying rationale of
the design of the mutations is also indicated.
[0174] Most of the substituting amino acids were selected either to
fill hydrophobic cavities in the core or to stabilize loops located
at the surface of the enzyme.
TABLE-US-00001 TABLE 1 Amino acid mutations of T7 RNA polymerase:
Design of an enzyme with increased thermostability Amino acid, WT
Position Mutation Rationale Ala 319 Ser, Val, Pro Fill cavity in
protein core Val 426 Leu, Ile, Phe, Trp Fill cavity in protein core
Val 629 Pro Stabilize loop Ser 633 Val, Leu, Met Stabilize loop Val
650 Leu Stabilize loop Thr 654 Leu Stabilize loop Ala 702 Val Fill
cavity in protein core Leu 705 Ile Fill cavity in protein core Leu
791 Phe Fill cavity in protein core Val 795 Ile Fill cavity in
protein core Leu 809 Phe Fill cavity in protein core Phe 814 Trp
Fill cavity in protein core Met 861 Trp Fill cavity in protein
core
[0175] In order to provide a coding sequence for any of the T7
mutants shown, the nucleotide sequence of SEQ ID NO: 1 encoding the
T7 wild type reference polypeptide was used as a basis. The
nucleotide codons corresponding to the amino acid residues at the
positions indicated in Table 1 were mutated, in order to encode the
changed amino acid at the respective position. Mutations were
preferably designed in accordance with the codon usage bias of E.
coli class II genes (Henaut, A., and Danchin, A., Analysis and
Predictions from Escherichia coli sequences. Escherichia coli and
Salmonella, Vol. 2, Ch. 114 (1996) 2047-2066, Neidhardt F C ed.,
ASM press, Washington, D.C.), as given in Table 2.
TABLE-US-00002 TABLE 2 Codon usage in E. coli Amino Class Amino
Class acid Codon I II III acid Codon I II III Phe T T T 55.09 29.08
67.14 Leu C T T 9.7 5.56 19 T T C 44.91 70.92 32.86 C T C 10.4 8.03
9.04 Leu T T A 10.99 3.44 20.09 C T A 3.09 0.83 6.81 T T G 13.02
5.47 15.05 C T G 52.79 76.67 29.99 Ser T C T 13.26 32.41 19.63 Pro
C C T 13.71 11.23 28.3 T C C 15.02 26.56 11.34 C C C 11.19 1.63
16.26 T C A 10.83 4.79 22.09 C C A 18.63 15.25 31.5 T C G 16.88
7.39 10.6 C C G 56.47 71.89 23.94 Tyr T A T 54.42 35.23 69.6 His C
A T 56.8 29.77 61.69 T A C 45.58 64.77 30.4 C A C 43.2 70.23 38.31
Stop T A A Gln C A A 33.4 18.65 37.06 T A G C A G 66.6 81.35 62.94
Cys T G T 40.9 38.85 55.71 Arg C G T 38.99 64.25 26.05 T G C 59.1
61.15 44.29 C G C 42.23 32.97 21.94 Stop T G A C G A 5.52 1.07 12.8
Trp T G G 100 100 100 C G G 8.97 0.8 13.62 Ile A T T 51.2 33.49
47.57 Val G T T 23.74 39.77 34.33 A T C 44.37 65.94 26.65 G T C
22.48 13.45 18.95 A T A 4.43 0.57 25.78 G T A 14.86 19.97 21.78 Met
A T G 100 100 100 G T G 38.92 26.81 24.94 Thr A C T 14.85 29.08
26.83 Ala G C T 14.52 27.54 22.86 A C C 46.83 53.6 24.45 G C C
27.62 16.14 23.67 A C A 10.52 4.67 27.93 G C A 19.63 24.01 31.27 A
C G 27.81 12.65 20.8 G C G 38.23 32.3 22.19 Asn A A T 40.87 17.25
64.06 Asp G A T 62.83 46.05 70.47 A A C 59.13 82.75 35.94 G A C
37.17 53.95 29.53 Lys A A A 75.44 78.55 72.21 Glu G A A 68.33 75.35
66.25 A A G 24.56 21.45 27.79 G A G 31.67 24.65 33.75 Ser A G T
13.96 4.52 18.73 Gly G G T 32.91 50.84 31.79 A G C 30.04 24.33
17.61 G G C 43.17 42.83 24.51 Arg A G A 1.75 0.62 15.63 G G A 9.19
1.97 24.75 A G G 1.54 0.29 9.96 G G G 14.74 4.36 18.95
[0176] The genes which served as the basis for the data in Table 2
were clustered by using factorial correspondence analysis into
three classes. Class I contains genes involved in most metabolic
processes. Class II genes correspond to genes highly and
continuously expressed during exponential growth. Class III genes
are implicated in horizontal transfer of DNA. One can see that the
distribution of codons in class III genes is more or less even,
whereas it is extremely biased in class II genes (in particular,
codons terminated in A are selected against).
[0177] The mutations on the codon level which were introduced in
the T7 coding sequence are shown in Table 3.
TABLE-US-00003 TABLE 3 T7 polymerase and variants thereof # T7
enzyme/variant WT codon Mutated codon SEQ ID NO: 1 Wild-type -- 1,
2 2 Ala319Ser GCG AGC 3, 4 3 Ala319Val GCG GTT 5, 6 4 Ala319Pro GCG
CCG 7, 8 5 Val426Leu GTT CTG 9, 10 6 Val426Ile GTT ATC 11, 12 7
Val426Phe GTT TTC 13, 14 8 Val426Trp GTT TGG 9 Val629Pro GTG CCG 10
Ser633Val TCA GTT 15, 16 11 Ser633Leu TCA CTG 17, 18 12 Ser633Met
TCA ATG 19, 20 13 Val650Leu GTG CTG 21, 22 14 Thr654Leu ACC CTG 23,
24 15 Ala702Val GCT GTT 25, 26 16 Leu705Ile CTG ATC 27, 28 17
Leu791Phe CTT TTC 18 Val795Ile GTA ATC 29, 30 19 Leu809Phe CTG TTC
31, 32 20 Phe814Trp TTC TGG 33, 34 21 Met861Trp ATG TGG 35, 36 22
Ala702Val GCT GTT 37, 38 Val795Ile GTA ATC 23 Val426Leu GTT CTG 39,
40 Ala702Val GCT GTT 24 Val426Leu GTT CTG 41, 42 Val795Ile GTA ATC
25 Val426Leu GTT CTG 43, 44 Ala702Val GCT GTT Val795Ile GTA ATC 26
Val426Leu GTT CTG 45, 46 Ser633Met TCA ATG Ala702Val GCT GTT
Val795Ile GTA ATC 27 Val426Leu GTT CTG 47, 48 Val650Leu GTG CTG
Ala702Val GCT GTT Val795Ile GTA ATC 28 Val426Leu GTT CTG 49, 50
Ser633Met TCA ATG Val650Leu GTG CTG Ala702Val GCT GTT Val795Ile GTA
ATC
[0178] On the amino acid level, the T7 variants are shown in SEQ ID
NOs: 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34,
36, 38, 40, 42, 44, 46, 48, and 50.
[0179] The nucleotide sequences encoding the mutated T7
polypeptides which were expressed in E. coli are shown in SEQ ID
NOs: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33,
35, 37, 39, 41, 43, 45, 47, and 49. The sequences are represented
including the start codons for N-terminal Methionine but without
any other additional artificial N-terminal structures such as
His-tags.
[0180] Well-known to the art, a His-tag (in the literature also
referred to as a polyHis-tag) is an amino acid motif in proteins
that typically consists of at least six consecutive His residues
(His6) (SEQ ID NO: 54). While the N-terminus of a T7 variant is
preferred for the addition of the His-tag, the C-terminus of the
polypeptide can serve as an alternative.
[0181] For clarification, a N-terminal His-tag can be located
between the Methionine at the N-terminus of the respective variant
T7 polypeptide and the subsequent amino acid according to the amino
acid sequence of SEQ ID NO: 2, i.e., Asn. Alternatively, the
His-tag can be appended to the N-terminal Methionine of the T7
variant. When appended at the C-terminus of the variant T7
polypeptide the His-tag forms the C-terminal amino acids.
[0182] The T7 variants were modified further such that each
polypeptide contained a His-tag at its N-terminus to facilitate
purification.
[0183] Apart from the Histidines the His-tag can additionally
comprise further amino acids depending on the design of the
nucleotide sequence encoding the His-tag. Thus, an oligonucleotide
linker with restriction sites typically adds 1 to 5 further amino
acids to the nucleotide fragment encoding the consecutive His
residues in the His-tag.
[0184] The amino acid sequences of the T7 variants of Table 1 and
the nucleic acid sequences encoding the T7 variants are shown in
the sequence listing of this disclosure. No His-tags are shown as
these may differ sequence-wise, depending on the particular cloning
vector used. However, the differences concerning the number of
Histidines and the linker sequence, according to the preferred
embodiments, are not expected to have a technical impact on the T7
variants according to the invention.
Example 2: Cloning of Nucleic Acids Encoding Variants of T7 RNA
Polymerase
[0185] All molecular biological procedures were performed according
to standard methods (Sambrook J., Fritsch E. F., Maniatis T.,
(1989) Molecular cloning: A Laboratory Manual second Edition, B.27
Cold Spring Harbor Laboratory Press NY (USA)). Nucleotide sequences
encoding the wild-type and the mutant T7 polypeptides were
synthesized by a combinatorial synthesis strategy as described (van
den Brulle, J., et al., Biotechniques 45(3) (2008) 340-343).
[0186] For expression of each of the T7 variants, the respective
coding DNA sequence was cloned in appropriate expression vectors in
such a way that the mutated T7 coding sequence is inserted in the
right orientation under the control of an appropriate promoter,
preferably an inducible promoter, particularly preferably the lac-,
lacUV5-, tac- or T5 promoter. Preferred expression vectors are pUC
plasmids with lac- or lacUV5 promoters or pKK plasmids. For
clarification an exemplary coding sequence comprises a DNA encoding
a polypeptide selected from any of SEQ ID NOs: 3, 5, 7, 9, 11, 13,
15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47,
and 49 which optionally include a further modification such as a
His-tag.
[0187] The synthesized genes were cloned in plasmid pUC18. The
recipient strain for transformations was E. coli XL-1 blue.
Transformed clones were grown at 37.degree. C. in LB media
containing ampicillin (100 .mu.g/ml). Plasmids were isolated and
digested using EcoRI and HindIII. The resulting fragments were
subjected to agarose gel electrophoresis and the respective band
corresponding to the variant T7 polymerase coding sequence was
extracted. The isolated fragments were ligated into the expression
plasmid pKKT5 (derived from pKK177-3 [Kopetzki, E., et al., Mol.
Gen. Genet. 216 (1989) 149-155] by exchanging the tac-promotors
with the T5-promoter derived from the plasmid pDS [Bujard, H., et
al., Methods Enzymol. 155 (1987) 416-433]) which was digested with
EcoRI and HindIII.
[0188] Plasmids were transformed into E. coli UT5600 (harboring
plasmid pUBS520). Clones were grown at 37.degree. C. in LB media
containing ampicillin (100 .mu.g/ml) and kanamycin (50
.mu.g/ml).
Example 3: Expression and Purification of Variant T7 Polymerase
Polypeptides
[0189] Transformed E. coli expression strains obtained as described
in Example 2 were cultivated at 37.degree. C. in LB media
containing ampicillin (100 .mu.g/ml) and kanamycin (50 .mu.g/ml).
Induction of recombinant expression was performed at an optical
density of 0.7 (measured at 578 nm) by adding IPTG in a final
concentration of 1 mM. After 5 hours the cells were harvested by
centrifugation and frozen at -20.degree. C.
[0190] His-tagged wild-type T7 polymerase and T7 variants were
purified separately to homogeneity using metal chelate affinity
matrix chromatography. Typically, wet frozen cells (2.1 g) were
suspended in 30 ml buffer A (50 mM Tris/HCl, pH 8.1 [pH adjusted at
room temperature], 1 M NaCl). To the suspension 315 .mu.l of a
lysozyme solution (10 mg/ml) were added. After incubation at room
temperature for 15 min, the cells were sonicated (6.times.2 min).
The cell debris was removed by centrifugation at 5,000 rpm for 15
min. A fraction of the supernatant (25 ml) was applied onto a
Ni-chelating Sepharose column (1 ml). The column was washed using
buffer A which additionally contained 10 mM imidazole. His-tagged
polypeptides were eluted in a linear gradient (10 mM-500 mM
imidazole in buffer A). Enzyme-containing fractions were pooled.
After dialysis against storage buffer (10 mM potassium phosphate,
200 mM KCl, 0.1 mM EDTA, 30 mM mercaptoethanol, 50% glycerol, 0.1%
Tween 20, pH 7.9) the pools were stored at -20.degree. C.
Example 4: Analysis of DNA-Dependent RNA Polymerase Activity
[0191] A transcription-based non-radioactive assay was used to
measure the activity of purified wild-type and variants of T7 RNA
polymerase obtained as described in Example 3. The enzyme activity
was measured in 40 .mu.l reaction buffer (40 mM Tris/HCl, 6 mM
MgCl.sub.2, 1 mM NTP (each), 10 mM DTE, 2 mM spermidine, pH 8.0, 1
.mu.g pSPT18 cleaved with SspI). T7 wild-type or T7 variant
polymerase enyzme was added in diluted form. After incubation at
37.degree. C. for 30 min. EDTA (0.4 M, 4 .mu.l) was added to stop
the reaction. RNA quantification was done using Quant-iT RNA Assay
(Invitrogen) on a LC480 Light Cycler platform (Roche Applied
Science, Roche Diagnostics GmbH, Mannheim). As a reference enzyme
commercially available T7 RNA polymerase was used (Roche Applied
Science, Roche Diagnostics GmbH, Mannheim).
Example 5: Analysis of Thermostability: Half-Life Time
[0192] To determine the stability of wild-type T7 polymerase and T7
variants the half-life time was determined as a first parameter.
Samples of wild-type enzyme and purified variants (see Example 3)
were incubated in storage buffer (10 mM potassium phosphate, 200 mM
KCl, 0.1 mM EDTA, 30 mM mercaptoethanol, 50% glycerol, 0.1% Tween
20, pH 7.9) at 50.degree. C. At different time points (10, 20 and
30 min) samples were taken and the residual enzyme activity was
measured as described in Example 3. The half-life time T.sub.1/2
expressed as a number of minutes [min] means that at this time
point the activity of the respective T7 variant is 50% of the
activity at the time point when the experiment was started, i.e.,
the exposure to 50.degree. C. was applied. Table 4 summarizes
results of the measurements.
TABLE-US-00004 TABLE 4 Half-life times at 50.degree. C. of
wild-type T7 RNA polymerase and T7 variants (single mutations and
combination mutations) # T7 enzyme T.sub.1/2 [min] Reference 1
Wild-type 6.0-9.7 Single amino acid substitution 2 Ala319Ser no
activity 3 Ala319Val 8.5 4 Ala319Pro no activity 5 Val426Leu 25.0 6
Val426Ile 17.0 7 Val426Phe 12.0 8 Val426Trp 0.4 9 Val629Pro no
activity 10 Ser633Val 7.3 11 Ser633Leu 5.3 12 Ser633Met 13.0 13
Val650Leu 13.0 14 Thr654Leu 13.0 15 Ala702Val 22.0 16 Leu705Ile
10.0 17 Leu791Phe no activity 18 Val795Ile 29.0 19 Leu809Phe 7.7 20
Phe814Trp 1.9 21 Met861Trp 5.5 Double amino acid substitution 22
Ala702Val 22.0 Val795Ile 23 Val426Leu 39.0 Ala702Val 24 Val426Leu
40.0 Val795Ile Triple amino acid substitution 25 Val426Leu 312.0
Ala702Val Val795Ile Quadruple amino acid substitution 26 Val426Leu
no activity Ser633Met Ala702Val Val795Ile 27 Val426Leu 64.0
Val650Leu Ala702Val Val795Ile Quintuple amino acid substitution 28
Val426Leu no activity Ser633Met Val650Leu Ala702Val Val795Ile
[0193] Designations of mutants are the same as in Table 3.
[0194] With regards to half-life times at 50.degree. C., the
inventors observed several surprising effects. Firstly, there were
single amino acid exchanges without noticeable impact on
thermostability, i.e., mutations which did not cause a substantial
difference compared to the wild-type reference (# 1). In this first
group all T7 variants with a T1/2 value between 5 and 12 (including
5 and 12) were combined. The first group comprises the mutations
according to ## 3, 7, 10, 11, 16, 19, and 21 as shown in Table
4.
[0195] A second group of T7 variants was found in which the mutants
had even shorter half-life times at 50.degree. C., compared to the
wild-type reference. Additionally, mutants which had lost enzymatic
activity completely were combined in the second group. The second
group comprises the mutations according to ## 2, 4, 8, 9, 17, 20,
26, and 28 as shown in Table 4.
[0196] A third group of amino acid exchange mutations was found to
enhance half-life time at 50.degree. C. over the values found for
the wild-type reference. A value greater than 12 was considered as
indicating a substantial increase of thermostability in the
respective T7 variant. The third group comprises the mutations
according to ## 5, 6, 12, 13, 14, 15, 18, 22, 23, 24, 25 and 27 as
shown in Table 4.
[0197] Surprisingly, some amino acid substitutions which, according
to theoretical predictions, were predicted to have a desired
positive effect on thermostability did not lead to the expected
results.
[0198] It was noted that mutations at the position Ala319 (see
Table 1) belonged to either the first or the second group.
According to the theory, these mutations were expected to fill a
hydrophobic cavity and thereby increase internal Van-der-Waals
forces within the core of the variant T7 polypeptide. Remarkably,
the mutation of Ala319 to Val (#3 in Table 4) leaves the RNA
polymerase activity intact but without having any noticeable impact
on the variant's half-life time at 50.degree. C. But at this
position changes to Ser or Pro (#2, #4 in Table 4) abolish
enzymatic activity. This finding was interpreted as an indication
of more complex intramolecular interactions which impact on the
structure and/or enzymatic function of T7 polymerase.
[0199] Further results were found at position Val426 which was
another candidate residue to be exchanged for generating increased
van-der-Waals forces in a cavity of the protein core. In line with
the prediction, changing Val426 to Leu, Ile or Phe (#5, #6, #7 in
Table 4) produced a variant with enhanced thermostability. But the
mutation Val426Trp (#8 in Table 4) unexpectedly destabilized the
polymerase enzyme. The same was observed for Leu791Phe (#17 in
Table 4) and, although to a somewhat lesser extent, for Phe814Trp
(#20in Table 4).
[0200] The results obtained for the further mutations Leu705Ile
(#16 in Table 4), Leu809Phe (#19 in Table 4) and Met861Trp (#21 in
Table 4) did not provide an indication for enhanced
thermostability.
[0201] Thus, with regards to the approach of increasing
van-der-Waals forces in internal hydrophobic cavities, the general
theoretical prediction of thermostabilization could not be verified
at this point. Rather, the results shown in Table 4 demonstrate
that (i) only a subgroup of the sites predicted to be suitable for
the desired class of mutations is amenable to amino acid
substitutions which actually lead to enhanced thermostability, (ii)
there is no means at hand to pre-select this subgroup, and (iii) no
prediction can be made which particular amino acid will be
effective and produce the desired technical effect.
[0202] The same appears to apply to mutations designed to stabilize
loop structures. Among six different mutations tested, one
(Val629Pro) (#9 in Table 4) completely abolishes enzymatic
activity, two (Ser633Val, Ser633Leu; #10 and #11 in Table 4) do not
show profound effects, and three (Ser633Met, Val650Leu, and
Thr654Leu; #12, #13 and #14 in Table 4) increase thermostability,
however not as markedly as mutations of the third group directed to
hydrophobic cavities. Again, one position (Ser633; #10, #11, and
#12) stands out in that mutations unexpectedly lead to different
effects.
[0203] Three mutations targeting stabilization of hydrophobic
cavities produced results which stood out: Val426Leu, Ala702Val,
and Val795Ile (#5, 15, and 18 in Table 4). The three mutations were
combined in double and triple mutant variants. Interestingly, all
combinations result in functional polymerase enzymes with increased
thermostablility when compared with the wild-type reference. Two
double mutant combinations (#23 and #24 in Table 4) showed an even
further increased thermostability which could be attributed to an
additive effect. One double mutant combination (#22 in Table 4)
suggests a polar mechanism in that the quantitative effect
Ala702Val mutation seems to override the effect of the Val795Ile
mutation. However, most strikingly, upon combination of the
mutations in a triple mutant a synergistic effect was observed in
that thermostability was increased by about one full order of
magnitude (#25 in Table 4).
[0204] The synergistic effect is further illustrated by the fact
that a variant with a quadruple mutant polypeptide which further
includes the Val650Leu exchange (#27 in Table 4) shows a further
additive effect. Regarding thermostability the effects seen with
the quadruple mutant exceeded each of the double mutants but they
fail to reach the high value observed with the triple mutant.
[0205] A further quadruple and a quintuple mutant variant (#26 and
#28 in Table 4) illustrate that it is not possible to deliberately
combine further mutations, i.e., that there are effects which
render some or all of the combined mutations incompatible.
Example 6: Protein Thermal Unfolding Assay
[0206] The stability of wild-type T7 polymerase enzyme and variants
of T7 RNA polymerase was further studied by way of determining
protein thermal unfolding. The assay was performed essentially as
by Yeh, A. P., et al., Acta Cryst. D62 (2006) 451-457, for
measuring the unfolding transition of membrane proteins by
fluorescent-probe binding in combination with real-time PCR
instrumentation and using SYPRO Orange as reporter dye.
[0207] Accordingly, all assays were performed using a LC480 LIGHT
CYCLER (Roche Applied Science). SYPRO Orange was obtained from
Molecular Probes Inc. (Eugene, Oreg., USA) and was diluted 1:10 in
DMSO. Protein samples (typically 2 .mu.g) were in Bis-Tris-propane
buffer (50-100 mM), pH 8.0 and contained diluted SYPRO Orange
(1:1430). Excitation wave length was 483 nm, emission was measured
at 568 nm.
[0208] Assays were performed in a temperature range starting from
37.degree. C. up to 94.degree. C. with a temperature ramp of
3.6.degree. C./min. Protein thermal unfolding was measured in the
absence (Buffer A) or presence (Buffer B) of 50% [v/v] glycerol
(see Table 5).
TABLE-US-00005 TABLE 5 Protein thermal unfolding, T.sub.m
determination # T7 enzyme T.sub.m [.degree. C.] in Buffer A T.sub.m
[.degree. C.] in Buffer B Reference 1 Wild-type 50.0 51.5 Single
amino acid exchange 5 Val426Leu 52.5 52.5 13 Val650Leu 51.0 n.d. 15
Ala702Val 51.0 n.d. 18 Val795Ile 51.5 n.d. Triple amino acid
exchange 25 Val426Leu n.d. (not determined) 55.0 Ala702Val n.d.
n.d. Val795Ile n.d. n.d.
[0209] Designations of T7 variants are the same as in Table 4.
[0210] The data again show that the triple mutant T7 variant #25
exhibits the highest increase of the T.sub.m, compared to the
wild-type reference.
Example 7: Determination of Protein Concentration in Solutions
[0211] Protein concentrations were determined by measuring the
optical density at 280 nm using a molar extinction coefficient of
E280 nm=1.4.times.10.sup.5 M.sup.-1 as described (He, B., et al.,
Protein Expr Purif 9 (1997) 142-151).
Sequence CWU 1
1
5412655DNAEscherichia coliPhage T7 1atgaacacga ttaacatcgc
taagaacgac ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca
ttacggtgag cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt
acgagatggg tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa
180gctggtgagg ttgcggataa cgctgccgcc aagcctctca tcactaccct
actccctaag 240atgattgcac gcatcaacga ctggtttgag gaagtgaaag
ctaagcgcgg caagcgcccg 300acagccttcc agttcctgca agaaatcaag
ccggaagccg tagcgtacat caccattaag 360accactctgg cttgcctaac
cagtgctgac aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg
ccattgagga cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag
480cacttcaaga aaaacgttga ggaacaactc aacaagcgcg tagggcacgt
ctacaagaaa 540gcatttatgc aagttgtcga ggctgacatg ctctctaagg
gtctactcgg tggcgaggcg 600tggtcttcgt ggcataagga agactctatt
catgtaggag tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt
tagcttacac cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta
tcgaactcgc acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg
780ctggctggca tctctccgat gttccaacct tgcgtagttc ctcctaagcc
gtggactggc 840attactggtg gtggctattg ggctaacggt cgtcgtcctc
tggcgctggt gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac
gtttacatgc ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc
atggaaaatc aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt
ggaagcattg tccggtcgag gacatccctg cgattgagcg tgaagaactc
1080ccgatgaaac cggaagacat cgacatgaat cctgaggctc tcaccgcgtg
gaaacgtgct 1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc
gccgtatcag ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac
cataaggcca tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta
cgctgtgtca atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc
ttacgctggc gaaaggtaaa ccaatcggta aggaaggtta ctactggctg
1380aaaatccacg gtgcaaactg tgcgggtgtc gataaggttc cgttccctga
gcgcatcaag 1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta
agtctccact ggagaacact 1500tggtgggctg agcaagattc tccgttctgc
ttccttgcgt tctgctttga gtacgctggg 1560gtacagcacc acggcctgag
ctataactgc tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc
agcacttctc cgcgatgctc cgagatgagg taggtggtcg cgcggttaac
1680ttgcttccta gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa
agtcaacgag 1740attctacaag cagacgcaat caatgggacc gataacgaag
tagttaccgt gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag
ctgggcacta aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg
cagtgtgact aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt
tcggcttccg tcaacaagtg ctggaagata ccattcagcc agctattgat
1980tccggcaagg gtctgatgtt cactcagccg aatcaggctg ctggatacat
ggctaagctg 2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg
aagcaatgaa ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc
aaagataaga agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg
ggtaactcct gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc
agacgcgctt gaacctgatg ttcctcggtc agttccgctt acagcctacc
2280attaacacca acaaagatag cgagattgat gcacacaaac aggagtctgg
tatcgctcct 2340aactttgtac acagccaaga cggtagccac cttcgtaaga
ctgtagtgtg ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt
cacgactcct tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc
agtgcgcgaa actatggttg acacatatga gtcttgtgat 2520gtactggctg
atttctacga ccagttcgct gaccagttgc acgagtctca attggacaaa
2580atgccagcac ttccggctaa aggtaacttg aacctccgtg acatcttaga
gtcggacttc 2640gcgttcgcgt aataa 26552883PRTEscherichia coliPhage T7
2Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1
5 10 15 Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg
Leu 20 25 30 Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu
Met Gly Glu 35 40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu
Lys Ala Gly Glu Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu
Ile Thr Thr Leu Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp
Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr
Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala
Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala
Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135
140 Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys
145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg
Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu
Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp
Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg
Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu
His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu
Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255
Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260
265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp
Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser
Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu
Val Tyr Lys Ala Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp
Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr
Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu
Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn
Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380
Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385
390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile
Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala
Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly
Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly
Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val
Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu
Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu
Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505
510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr
515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly
Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly
Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp
Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln
Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val
Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu
Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val
Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630
635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile
Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln
Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu
Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn
Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys
Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala
Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu
Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750
Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755
760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val
His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp
Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile
His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu
Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser
Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln
Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala
Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875
880 Ala Phe Ala 32655DNAArtificial SequenceDescription of
Artificial Sequence Synthetic coding polynucleotide 3atgaacacga
ttaacatcgc taagaacgac ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc
tggctgacca ttacggtgag cgtttagctc gcgaacagtt ggcccttgag
120catgagtctt acgagatggg tgaagcacgc ttccgcaaga tgtttgagcg
tcaacttaaa 180gctggtgagg ttgcggataa cgctgccgcc aagcctctca
tcactaccct actccctaag 240atgattgcac gcatcaacga ctggtttgag
gaagtgaaag ctaagcgcgg caagcgcccg 300acagccttcc agttcctgca
agaaatcaag ccggaagccg tagcgtacat caccattaag 360accactctgg
cttgcctaac cagtgctgac aatacaaccg ttcaggctgt agcaagcgca
420atcggtcggg ccattgagga cgaggctcgc ttcggtcgta tccgtgacct
tgaagctaag 480cacttcaaga aaaacgttga ggaacaactc aacaagcgcg
tagggcacgt ctacaagaaa 540gcatttatgc aagttgtcga ggctgacatg
ctctctaagg gtctactcgg tggcgaggcg 600tggtcttcgt ggcataagga
agactctatt catgtaggag tacgctgcat cgagatgctc 660attgagtcaa
ccggaatggt tagcttacac cgccaaaatg ctggcgtagt aggtcaagac
720tctgagacta tcgaactcgc acctgaatac gctgaggcta tcgcaacccg
tgcaggtgcg 780ctggctggca tctctccgat gttccaacct tgcgtagttc
ctcctaagcc gtggactggc 840attactggtg gtggctattg ggctaacggt
cgtcgtcctc tggcgctggt gcgtactcac 900agtaagaaag cactgatgcg
ctacgaagac gtttacatgc ctgaggtgta caaaagcatt 960aacattgcgc
aaaacaccgc atggaaaatc aacaagaaag tcctagcggt cgccaacgta
1020atcaccaagt ggaagcattg tccggtcgag gacatccctg cgattgagcg
tgaagaactc 1080ccgatgaaac cggaagacat cgacatgaat cctgaggctc
tcaccgcgtg gaaacgtgct 1140gccgctgctg tgtaccgcaa ggacaaggct
cgcaagtctc gccgtatcag ccttgagttc 1200atgcttgagc aagccaataa
gtttgctaac cataaggcca tctggttccc ttacaacatg 1260gactggcgcg
gtcgtgttta cgctgtgtca atgttcaacc cgcaaggtaa cgatatgacc
1320aaaggactgc ttacgctggc gaaaggtaaa ccaatcggta aggaaggtta
ctactggctg 1380aaaatccacg gtgcaaactg tgcgggtgtc gataaggttc
cgttccctga gcgcatcaag 1440ttcattgagg aaaaccacga gaacatcatg
gcttgcgcta agtctccact ggagaacact 1500tggtgggctg agcaagattc
tccgttctgc ttccttgcgt tctgctttga gtacgctggg 1560gtacagcacc
acggcctgag ctataactgc tcccttccgc tggcgtttga cgggtcttgc
1620tctggcatcc agcacttctc cgcgatgctc cgagatgagg taggtggtcg
cgcggttaac 1680ttgcttccta gtgaaaccgt tcaggacatc tacgggattg
ttgctaagaa agtcaacgag 1740attctacaag cagacgcaat caatgggacc
gataacgaag tagttaccgt gaccgatgag 1800aacactggtg aaatctctga
gaaagtcaag ctgggcacta aggcactggc tggtcaatgg 1860ctggcttacg
gtgttactcg cagtgtgact aagcgttcag tcatgacgct ggcttacggg
1920tccaaagagt tcggcttccg tcaacaagtg ctggaagata ccattcagcc
agctattgat 1980tccggcaagg gtctgatgtt cactcagccg aatcaggctg
ctggatacat ggctaagctg 2040atttgggaat ctgtgagcgt gacggtggta
gctgcggttg aagcaatgaa ctggcttaag 2100tctgctgcta agctgctggc
tgctgaggtc aaagataaga agactggaga gattcttcgc 2160aagcgttgcg
ctgtgcattg ggtaactcct gatggtttcc ctgtgtggca ggaatacaag
2220aagcctattc agacgcgctt gaacctgatg ttcctcggtc agttccgctt
acagcctacc 2280attaacacca acaaagatag cgagattgat gcacacaaac
aggagtctgg tatcgctcct 2340aactttgtac acagccaaga cggtagccac
cttcgtaaga ctgtagtgtg ggcacacgag 2400aagtacggaa tcgaatcttt
tgcactgatt cacgactcct tcggtaccat tccggctgac 2460gctgcgaacc
tgttcaaagc agtgcgcgaa actatggttg acacatatga gtcttgtgat
2520gtactggctg atttctacga ccagttcgct gaccagttgc acgagtctca
attggacaaa 2580atgccagcac ttccggctaa aggtaacttg aacctccgtg
acatcttaga gtcggacttc 2640gcgttcgcgt aataa 26554883PRTArtificial
SequenceDescription of Artificial Sequence Synthetic variant
polypeptide 4Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp
Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His
Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu Ala Leu Glu His
Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe Arg Lys Met Phe
Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala Asp Asn Ala Ala
Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70 75 80 Met Ile Ala
Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95 Gly
Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105
110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser
115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly
Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp
Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln
Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met
Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly
Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His
Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly
Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230
235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala
Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln
Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly
Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val
Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val
Tyr Met Pro Glu Val Tyr Lys Ser Ile 305 310 315 320 Asn Ile Ala Gln
Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala
Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350
Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355
360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala
Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser
Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn
His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly
Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp
Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile
Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn
Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475
480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro
485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys
Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His
Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly
Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp
Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser
Glu
Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn
Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu
Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600
605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly
610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala
Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu
Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu
Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys
Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val
Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala
Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720
Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725
730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe
Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys
Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala
Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys
Thr Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser
Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp
Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp
Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845
Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850
855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp
Phe 865 870 875 880 Ala Phe Ala 52655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 5atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagttatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgctgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
26556883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 6Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Val Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp
Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
72655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 7atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaaccgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 26558883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 8Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220
Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225
230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile
Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe
Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr
Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu
Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp
Val Tyr Met Pro Glu Val Tyr Lys Pro Ile 305 310 315 320 Asn Ile Ala
Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val
Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345
350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp
355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala
Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile
Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala
Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg
Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn
Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro
Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala
Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470
475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser
Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe
Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His
His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp
Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg
Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser
Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val
Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590
Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595
600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr
Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu
Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val
Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly
Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala
Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala
Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu
Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715
720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp
725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met
Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn
Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile
Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg
Lys Thr Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu
Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala
Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val
Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840
845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu
850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser
Asp Phe 865 870 875 880 Ala Phe Ala 92655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 9atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgctgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265510883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 10Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp
Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
112655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 11atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtatcta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat
ggctaagctg 2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg
aagcaatgaa ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc
aaagataaga agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg
ggtaactcct gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc
agacgcgctt gaacctgatg ttcctcggtc agttccgctt acagcctacc
2280attaacacca acaaagatag cgagattgat gcacacaaac aggagtctgg
tatcgctcct 2340aactttgtac acagccaaga cggtagccac cttcgtaaga
ctgtagtgtg ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt
cacgactcct tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc
agtgcgcgaa actatggttg acacatatga gtcttgtgat 2520gtactggctg
atttctacga ccagttcgct gaccagttgc acgagtctca attggacaaa
2580atgccagcac ttccggctaa aggtaacttg aacctccgtg acatcttaga
gtcggacttc 2640gcgttcgcgt aataa 265512883PRTArtificial
SequenceDescription of Artificial Sequence Synthetic variant
polypeptide 12Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp
Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His
Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu Ala Leu Glu His
Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe Arg Lys Met Phe
Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala Asp Asn Ala Ala
Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70 75 80 Met Ile Ala
Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95 Gly
Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105
110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser
115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly
Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp
Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln
Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met
Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly
Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His
Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly
Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230
235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala
Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln
Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly
Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val
Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val
Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315 320 Asn Ile Ala Gln
Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala
Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350
Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355
360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala
Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser
Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn
His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly
Arg Ile Tyr Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp
Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile
Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn
Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475
480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro
485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys
Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His
Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly
Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp
Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu
Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn
Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu
Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600
605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly
610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala
Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu
Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu
Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys
Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val
Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala
Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720
Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725
730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe
Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys
Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala
Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys
Thr Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser
Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp
Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp
Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845
Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850
855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp
Phe 865 870 875 880 Ala Phe Ala 132655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 13atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtttcta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgctgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265514883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 14Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Phe Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
152655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 15atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgtgttg tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265516883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 16Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Val Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
172655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 17atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgtctgg tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265518883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 18Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525
Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530
535 540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val
Asn 545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly
Ile Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala
Ile Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu
Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys
Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser
Val Thr Lys Arg Leu Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser
Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650
655 Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln
660 665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser
Val Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys
Ser Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys
Thr Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp
Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys
Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe
Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile
Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775
780 Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu
785 790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser
Phe Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala
Val Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val
Leu Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu
Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn
Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe
Ala 192655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 19atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgtatgg tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265520883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 20Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Met Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
212655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 21atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaactg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265522883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 22Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180
185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu
Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile
Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly
Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro
Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala
Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys
Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly
Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300
Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305
310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val
Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro
Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met
Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala
Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala
Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu
Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro
Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425
430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys
435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile
His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro
Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile
Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala
Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu
Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser
Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His
Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550
555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala
Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly
Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly
Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala
Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys
Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe
Gly Phe Arg Gln Gln Leu Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala
Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670
Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675
680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala
Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu
Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro
Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln
Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln
Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His
Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln
Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785 790 795
800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr
805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu
Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp
Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu
Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu
Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
232655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 23atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagatc tgattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265524883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 24Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Leu Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
252655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 25atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgttgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265526883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 26Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Val Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
272655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 27atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agatcctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265528883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 28Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Ala Ala Lys 690 695 700 Ile Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825
830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln
835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro
Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu
Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala 292655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 29atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgctgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctatcgtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265530883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 30Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp
Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
312655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 31atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctgtagtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcattcatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265532883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 32Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485
490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe
Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly
Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser
Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp Glu
Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu Thr
Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn Glu
Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu Val
Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605
Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610
615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala Tyr
Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu Glu
Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu Met
Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys Leu
Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val Glu
Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala Ala
Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720 Lys
Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730
735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu
740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp
Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro
Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys Thr
Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser Phe
Ala Phe Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp Ala
Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp Thr
Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845 Phe
Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855
860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe
865 870 875 880 Ala Phe Ala 332655DNAArtificial SequenceDescription
of Artificial Sequence Synthetic coding polynucleotide 33atgaacacga
ttaacatcgc taagaacgac ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc
tggctgacca ttacggtgag cgtttagctc gcgaacagtt ggcccttgag
120catgagtctt acgagatggg tgaagcacgc ttccgcaaga tgtttgagcg
tcaacttaaa 180gctggtgagg ttgcggataa cgctgccgcc aagcctctca
tcactaccct actccctaag 240atgattgcac gcatcaacga ctggtttgag
gaagtgaaag ctaagcgcgg caagcgcccg 300acagccttcc agttcctgca
agaaatcaag ccggaagccg tagcgtacat caccattaag 360accactctgg
cttgcctaac cagtgctgac aatacaaccg ttcaggctgt agcaagcgca
420atcggtcggg ccattgagga cgaggctcgc ttcggtcgta tccgtgacct
tgaagctaag 480cacttcaaga aaaacgttga ggaacaactc aacaagcgcg
tagggcacgt ctacaagaaa 540gcatttatgc aagttgtcga ggctgacatg
ctctctaagg gtctactcgg tggcgaggcg 600tggtcttcgt ggcataagga
agactctatt catgtaggag tacgctgcat cgagatgctc 660attgagtcaa
ccggaatggt tagcttacac cgccaaaatg ctggcgtagt aggtcaagac
720tctgagacta tcgaactcgc acctgaatac gctgaggcta tcgcaacccg
tgcaggtgcg 780ctggctggca tctctccgat gttccaacct tgcgtagttc
ctcctaagcc gtggactggc 840attactggtg gtggctattg ggctaacggt
cgtcgtcctc tggcgctggt gcgtactcac 900agtaagaaag cactgatgcg
ctacgaagac gtttacatgc ctgaggtgta caaagcgatt 960aacattgcgc
aaaacaccgc atggaaaatc aacaagaaag tcctagcggt cgccaacgta
1020atcaccaagt ggaagcattg tccggtcgag gacatccctg cgattgagcg
tgaagaactc 1080ccgatgaaac cggaagacat cgacatgaat cctgaggctc
tcaccgcgtg gaaacgtgct 1140gccgctgctg tgtaccgcaa ggacaaggct
cgcaagtctc gccgtatcag ccttgagttc 1200atgcttgagc aagccaataa
gtttgctaac cataaggcca tctggttccc ttacaacatg 1260gactggcgcg
gtcgtgttta cgctgtgtca atgttcaacc cgcaaggtaa cgatatgacc
1320aaaggactgc ttacgctggc gaaaggtaaa ccaatcggta aggaaggtta
ctactggctg 1380aaaatccacg gtgcaaactg tgcgggtgtc gataaggttc
cgttccctga gcgcatcaag 1440ttcattgagg aaaaccacga gaacatcatg
gcttgcgcta agtctccact ggagaacact 1500tggtgggctg agcaagattc
tccgttctgc ttccttgcgt tctgctttga gtacgctggg 1560gtacagcacc
acggcctgag ctataactgc tcccttccgc tggcgtttga cgggtcttgc
1620tctggcatcc agcacttctc cgcgatgctc cgagatgagg taggtggtcg
cgcggttaac 1680ttgcttccta gtgaaaccgt tcaggacatc tacgggattg
ttgctaagaa agtcaacgag 1740attctacaag cagacgcaat caatgggacc
gataacgaag tagttaccgt gaccgatgag 1800aacactggtg aaatctctga
gaaagtcaag ctgggcacta aggcactggc tggtcaatgg 1860ctggcttacg
gtgttactcg cagtgtgact aagcgttcag tcatgacgct ggcttacggg
1920tccaaagagt tcggcttccg tcaacaagtg ctggaagata ccattcagcc
agctattgat 1980tccggcaagg gtctgatgtt cactcagccg aatcaggctg
ctggatacat ggctaagctg 2040atttgggaat ctgtgagcgt gacggtggta
gctgcggttg aagcaatgaa ctggcttaag 2100tctgctgcta agctgctggc
tgctgaggtc aaagataaga agactggaga gattcttcgc 2160aagcgttgcg
ctgtgcattg ggtaactcct gatggtttcc ctgtgtggca ggaatacaag
2220aagcctattc agacgcgctt gaacctgatg ttcctcggtc agttccgctt
acagcctacc 2280attaacacca acaaagatag cgagattgat gcacacaaac
aggagtctgg tatcgctcct 2340aactttgtac acagccaaga cggtagccac
cttcgtaaga ctgtagtgtg ggcacacgag 2400aagtacggaa tcgaatcttt
tgcactgatt cacgactcct ggggtaccat tccggctgac 2460gctgcgaacc
tgttcaaagc agtgcgcgaa actatggttg acacatatga gtcttgtgat
2520gtactggctg atttctacga ccagttcgct gaccagttgc acgagtctca
attggacaaa 2580atgccagcac ttccggctaa aggtaacttg aacctccgtg
acatcttaga gtcggacttc 2640gcgttcgcgt aataa 265534883PRTArtificial
SequenceDescription of Artificial Sequence Synthetic variant
polypeptide 34Met Asn Thr Ile Asn Ile Ala Lys Asn Asp Phe Ser Asp
Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr Leu Ala Asp His
Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu Ala Leu Glu His
Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe Arg Lys Met Phe
Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala Asp Asn Ala Ala
Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70 75 80 Met Ile Ala
Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys Arg 85 90 95 Gly
Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile Lys Pro Glu 100 105
110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser
115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly
Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp
Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln
Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met
Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly
Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His
Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly
Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230
235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala
Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln
Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly
Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val
Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val
Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315 320 Asn Ile Ala Gln
Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala
Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350
Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355
360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala
Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser
Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn
His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly
Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp
Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile
Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn
Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475
480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro
485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys
Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His
Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly
Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp
Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu
Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn
Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu
Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600
605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly
610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala
Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu
Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu
Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys
Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val
Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala
Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720
Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725
730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe
Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys
Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala
Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys
Thr Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser
Phe Ala Leu Ile His Asp Ser Trp Gly Thr 805 810 815 Ile Pro Ala Asp
Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp
Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845
Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850
855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp
Phe 865 870 875 880 Ala Phe Ala 352655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 35atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgctgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580tggccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265536883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 36Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp
Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln
Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met
Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly
Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His
Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly
Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230
235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala
Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln
Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly
Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val
Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val
Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315 320 Asn Ile Ala Gln
Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala
Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350
Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355
360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala
Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser
Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn
His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly
Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp
Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile
Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn
Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475
480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro
485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys
Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His
Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly
Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp
Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu
Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn
Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu
Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600
605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly
610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser Val Met Thr Leu Ala
Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Val Leu
Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu
Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys
Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val
Glu Ala Met Asn Trp Leu Lys Ser Ala Ala Lys 690 695 700 Leu Leu Ala
Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720
Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725
730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe
Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys
Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala
Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys
Thr Val Val Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser
Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp
Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp
Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845
Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys Trp Pro Ala Leu 850
855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp
Phe 865 870 875 880 Ala Phe Ala 372655DNAArtificial
SequenceDescription of Artificial Sequence Synthetic coding
polynucleotide 37atgaacacga ttaacatcgc taagaacgac ttctctgaca
tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag cgtttagctc
gcgaacagtt ggcccttgag 120catgagtctt acgagatggg tgaagcacgc
ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg ttgcggataa
cgctgccgcc aagcctctca tcactaccct actccctaag 240atgattgcac
gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg caagcgcccg
300acagccttcc agttcctgca agaaatcaag ccggaagccg tagcgtacat
caccattaag 360accactctgg cttgcctaac cagtgctgac aatacaaccg
ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga cgaggctcgc
ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga aaaacgttga
ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa 540gcatttatgc
aagttgtcga ggctgacatg ctctctaagg gtctactcgg tggcgaggcg
600tggtcttcgt ggcataagga agactctatt catgtaggag tacgctgcat
cgagatgctc 660attgagtcaa ccggaatggt tagcttacac cgccaaaatg
ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc acctgaatac
gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca tctctccgat
gttccaacct tgcgtagttc ctcctaagcc gtggactggc 840attactggtg
gtggctattg ggctaacggt cgtcgtcctc tggcgctggt gcgtactcac
900agtaagaaag cactgatgcg ctacgaagac gtttacatgc ctgaggtgta
caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc aacaagaaag
tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg tccggtcgag
gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac cggaagacat
cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct 1140gccgctgctg
tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag ccttgagttc
1200atgcttgagc aagccaataa gtttgctaac cataaggcca tctggttccc
ttacaacatg 1260gactggcgcg gtcgtgttta cgctgtgtca atgttcaacc
cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc gaaaggtaaa
ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg gtgcaaactg
tgcgggtgtc gataaggttc cgttccctga gcgcatcaag 1440ttcattgagg
aaaaccacga gaacatcatg gcttgcgcta agtctccact ggagaacact
1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgttgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctatcgtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265538883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 38Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Val Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Val Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp
Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
392655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 39atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga
gtacgctggg 1560gtacagcacc acggcctgag ctataactgc tcccttccgc
tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc cgcgatgctc
cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta gtgaaaccgt
tcaggacatc tacgggattg ttgctaagaa agtcaacgag 1740attctacaag
cagacgcaat caatgggacc gataacgaag tagttaccgt gaccgatgag
1800aacactggtg aaatctctga gaaagtcaag ctgggcacta aggcactggc
tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact aagcgttcag
tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg tcaacaagtg
ctggaagata ccattcagcc agctattgat 1980tccggcaagg gtctgatgtt
cactcagccg aatcaggctg ctggatacat ggctaagctg 2040atttgggaat
ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa ctggcttaag
2100tctgttgcta agctgctggc tgctgaggtc aaagataaga agactggaga
gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct gatggtttcc
ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt gaacctgatg
ttcctcggtc agttccgctt acagcctacc 2280attaacacca acaaagatag
cgagattgat gcacacaaac aggagtctgg tatcgctcct 2340aactttgtac
acagccaaga cggtagccac cttcgtaaga ctgtagtgtg ggcacacgag
2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct tcggtaccat
tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa actatggttg
acacatatga gtcttgtgat 2520gtactggctg atttctacga ccagttcgct
gaccagttgc acgagtctca attggacaaa 2580atgccagcac ttccggctaa
aggtaacttg aacctccgtg acatcttaga gtcggacttc 2640gcgttcgcgt aataa
265540883PRTArtificial SequenceDescription of Artificial Sequence
Synthetic variant polypeptide 40Met Asn Thr Ile Asn Ile Ala Lys Asn
Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala Ile Pro Phe Asn Thr
Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30 Ala Arg Glu Gln Leu
Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35 40 45 Ala Arg Phe
Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu Val 50 55 60 Ala
Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu Leu Pro Lys 65 70
75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu Val Lys Ala Lys
Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe Leu Gln Glu Ile
Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile Lys Thr Thr Leu
Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr Val Gln Ala Val
Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp Glu Ala Arg Phe
Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160 His Phe Lys Lys
Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165 170 175 Val Tyr
Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu Ser 180 185 190
Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His Lys Glu Asp 195
200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met Leu Ile Glu Ser
Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn Ala Gly Val Val
Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu Ala Pro Glu Tyr
Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala Leu Ala Gly Ile
Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro Pro Lys Pro Trp
Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285 Asn Gly Arg Arg
Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290 295 300 Leu Met
Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala Ile 305 310 315
320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys Lys Val Leu Ala
325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His Cys Pro Val Glu
Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu Pro Met Lys Pro
Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu Thr Ala Trp Lys
Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp Lys Ala Arg Lys
Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met Leu Glu Gln Ala
Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410 415 Pro Tyr Asn
Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe 420 425 430 Asn
Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu Ala Lys 435 440
445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu Lys Ile His Gly
450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro Phe Pro Glu Arg
Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu Asn Ile Met Ala
Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp Trp Ala Glu Gln
Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys Phe Glu Tyr Ala
Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn Cys Ser Leu Pro
Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535 540 His Phe Ser
Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn 545 550 555 560
Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile Val Ala Lys 565
570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile Asn Gly Thr Asp
Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn Thr Gly Glu Ile
Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala Leu Ala Gly Gln
Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val Thr Lys Arg Ser
Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys Glu Phe Gly Phe
Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655 Pro Ala Ile Asp
Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660 665 670 Ala Ala
Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val Thr 675 680 685
Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser Val Ala Lys 690
695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr Gly Glu Ile Leu
Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val Thr Pro Asp Gly
Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro Ile Gln Thr Arg
Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg Leu Gln Pro Thr
Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp Ala His Lys Gln
Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780 Ser Gln Asp Gly
Ser His Leu Arg Lys Thr Val Val Trp Ala His Glu 785 790 795 800 Lys
Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe Gly Thr 805 810
815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val Arg Glu Thr Met
820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu Ala Asp Phe Tyr
Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser Gln Leu Asp Lys
Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu Asn Leu Arg Asp
Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
412655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 41atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgctgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctatcgtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265542883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 42Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Ala Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu
785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
432655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 43atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgttgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctatcgtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265544883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 44Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Val Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
452655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 45atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgtatgg tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaagtg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgttgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctatcgtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265546883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 46Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp
Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Met Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Val Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Val Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
472655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 47atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgttcag tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaactg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgttgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctatcgtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265548883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 48Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val Ala Tyr Ile Thr Ile
Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125 Ala Asp Asn Thr Thr
Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130 135 140 Ile Glu Asp
Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala Lys 145 150 155 160
His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys Arg Val Gly His 165
170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val Glu Ala Asp Met Leu
Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala Trp Ser Ser Trp His
Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val Arg Cys Ile Glu Met
Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser Leu His Arg Gln Asn
Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser Glu Thr Ile Glu Leu
Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250 255 Arg Ala Gly Ala
Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val 260 265 270 Val Pro
Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr Trp Ala 275 280 285
Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His Ser Lys Lys Ala 290
295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro Glu Val Tyr Lys Ala
Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala Trp Lys Ile Asn Lys
Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile Thr Lys Trp Lys His
Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile Glu Arg Glu Glu Leu
Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met Asn Pro Glu Ala Leu
Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375 380 Tyr Arg Lys Asp
Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe 385 390 395 400 Met
Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala Ile Trp Phe 405 410
415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Leu Tyr Ala Val Ser Met Phe
420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys Gly Leu Leu Thr Leu
Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu Gly Tyr Tyr Trp Leu
Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly Val Asp Lys Val Pro
Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile Glu Glu Asn His Glu
Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495 Leu Glu Asn Thr Trp
Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500 505 510 Ala Phe Cys
Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser Tyr 515 520 525 Asn
Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser Gly Ile Gln 530 535
540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly Gly Arg Ala Val Asn
545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln Asp Ile Tyr Gly Ile
Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu Gln Ala Asp Ala Ile
Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr Val Thr Asp Glu Asn
Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys Leu Gly Thr Lys Ala
Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620 Val Thr Arg Ser Val
Thr Lys Arg Ser Val Met Thr Leu Ala Tyr Gly 625 630 635 640 Ser Lys
Glu Phe Gly Phe Arg Gln Gln Leu Leu Glu Asp Thr Ile Gln 645 650 655
Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr Gln Pro Asn Gln 660
665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp Glu Ser Val Ser Val
Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met Asn Trp Leu Lys Ser
Val Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val Lys Asp Lys Lys Thr
Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys Ala Val His Trp Val
Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln Glu Tyr Lys Lys Pro
Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745 750 Gly Gln Phe Arg
Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu 755 760 765 Ile Asp
Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe Val His 770 775 780
Ser Gln Asp Gly Ser His Leu Arg Lys Thr Ile Val Trp Ala His Glu 785
790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu Ile His Asp Ser Phe
Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn Leu Phe Lys Ala Val
Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu Ser Cys Asp Val Leu
Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp Gln Leu His Glu Ser
Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro Ala Lys Gly Asn Leu
Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870 875 880 Ala Phe Ala
492655DNAArtificial SequenceDescription of Artificial Sequence
Synthetic coding polynucleotide 49atgaacacga ttaacatcgc taagaacgac
ttctctgaca tcgaactggc tgctatcccg 60ttcaacactc tggctgacca ttacggtgag
cgtttagctc gcgaacagtt ggcccttgag 120catgagtctt acgagatggg
tgaagcacgc ttccgcaaga tgtttgagcg tcaacttaaa 180gctggtgagg
ttgcggataa cgctgccgcc aagcctctca tcactaccct actccctaag
240atgattgcac gcatcaacga ctggtttgag gaagtgaaag ctaagcgcgg
caagcgcccg 300acagccttcc agttcctgca agaaatcaag ccggaagccg
tagcgtacat caccattaag 360accactctgg cttgcctaac cagtgctgac
aatacaaccg ttcaggctgt agcaagcgca 420atcggtcggg ccattgagga
cgaggctcgc ttcggtcgta tccgtgacct tgaagctaag 480cacttcaaga
aaaacgttga ggaacaactc aacaagcgcg tagggcacgt ctacaagaaa
540gcatttatgc aagttgtcga ggctgacatg ctctctaagg gtctactcgg
tggcgaggcg 600tggtcttcgt ggcataagga agactctatt catgtaggag
tacgctgcat cgagatgctc 660attgagtcaa ccggaatggt tagcttacac
cgccaaaatg ctggcgtagt aggtcaagac 720tctgagacta tcgaactcgc
acctgaatac gctgaggcta tcgcaacccg tgcaggtgcg 780ctggctggca
tctctccgat gttccaacct tgcgtagttc ctcctaagcc gtggactggc
840attactggtg gtggctattg ggctaacggt cgtcgtcctc tggcgctggt
gcgtactcac 900agtaagaaag cactgatgcg ctacgaagac gtttacatgc
ctgaggtgta caaagcgatt 960aacattgcgc aaaacaccgc atggaaaatc
aacaagaaag tcctagcggt cgccaacgta 1020atcaccaagt ggaagcattg
tccggtcgag gacatccctg cgattgagcg tgaagaactc 1080ccgatgaaac
cggaagacat cgacatgaat cctgaggctc tcaccgcgtg gaaacgtgct
1140gccgctgctg tgtaccgcaa ggacaaggct cgcaagtctc gccgtatcag
ccttgagttc 1200atgcttgagc aagccaataa gtttgctaac cataaggcca
tctggttccc ttacaacatg 1260gactggcgcg gtcgtctgta cgctgtgtca
atgttcaacc cgcaaggtaa cgatatgacc 1320aaaggactgc ttacgctggc
gaaaggtaaa ccaatcggta aggaaggtta ctactggctg 1380aaaatccacg
gtgcaaactg tgcgggtgtc gataaggttc cgttccctga gcgcatcaag
1440ttcattgagg aaaaccacga gaacatcatg gcttgcgcta agtctccact
ggagaacact 1500tggtgggctg agcaagattc tccgttctgc ttccttgcgt
tctgctttga gtacgctggg 1560gtacagcacc acggcctgag ctataactgc
tcccttccgc tggcgtttga cgggtcttgc 1620tctggcatcc agcacttctc
cgcgatgctc cgagatgagg taggtggtcg cgcggttaac 1680ttgcttccta
gtgaaaccgt tcaggacatc tacgggattg ttgctaagaa agtcaacgag
1740attctacaag cagacgcaat caatgggacc gataacgaag tagttaccgt
gaccgatgag 1800aacactggtg aaatctctga gaaagtcaag ctgggcacta
aggcactggc tggtcaatgg 1860ctggcttacg gtgttactcg cagtgtgact
aagcgtatgg tcatgacgct ggcttacggg 1920tccaaagagt tcggcttccg
tcaacaactg ctggaagata ccattcagcc agctattgat 1980tccggcaagg
gtctgatgtt cactcagccg aatcaggctg ctggatacat ggctaagctg
2040atttgggaat ctgtgagcgt gacggtggta gctgcggttg aagcaatgaa
ctggcttaag 2100tctgttgcta agctgctggc tgctgaggtc aaagataaga
agactggaga gattcttcgc 2160aagcgttgcg ctgtgcattg ggtaactcct
gatggtttcc ctgtgtggca ggaatacaag 2220aagcctattc agacgcgctt
gaacctgatg ttcctcggtc agttccgctt acagcctacc 2280attaacacca
acaaagatag cgagattgat gcacacaaac aggagtctgg tatcgctcct
2340aactttgtac acagccaaga cggtagccac cttcgtaaga ctatcgtgtg
ggcacacgag 2400aagtacggaa tcgaatcttt tgcactgatt cacgactcct
tcggtaccat tccggctgac 2460gctgcgaacc tgttcaaagc agtgcgcgaa
actatggttg acacatatga gtcttgtgat 2520gtactggctg atttctacga
ccagttcgct gaccagttgc acgagtctca attggacaaa 2580atgccagcac
ttccggctaa aggtaacttg aacctccgtg acatcttaga gtcggacttc
2640gcgttcgcgt aataa 265550883PRTArtificial SequenceDescription of
Artificial Sequence Synthetic variant polypeptide 50Met Asn Thr Ile
Asn Ile Ala Lys Asn Asp Phe Ser Asp Ile Glu Leu 1 5 10 15 Ala Ala
Ile Pro Phe Asn Thr Leu Ala Asp His Tyr Gly Glu Arg Leu 20 25 30
Ala Arg Glu Gln Leu Ala Leu Glu His Glu Ser Tyr Glu Met Gly Glu 35
40 45 Ala Arg Phe Arg Lys Met Phe Glu Arg Gln Leu Lys Ala Gly Glu
Val 50 55 60 Ala Asp Asn Ala Ala Ala Lys Pro Leu Ile Thr Thr Leu
Leu Pro Lys 65 70 75 80 Met Ile Ala Arg Ile Asn Asp Trp Phe Glu Glu
Val Lys Ala Lys Arg 85 90 95 Gly Lys Arg Pro Thr Ala Phe Gln Phe
Leu Gln Glu Ile Lys Pro Glu 100 105 110 Ala Val
Ala Tyr Ile Thr Ile Lys Thr Thr Leu Ala Cys Leu Thr Ser 115 120 125
Ala Asp Asn Thr Thr Val Gln Ala Val Ala Ser Ala Ile Gly Arg Ala 130
135 140 Ile Glu Asp Glu Ala Arg Phe Gly Arg Ile Arg Asp Leu Glu Ala
Lys 145 150 155 160 His Phe Lys Lys Asn Val Glu Glu Gln Leu Asn Lys
Arg Val Gly His 165 170 175 Val Tyr Lys Lys Ala Phe Met Gln Val Val
Glu Ala Asp Met Leu Ser 180 185 190 Lys Gly Leu Leu Gly Gly Glu Ala
Trp Ser Ser Trp His Lys Glu Asp 195 200 205 Ser Ile His Val Gly Val
Arg Cys Ile Glu Met Leu Ile Glu Ser Thr 210 215 220 Gly Met Val Ser
Leu His Arg Gln Asn Ala Gly Val Val Gly Gln Asp 225 230 235 240 Ser
Glu Thr Ile Glu Leu Ala Pro Glu Tyr Ala Glu Ala Ile Ala Thr 245 250
255 Arg Ala Gly Ala Leu Ala Gly Ile Ser Pro Met Phe Gln Pro Cys Val
260 265 270 Val Pro Pro Lys Pro Trp Thr Gly Ile Thr Gly Gly Gly Tyr
Trp Ala 275 280 285 Asn Gly Arg Arg Pro Leu Ala Leu Val Arg Thr His
Ser Lys Lys Ala 290 295 300 Leu Met Arg Tyr Glu Asp Val Tyr Met Pro
Glu Val Tyr Lys Ala Ile 305 310 315 320 Asn Ile Ala Gln Asn Thr Ala
Trp Lys Ile Asn Lys Lys Val Leu Ala 325 330 335 Val Ala Asn Val Ile
Thr Lys Trp Lys His Cys Pro Val Glu Asp Ile 340 345 350 Pro Ala Ile
Glu Arg Glu Glu Leu Pro Met Lys Pro Glu Asp Ile Asp 355 360 365 Met
Asn Pro Glu Ala Leu Thr Ala Trp Lys Arg Ala Ala Ala Ala Val 370 375
380 Tyr Arg Lys Asp Lys Ala Arg Lys Ser Arg Arg Ile Ser Leu Glu Phe
385 390 395 400 Met Leu Glu Gln Ala Asn Lys Phe Ala Asn His Lys Ala
Ile Trp Phe 405 410 415 Pro Tyr Asn Met Asp Trp Arg Gly Arg Leu Tyr
Ala Val Ser Met Phe 420 425 430 Asn Pro Gln Gly Asn Asp Met Thr Lys
Gly Leu Leu Thr Leu Ala Lys 435 440 445 Gly Lys Pro Ile Gly Lys Glu
Gly Tyr Tyr Trp Leu Lys Ile His Gly 450 455 460 Ala Asn Cys Ala Gly
Val Asp Lys Val Pro Phe Pro Glu Arg Ile Lys 465 470 475 480 Phe Ile
Glu Glu Asn His Glu Asn Ile Met Ala Cys Ala Lys Ser Pro 485 490 495
Leu Glu Asn Thr Trp Trp Ala Glu Gln Asp Ser Pro Phe Cys Phe Leu 500
505 510 Ala Phe Cys Phe Glu Tyr Ala Gly Val Gln His His Gly Leu Ser
Tyr 515 520 525 Asn Cys Ser Leu Pro Leu Ala Phe Asp Gly Ser Cys Ser
Gly Ile Gln 530 535 540 His Phe Ser Ala Met Leu Arg Asp Glu Val Gly
Gly Arg Ala Val Asn 545 550 555 560 Leu Leu Pro Ser Glu Thr Val Gln
Asp Ile Tyr Gly Ile Val Ala Lys 565 570 575 Lys Val Asn Glu Ile Leu
Gln Ala Asp Ala Ile Asn Gly Thr Asp Asn 580 585 590 Glu Val Val Thr
Val Thr Asp Glu Asn Thr Gly Glu Ile Ser Glu Lys 595 600 605 Val Lys
Leu Gly Thr Lys Ala Leu Ala Gly Gln Trp Leu Ala Tyr Gly 610 615 620
Val Thr Arg Ser Val Thr Lys Arg Met Val Met Thr Leu Ala Tyr Gly 625
630 635 640 Ser Lys Glu Phe Gly Phe Arg Gln Gln Leu Leu Glu Asp Thr
Ile Gln 645 650 655 Pro Ala Ile Asp Ser Gly Lys Gly Leu Met Phe Thr
Gln Pro Asn Gln 660 665 670 Ala Ala Gly Tyr Met Ala Lys Leu Ile Trp
Glu Ser Val Ser Val Thr 675 680 685 Val Val Ala Ala Val Glu Ala Met
Asn Trp Leu Lys Ser Val Ala Lys 690 695 700 Leu Leu Ala Ala Glu Val
Lys Asp Lys Lys Thr Gly Glu Ile Leu Arg 705 710 715 720 Lys Arg Cys
Ala Val His Trp Val Thr Pro Asp Gly Phe Pro Val Trp 725 730 735 Gln
Glu Tyr Lys Lys Pro Ile Gln Thr Arg Leu Asn Leu Met Phe Leu 740 745
750 Gly Gln Phe Arg Leu Gln Pro Thr Ile Asn Thr Asn Lys Asp Ser Glu
755 760 765 Ile Asp Ala His Lys Gln Glu Ser Gly Ile Ala Pro Asn Phe
Val His 770 775 780 Ser Gln Asp Gly Ser His Leu Arg Lys Thr Ile Val
Trp Ala His Glu 785 790 795 800 Lys Tyr Gly Ile Glu Ser Phe Ala Leu
Ile His Asp Ser Phe Gly Thr 805 810 815 Ile Pro Ala Asp Ala Ala Asn
Leu Phe Lys Ala Val Arg Glu Thr Met 820 825 830 Val Asp Thr Tyr Glu
Ser Cys Asp Val Leu Ala Asp Phe Tyr Asp Gln 835 840 845 Phe Ala Asp
Gln Leu His Glu Ser Gln Leu Asp Lys Met Pro Ala Leu 850 855 860 Pro
Ala Lys Gly Asn Leu Asn Leu Arg Asp Ile Leu Glu Ser Asp Phe 865 870
875 880 Ala Phe Ala 5133DNAArtificial SequenceDescription of
Artificial Sequence Synthetic oligonucleotide 51atgcatcacc
atcatcacca cggatccatg aac 335211PRTArtificial SequenceDescription
of Artificial Sequence Synthetic peptide 52Met His His His His His
His Gly Ser Met Asn 1 5 10 537PRTArtificial SequenceDescription of
Artificial Sequence Synthetic His tag 53His His His His His His His
1 5 546PRTArtificial SequenceDescription of Artificial Sequence
Synthetic 6xHis tag 54His His His His His His 1 5
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