Method for zymotic production of fine chemicals containing sulphur (metA)

Abstract

The invention relates to methods for the fermentative production of sulfur-containing fine chemicals, in particular L-methionine, by using bacteria which express a nucleotide sequence coding for a methionine synthase (meta) gene.

Description

RELATED APPLICATIONS

[0001] The application is a divisional of application Ser. No. 10/525,674 filed Feb. 24, 2005, which is a national stage application (under 35 U.S.C. 371) of PCT/EP2003/009452 filed Aug. 26, 2003, which claims benefit of German application 102 39 073.8 filed Aug. 26, 2002. The entire content of each above-mentioned application is hereby incorporated by reference in entirety.

SUBMISSION ON COMPACT DISC

[0002] The contents of the following submission on compact discs are incorporated herein by reference in its entirety: two copies of the Sequence Listing (COPY 1 and COPY 2) and a computer readable form copy of the Sequence Listing (CRF COPY), all on compact disc, each containing: file name: Sequence Listing-13111-00055-DIV, date recorded: May 4, 2007, size: 237 KB.

BACKGROUND OF THE INVENTION

[0003] 1. Field of the Invention

[0004] The invention relates to a method for the fermentative production of sulfur-containing fine chemicals, in particular L-methionine, by using bacteria which express a nucleotide sequence coding for a homoserine O-acetyltransferase (meta) gene.

[0005] 2. Background Art

[0006] Sulfur-containing fine chemicals such as, for example, methionine, homocysteine, S-adenosylmethionine, glutathione, cysteine, biotin, thiamine, lipoic acid are produced in cells via natural metabolic processes and are used in many branches of industry, including the food, animal feed, cosmetics and pharmaceutical industries. These substances which are collectively referred to "sulfur-containing fine chemicals" include organic acids, both proteinogenic and non-proteinogenic amino acids, vitamins and cofactors. They are most expediently produced on a large scale by means of cultivating bacteria which have been developed in order to produce and secrete large amounts of the substance desired in each case. Organisms which are particularly suitable for this purpose are coryneform bacteria, Gram-positive nonpathogenic bacteria.

[0007] It is known that amino acids are produced by fermentation of strains of coryneform bacteria, in particular Corynebacterium glutamicum. Due to the great importance, the production processes are constantly improved. Process improvements can relate to measures regarding technical aspects of the fermentation, such as, for example, stirring and oxygen supply, or to the nutrient media composition such as, for example, sugar concentration during fermentation or to the work-up to give the product, for example by ion exchange chromatography, or to the intrinsic performance properties of the microorganism itself.

[0008] A number of mutant strains which produce an assortment of desirable compounds from the group of sulfur-containing fine chemicals have been developed via strain selection. The performance properties of said microorganisms are improved with respect to the production of a particular molecule by applying methods of mutagenesis, selection and mutant selection. However, this is a time-consuming and difficult process. In this way strains are obtained, for example, which are resistant to antimetabolites such as, for example, the methionine analogs .alpha.-methylmethionine, ethionine, norleucine, n-acetylnorleucine, S-trifluoromethylhomocysteine, 2-amino-5-heprenoitic acid, selenomethionine, methioninesulfoximine, methoxine, 1-aminocyclopentanecarboxylic acid or which are auxotrophic for metabolites important for regulation and which produce sulfur-containing fine chemicals such as, for example, L-methionine.

[0009] Methods of recombinant DNA technology have also been used for some years to improve Corynebacterium strains producing L-amino acids by amplifying individual amino-acid biosynthesis genes and investigating the effect on amino acid production.

BRIEF SUMMARY OF THE INVENTION

[0010] It is an object of the present invention to provide a novel method for the improved fermentative production of sulfur-containing fine chemicals, in particular L-methionine.

[0011] We have found that this object is achieved by providing a method for the fermentative production of a sulfur-containing fine chemical, comprising the expression of a heterologous nucleotide sequence coding for a protein with meta activity in a coryneform bacterium.

[0012] The invention firstly relates to a method for the fermentative production of at least one sulfur-containing fine chemical, which comprises the following steps: [0013] a) fermentation of a coryneform bacteria culture producing the desired sulfur-containing fine chemical, the coryneform bacteria expressing at least one heterologous nucleotide sequence which codes for a protein with homoserine O-acetyltransferase (meta) activity; [0014] b) concentration of the sulfur-containing fine chemical in the medium or in the bacterial cells, and [0015] c) isolation of the sulfur-containing fine chemical, which preferably comprises L-methionine.

[0016] The above heterologous metA-encoding nucleotide sequence is preferably less than 100% and preferably more than 70%, homologous to the metA-encoding sequence from Corynebacterium glutamicum ATCC 13032. The metA-encoding sequence is derived preferably from any of the following organisms of list I: TABLE-US-00001 List I Corynebacterium diphteriae ATCC 14779 Mycobacterium leprae ATCC 43910 Mycobacterium tuberculosis CDC1551 ATCC 25584 Chlorobium tepidum ATCC 49652 Pseudomonas aeruginosa ATCC 17933 Caulobacter crescentus ATCC 19089 Neisseria gonorrhoeae ATCC 53420 Neisseria meningitidis ATCC 53414 Pseudomonas fluorescens ATCC 13525 Burkholderia cepacia ATCC 25416 Nitrosomonas europaea ATCC 19718 Haemophilus influenzae ATCC 51907 Halobacterium sp NRC1 ATCC 33170 Thermus thermophilus ATCC 27634 Deinococcus radiodurans ATCC 13939 Saccharomyces cerevisiae ATCC 10751 Schizosaccharomyces pombe ATCC 24969 Xylella fastidiosa ATCC 35881 Emericella nidulans ATCC 36104 Mesorhizobium loti ATCC 35173 Acremonium crysogenum ATCC 11550 Pseudomonas putida ATCC 47054 Staphylococcus aureus ATCC 35556 ATCC: American Type Culture Collection, Rockville, MD, USA

[0017] The metA-encoding sequence used according to the invention preferably comprises a coding sequence according to SEQ ID NO:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43 and 45 or a nucleotide sequence homologous thereto which codes for a protein with metA activity.

[0018] Moreover, the metA-encoding sequence used according to the invention preferably codes for a protein with metA activity, said protein comprising an amino acid sequence according to SEQ ID NO:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44 and 46 or an amino acid sequence homologous thereto which represents a protein with metA activity.

[0019] The coding meta sequence is preferably a DNA or an RNA which can be replicated in coryneform bacteria or is stably integrated into the chromosome.

[0020] According to a preferred embodiment, the method of the invention is carried out by [0021] a) using a bacterial strain transformed with a plasmid vector which carries at least one copy of the coding meta sequence under the control of regulatory sequences or [0022] b) using a strain in which the coding metA sequence has been integrated into the bacterial chromosome.

[0023] Furthermore, preference is given to overexpressing the coding meta sequence for the fermentation.

[0024] It may also be desirable to ferment bacteria in which additionally at least one further gene of the biosynthetic pathway of the desired sulfur-containing fine chemical has been amplified; and/or in which at least one metabolic pathway, which reduces production of the desired sulfur-containing fine chemical has, at least partially, been switched off.

[0025] It may also be desirable to ferment bacteria in which additionally the activity of at least one further gene of the biosynthetic pathway of the desired sulfur-containing fine chemical is not undesirably influenced by metabolic metabolites.

[0026] Therefore, according to another embodiment of the method of the invention, coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among [0027] a) the gene lysC, which encodes an aspartate kinase, [0028] b) the gene asd, which encodes an aspartate-semialdehyde dehydrogenase, [0029] c) the glyceraldehyde-3-phosphate dehydrogenase-encoding gene gap, [0030] d) the 3-phosphoglycerate kinase-encoding gene pgk, [0031] e) the pyruvate carboxylase-encoding gene pyc, [0032] f) the triose phosphate isomerase-encoding gene tpi, [0033] g) the methionine synthase-encoding gene metH, [0034] h) the cystathionine gamma-synthase-encoding gene metB, [0035] i) the cystathionine gamma-lyase-encoding gene metC, [0036] j) the serine hydroxymethyltransferase-encoding gene glyA, [0037] k) the O-acetylhomoserine sulfhydrylase-encoding gene metY, [0038] l) the methylene tetrahydrofolate reductase-encoding gene metF, [0039] m) the phosphoserine aminotransferase-encoding gene serC, [0040] n) the phosphoserine phosphatase-encoding gene serB, [0041] o) the serine acetyl transferase-encoding gene cysE, [0042] p) the homoserine dehydrogenase-encoding gene hom is overexpressed.

[0043] According to another embodiment of the method of the invention, coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among genes of the abovementioned group a) to p) is mutated in such a way that the activity of the corresponding proteins is influenced by metabolic metabolites to a smaller extent, if at all, compared to nonmutated proteins and that in particular the inventive production of the fine chemical is not adversely affected.

[0044] According to another embodiment of the method of the invention, coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among [0045] q) the homoserine kinase-encoding gene thrB, [0046] r) the threonine dehydratase-encoding gene ilvA, [0047] s) the threonine synthase-encoding gene thrC, [0048] t) the meso-diaminopimelate D-dehydrogenase-encoding gene ddh, [0049] u) the phosphoenolpyruvate carboxykinase-encoding gene pck, [0050] v) the glucose-6-phosphate 6-isomerase-encoding gene pgi, [0051] w) the pyruvate oxidase-encoding gene poxB, [0052] x) the dihydrodipicolinate synthase-encoding gene dapA, [0053] y) the dihydrodipicolinate reductase-encoding gene dapB; or [0054] z) the diaminopicolinate decarboxylase-encoding gene lysA is attenuated, in particular by reducing the rate of expression of the corresponding gene.

[0055] According to another embodiment of the method of the invention, coryneform bacteria are fermented in which, at the same time, at least one of the genes of the above groups q) to z) is mutated in such a way that the enzymic activity of the corresponding protein is partially or completely reduced.

[0056] Preference is given to using, in the method of the invention, microorganisms of the species Corynebacterium glutamicum.

[0057] The invention further relates to a method for producing an L-methionine-containing animal feed additive from fermentation broths, which comprises the following steps: [0058] a) culturing and fermentation of an L-methionine-producing microorganism in a fermentation medium; [0059] b) removal of water from the L-methionine-containing fermentation broth; [0060] c) removal of from 0 to 100% by weight of the biomass formed during fermentation; and [0061] d) drying of the fermentation broth obtained according to b) and/or c), in order to obtain the animal feed additive in the desired powder or granule form.

[0062] The invention likewise relates to the coding meta sequences isolated from the above microorganisms for the first time, to the homoserine 0-acetyltransferase encoded thereby and to the functional homologs of these polynucleotides and proteins, respectively.

BRIEF DESCRIPTION OF THE DRAWINGS

[0063] FIG. 1 shows the plasmid map for plasmid pClysC;

[0064] FIG. 2 shows the plasmid map for plasmid pClSlysCthr311ile;

[0065] FIG. 3 shows the plasmid map for plasmid pC_metA_Cd.

[0066] Restriction cleavage sites with their respective positions in brackets are shown in the plasmid maps. Essential sequence segments are printed in bold. KanR means kanamycin resistance gene; ask means aspartate kinase gene.

DETAILED DESCRIPTION OF THE INVENTION

a) General Terms

[0067] Proteins with homoserine O-acetyltransferase activity, also referred to as meta (EC 2.3.1.31), are described as being proteins which are capable of converting homoserine and acetyl coenzyme A into O-acetylhomoserine. The skilled worker distinguishes between the activity of homoserine O-acetyltransferase and that of homoserine O-succinyltransferase, but also referred to as meta in the literature. In the latter enzyme, succinyl coenzyme A and not acetyl coenzyme A acts as the substrate for the reaction. The skilled worker can detect the enzymatic activity of homoserine O-acetyltransferase by means of enzyme assays, protocols for which may be: Park S D. Lee J Y. Kim Y. Kim J H. Lee H S. Molecules & Cells. 8(3): 286-94, 1998.

[0068] Within the scope of the present invention, the term "sulfur-containing fine chemical" includes any chemical compound which contains at least one covalently bound sulfur atom and is accessible by a fermentation method of the invention. Nonlimiting examples thereof are methionine, homocysteine, S-adenosylmethionine, in particular methionine and S-adenosyl-methionine.

[0069] Within the scope of the present invention, the terms "L-methionine", "methionine", homocysteine and S-adenosylmethionine also include the corresponding salts such as, for example, methionine hydrochloride or methionine sulfate.

[0070] "Polynucleotides" in general refers to polyribonucleotides (RNA) and polydeoxyribonucleotides (DNA) which may be unmodified RNA and DNA respectively, or modified RNA and DNA, respectively.

[0071] According to the invention, "polypeptides" means peptides or proteins which contain two or more amino acids linked via peptide bonds.

[0072] The term "metabolic metabolite" refers to chemical compounds which occur in the metabolism of organisms as intermediates or else final products and which, apart from their property as chemical building blocks, may also have a modulating effect on enzymes and on their catalytic activity. It is known from the literature that such metabolic metabolites may act on the activity of enzymes in both an inhibiting and a stimulating manner (Biochemistry, Stryer, Lubert, 1995 W. H. Freeman & Company, New York, N.Y.). The possibility of producing in organisms enzymes in which the influence of metabolic metabolites has been modified by measures such as mutation of the genomic DNA by UV radiation, ionizing radiation or mutagenic substances and subsequent selection for particular phenotypes has also been described in the literature (Sahm H., Eggeling L., de Graaf A A., Biological Chemistry 381(9-10):899-910, 2000; Eikmanns B J., Eggeling L., Sahm H., Antonie van Leeuwenhoek., 64:145-63,1993-94). These altered properties may also be achieved by specific measurements. The skilled worker knows that it is also possible specifically to modify in enzyme genes particular nucleotides of the DNA coding for the protein in such a way that the protein resulting from the expressed DNA sequence has certain new properties, for example that the modulating effect of metabolic metabolites on the unmodified protein has changed.

[0073] The terms "express" and "amplification" or "overexpression" describe in the context of the invention the production of or increase in intracellular activity of one or more enzymes encoded by the corresponding DNA in a microorganism. For this purpose, for example, it is possible to introduce a gene into an organism, to replace an existing gene by another gene, to increase the copy number of the gene or genes, to use a strong promoter or to use a gene which codes for a corresponding enzyme having a high activity, and these measures can be combined, where appropriate.

b) MetA Proteins of the Invention

[0074] The invention likewise includes "functional equivalents" of the specifically disclosed meta enzymes of organisms in the above list I.

[0075] Within the scope of the present invention, "functional equivalents" or analogs of the specifically disclosed polypeptides are polypeptides different therefrom, which furthermore have the desired biological activity such as, for example, substrate specificity.

[0076] According to the invention, "functional equivalents" means in particular mutants which have in at least one of the abovementioned sequence positions an amino acid other than the specifically mentioned amino acid, but which have nevertheless one of the abovementioned biological activities. "Functional equivalents" thus also include the mutants obtainable by one or more amino acid additions, substitutions, deletions and/or inversions, it being possible for said modifications to occur at any position in the sequence as long as they result in a mutant having the property profile of the invention. There is functional equivalence in particular also when the reaction patterns of mutant and unmodified polypeptide match qualitatively, i.e. identical substrates are converted with different rates, for example.

[0077] "Functional equivalents" naturally also comprise polypeptides which are obtainable from other organisms, and naturally occurring variants. For example, homologous sequence regions can be found by sequence comparison, and equivalent enzymes can be established following the specific guidelines of the invention.

[0078] "Functional equivalents" likewise comprise fragments, preferably individual domains or sequence motifs, of the polypeptides of the invention, which have the desired biological function, for example.

[0079] "Functional equivalents" are also fusion proteins which have one of the abovementioned polypeptide sequences or functional equivalents derived therefrom and at least one further heterologous sequence functionally different therefrom in functional N-- or C-terminal linkage (i.e. with negligible functional impairment of the functions of the fusion protein parts). Nonlimiting examples of such heterologous sequences are, for example, signal peptides, enzymes, immunoglobulins, surface antigens, receptors or receptor ligands.

[0080] According to the invention, "functional equivalents" include homologs of the specifically disclosed proteins. These have at least 20%, or about 30%, 40%, 50%, preferably at least about 60%, 65%, 70%, or 75%, in particular at least 85%, such as, for example, 90%, 95% or 99%, homology to one of the specifically disclosed sequences, calculated by the algorithm of Pearson and Lipman, Proc. Natl. Acad., Sci. (USA) 85(8), 1988, 2444-2448.

[0081] Homologs of the proteins or polypeptides of the invention can be generated by mutagenesis, for example by point mutation or truncation of the protein. The term "homolog", as used herein, relates to a variant form of the protein, which acts as agonist or antagonist of the protein activity.

[0082] Homologs of the proteins of the invention can be identified by screening combinatorial libraries of mutants such as, for example, truncation mutants. It is possible, for example, to generate a variegated library of protein variants by combinatory mutagenesis at the nucleic acid level, for example by enzymatic ligation of a mixture of synthetic oligonucleotides. There is a multiplicity of methods which can be used for preparing libraries of potential homologs from a degenerate oligonucleotide sequence. Chemical synthesis of a degenerate gene sequence can be carried out in an automatic DNA synthesizer, and the synthetic gene can then be ligated into a suitable expression vector. The use of a degenerate set of genes makes it possible to provide whole sequences which encode the desired set of potential protein sequences in one mixture. Methods for synthesizing degenerate oligonucleotides are known to the skilled worker (for example, Narang, S. A., (1983) Tetrahedron 39:3; Itakura et al., (1984) Annu. Rev. Biochem. 53:323; Itakura et al., (1984) Science 198:1056; Ike et al., (1983) Nucleic Acids Res. 11:477).

[0083] In addition, libraries of fragments of the protein codon can be used to generate a variegated population of protein fragments for screening and for subsequent selection of homologs of a protein of the invention. In one embodiment, a library of coding sequence fragments can be generated by treating a double-stranded PCR fragment of a coding sequence with a nuclease under conditions under which nicking occurs only about once per molecule, denaturing the double-stranded DNA, renaturing the DNA to form double-stranded DNA which may comprise sense/antisense pairs of various nicked products, removing single-stranded sections from newly formed duplexes by treatment with S1 nuclease and ligating the resulting fragment library into an expression vector. It is possible by this method to devise an expression library which encodes N-terminal, C-terminal and internal fragments of the protein of the invention, which has different sizes.

[0084] Several techniques are known in the prior art for screening gene products from combinatorial libraries which have been produced by point mutations or truncation and for screening DNA libraries for gene products with a selected property. These techniques can be adapted to rapid screening of gene libraries which have been generated by combinatorial mutagenesis of homologs of the invention. The most frequently used techniques for screening large gene libraries undergoing high-throughput analysis comprise the cloning of the gene library into replicable expression vectors, transformation of suitable cells with the resulting vector library and expression of the combinatorial genes under conditions under which detection of the desired activity facilitates isolation of the vector encoding the gene whose product has been detected. Recursive ensemble mutagenesis (REM), a technique which increases the frequency of functional mutants in the libraries, can be used in combination with the screening tests in order to identify homologs (Arkin und Yourvan (1992) PNAS 89:7811-7815; Delgrave et al. (1993) Protein Engineering 6(3):327-331.

c) Polynucleotides of the Invention

[0085] The invention also relates to nucleic acid sequences (single- and double-stranded DNA and RNA sequences such as, for example cDNA and mRNA) coding for one of the above meta enzymes and the functional equivalents thereof which are obtainable, for example, also by use of artificial nucleotide analogs.

[0086] The invention relates both to isolated nucleic acid molecules which code for polypeptides or proteins of the invention or for biologically active sections thereof, and to nucleic acid fragments which can be used, for example, for use as hybridization probes or primers for identifying or amplifying coding nucleic acids of the invention.

[0087] Moreover, the nucleic acid molecules of the invention may contain untranslated sequences from the 3' and/or 5' ends of the coding region of the gene.

[0088] An "isolated" nucleic acid molecule is separated from other nucleic acid molecules which are present in the natural source of the nucleic acid and may moreover be essentially free of other cellular material or culture medium if it is prepared by recombinant techniques, or free of chemical precursors or other chemicals if it is chemically synthesized.

[0089] The invention furthermore comprises the nucleic acid molecules complementary to the specifically described nucleotide sequences or a section thereof.

[0090] The nucleotide sequences of the invention make it possible to generate probes and primers which can be used for identifying and/or cloning homologous sequences in other cell types and organisms. Such probes and primers usually complete a nucleotide sequence region which hybridizes under stringent conditions to at least about 12, preferably at least about 25, such as, for example 40, 50 or 75, consecutive nucleotides of a sense strand of a nucleic acid sequence of the invention or of a corresponding antisense strand.

[0091] Further nucleic acid sequences of the invention are derived from SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43 or 45 and differ therefrom through addition, substitution, insertion or deletion of one or more nucleotides, but still code for polypeptides having the desired profile of properties. These may be polynucleotides which are identical to above sequences in at least about 50%, 55%, 60%, 65%, 70%, 80% or 90%, preferably in at least about 95%, 96%, 97%, 98% or 99%, of the sequence positions.

[0092] The invention also includes those nucleic acid sequences which comprise "silent" mutations or are modified, by comparison with a specifically mentioned sequence, in accordance with the codon usage of a specific source or host organism, as well as naturally occurring variants such as, for example, splice variants or allelic variants. The invention likewise relates to sequences which are obtainable by conservative nucleotide substitutions (i.e. the relevant amino acid is replaced by an amino acid of the same charge, size, polarity and/or solubility).

[0093] The invention also relates to molecules derived from specifically disclosed nucleic acids through sequence polymorphisms. These genetic polymorphisms may exist because of the natural variation between individuals within a population. These natural variations usually result in a variance of from 1 to 5% in the nucleotide sequence of a gene.

[0094] The invention furthermore also comprises nucleic acid sequences which hybridize with or are complementary to the abovementioned coding sequences. These polynucleotides can be found on screening of genomic or cDNA libraries, and where appropriate, be amplified therefrom by means of PCR using suitable primers, and then, for example, be isolated with suitable probes. Another possibility is to transform suitable microorganisms with polynucleotides or vectors of the invention, multiply the microorganisms and thus the polynucleotides, and then isolate them. An additional possibility is to synthesize polynucleotides of the invention by chemical routes.

[0095] The property of being able to "hybridize" to polynucleotides means the ability of a polynucleotide or oligonucleotide to bind under stringent conditions to an almost complementary sequence, while there are no unspecific bindings between noncomplementary partners under these conditions. For this purpose, the sequences should be 70-100%, preferably 90-100%, complementary. The property of complementary sequences being able to bind specifically to one another is made use of, for example, in the Northern or Southern blot technique or in PCR or RT-PCR in the case of primer binding. Oligonucleotides with a length of 30 base pairs or more are usually employed for this purpose. Stringent conditions means, for example, in the Northern blot technique the use of a washing solution at 50-70.degree. C., preferably 60-65.degree. C., for example 0.1.times.SSC buffer with 0.1% SDS (20.times.SSC; 3M NaCl, 0.3M Na citrate, pH 7.0) for eluting nonspecifically hybridized cDNA probes or oligonucleotides. In this case, as mentioned above, only nucleic acids with a high degree of complementarity remain bound to one another. The setting up of stringent conditions is known to the skilled worker and is described, for example, in Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1-6.3.6.

d) Isolation of the Coding MetA Gene

[0096] The meta genes coding for the enzyme homoserine O-acetyltransferase can be isolated from the organisms of the above list I in a manner known per se.

[0097] In order to isolate the meta genes or else other genes of the organisms of the above list 1, first a gene library of this organism is generated in Escherichia coli (E. coli). The generation of gene libraries is described in detail in generally known textbooks and manuals. Examples which may be mentioned are the textbook by Winnacker: Gene und Klone, Eine Einfuchrung in die Gentechnologie (Verlag Chemie, Weinheim, Germany, 1990), and the manual by Sambrook et al.: Molecular Cloning, A Laboratory Manual (Cold Spring Harbor Laboratory Press, 1989). A very well-known gene library is that of E. coli K-12 strain W3110, which was generated in .lamda. vectors by Kohara et al. (Cell50, 495-508 (198)).

[0098] In order to produce a gene library from organisms of list I in E. coli, cosmids such as the cosmid vector SuperCos I (Wahl et al., 1987, Proceedings of the National Academy of Sciences USA, 84: 2160-2164), or else plasmids such as pBR322 (BoliVal; Life Sciences, 25, 807-818 (1979)) or pUC9 (Vieira et al., 1982, Gene, 19: 259-268) can be used. Suitable hosts are in particular those E. coli strains which are restriction and recombination defective. An example of this is the strain DH5.alpha.mcr which has been described by Grant et al. (Proceedings of the National Academy of Sciences USA, 87 (1990) 4645-4649). The long DNA fragments cloned with the aid of cosmids may then in turn be subcloned into common vectors suitable for sequencing and subsequently be sequenced, as described, for example, in Sanger et al. (proceedings of the National Academy of Sciences of the United States of America, 74: 5463-5467, 1977).

[0099] The DNA sequences obtained can then be studied using known algorithms or sequence analysis programs such as, for example, those by Staden (Nucleic Acids Research 14, 217-232(1986)), by Marck (Nucleic Acids Research 16, 1829-1836 (1988)) or the GCG program by Butler (Methods of Biochemical Analysis 39, 74-97 (1998)).

[0100] The meta-encoding DNA sequences from organisms according to the above list I were found. In particular, DNA sequences according to SEQ ID NO:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43 and 45, were found. Furthermore, the amino acid sequences of the corresponding proteins were derived from said DNA sequences present, using the above-described methods. SEQ ID NO:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44 and 46 depict the resulting amino acid sequences of the meta gene products.

[0101] Coding DNA sequences which result from the sequences according to SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43 and 45 due to the degeneracy of the genetic code are likewise subject of the invention. In the same way, the invention relates to DNA sequences which hybridize with said sequences or parts of sequences derived therefrom.

[0102] Instructions for identifying DNA sequences by means of hybridization can be found by the skilled worker, inter alia, in the manual "The DIG System Users Guide fur Filter Hybridization" from Boehringer Mannheim GmbH (Mannheim, Germany, 1993) and in Liebl et al. (International Journal of Systematic Bacteriology (1991) 41: 255-260). Instructions for amplifying DNA sequences with the aid of the polymerase chain reaction (PCR) can be found by the skilled worker, inter alia, in the manual by Gait: Oligonucleotide synthesis: A Practical Approach (IRL Press, Oxford, UK, 1984) and in Newton and Graham: PCR (Spektrum Akademischer Verlag, Heidelberg, Germany, 1994).

[0103] It is furthermore known that changes at the N-- and/or C-terminus of a protein do not substantially impair its function or may even stabilize said function. Information on this can be found by the skilled worker, inter alia, in Ben-Bassat et al. (Journal of Bacteriology 169: 751-757 (1987)), in O'Regan et al. (Gene 77: 237-251 (1989), in Sahin-Toth et al. (Protein Sciences 3: 240-247 (1994)), in Hochuli et al. (Biotechnology 6: 1321-1325 (1988)) and in known textbooks of genetics and molecular biology.

[0104] Amino acid sequences which result accordingly from SEQ ID NO:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44 and 46 are likewise part of the invention.

e) Host Cells Used According to the Invention

[0105] The invention further relates to microorganisms serving as host cells, in particular coryneform bacteria, which contain a vector, in particular a shuttle vector or plasmid vector, carrying at least one meta gene as defined by the invention or in which a meta gene of the invention is expressed or amplified.

[0106] These microorganisms can produce sulfur-containing fine chemicals, in particular L-methionine, from glucose, sucrose, lactose, fructose, maltose, molasses, starch, cellulose or from glycerol and ethanol. Said microorganisms are preferably coryneform bacteria, in particular of the genus Corynebacterium. Of the genus Corynebacterium, mention must be made in particular of the species Corynebacterium glutamicum which is known in the art for its ability to produce L-amino acids.

[0107] Examples of suitable strains of coryneform bacteria, which may be mentioned, are those of the genus Corynebacterium, in particular of the species Corynebacterium glutamicum [0108] (C. glutamicum), such as [0109] Corynebacterium glutamicum ATCC 13032, [0110] Corynebacterium acetoglutamicum ATCC 15806, [0111] Corynebacterium acetoacidophilum ATCC 13870, [0112] Corynebacterium thermoaminogenes FERM BP-1539, [0113] Corynebacterium melassecola ATCC 17965 [0114] or [0115] of the genus Brevibacterium, such as [0116] Brevibacterium flavum ATCC 14067 [0117] Brevibacterium lactofermentum ATCC 13869 and [0118] Brevibacterium divaricatum ATCC 14020; [0119] Or strains derived therefrom such as [0120] Corynebacterium glutamicum KFCC10065 [0121] Corynebacterium glutamicum ATCC21608 which likewise produce the desired fine chemical or the precursor(s) thereof.

[0122] The abbreviation KFCC means the Korean Federation of Culture Collection, the abbreviation ATCC means the American Type Strain Culture Collection, the abbreviation FERM BP refers to the Collection of the National Institute of Bioscience and Human Technology, Agency of Industrial Science and Technology, Japan.

f) Carrying Out the Fermentation of the Invention

[0123] According to the invention, it was found that coryneform bacteria, after overexpression of a meta gene from organisms of the list I, produce sulfur-containing fine chemicals, in particular L-methionine, in an advantageous manner.

[0124] To achieve overexpression, the skilled worker can take different measures individually or in combination. Thus it is possible to increase the copy number of the appropriate genes or to mutate the promoter and regulatory region or the ribosomal binding site which is located upstream of the structural gene. Expression cassettes which are incorporated upstream of the structural gene act in the same way. Inducible promoters make it additionally possible to increase expression during the course of the fermentative L-methionine production. Expression is likewise improved by measures which extend the life span of the mRNA. Furthermore, the enzymic activity is likewise enhanced by preventing degradation of the enzyme protein. The genes or gene constructs may be either present in plasmids with varying copy number or integrated and amplified in the chromosome. A further possible alternative is to achieve overexpression of the relevant genes by changing the media composition and management of the culture.

[0125] Instructions for this can be found by the skilled worker, inter alia, in Martin et al. (Biontechnology 5,137-146 (1987)), in Guerrero et al. (Gene 138, 35-41 (1994)), Tsuchiya and Morinaga (Bio/Technology 6, 428-430 (1988)), in Eikmanns et al. (Gene 102, 93-98 (1991)), in the European patent 0472869, in U.S. Pat. No. 4,601,893, in Schwarzer and Puhler (Biotechnology 9, 84-87 (1991), in Remscheid et al. (Applied and Environmental Microbiology 60, 126-132 (1994), in LaBarre et al. (Journal of Bacteriology 175, 1001-1007 (1993)), in the patent application WO 96/15246, in Malumbres et al. (Gene 134, 15-24 (1993)), in the Japanese published specification JP-A-10-229891, in Jensen und Hammer (Biotechnology and Bioengineering 58, 191-195 (1998)), in Makrides (Microbiological Reviews 60: 512-538 (1996) and in known textbooks of genetics and molecular biology.

[0126] The invention therefore also relates to expression constructs comprising a nucleic acid sequence coding for a polypeptide of the invention under the genetic control of regulatory nucleic acid sequences; and to vectors comprising at least one of said expression constructs. Such constructs of the invention preferably include a promoter 5' upstream of the particular coding sequence and a terminator sequence 3' downstream and also, where appropriate, further regulatory elements, in each case operatively linked to the coding sequence. An "operative linkage" means the sequential arrangement of promoter, coding sequence, terminator and, where appropriate, further regulatory elements such that each of the regulatory elements can properly carry out its function in the expression of the coding sequence. Examples of operatively linkable sequences are activating sequences and enhancers and the like. Further regulatory elements include selectable markers, amplification signals, origins of replication and the like. Suitable regulatory sequences are described, for example in Goeddel, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990).

[0127] In addition to the artificial regulatory sequences, the natural regulatory sequence may still be present upstream of the actual structural gene. Genetic modification can, where appropriate, switch off this natural regulation and increase or decrease expression of the genes. However, the gene construct may also have a simpler design, i.e. no additional regulatory signals are inserted upstream of the structural gene and the natural promoter with its regulation is not removed. Instead, the natural regulatory sequence is mutated such that regulation no longer takes place and gene expression is increased or reduced. The gene construct may contain one or more copies of the nucleic acid sequences.

[0128] Examples of useful promoters are: ddh, amy, lysC, dapA, lysA from Corynebacterium glutamicum promoters, but also Gram-positive promoters SP02, as are described in Bacillus Subtilis and Its Closest Relatives, Sonenshein, Abraham L., Hoch, James A., Losick, Richard; ASM Press, D.C., Washington and Patek M. Eikmanns B J., Patek J., Sahm H., Microbiology. 142 1297-309, 1996 or else the cos, tac, trp, tet, trp-tet, lpp, lac, lpp-lac, lacIq, T7, T5, T3, gal, trc, ara, SP6, .lamda.-PR and .lamda.-PL promoters which are advantageously applied in Gram-negative bacteria. Preference is also give to using inducible promoters such as, for example light- and, in particular, temperature-inducible promoters such as the P.sub.rP.sub.l promoter. It is in principle possible to use all natural promoters with their regulatory sequences. In addition, it is also possible to use advantageously synthetic promoters.

[0129] The regulatory sequences mentioned are intended to make specific expression of the nucleic acid sequences possible. Depending on the host organism, this may mean, for example, that the gene is expressed or overexpressed only after induction, or that it is immediately expressed and/or overexpressed.

[0130] In this connection, the regulatory sequences and factors may preferably have a beneficial effect on, and thus increase or decrease, expression. Thus, it is possible and advantageous to enhance the regulatory elements at the transcriptional level by using strong transcription signals such as promoters and/or enhancers. However, it is also possible besides this to enhance translation by, for example, improving the stability of the mRNA.

[0131] An expression cassette is prepared by fusing a suitable promoter, a suitable Shine-Dalgarno sequence, to a meta nucleotide sequence and a suitable termination signal. For this purpose, common recombination and cloning techniques are used, such as those described, for example, in Current Protocols in Molecular Biology, 1993, John Wiley & Sons, Incorporated, New York, N.Y., PCR Methods, Gelfand, David H., Innis, Michael A., Sninsky, John J., 1999, Academic Press, Incorporated, Calif., San Diego, PCR Cloning Protocols, Methods in Molecular Biology Ser., Vol. 192, 2nd ed., Humana Press, N.J., Totowa. T. Maniatis, E. F. Fritsch and J. Sambrook, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. (1989) and in T. J. Silhavy, M. L. Berman und L. W. Enquist, Experiments with Gene Fusions, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. (1984) and in Ausubel, F. M. et al., Current Protocols in Molecular Biology, Greene Publishing Assoc. and Wiley Interscience (1987).

[0132] The recombinant nucleic acid construct or gene construct is expressed in a suitable host organism by inserting it advantageously into a host-specific vector which makes optimal expression of the genes in the host possible. Vectors are well known to the skilled worker and can be found, for example, in "Cloning Vectors" (Pouwels P. H. et al., Hrsg, Elsevier, Amsterdam-New York-Oxford, 1985). The term "vectors" means, apart from plasmids, also all other vectors known to the skilled worker, such as, for example, phages, transposons, IS elements, plasmids, cosmids and linear or circular DNA. These vectors can replicate autonomously in the host organism or are replicated chromosomally.

[0133] MetA genes of the invention were amplified by overexpressing them by way of example with the aid of episomal plasmids. Suitable plasmids are those which are replicated in coryneform bacteria. Numerous known plasmid vectors such as, for example, pZ1 (Menkel et al., Applied and Environmental Microbiology (1989) 64: 549-554), pEKEx1 (Eikmanns et al., Gene 102: 93-98 (1991)) or pHS2-1 (Sonnen et al., Gene 107: 69-74 (1991)) are based on the cryptic plasmids pHM1519, pBL1 or pGA1. Other plasmid vectors such as, for example, pCLiK5MCS, or those based on pCG4 (U.S. Pat. No. 4,489,160) or pNG2 (Serwold-Davis et al., FEMS Microbiology Letters 66, 119-124 (1990)) or pAG1 (U.S. Pat. No. 5,158,891) may be used in the same way.

[0134] Suitable plasmid vectors are furthermore also those with the aid of which it is possible to apply the method of gene amplification by integration into the chromosome, as has been described, for example, by Remscheid et al. (Applied and Environmental Microbiology 60,126-132 (1994)) for the duplication and amplification of the hom-thrB operon. In this method, the complete gene is cloned into a plasmid vector which can replicate in a host (typically E. coli) but not in C. glutamicum. Suitable vectors are, for example, pSUP301 (Sirnon et al., Bio/Technology 1, 784-791 (1983)), pK18mob or pK19mob (Schafer et al., Gene 145, 69-73 (1994)), Bernard et al., Journal of Molecular Biology, 234: 534-541 (1993)), pEM1 (Schrumpf et al., 1991, Journal of Bacteriology 173: 4510-4516) or pBGS8 (Spratt et al.,1986, Gene 41: 337-342). The plasmid vector containing the gene to be amplified is then transferred into the desired C. glutamicum strain via transformation. Methods for transformation are described, for example, in Thierbach et al. (Applied Microbiology and Biotechnology 29, 356-362 (1988)), Dunican and Shivnan (Biotechnology 7, 1067-1070 (1989)) and Tauch et al. (FEMS Microbiological Letters 123, 343-347 (1994)).

[0135] The activity of enzymes can be influenced by mutations in the corresponding genes in such a way that the rate of the enzymic reaction is partly or completely reduced. Examples of such mutations are known to the skilled worker (Motoyama H., Yano H., Terasaki Y., Anazawa H., Applied & Environmental Microbiology. 67:3064-70, 2001, Eikmanns B J., Eggeling L., Sahm H., Antonie van Leeuwenhoek. 64:145-63, 1993-94.)

[0136] Additionally, it may be advantageous for the production of sulfur-containing fine chemicals, in particular L-methionine, to amplify, in addition to expression and amplification of a meta gene of the invention, one or more enzymes of the respective biosynthetic pathway, the cysteine pathway, of aspartate-semialdehyde synthesis, of glycolysis, of anaplerosis, of the pentose phosphate metabolism, the citrate acid cycle or the amino acid export.

[0137] Thus, one or more of the following genes can be amplified to produce sulfur-containing fine chemicals, in particular L-methionine: [0138] the gene lysC, which encodes an aspartate kinase (EP 1 108 790 A2; DNA-SEQ NO. 281), [0139] the gene asd, which encodes an aspartate-semialdehyde dehydrogenase (EP 1 108 790 A2; DNA-SEQ NO. 282), [0140] the glyceraldehyde-3-phosphate dehydrc ogenase-encoding gene gap (Eikmanns (1992), Journal of Bacteriology 174: 6076-6086), [0141] the 3-phosphoglycerate kinase-encoding gene pgk (Eikmanns (1992), Journal of Bacteriology 174: 6076-6086), [0142] the pyruvate carboxylase-encoding gene pyc (Eikmanns (1992), Journal of Bacteriology 174: 6076-6086), [0143] the triose phosphate isomerase-encoding gene tpi (Eikmanns (1992), Journal of Bacteriology 174: 6076-6086), [0144] the methionine synthase-encoding gene metH (EP 1 108 790 A2), [0145] the cystathionine gamma-synthase-encoding gene metB (EP 1 108 790 A2; DNA-SEQ NO. 3491), [0146] the cystathionine gamma-lyase-encoding gene metC (EP 1 108 790 A2; DNA-SEQ NO. 3061), [0147] the serine hydroxymethyltransferase-encoding gene glyA (EP 1 108 790 A2; DNA-SEQ NO. 1110), [0148] the O-acetylhomoserine sulfhydrylase-encoding gene metY (EP 1 108 790 A2; DNA-SEQ NO. 726), [0149] the methylene tetrahydrofolate reductase-encoding gene metF (EP 1 108 790 A2; DNA-SEQ NO. 2379), [0150] the phosphoserine aminotransferase-encoding gene serC (EP 1 108 790 A2; DNA-SEQ NO. 928), [0151] a phosphoserine phosphatase-encoding gene serB (EP 1 108 790 A2; DNA-SEQ NO. 334, DNA-SEQ NO. 467, DNA-SEQ NO. 2767), [0152] the serine acetyltransferase-encoding gene cysE (EP 1 108 790 A2; DNA-SEQ NO. 2818), [0153] the gene hom, which encodes a homoserine dehydrogenase (EP 1 108 790 A2; DNA-SEQ NO. 1306)

[0154] Thus, it may be advantageous for the production of sulfur-containing fine chemicals, in particular L-methionine, in coryneform bacteria to mutate, at the same time, at least one of the genes below, so that the activity of the corresponding proteins, compared to that of unmutated proteins, is influenced by a metabolic metabolite to a lesser extent or not at all: [0155] the gene lysC, which encodes an aspartate kinase (EP 1 108 790 A2; DNA-SEQ NO. 281), [0156] the pyruvate carboxylase-encoding gene pyc (Eikmanns (1992), Journal of Bacteriology 174: 6076-6086), [0157] the methionine-synthase-encoding gene metH (EP 1 108 790 A2), [0158] the cystathionine gamma-synthase-encoding gene metB (EP 1 108 790 A2; DNA-SEQ NO. 3491), [0159] the cystathionine gamma-lyase-encoding gene metC (EP 1 108 790 A2; DNA-SEQ NO. 3061), [0160] the serine hydroxymethyltransferase-encoding gene glyA (EP 1 108 790 A2; DNA-SEQ NO. 1110), [0161] the O-acetyl-homoserine-sulfhydrylase-encoding gene metY (EP 1 108 790 A2; DNA-SEQ NO. 726), [0162] the methylene tetrahydrofolate reductase-encoding gene metF (EP 1 108 790 A2; DNA-SEQ NO. 2379), [0163] the phosphoserine aminotransferase-encoding gene serC (EP 1 108 790 A2; DNA-SEQ NO. 928), [0164] a phosphoserine phosphatase-encoding gene serB (EP 1 108 790 A2; DNA-SEQ NO. 334, DNA-SEQ NO. 467, DNA-SEQ NO. 2767), [0165] the serine acetyl transferase-encoding gene cysE (EP 1 108 790 A2; DNA-SEQ NO. 2818), [0166] the gene hom, which encodes a homoserine dehydrogenase (EP 1 108 790 A2; DNA-SEQ NO. 1306).

[0167] It may be furthermore advantageous for the production of sulfur-containing fine chemicals, in particular L-methionine, in addition to expression and amplification of one of the meta genes of the invention, to attenuate one or more of the following genes, in particular to reduce expression thereof, or to switch them off: [0168] the homoserine kinase-encoding gene thrB (EP 1 108 790 A2; DNA-SEQ NO. 3453), [0169] the threonine dehydratase-encoding gene ilvA (EP 1 108 790 A2; DNA-SEQ NO. 2328), [0170] the threonine synthase-encoding gene thrC (EP 1 108 790 A2; DNA-SEQ NO. 3486), [0171] the meso-diaminopimelate D-dehydrogenase-encoding gene ddh (EP 1 108 790 A2; DNA-SEQ NO. 3494), [0172] the phosphoenolpyruvate carboxykinase-encoding gene pck (EP 1 108 790 A2; DNA-SEQ NO. 3157), [0173] the glucose-6-phosphate 6-isomerase-encoding gene pgi (EP 1 108 790 A2; DNA-SEQ NO. 950), [0174] the pyruvate oxidase-encoding gene poxB (EP 1 108 790 A2; DNA-SEQ NO. 2873), [0175] the dihydrodipicolinate synthase-encoding gene dapA (EP 1 108 790 A2; DNA-SEQ NO. 3476), [0176] the dihydrodipicolinate reductase-encoding gene dapB (EP 1 108 790 A2; DNA-SEQ NO. 3477) [0177] the diaminopicolinate decarboxylase-encoding gene lysA (EP 1 108 790 A2; DNA-SEQ NO. 3451).

[0178] It may be furthermore advantageous for the production of sulfur-containing fine chemicals, in particular L-methionine, in addition to expression and amplification of one of the metA genes of the invention in coryneform bacteria, to mutate, at the same time, at least one of the following genes in such a way that the enzymic activity of the corresponding protein is partly or completely reduced: [0179] the homoserine kinase-encoding gene thrB (EP 1 108 790 A2; DNA-SEQ NO. 3453), [0180] the threonine dehydratase-encoding gene ilvA (EP 1 108 790 A2; DNA-SEQ NO. 2328), [0181] the threonine synthase-encoding gene thrC (EP 1 108 790 A2; DNA-SEQ NO. 3486), [0182] the meso-diaminopimelate D-dehydrogenase-encoding gene ddh (EP 1 108 790 A2; DNA-SEQ NO. 3494), [0183] the phosphoenolpyruvate carboxykinase-encoding gene pck (EP 1 108 790 A2; DNA-SEQ NO. 3157), [0184] the glucose-6-phosphate 6-isomerase-encoding gene pgi (EP 1 108 790 A2; DNA-SEQ NO. 950), [0185] the pyruvate oxidase-encoding gene poxB (EP 1 108 790 A2; DNA-SEQ NO. 2873), [0186] the dihydrodipicolinate synthase-encoding gene dapA (EP 1 108 790 A2; DNA-SEQ NO. 3476), [0187] the dihydrodipicolinate reductase-encoding gene dapB (EP 1 108 790 A2; DNA-SEQ NO. 3477) [0188] the diaminopicolinate decarboxylase-encoding gene lysA (EP 1 108 790 A2; DNA-SEQ NO. 3451).

[0189] It may be furthermore advantageous for the production of sulfur-containing fine chemicals, in particular L-methionine, apart from expression and amplification of a meta gene of the invention, to eliminate unwanted secondary reactions (Nakayama: "Breeding of Amino Acid Producing Microorganisms", in: Overproduction of Microbial Products, Krumphanzl, Sikyta, Vanek (eds.), Academic Press, London, UK, 1982).

[0190] The microorganisms produced according to the invention may be cultured continuously or batchwise or in a fed batch or repeated fed batch process to produce sulfur-containing fine chemicals, in particular L-methionine. An overview of known cultivation methods can be found in the textbook by Chmiel (Bioproze.beta.technik 1. Einfuhrung in die Bioverfahrenstechnik (Gustav Fischer Verlag, Stuttgart, 1991)) or in the textbook by Storhas (Bioreaktoren und periphere Einrichtungen (Vieweg Verlag, Braunschweig/Wiesbaden, 1994)).

[0191] The culture medium to be used must satisfy the demands of the particular strains in a suitable manner. The textbook "Manual of Methods fur General Bacteriology" by the American Society for Bacteriology (Washington D.C., USA, 1981) contains descriptions of culture media for various microorganisms.

[0192] Said media which can be used according to the invention usually comprise one or more carbon sources, nitrogen sources, inorganic salts, vitamins and/or trace elements.

[0193] Preferred carbon sources are sugars such as mono-, di- or polysaccharides. Examples of very good carbon sources are glucose, fructose, mannose, galactose, ribose, sorbose, ribulose, lactose, maltose, sucrose, raffinose, starch and cellulose. Sugars may also be added to the media via complex compounds such as molasses or other byproducts of sugar refining. It may also be advantageous to add mixtures of different carbon sources. Other possible carbon sources are oils and fats such as, for example, soybean oil, sunflower oil, peanut oil and coconut fat, fatty acids such as, for example, palmitic acid, stearic acid and linoleic acid, alcohols such as, for example, glycerol, methanol and ethanol and organic acids such as, for example acetic acid and lactic acid.

[0194] Nitrogen sources are usually organic or inorganic hydrogen compounds or materials containing said compounds. Examples of nitrogen sources include ammonia gas or ammonium salts such as ammonium sulfate, ammonium chloride, ammonium phosphate, ammonium carbonate and ammonium nitrate, nitrates, urea, amino acids and complex nitrogen sources such as cornsteep liquor, soybean flour, soybean protein, yeast extract, meat extract and others. The nitrogen sources may be used singly or as mixture.

[0195] Inorganic salt compounds which may be included in the media comprise the chloride, phosphorus or sulfate salts of calcium, magnesium, sodium, cobalt, molybdenum, potassium, manganese, zinc, copper and iron.

[0196] Inorganic sulfur-containing compounds such as, for example, sulfates, sulfites, dithionites, tetrathionates, thiosulfates, sulfides, or else organic sulfur compounds such as mercaptans and thiols may be used as sources of sulfur for the production of sulfur-containing fine chemicals, in particular of methionine.

[0197] Phosphoric acid, potassium dihydrogen phosphate or dipotassium hydrogen phosphate or the corresponding sodium-containing salts may be used as sources of phosphorus.

[0198] Chelating agents may be added to the medium in order to keep the metal ions in solution. Particularly suitable chelating agents include dihydroxyphenols such as catechol or protocatechuate and organic acids such as citric acid.

[0199] The fermentation media used according to the invention usually also contain other growth factors such as vitamins or growth promoters, which include, for example, biotin, riboflavin, thiamine, folic acid, nicotinic acid, panthothenate and pyridoxine. Growth factors and salts are frequently derived from complex media components such as yeast extract, molasses, cornsteep liquor and the like. It is moreover possible to add suitable precursors to the culture medium. The exact composition of the media heavily depends on the particular experiment and is decided upon individually for each specific case. Information on the optimization of media can be found in the textbook "Applied Microbiol. Physiology, A Practical Approach" (Editors. P. M. Rhodes, P. F. Stanbury, IRL Press (1997) pp. 53-73, ISBN 0 19 963577 3). Growth media can also be obtained from commercial suppliers, for example Standard 1 (Merck) or BHI (brain heart infusion, DIFCO) and the like.

[0200] All media components are sterilized, either by heat (20 min at 1.5 bar and 121.degree. C.) or by sterile filtration. The components may be sterilized either together or, if required, separately. All media components may be present at the start of the cultivation or added continuously or batchwise, as desired.

[0201] The culture temperature is normally between 15.degree. C. and 45.degree. C., preferably at from 25.degree. C. to 40.degree. C., and may be kept constant or may be altered during the experiment. The pH of the medium should be in the range from 5 to 8.5, preferably around 7.0. The pH for cultivation can be controlled during cultivation by adding basic compounds such as sodium hydroxide, potassium hydroxide, ammonia and aqueous ammonia or acidic compounds such as phosphoric acid or sulfuric acid. Foaming can be controlled by employing antifoams such as, for example, fatty acid polyglycol esters. To maintain the stability of plasmids it is possible to add to the medium suitable substances having a selective effect, for example antibiotics. Aerobic conditions are maintained by introducing oxygen or oxygen-containing gas mixtures such as, for example, air into the culture. The temperature of the culture is normally 20.degree. C. to 45.degree. C. The culture is continued until formation of the desired product is at a maximum. This aim is normally achieved within 10 to 160 hours.

[0202] The fermentation broths obtained in this way, in particular those containing L-methionine, usually contain a dry biomass of from 7.5 to 25% by weight.

[0203] An additional advantage is to carry out the fermentation under sugar limitation, at least at the end, but in particular over at least 30% of the fermentation period. This means that during this time the concentration of utilizable sugar in the fermentation medium is maintained at or reduced to .gtoreq.0 to 3 g/l.

[0204] The fermentation broth is then processed further. The biomass may, according to requirement, be removed completely or partially from the fermentation broth by separation methods such as, for example, centrifugation, filtration, decanting or a combination of these methods or be left completely in said broth.

[0205] Subsequently, the fermentation broth may be thickened or concentrated using known methods such as, for example, with the aid of a rotary evaporator, thin film evaporator, falling film evaporator, by reverse osmosis, or by nanofiltration. This concentrated fermentation broth can then be worked up by freeze drying, spray drying, spray granulation or by other methods.

[0206] However, it is also possible to further purify the sulfur-containing fine chemicals, in particular L-methionine. To this end, the product-containing broth, after removing the biomass, is subjected to a chromatography using a suitable resin, the desired product or the contaminations being retained completely or partially on the chromatographic resin. These chromatographic steps can be repeated, if necessary, using the same or different chromatographic resin. The skilled worker is familiar with the selection of suitable chromatographic resins and their most effective application. The purified product can be concentrated by filtration or ultrafiltration and stored at a temperature at which the stability of the product is greatest.

[0207] The identity and purity of the isolated compound(s) can be determined by techniques of the art. These include high performance liquid chromatography (HPLC), spectroscopic methods, staining methods, thin-layer chromatography, NIRS, enzyme assay or microbiological assays. These analytic methods are summarized in: Patek et al. (1994) Appl. Environ. Microbiol. 60:133-140; Malakhova et al. (1996) Biotekhnologiya 11 27-32; and Schmidt et al. (1998) Bio-process Engineer. 19:67-70. Ulmann's Encyclopedia of Industrial Chemistry (1996) Bd. A27, VCH: Weinheim, pp. 89-90, pp. 521-540, pp. 540-547, pp.559-566, 575-581 and pp. 581-587; Michal, G., (1999) Biochemical Pathways: An Atlas of Biochemistry and Molecular Biology, John Wiley and Sons; Fallon, A. et al. (1987) Applications of HPLC in Biochemistry in: Laboratory Techniques in Biochemistry and Molecular Biology, Volume 17.

[0208] The following nonlimiting examples and attached figures describe the invention in more detail:

EXAMPLE 1

Construction of pCLiK5MCS

[0209] First, ampicillin resistance and origin of replication of the vector pBR322 were amplified using the oligonucleotides p1.3 (SEQ ID NO:47) and p2.3 (SEQ ID NO:48) with the aid of the polymerase chain reaction (PCR). TABLE-US-00002 p1.3 (SEQ ID NO:47) 5'-CCCGGGATCCGCTAGCGGCGCGCCGGCCGGCCCGGTGTGAAATACCG CACAG-3' p2.3 (SEQ ID NO:48) 5'-TCTAGACTCGAGCGGCCGCGGCCGGCCTTTAAATTGAAGACGAAAGG GCCTCG-3'

[0210] In addition to sequences complementary to pBR322, the oligonucleotide p1.3 (SEQ ID NO:47) contains in 5'-3' direction the cleavage sites for the restriction nucleases SmaI, BamHI, NheI and AscI and the oligonucleotide p2.3. (SEQ ID NO:48) contains in 5'-3' direction the cleavage sites for the restriction endonucleases XbaI, XhoI, NotI and DraI. The PCR reaction was carried out according to a standard method such as that by Innis et al. (PCR Protocols. A Guide to Methods and Applications, Academic Press (1990)) using PfuTurbo polymerase (Stratagene, La Jolla, USA). The DNA fragment obtained of approximately 2.1 kb in size was purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The blunt ends of the DNA fragment were ligated to one another using the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1 Blue (Stratagene, La Jolla, USA) according to standard methods, as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). Plasmid-carrying cells were selected for by plating out onto ampicillin (50 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0211] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK1.

[0212] Starting from plasmid pWLT1 (Liebl et al., 1992) as template for a PCR reaction, a kanamycin resistance cassette was amplified using the oligonucleotides neo1 (SEQ ID NO:49) and neo2 (SEQ ID NO:50). TABLE-US-00003 neo1 (SEQ ID NO:49): 5'-GAGATCTAGACCCGGGGATCCGCTAGCGGGCTGCTAAAGGAAGCGG A-3' neo2 (SEQ ID NO:50): 5'-GAGAGGCGCGCCGCTAGCGTGGGCGAAGAACTCCAGCA-3'

[0213] Apart from the sequences complementary to pWLT1, the oligonucleotide neo1 contains in 5'-3' direction the cleavage sites for the restriction endonucleases XbaI, SmaI, BamHI, NheI and the oligonucleotide neo2 (SEQ ID NO:50) contains in 5'-3' direction the cleavage sites for the restriction endonucleases AscI and NheI. The PCR reaction was carried out using PfuTurbo polymerase (Stratagene, La Jolla, USA) according to a standard method such as that of Innis et al. (PCR Protocols. A Guide to Methods and Applications, Academic Press (1990)). The DNA fragment obtained was approximately 1.3 kb in size was purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The DNA fragment was cleaved with restriction endonucleases XbaI and AscI (New England Biolabs, Beverly, USA) and, following that, again purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The vector pCLiK1 was likewise cleaved with the restriction endonucleases XbaI and AscI and dephosphorylated using alkaline phosphatase (Roche Diagnostics, Mannheim) according to the manufacturer's instructions. After electrophoresis in a 0.8% strength agarose gel, the linearized vector (approx. 2.1 kb) was isolated using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. This vector fragment was ligated with the cleaved PCR fragment with the aid of the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1 Blue (Stratagene, La Jolla, USA) according to standard methods, as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). Plasmid-carrying cells were selected for by plating out onto ampicillin (50 .mu.g/ml)- and kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0214] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK2.

[0215] The vector pCLiK2 was cleaved with the restriction endonuclease DraI (New England Biolabs, Beverly, USA). After electrophoresis in 0.8% strength agarose gel, an approx. 2.3 kb vector fragment was isolated using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. This vector fragment was religated with the aid of the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1 Blue (Stratagene, La Jolla, USA) according to standard methods, as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). Plasmid-carrying cells were selected for by plating out onto kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0216] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK3.

[0217] Starting from plasmid pWLQ2 (Liebl et al., 1992) as template for a PCR reaction, the origin of replication pHM1519 was amplified using the oligonucleotides cg1 (SEQ ID NO:51) and cg2 (SEQ ID NO:52). TABLE-US-00004 cg1 (SEQ ID NO:51): 5'-GAGAGGGCGGCCGCGCAAAGTCCCGCTTCGTGAA-3' cg2 (SEQ ID NO:52): 5'-GAGAGGGCGGCCGCTCAAGTCGGTCAAGCCACGC-3'

[0218] Apart from the sequences complementary to pWLQ2, the oligonucleotides cg1 (SEQ ID NO:51) and cg2 (SEQ ID NO:52) contain cleavage sites for the restriction endonuclease NotI. The PCR reaction was carried out using PfuTurbo polymerase (Stratagene, La Jolla, USA) according to a standard method such as that of Innis et al. (PCR Protocols. A Guide to Methods and Applications, Academic Press (1990)). The DNA fragment obtained was approximately 2.7 kb in size and was purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The DNA fragment was cleaved with restriction endonuclease NotI (New England Biolabs, Beverly, USA) and, following that, again purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The vector pCLiK3 was likewise cleaved with the restriction endonuclease NotI and dephosphorylated using alkaline phosphatase (Roche Diagnostics, Mannheim) according to the manufacturer's instructions. After electrophoresis in a 0.8% strength agarose gel, the linearized vector (approx. 2.3 kb) was isolated using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. This vector fragment was ligated with the cleaved PCR fragment with the aid of the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1Blue (Stratagene, La Jolla, USA) according to standard methods, as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). Plasmid-carrying cells were selected for by plating out onto kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0219] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK5.

[0220] PCLik5 was extended by a multiple cloning site (MCS) by combining the two synthetic essentially complementary oligonucleotides HS445 ((SEQ ID NO:53) and HS446 (SEQ ID NO:54)) which contain cleavage sites for the restriction endonucleases SwaI, XhoI, AatI, ApaI, Asp718, MluI, NdeI, SpeI, EcoRV, SalI, ClaI, BamHI, XbaI and SmaI to give a double-stranded DNA fragment by heating them together to 95.degree. C. followed by slow cooling. TABLE-US-00005 HS445 (SEQ ID NO:53): 5'-TCGAATTTAAATCTCGAGAGGCCTGACGTCGGGCCCGGTACCACGCG TCATATGACTAGTTCGGACCTAGGGATATCGTCGACATCGATGCTCTTCT GCGTTAATTAACAATTGGGATCCTCTAGACCCGGGATTTAAAT-3' HS446 (SEQ ID NO:54): 5'-GATCATTTAAATCCCGGGTCTAGAGGATCCCAATTGTTAATTAACGC AGAAGAGCATCGATGTCGACGATATCCCTAGGTCCGAACTAGTCATATGA CGCGTGGTACCGGGCCCGACGTCAGGCCTCTCGAGATTTAAAT-3'

[0221] The vector pCLiK5 was cleaved with the restriction endonucleases XhoI and BamHI (New England Biolabs, Beverly, USA) and dephosphorylated using alkaline phosphatase (I (Roche Diagnostics, Mannheim)) according to the manufacturer's instructions. After electrophoresis in a 0.8% strength agarose gel, the linearized vector (approx. 5.0 kb) was isolated using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. This vector fragment was ligated with the synthetic double-stranded DNA fragment with the aid of the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1 Blue (Stratagene, La Jolla, USA) according to standard methods as described Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor (1989)). Plasmid-carrying cells were selected for by plating out onto kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0222] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK5MCS.

[0223] Sequencing reactions were carried out according to Sanger et al. (1977) Proceedings of the National Academy of Sciences USA 74:5463-5467. The sequencing reactions were fractionated and analyzed by means of ABI Prism 377 (PE Applied Biosystems, Weiterstadt).

[0224] The resultant plasmid pCLiK5MCS is listed as SEQ ID NO: 57.

EXAMPLE 2

Construction of pCLiK5MCS Integrativ sacB

[0225] Starting from the plasmid pK19mob (Schafer et al., Gene 145,69-73(1994)) as template for a PCR reaction, the Bacillus subtilis sacB gene (coding for levan sucrase) was amplified using the oligonucleotides BK1732 and BK1733. TABLE-US-00006 BK1732 (SEQ ID NO:55): 5'-GAGAGCGGCCGCCGATCCTTTTTAACCCATCAC-3' BK1733 (SEQ ID NO:56): 5'-AGGAGCGGCCGCCATCGGCATTTTCTTTTGCG-3'

[0226] Apart from the sequences complementary to pEK19mobsac, the oligonucleotides BK1732 and BK1733 contain cleavage sites for the restriction endonuclease NotI. The PCR reaction was carried out using PfuTurbo polymerase (Stratagene, La Jolla, USA) using a standard method like that of Innis et al. (PCR Protocols. A Guide to Methods and Applications, Academic Press (1990)). The DNA fragment obtained of approximately 1.9 kb in size was purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. The DNA fragment was cleaved with the restriction endonuclease NotI (New England Biolabs, Beverly, USA) and, following that, again purified using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions.

[0227] The vector pCLiK5MCS (prepared according to example 1) was likewise cleaved with the restriction endonuclease NotI and dephosphorylated using alkali phosphatase (I (Roche Diagnostics, Mannheim)) according to the manufacturer's instructions. After electrophoresis in a 0.8% strength agarose gel, an approximately 2.4 kb in size vector fragment was isolated using the GFX.TM.PCR, DNA and gel band purification kit (Amersham Pharmacia, Freiburg) according to the manufacturer's instructions. This vector fragment was ligated with the cleaved PCR fragment with the aid of the rapid DNA ligation kit (Roche Diagnostics, Mannheim) according to the manufacturer's instructions and the ligation mixture was transformed into competent E. coli XL-1Blue (Stratagene, La Jolla, USA) according to standard methods, as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). Plasmid-carrying cells were selected for by plating out onto kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0228] The plasmid DNA of an individual clone was isolated using the Qiaprep spin miniprep kit (Qiagen, Hilden) according to the manufacturer's instructions and checked by restriction digests. The plasmid obtained in this way is denoted pCLiK5MCS integrativ sacB.

[0229] Sequencing reactions were carried out according to Sanger et al. (1977) Proceedings of the National Academy of Sciences USA 74:5463-5467. The sequencing reactions were fractionated and analyzed by means of ABI Prism 377 (PE Applied Biosystems, Weiterstadt).

[0230] The resultant plasmid pCLiK5MCS integrativ sacB is listed as SEQ ID NO: 58.

[0231] It is possible to prepare in an analog manner further vectors which are suitable for the inventive expression or overproduction of meta genes.

EXAMPLE 3

Isolation of the lysC gene from C. glutamicum strain LU1479

[0232] The first step of the strain construction is intended as an allelic substitution of the lysC wild-type gene encoding the enzyme aspartate kinase in C. glutamicum ATCC13032, hereinbelow referred to as LU1479. It is intended to carry out a nucleotide substitution in the LysC gene so that, in the resulting protein, the amino acid Thr at position 311 is exchanged for the amino acid lie.

[0233] Starting with the chromosomal DNA from LU1479 as the template for a PCR reaction, an amplification was performed with the oligonucleotide primers SEQ ID NO:59 and SEQ ID NO:60 lysC with the aid of the Pfu-Turbo PCR system (Stratagene USA) following the manufacturer's instructions. Chromosomal DNA from C. glutamicum ATCC 13032 was prepared as described by Tauch et al. (1995) Plasmid 33:168-179 or Eikmanns et al. (1994) Microbiology 140:1817-1828. The amplified fragment is flanked at its 5' end by an SalI restriction cleavage and at its 3' end by an MluI restriction cleavage. Prior to cloning, the amplified fragment was digested by these two restriction enzymes and purified with GFX.TM.PCR, DNA and Gel Band Purification Kit (Amersham Pharmacia, Freiburg). TABLE-US-00007 SEQ ID NO:59 5'-GAGAGAGAGACGCGTCCCAGTGGCTGAGACGCATC-3' SEQ ID NO:60 5'-CTCTCTCTGTCGACGAATTCAATCTTACGGCCTG-3'

[0234] The resulting polynucleotide was cloned into pCLIK5 MCS integrativ SacB (hereinbelow referred to as pCIS; SEQ ID NO: 58 of Example 2) via the SalI and MluI restriction cleavages and transformed into E. coli XL-1 blue. Selection for plasmid-bearing cells was achieved by plating on kanamycin (20 .mu.g/ml)-containing LB Agar (Lennox, 1955, Virology, 1:190). The plasmid was isolated and the expected nucleotide sequence was verified by sequencing. The plasmid DNA was prepared by methods of, and using materials from, Quiagen. Sequencing reactions were carried out as described by Sanger et al. (1977) Proceedings of the National Academy of Sciences USA 74:5463-5467. The sequencing reactions were separated using ABI Prism 377 (PE Applied Biosystems, Weiterstadt) and evaluated. The resulting plasmid pCIS lysC is shown as SEQ ID NO:61. The corresponding plasmid map is shown in FIG. 1.

[0235] Sequence SEQ ID NO:61 encompasses the following essential part-regions: TABLE-US-00008 LOCUS pCIS\lysC 5860 bp DNA circular FEATURES Location/Qualifiers CDS.sup.1) 155..1420 /vntifkey="4" /label=lysC CDS complement.sup.2)(3935..5356) /vntifkey="4" /label=sacB\(Bacillus\subtilis) promoter complement(5357..5819) /vntifkey="30" /label=Promotor\sacB C_region complement(3913..3934) /vntifkey="2" /label=sacB\downstream region CDS 1974..2765 /vntifkey="4" /label=Kan\R CDS complement(3032..3892) /vntifkey="4" /label=Ori\-EC\(pMB) .sup.1)coding sequence .sup.2)on the complementary strand

EXAMPLE 4

Mutagenesis of the C. glutamicum lysC Gene

[0236] Site-specific mutagenesis of the C. glutamicum lysC gene (example 3) was carried out using the QuickChange Kit (Stratagene/USA) following the manufacturer's instructions. The mutagenesis was carried out in the plasmid pCIS lysC, SEQ ID NO:61. The following oligonucleotide primers were synthesized for the exchange of thr311 for 311ile with the aid of the Quick-change method (Stratagene): TABLE-US-00009 SEQ ID NO:62 5'-CGGCACCACCGACATCATCTTCACCTGCCCTCGTTCCG-3' SEQ ID NO:63 5'-CGGAACGAGGGCAGGTGAAGATGATGTCGGTGGTGCCG-3'

[0237] The use of these oligonucleotide primers in the Quickchange reaction leads to a substitution of the nucleotide in position 932 (T being substituted for C) in the lysC gene (cf. SEQ ID NO:64) and to an amino acid substitution in position 311 (Thr.fwdarw.Ile) in the corresponding enzyme (cf. SEQ ID NO:65). The resulting amino acid substitution Thr311Ile in the lysC gene was verified by sequencing after transformation into E. coli XL1-blue and plasmid preparation. The plasmid was named pCIS lysC thr311ile and is shown as SEQ ID NO:66. The corresponding plasmid map is shown in FIG. 2.

[0238] Sequence SEQ ID NO:66 encompasses the following essential part regions: TABLE-US-00010 LOCUS pCIS\lysC\thr311ile 5860 bp DNA circular FEATURES Location/Qualifiers CDS.sup.1) 155..1420 /vntifkey="4" /label=lysC CDS complement.sup.2)(3935..5356) /vntifkey="4" /label=sacB\(Bacillus\subtilis) promoter complement(5357..5819) /vntifkey="30" /label=Promotor\sacB C_region complement(3913..3934) /vntifkey="2" /label=sacB\downstream region CDS 1974..2765 /vntifkey="4" /label=Kan\R CDS complement(3032..3892) /vntifkey="4" /label=Ori\-EC\(pMB) .sup.1)coding sequence .sup.2)on the complementary strand

[0239] The plasmid pCIS lysC thr311ile was transformed into C. glutamicum LU1479 by means of electroporation as described by Liebl, et al. (1989) FEMS Microbiology Letters 53:299-303. Modifications of the protocol are described in DE-A-10046870. The chromosomal arrangement of the lysC locus of individual transformants was checked using standard methods by Southern blot and hybridization as described by Sambrook et al. (1989), Molecular Cloning. A Laboratory Manual, Cold Spring Harbor. It was thus ensured that the transformants were those which have the transformed plasmid integrated at the lysC locus by homologous recombination. After such colonies had grown overnight in media without antibiotic, the cells were plated onto a sucrose --CM agar medium (10% sucrose) and incubated for 24 hours at 30.degree. C.

[0240] Since the sacB gene which is present in the vector pCIS lysC thr311ile converts sucrose into a toxic product, only those colonies which have the sacB gene deleted between the wild-type lysC gene and the mutated gene lysC thr311ile by a second homologous recombination step are capable of growing. Either the wild-type gene or the mutated gene together with the sacB gene can be deleted during homologous recombination. When the sacB gene is removed together with the wild-type gene, a mutated transformant results.

[0241] Growing colonies were picked and examined for a kanamycin-sensitive phenotype. Clones with deleted SacB gene must simultaneously show kanamycin-sensitive growth behavior. Such Kan-sensitive clones were studied in a shake flask for their lysine productivity (see example 6). The untreated strain LU1479 was grown for comparison purposes. Clones whose lysin production was increased over that of the control were selected, chromosomal DNA was obtained, and the corresponding region of the lysC gene was amplified by PCR reaction and sequenced. One such clone with the property of an increased lysine synthesis and confirmed mutation in lysC at position 932 was named LU1479 lysC 311ile.

EXAMPLE 5

Preparation of Ethionine-Resistant C. glutamicum Strains

[0242] In the second strain construction step, the resulting strain LU1479 lysC 311ile (example 4) was treated in order to induce resistance to ethionine (Kase, H. Nakayama K.Agr. Biol. Chem. 39 153-106 1975 L-methionine production by methionine analog-resistant mutants of Corynebacterium glutamicum): an overnight culture in BHI medium (Difco) was washed in citrate buffer (50 mM pH 5.5) and treated for 20 min at 30.degree. C. with N-methylnitrosoguanidine (10 mg/ml in 50 mM citrate pH 5.5). After treatment with the chemical mutagen N-methylnitrosoguanidine, the cells were washed (citrate buffer 50 mM pH 5.5) and plated out on a medium composed of the following components, based on 500 ml: 10 g (NH.sub.4).sub.2SO.sub.4, 0.5 g KH.sub.2PO.sub.4, 0.5 g K.sub.2HPO.sub.4, 0.125 g MgSO.sub.4.7H.sub.2O, 21 g MOPS, 50 mg CaCl.sub.2, 15 mg proteocatechuate, 0.5 mg biotin, 1 mg thiamine, 5 g/l D,L-ethionine (Sigma Chemicals Deutschland), pH 7.0. In addition, the medium comprised 0.5 ml of a microsalt solution of: 10 g/l FeSO.sub.4.7H.sub.2O, 1 g/l MnSO.sub.4*H.sub.2O, 0.1 g/l ZnSO.sub.4*7H.sub.2O, 0.02 g/l CuSO.sub.4, 0.002 g/l NiCl.sub.2*6H.sub.2O; all salts were dissolved in 0.1 M HCl. The finished medium was filter-sterilized and, after addition of 40 ml of sterile 50% glucose solution, liquid sterile agar was added in a final concentration of 1.5% agar and the mixture was poured into culture dishes.

[0243] Cells which had undergone mutagenic treatment were applied to plates containing the above-described medium and incubated for 3-7 days at 30.degree. C. Resulting clones were isolated, and individual clones were isolated at least once on the selection medium and then analyzed for their methionine productivity in a shake flask in medium II (see example 6

EXAMPLE 6

Preparation of Methionine Using the Strain LU1479 lysC 311ile ET-16

[0244] The strains generated in Example 5 were grown for 2 days at 30.degree. C. on an agar plate comprising CM medium.

[0245] CM agar: 10.0 g/l D-glucose, 2.5 g/l NaCl, 2.0 g/l urea, 10.0 g/l Bacto peptone (Difco), 5.0 g/l yeast extract (Difco), 5.0 g/l beef extract (Difco), 22.0 g/l agar (Difco), autoclaved (20 min., 121.degree. C.)

[0246] The cells were subsequently scraped from the plate and resuspended in saline. For the main culture, 10 ml of medium II and 0.5 g of autoclaved CaCO.sub.3 (Riedel de Haen) in a 100 ml Erlenmeyer flask were inoculated with the cell suspension to an OD 600 nm of 1.5 and incubated for 72 h at 30.degree. C. on an orbital shaker at 200 rpm. TABLE-US-00011 Medium II: 40 g/l sucrose 60 g/l molasses (based on 100% sugar content) 10 g/l (NH.sub.4).sub.2SO.sub.4 0.4 g/l MgSO.sub.4*7H.sub.2O 0.6 g/l KH.sub.2PO.sub.4 0.3 mg/l thiamine*HCl 1 mg/l biotin (from a 1 mg/ml filter-sterilized stock solution brought to pH 8.0 with NH.sub.4OH) 2 mg/l FeSO.sub.4 2 mg/l MnSO.sub.4

a pH of 7.8 was established with NH.sub.4OH and the mixture was autoclaved (121.degree. C., 20 min). In addition, vitamin B12 (hydroxycobalamin Sigma Chemicals) was added from a stock solution (200 .mu.g/ml, filter-sterilized) to a final concentration of 100 .mu.g/l.

[0247] Methionine formed, as well as other amino acids in the culture broth, were with the aid of the Agilent amino acid acid determination method on an Agilent 1100 Series LC System HPLC. Pre-column derivatization with ortho-phtalaldehyde allowed the amount of the amino acid formed to be determined. The amino acid mixture was separated on a column. The amino acid mixture was separated on a Hypersil AA column (Agilent).

[0248] Clones whose methionine productivity was at least twice as high as that of the original strain LU1479 lysC 311ile were isolated. One such clone was employed in the further experiment and was named LU1479 lysC 311ile ET-16.

EXAMPLE 7

Cloning meta from Corynebacterium diphtheriae and Cloning into the Plasmid pC metA_Cd

[0249] Chromosomal DNA of Corynebacterium diphtheriae was obtained from the American Type Strain Culture Collection (ATCC, Atlanta-USA), Cat. No. 700971D from strain ATCC 700971.

[0250] Using the oligonucleotide primers SEQ ID NO:67 and SEQ ID NO:68, the C. diphtheriae chromosomal DNA as template and Pfu Turbo polymerase (Stratagene), a DNA fragment of approx. 1.4 kb, which comprises the metA gene including a noncoding 5' region (promoter region), was amplified with the aid of the polymerase chain reaction (PCR) following standard methods such as Innis et al. (1990) PCR Protocols. A Guide to Methods and Applications, Academic Press. The amplified fragment is flanked at its 5' end by an XhoI restriction cleavage site and at the 3' end by an NdeI restriction cleavage site, which had been introduced via the oligonucleotide primers. TABLE-US-00012 SEQ ID NO:67 5'-GAGACTCGAGGTTGGCTGGTCATCATAGG-3' and SEQ ID NO:76 5' GAAGAGAGCATATGTCAGCGCTCTAGTTTGGTTC-3'

[0251] The resulting DNA fragment was purified with the GFX.TM.PCR, DNA and Gel Band Purification Kit (Amersham Pharmacia, Freiburg) following the manufacturer's instructions. Thereafter, it was cleaved with the restriction enzymes XhoI and NdeI (Roche Diagnostics, Mannheim) and separated by gel electrophoresis. The approximately 1.4 kb DNA fragment was subsequently isolated from the agarose using the GFX.TM.PCR, DNA and Gel Band Purification Kit (Amersham Pharmacia, Freiburg).

[0252] The vector pClik5MCS SEQ ID NO: 57, hereinbelow referred to as pC, was cut with the restriction enzymes XhoI and NdeI (Roche Diagnostics, Mannheim), and an approximately 5 kb fragment was separated by electrophoresis and then isolated using the GFX.TM.PCR, DNA and Gel Band Purification Kit.

[0253] The vector fragment was ligated together with the PCR fragment with the aid of the Rapid DNA Ligation Kit (Roche Diagnostics, Mannheim) following the manufacturer's instructions, and the ligation reaction was transformed into competent E. coli XL-1 Blue (Stratagene, La Jolla, USA) using standard methods as described in Sambrook et al. (Molecular Cloning. A Laboratory Manual, Cold Spring Harbor, (1989)). A selection for plasmid-bearing cells was achieved by plating onto kanamycin (20 .mu.g/ml)-containing LB agar (Lennox, 1955, Virology, 1:190).

[0254] The plasmid DNA was prepared using methods of, and materials from, Quiagen. Sequencing reactions were carried out as described by Sanger et al. (1977) Proceedings of the National Academy of Sciences USA 74:5463-5467. The sequencing reactions were separated and evaluated by means of ABI Prism 377 (PE Applied Biosystems, Weiterstadt).

[0255] The resulting plasmid pC metA_Cd (Corynebacterium diphtheriae) is shown as SEQ ID NO:69. The corresponding plasmid map is shown in FIG. 3. TABLE-US-00013 LOCUS pC_metA_Cd 6472 bp DNA circular FEATURES Location/Qualifiers CDS 313..1416 /vntifkey="4" /label=metA\Corynebacterium\diphtheriae CDS 1838..2629 /vntifkey="4" /label=Kan\R CDS 4910..6031 /vntifkey="4" /label=Rep\Protein CDS 3902..4576 /vntifkey="4" /label=ORF\1 CDS complement(2896..3756) /vntifkey="4" /label=Ori\-EC\(pMB)

EXAMPLE 8

Transformation of the Strain LU1479 lysC 311ile ET-16 with the Plasmid pC metA_Cd

[0256] The strain LU 1479 lysC 311ile ET-16 was transformed with the plasmid pC metA_Cd by the above-described method (Liebl, et al. (1989) FEMS Microbiology Letters 53:299-303). The transformation mixture was plated onto CM plates which additionally comprise 20 mg/l kanamycin in order to obtain a selection for plasmid-containing cells. Resulting Kan-resistant clones were picked and individual clones were isolated. The methionine productivity of the clones was studied in a shake-flask experiment (see Example 6). The strain LU1479 lysC 311ile ET-16 pC metA_Cd produced significantly more methionine in comparison with LU1479 lysC 311ile ET-16.

Sequence CWU 1

1

69 1 1104 DNA Corynebacterium diphteriae CDS (1)..(1101) RDI00386 1 atg ctc acc acc aca ggg acg ctc acg cac caa aaa atc gga gac ttt 48 Met Leu Thr Thr Thr Gly Thr Leu Thr His Gln Lys Ile Gly Asp Phe 1 5 10 15 tac acc gaa gcc gga gcg acg ctt cac gac gta acc atc gcc tac caa 96 Tyr Thr Glu Ala Gly Ala Thr Leu His Asp Val Thr Ile Ala Tyr Gln 20 25 30 gca tgg ggc cac tac acc ggc acc aat ctc atc gtt ctc gaa cat gcc 144 Ala Trp Gly His Tyr Thr Gly Thr Asn Leu Ile Val Leu Glu His Ala 35 40 45 ctg acc ggc gac tct aac gct att tca tgg tgg gac gga ctg att ggc 192 Leu Thr Gly Asp Ser Asn Ala Ile Ser Trp Trp Asp Gly Leu Ile Gly 50 55 60 cct ggc aaa gca ctc gac acc aac cgc tac tgc atc cta tgc acc aac 240 Pro Gly Lys Ala Leu Asp Thr Asn Arg Tyr Cys Ile Leu Cys Thr Asn 65 70 75 80 gtg ctc gga gga tgc aaa gga tcc acc gga ccg agc agt cca cac cca 288 Val Leu Gly Gly Cys Lys Gly Ser Thr Gly Pro Ser Ser Pro His Pro 85 90 95 gac gga aaa cca tgg gga tcc aga ttt cca gcc ctt tca atc cgt gac 336 Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Ala Leu Ser Ile Arg Asp 100 105 110 ctt gtc aat gcc gaa aaa caa ctt ttc gac cac ctc ggc atc aat aaa 384 Leu Val Asn Ala Glu Lys Gln Leu Phe Asp His Leu Gly Ile Asn Lys 115 120 125 att cac gca atc atc ggc gga tcc atg gga ggc gca cgc acc ctc gaa 432 Ile His Ala Ile Ile Gly Gly Ser Met Gly Gly Ala Arg Thr Leu Glu 130 135 140 tgg gct gca ctc cac cca cac atg atg acg act gga ttc gtc ata gca 480 Trp Ala Ala Leu His Pro His Met Met Thr Thr Gly Phe Val Ile Ala 145 150 155 160 gtc tca gca cgc gca agc gct tgg caa atc ggt att caa act gca caa 528 Val Ser Ala Arg Ala Ser Ala Trp Gln Ile Gly Ile Gln Thr Ala Gln 165 170 175 atc agc gcc ata gaa ctc gac ccc cac tgg aac ggc ggc gat tac tac 576 Ile Ser Ala Ile Glu Leu Asp Pro His Trp Asn Gly Gly Asp Tyr Tyr 180 185 190 agc ggt cac gca cca tgg gaa gga atc gcc gcc gct cgc cgg atc gcc 624 Ser Gly His Ala Pro Trp Glu Gly Ile Ala Ala Ala Arg Arg Ile Ala 195 200 205 cac ctc acc tat cgc ggc gaa cta gaa ata gac gaa cga ttc ggc act 672 His Leu Thr Tyr Arg Gly Glu Leu Glu Ile Asp Glu Arg Phe Gly Thr 210 215 220 tcc gca caa cac ggt gaa aac cca ctc ggc ccc ttc cga gat cca cat 720 Ser Ala Gln His Gly Glu Asn Pro Leu Gly Pro Phe Arg Asp Pro His 225 230 235 240 caa cgt ttt gcg gtc acg agc tac ctc caa cac caa ggc atc aaa ctc 768 Gln Arg Phe Ala Val Thr Ser Tyr Leu Gln His Gln Gly Ile Lys Leu 245 250 255 gct caa cga ttc gat gca ggt agt tac gtc gtg ctt acc gaa gcc ctc 816 Ala Gln Arg Phe Asp Ala Gly Ser Tyr Val Val Leu Thr Glu Ala Leu 260 265 270 aat cgt cat gac atc gga cgc ggc cga ggc gga ctc aac aaa gcc ctc 864 Asn Arg His Asp Ile Gly Arg Gly Arg Gly Gly Leu Asn Lys Ala Leu 275 280 285 agc gca atc aca gtc ccc atc atg att gct ggc gtt gat acc gat att 912 Ser Ala Ile Thr Val Pro Ile Met Ile Ala Gly Val Asp Thr Asp Ile 290 295 300 ctc tac ccc tat cac cag caa gaa cac cta tca cga aat cta ggc aac 960 Leu Tyr Pro Tyr His Gln Gln Glu His Leu Ser Arg Asn Leu Gly Asn 305 310 315 320 cta ctc gct atg gca aaa atc agc tca cca gta ggc cac gac gct ttc 1008 Leu Leu Ala Met Ala Lys Ile Ser Ser Pro Val Gly His Asp Ala Phe 325 330 335 ctc aca gaa ttc cga caa atg gag cga atc cta aga cat ttc atg gag 1056 Leu Thr Glu Phe Arg Gln Met Glu Arg Ile Leu Arg His Phe Met Glu 340 345 350 ctt tcg gaa gga atc gac gat tcc ttc cga acc aaa cta gag cgc 1101 Leu Ser Glu Gly Ile Asp Asp Ser Phe Arg Thr Lys Leu Glu Arg 355 360 365 tga 1104 2 367 PRT Corynebacterium diphteriae 2 Met Leu Thr Thr Thr Gly Thr Leu Thr His Gln Lys Ile Gly Asp Phe 1 5 10 15 Tyr Thr Glu Ala Gly Ala Thr Leu His Asp Val Thr Ile Ala Tyr Gln 20 25 30 Ala Trp Gly His Tyr Thr Gly Thr Asn Leu Ile Val Leu Glu His Ala 35 40 45 Leu Thr Gly Asp Ser Asn Ala Ile Ser Trp Trp Asp Gly Leu Ile Gly 50 55 60 Pro Gly Lys Ala Leu Asp Thr Asn Arg Tyr Cys Ile Leu Cys Thr Asn 65 70 75 80 Val Leu Gly Gly Cys Lys Gly Ser Thr Gly Pro Ser Ser Pro His Pro 85 90 95 Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Ala Leu Ser Ile Arg Asp 100 105 110 Leu Val Asn Ala Glu Lys Gln Leu Phe Asp His Leu Gly Ile Asn Lys 115 120 125 Ile His Ala Ile Ile Gly Gly Ser Met Gly Gly Ala Arg Thr Leu Glu 130 135 140 Trp Ala Ala Leu His Pro His Met Met Thr Thr Gly Phe Val Ile Ala 145 150 155 160 Val Ser Ala Arg Ala Ser Ala Trp Gln Ile Gly Ile Gln Thr Ala Gln 165 170 175 Ile Ser Ala Ile Glu Leu Asp Pro His Trp Asn Gly Gly Asp Tyr Tyr 180 185 190 Ser Gly His Ala Pro Trp Glu Gly Ile Ala Ala Ala Arg Arg Ile Ala 195 200 205 His Leu Thr Tyr Arg Gly Glu Leu Glu Ile Asp Glu Arg Phe Gly Thr 210 215 220 Ser Ala Gln His Gly Glu Asn Pro Leu Gly Pro Phe Arg Asp Pro His 225 230 235 240 Gln Arg Phe Ala Val Thr Ser Tyr Leu Gln His Gln Gly Ile Lys Leu 245 250 255 Ala Gln Arg Phe Asp Ala Gly Ser Tyr Val Val Leu Thr Glu Ala Leu 260 265 270 Asn Arg His Asp Ile Gly Arg Gly Arg Gly Gly Leu Asn Lys Ala Leu 275 280 285 Ser Ala Ile Thr Val Pro Ile Met Ile Ala Gly Val Asp Thr Asp Ile 290 295 300 Leu Tyr Pro Tyr His Gln Gln Glu His Leu Ser Arg Asn Leu Gly Asn 305 310 315 320 Leu Leu Ala Met Ala Lys Ile Ser Ser Pro Val Gly His Asp Ala Phe 325 330 335 Leu Thr Glu Phe Arg Gln Met Glu Arg Ile Leu Arg His Phe Met Glu 340 345 350 Leu Ser Glu Gly Ile Asp Asp Ser Phe Arg Thr Lys Leu Glu Arg 355 360 365 3 1149 DNA Mycobacterium leprae CDS (1)..(1146) RML02951 unsure 224 .. 224 All occurrences of n indicate any nucleotide 3 atg aca atc tcc aag gtc cct acc cag aag ctg ccg gcc gaa ggc gag 48 Met Thr Ile Ser Lys Val Pro Thr Gln Lys Leu Pro Ala Glu Gly Glu 1 5 10 15 gtc ggc ttg gtc gac atc ggc tca ctt acc acc gaa agc ggt gcc gtc 96 Val Gly Leu Val Asp Ile Gly Ser Leu Thr Thr Glu Ser Gly Ala Val 20 25 30 atc gac gat gtc tgc atc gcc gtt cag cgc tgg ggg gaa ttg tcg ccc 144 Ile Asp Asp Val Cys Ile Ala Val Gln Arg Trp Gly Glu Leu Ser Pro 35 40 45 acg cga gac aac gta gtg atg gta ctg cat gca ctc acc ggt gac tcg 192 Thr Arg Asp Asn Val Val Met Val Leu His Ala Leu Thr Gly Asp Ser 50 55 60 cac atc acc ggg ccc gcc gga ccg gga cat cnc aca ccc ggc tgg tgg 240 His Ile Thr Gly Pro Ala Gly Pro Gly His Xaa Thr Pro Gly Trp Trp 65 70 75 80 gac tgg ata gct gga ccg ggt gca cca atc gac acc aac cgc tgg tgc 288 Asp Trp Ile Ala Gly Pro Gly Ala Pro Ile Asp Thr Asn Arg Trp Cys 85 90 95 gcg ata gcc acc aac gtg ctg ggc ggt tgc cgt ggc tcc acc ggc cct 336 Ala Ile Ala Thr Asn Val Leu Gly Gly Cys Arg Gly Ser Thr Gly Pro 100 105 110 agt tcg ctt gcc cgc gac gga aag cct tgg ggt tca aga ttt ccg ctg 384 Ser Ser Leu Ala Arg Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Leu 115 120 125 ata tct ata cgc gac cag gta gag gca gat atc gct gca ctg gcc gcc 432 Ile Ser Ile Arg Asp Gln Val Glu Ala Asp Ile Ala Ala Leu Ala Ala 130 135 140 atg gga att aca aag gtt gcc gcc gtc gtt gga gga tct atg ggc ggg 480 Met Gly Ile Thr Lys Val Ala Ala Val Val Gly Gly Ser Met Gly Gly 145 150 155 160 gcg cgt gca ctg gaa tgg atc atc ggc cac ccg gac caa gtc cgg gcc 528 Ala Arg Ala Leu Glu Trp Ile Ile Gly His Pro Asp Gln Val Arg Ala 165 170 175 ggg ctg ttg ctg gcg gtc ggt gtg cgc gcc acc gcc gac cag atc ggc 576 Gly Leu Leu Leu Ala Val Gly Val Arg Ala Thr Ala Asp Gln Ile Gly 180 185 190 acc caa acc acc caa atc gca gcc atc aag aca gac ccg aac tgg caa 624 Thr Gln Thr Thr Gln Ile Ala Ala Ile Lys Thr Asp Pro Asn Trp Gln 195 200 205 ggc ggt gac tac tac gag aca ggg agg gca cca gag aac ggc ttg aca 672 Gly Gly Asp Tyr Tyr Glu Thr Gly Arg Ala Pro Glu Asn Gly Leu Thr 210 215 220 att gcc cgc cgc ttc gcc cac ctg acc tac cgc agc gag gtc gag ctc 720 Ile Ala Arg Arg Phe Ala His Leu Thr Tyr Arg Ser Glu Val Glu Leu 225 230 235 240 gac acc cgg ttt gcc aac aac aac caa ggc aat gag gac ccg gcg acg 768 Asp Thr Arg Phe Ala Asn Asn Asn Gln Gly Asn Glu Asp Pro Ala Thr 245 250 255 ggc ggg cgt tac gca gtg cag agt tac cta gag cac cag ggt gac aag 816 Gly Gly Arg Tyr Ala Val Gln Ser Tyr Leu Glu His Gln Gly Asp Lys 260 265 270 cta ttg gcc cgc ttt gac gca ggc agc tac gtg gtc ttg acc gaa acg 864 Leu Leu Ala Arg Phe Asp Ala Gly Ser Tyr Val Val Leu Thr Glu Thr 275 280 285 ctg aac agc cac gac gtt ggc cgg ggc cgc gga ggg atc ggt aca gcg 912 Leu Asn Ser His Asp Val Gly Arg Gly Arg Gly Gly Ile Gly Thr Ala 290 295 300 ctg cgc ggg tgc ccg gta ccg gtg gtg gtg ggt ggc att acc tcg gat 960 Leu Arg Gly Cys Pro Val Pro Val Val Val Gly Gly Ile Thr Ser Asp 305 310 315 320 cgg ctc tac cca ctg cgc ttg cag cag gag ctg gcc gag atg ctg ccg 1008 Arg Leu Tyr Pro Leu Arg Leu Gln Gln Glu Leu Ala Glu Met Leu Pro 325 330 335 ggc tgc acc ggg ctg cag gtt gta gac tcc acc tac ggg cac gac ggc 1 056 Gly Cys Thr Gly Leu Gln Val Val Asp Ser Thr Tyr Gly His Asp Gly 340 345 350 ttc ctg gtg gaa tcc gag gcc gtc ggc aaa ttg atc cgt caa acc ctc 1104 Phe Leu Val Glu Ser Glu Ala Val Gly Lys Leu Ile Arg Gln Thr Leu 355 360 365 gaa ttg gcc gac gtg ggt tcc aag gaa gac gcg tgt tcg caa 1146 Glu Leu Ala Asp Val Gly Ser Lys Glu Asp Ala Cys Ser Gln 370 375 380 tga 1149 4 382 PRT Mycobacterium leprae unsure 75 .. 75 All occurrences of Xaa indicate any amino acid 4 Met Thr Ile Ser Lys Val Pro Thr Gln Lys Leu Pro Ala Glu Gly Glu 1 5 10 15 Val Gly Leu Val Asp Ile Gly Ser Leu Thr Thr Glu Ser Gly Ala Val 20 25 30 Ile Asp Asp Val Cys Ile Ala Val Gln Arg Trp Gly Glu Leu Ser Pro 35 40 45 Thr Arg Asp Asn Val Val Met Val Leu His Ala Leu Thr Gly Asp Ser 50 55 60 His Ile Thr Gly Pro Ala Gly Pro Gly His Xaa Thr Pro Gly Trp Trp 65 70 75 80 Asp Trp Ile Ala Gly Pro Gly Ala Pro Ile Asp Thr Asn Arg Trp Cys 85 90 95 Ala Ile Ala Thr Asn Val Leu Gly Gly Cys Arg Gly Ser Thr Gly Pro 100 105 110 Ser Ser Leu Ala Arg Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Leu 115 120 125 Ile Ser Ile Arg Asp Gln Val Glu Ala Asp Ile Ala Ala Leu Ala Ala 130 135 140 Met Gly Ile Thr Lys Val Ala Ala Val Val Gly Gly Ser Met Gly Gly 145 150 155 160 Ala Arg Ala Leu Glu Trp Ile Ile Gly His Pro Asp Gln Val Arg Ala 165 170 175 Gly Leu Leu Leu Ala Val Gly Val Arg Ala Thr Ala Asp Gln Ile Gly 180 185 190 Thr Gln Thr Thr Gln Ile Ala Ala Ile Lys Thr Asp Pro Asn Trp Gln 195 200 205 Gly Gly Asp Tyr Tyr Glu Thr Gly Arg Ala Pro Glu Asn Gly Leu Thr 210 215 220 Ile Ala Arg Arg Phe Ala His Leu Thr Tyr Arg Ser Glu Val Glu Leu 225 230 235 240 Asp Thr Arg Phe Ala Asn Asn Asn Gln Gly Asn Glu Asp Pro Ala Thr 245 250 255 Gly Gly Arg Tyr Ala Val Gln Ser Tyr Leu Glu His Gln Gly Asp Lys 260 265 270 Leu Leu Ala Arg Phe Asp Ala Gly Ser Tyr Val Val Leu Thr Glu Thr 275 280 285 Leu Asn Ser His Asp Val Gly Arg Gly Arg Gly Gly Ile Gly Thr Ala 290 295 300 Leu Arg Gly Cys Pro Val Pro Val Val Val Gly Gly Ile Thr Ser Asp 305 310 315 320 Arg Leu Tyr Pro Leu Arg Leu Gln Gln Glu Leu Ala Glu Met Leu Pro 325 330 335 Gly Cys Thr Gly Leu Gln Val Val Asp Ser Thr Tyr Gly His Asp Gly 340 345 350 Phe Leu Val Glu Ser Glu Ala Val Gly Lys Leu Ile Arg Gln Thr Leu 355 360 365 Glu Leu Ala Asp Val Gly Ser Lys Glu Asp Ala Cys Ser Gln 370 375 380 5 1140 DNA Mycobacterium tuberculosis CDS (1)..(1137) RMTB03565 5 atg acg atc tcc gat gta ccc acc cag acg ctg ccc gcc gaa ggc gaa 48 Met Thr Ile Ser Asp Val Pro Thr Gln Thr Leu Pro Ala Glu Gly Glu 1 5 10 15 atc ggc ctg ata gac gtc ggc tcg ctg caa ctg gaa agc ggg gcg gtg 96 Ile Gly Leu Ile Asp Val Gly Ser Leu Gln Leu Glu Ser Gly Ala Val 20 25 30 atc gac gat gtc tgt atc gcc gtg caa cgc tgg ggc aaa ttg tcg ccc 144 Ile Asp Asp Val Cys Ile Ala Val Gln Arg Trp Gly Lys Leu Ser Pro 35 40 45 gca cgg gac aac gtg gtg gtg gtc ttg cac gcg ctc acc ggc gac tcg 192 Ala Arg Asp Asn Val Val Val Val Leu His Ala Leu Thr Gly Asp Ser 50 55 60 cac atc act gga ccc gcc gga ccc ggc cac ccc acc ccc ggc tgg tgg 240 His Ile Thr Gly Pro Ala Gly Pro Gly His Pro Thr Pro Gly Trp Trp 65 70 75 80 gac ggg gtg gcc ggg ccg agt gcg ccg att gac acc acc cgc tgg tgc 288 Asp Gly Val Ala Gly Pro Ser Ala Pro Ile Asp Thr Thr Arg Trp Cys 85 90 95 gcg gta gct acc aat gtg ctc ggc ggc tgc cgc ggc tcc acc ggg ccc 336 Ala Val Ala Thr Asn Val Leu Gly Gly Cys Arg Gly Ser Thr Gly Pro 100 105 110 agc tcg ctt gcc cgc gac gga aag cct tgg ggc tca aga ttt ccg ctg 384 Ser Ser Leu Ala Arg Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Leu 115 120 125 atc tcg ata cgt gac cag gtg cag gcg gac gtc gcg gcg ctg gcc gcg 432 Ile Ser Ile Arg Asp Gln Val Gln Ala Asp Val Ala Ala Leu Ala Ala 130 135 140 ctg ggc atc acc gag gtc gcc gcc gtc gtc ggc ggc tcc atg ggc ggc 480 Leu Gly Ile Thr Glu Val Ala Ala Val Val Gly Gly Ser Met Gly Gly 145 150 155 160 gcc cgg gcc ctg gaa tgg gtg gtc ggc tac ccg gat cgg gtc cga gcc 528 Ala Arg Ala Leu Glu Trp Val Val Gly Tyr Pro Asp Arg Val Arg Ala 165 170 175 gga ttg ctg ctg gcg gtc ggt gcg cgt gcc acc gca gac cag atc ggc 576 Gly Leu Leu Leu Ala Val Gly Ala Arg Ala Thr Ala Asp Gln Ile Gly 180 185 190 acg cag aca acg caa atc gcg gcc atc aaa gcc gac ccg gac tgg cag 624 Thr Gln Thr Thr Gln Ile Ala Ala Ile Lys Ala Asp Pro Asp Trp Gln 195 200 205 agc ggc gac tac cac gag acg ggg agg gca cca gac gcc ggg ctg cga 672 Ser Gly Asp Tyr His Glu Thr Gly Arg Ala Pro Asp Ala Gly Leu Arg 210 215 220 ctc gcc cgc cgc ttc gcg cac ctc acc tac cgc ggc gag atc gag ctc 720 Leu Ala Arg Arg Phe Ala His Leu Thr Tyr Arg Gly Glu Ile Glu Leu 225 230 235 240 gac acc cgg ttc gcc aac cac aac cag ggc aac gag gat ccg acg gcc 768 Asp Thr Arg Phe Ala Asn His Asn Gln Gly Asn Glu Asp Pro Thr Ala 245 250 255 ggc ggg cgc tac gcg gtg caa agt tat ctg gaa cac caa gga gac aaa 816 Gly

Gly Arg Tyr Ala Val Gln Ser Tyr Leu Glu His Gln Gly Asp Lys 260 265 270 ctg tta tcc cgg ttc gac gcc ggc agc tac gtg att ctc acc gag gcg 864 Leu Leu Ser Arg Phe Asp Ala Gly Ser Tyr Val Ile Leu Thr Glu Ala 275 280 285 ctc aac agc cac gac gtc ggc cgc ggc cgc ggc ggg gtc tcc gcg gct 912 Leu Asn Ser His Asp Val Gly Arg Gly Arg Gly Gly Val Ser Ala Ala 290 295 300 ctg cgc gcc tgc ccg gtg ccg gtg gtg gtg ggc ggc atc acc tcc gac 960 Leu Arg Ala Cys Pro Val Pro Val Val Val Gly Gly Ile Thr Ser Asp 305 310 315 320 cgg ctc tac ccg ctg cgc ctg cag cag gag ctg gcc gac ctg ctg ccg 1008 Arg Leu Tyr Pro Leu Arg Leu Gln Gln Glu Leu Ala Asp Leu Leu Pro 325 330 335 ggc tgc gcc ggg ctg cga gtc gtc gag tcg gtc tac gga cac gac ggc 1056 Gly Cys Ala Gly Leu Arg Val Val Glu Ser Val Tyr Gly His Asp Gly 340 345 350 ttc ctg gtg gaa acc gag gcc gtg ggc gaa ttg atc cgc cag aca ctg 1104 Phe Leu Val Glu Thr Glu Ala Val Gly Glu Leu Ile Arg Gln Thr Leu 355 360 365 gga ttg gct gat cgt gaa ggc gcg tgt cgg cgg tga 1140 Gly Leu Ala Asp Arg Glu Gly Ala Cys Arg Arg 370 375 6 379 PRT Mycobacterium tuberculosis 6 Met Thr Ile Ser Asp Val Pro Thr Gln Thr Leu Pro Ala Glu Gly Glu 1 5 10 15 Ile Gly Leu Ile Asp Val Gly Ser Leu Gln Leu Glu Ser Gly Ala Val 20 25 30 Ile Asp Asp Val Cys Ile Ala Val Gln Arg Trp Gly Lys Leu Ser Pro 35 40 45 Ala Arg Asp Asn Val Val Val Val Leu His Ala Leu Thr Gly Asp Ser 50 55 60 His Ile Thr Gly Pro Ala Gly Pro Gly His Pro Thr Pro Gly Trp Trp 65 70 75 80 Asp Gly Val Ala Gly Pro Ser Ala Pro Ile Asp Thr Thr Arg Trp Cys 85 90 95 Ala Val Ala Thr Asn Val Leu Gly Gly Cys Arg Gly Ser Thr Gly Pro 100 105 110 Ser Ser Leu Ala Arg Asp Gly Lys Pro Trp Gly Ser Arg Phe Pro Leu 115 120 125 Ile Ser Ile Arg Asp Gln Val Gln Ala Asp Val Ala Ala Leu Ala Ala 130 135 140 Leu Gly Ile Thr Glu Val Ala Ala Val Val Gly Gly Ser Met Gly Gly 145 150 155 160 Ala Arg Ala Leu Glu Trp Val Val Gly Tyr Pro Asp Arg Val Arg Ala 165 170 175 Gly Leu Leu Leu Ala Val Gly Ala Arg Ala Thr Ala Asp Gln Ile Gly 180 185 190 Thr Gln Thr Thr Gln Ile Ala Ala Ile Lys Ala Asp Pro Asp Trp Gln 195 200 205 Ser Gly Asp Tyr His Glu Thr Gly Arg Ala Pro Asp Ala Gly Leu Arg 210 215 220 Leu Ala Arg Arg Phe Ala His Leu Thr Tyr Arg Gly Glu Ile Glu Leu 225 230 235 240 Asp Thr Arg Phe Ala Asn His Asn Gln Gly Asn Glu Asp Pro Thr Ala 245 250 255 Gly Gly Arg Tyr Ala Val Gln Ser Tyr Leu Glu His Gln Gly Asp Lys 260 265 270 Leu Leu Ser Arg Phe Asp Ala Gly Ser Tyr Val Ile Leu Thr Glu Ala 275 280 285 Leu Asn Ser His Asp Val Gly Arg Gly Arg Gly Gly Val Ser Ala Ala 290 295 300 Leu Arg Ala Cys Pro Val Pro Val Val Val Gly Gly Ile Thr Ser Asp 305 310 315 320 Arg Leu Tyr Pro Leu Arg Leu Gln Gln Glu Leu Ala Asp Leu Leu Pro 325 330 335 Gly Cys Ala Gly Leu Arg Val Val Glu Ser Val Tyr Gly His Asp Gly 340 345 350 Phe Leu Val Glu Thr Glu Ala Val Gly Glu Leu Ile Arg Gln Thr Leu 355 360 365 Gly Leu Ala Asp Arg Glu Gly Ala Cys Arg Arg 370 375 7 972 DNA Chlorobium tepidum CDS (1)..(969) RCL01447 7 gtg agg gtc gct tac cgt acc tgg ggt acg cta aac gca gag aaa agc 48 Val Arg Val Ala Tyr Arg Thr Trp Gly Thr Leu Asn Ala Glu Lys Ser 1 5 10 15 aac gtg att ctg gtc tgc cac gcg ctg acc ggc aac gcc gac gcc gac 96 Asn Val Ile Leu Val Cys His Ala Leu Thr Gly Asn Ala Asp Ala Asp 20 25 30 agc tgg tgg tgc ggc atg ttc ggt gag gga cgg gcg ttc gac gag act 144 Ser Trp Trp Cys Gly Met Phe Gly Glu Gly Arg Ala Phe Asp Glu Thr 35 40 45 cgg gac ttc atc gta tgc agc aac gtg ctt gga agc tgc tac gga acg 192 Arg Asp Phe Ile Val Cys Ser Asn Val Leu Gly Ser Cys Tyr Gly Thr 50 55 60 acc ggg ccg atg tcg gtg aat ccg ctg agt ggc agg cac tac ggt ccc 240 Thr Gly Pro Met Ser Val Asn Pro Leu Ser Gly Arg His Tyr Gly Pro 65 70 75 80 gat ttt ccg cgc att acc att cgc gac atg gtg aat gtt cag cga tta 288 Asp Phe Pro Arg Ile Thr Ile Arg Asp Met Val Asn Val Gln Arg Leu 85 90 95 ttg ctt cgt tcg ctc ggc atc gac cgg atc cgg ctc atc gtt ggt gca 336 Leu Leu Arg Ser Leu Gly Ile Asp Arg Ile Arg Leu Ile Val Gly Ala 100 105 110 tcg ctt ggc ggg atg cag gtg ctc gaa tgg ggc gca atg tat ccc gaa 384 Ser Leu Gly Gly Met Gln Val Leu Glu Trp Gly Ala Met Tyr Pro Glu 115 120 125 atg gcc ggg gcg ctg atg ccg atg ggc gtt tcg ggt cgt cat tcg gcg 432 Met Ala Gly Ala Leu Met Pro Met Gly Val Ser Gly Arg His Ser Ala 130 135 140 tgg tgc atc gcg cag agc gag gcg cag cgg cag gct atc gcc gcc gat 480 Trp Cys Ile Ala Gln Ser Glu Ala Gln Arg Gln Ala Ile Ala Ala Asp 145 150 155 160 gcg gag tgg caa gat ggc tgg tat gat ccg gag gtg cag cca cgc aaa 528 Ala Glu Trp Gln Asp Gly Trp Tyr Asp Pro Glu Val Gln Pro Arg Lys 165 170 175 gga ctt gcc gcc gcg cgg atg atg gcg atg tgc acc tac cgc tgc ttc 576 Gly Leu Ala Ala Ala Arg Met Met Ala Met Cys Thr Tyr Arg Cys Phe 180 185 190 gag aac tac cag caa cgc ttt ggc cgc aag cag cgc gag gac ggc ttg 624 Glu Asn Tyr Gln Gln Arg Phe Gly Arg Lys Gln Arg Glu Asp Gly Leu 195 200 205 ttc gaa gcc gaa agc tac gtg cgt cac cag ggc gac aag ctg gtt ggg 672 Phe Glu Ala Glu Ser Tyr Val Arg His Gln Gly Asp Lys Leu Val Gly 210 215 220 cgc ttt gat gca aac acc tat atc acg ctc acc aga gcg atg gac atg 720 Arg Phe Asp Ala Asn Thr Tyr Ile Thr Leu Thr Arg Ala Met Asp Met 225 230 235 240 cac gac ctc ggg cgc gga cgc gac tcc tac gaa gcg gcg ctc gga gcg 768 His Asp Leu Gly Arg Gly Arg Asp Ser Tyr Glu Ala Ala Leu Gly Ala 245 250 255 ctg aag atg ccg gtc gag att ctc tcc atc gac tcg gac gtg ctc tat 816 Leu Lys Met Pro Val Glu Ile Leu Ser Ile Asp Ser Asp Val Leu Tyr 260 265 270 ccc agg cag gag cag gag gaa ctt gcc cgc ctc att ccc ggc tca cgc 864 Pro Arg Gln Glu Gln Glu Glu Leu Ala Arg Leu Ile Pro Gly Ser Arg 275 280 285 ctg ctt ttc ctt gac gaa ccc tat ggc cac gac gcc ttt ctt atc gac 912 Leu Leu Phe Leu Asp Glu Pro Tyr Gly His Asp Ala Phe Leu Ile Asp 290 295 300 acc gag acc gtc agc cgc atg gtc tgc gag ttc aag agg cag ttg ata 960 Thr Glu Thr Val Ser Arg Met Val Cys Glu Phe Lys Arg Gln Leu Ile 305 310 315 320 gtt gac aat tga 972 Val Asp Asn 8 323 PRT Chlorobium tepidum 8 Val Arg Val Ala Tyr Arg Thr Trp Gly Thr Leu Asn Ala Glu Lys Ser 1 5 10 15 Asn Val Ile Leu Val Cys His Ala Leu Thr Gly Asn Ala Asp Ala Asp 20 25 30 Ser Trp Trp Cys Gly Met Phe Gly Glu Gly Arg Ala Phe Asp Glu Thr 35 40 45 Arg Asp Phe Ile Val Cys Ser Asn Val Leu Gly Ser Cys Tyr Gly Thr 50 55 60 Thr Gly Pro Met Ser Val Asn Pro Leu Ser Gly Arg His Tyr Gly Pro 65 70 75 80 Asp Phe Pro Arg Ile Thr Ile Arg Asp Met Val Asn Val Gln Arg Leu 85 90 95 Leu Leu Arg Ser Leu Gly Ile Asp Arg Ile Arg Leu Ile Val Gly Ala 100 105 110 Ser Leu Gly Gly Met Gln Val Leu Glu Trp Gly Ala Met Tyr Pro Glu 115 120 125 Met Ala Gly Ala Leu Met Pro Met Gly Val Ser Gly Arg His Ser Ala 130 135 140 Trp Cys Ile Ala Gln Ser Glu Ala Gln Arg Gln Ala Ile Ala Ala Asp 145 150 155 160 Ala Glu Trp Gln Asp Gly Trp Tyr Asp Pro Glu Val Gln Pro Arg Lys 165 170 175 Gly Leu Ala Ala Ala Arg Met Met Ala Met Cys Thr Tyr Arg Cys Phe 180 185 190 Glu Asn Tyr Gln Gln Arg Phe Gly Arg Lys Gln Arg Glu Asp Gly Leu 195 200 205 Phe Glu Ala Glu Ser Tyr Val Arg His Gln Gly Asp Lys Leu Val Gly 210 215 220 Arg Phe Asp Ala Asn Thr Tyr Ile Thr Leu Thr Arg Ala Met Asp Met 225 230 235 240 His Asp Leu Gly Arg Gly Arg Asp Ser Tyr Glu Ala Ala Leu Gly Ala 245 250 255 Leu Lys Met Pro Val Glu Ile Leu Ser Ile Asp Ser Asp Val Leu Tyr 260 265 270 Pro Arg Gln Glu Gln Glu Glu Leu Ala Arg Leu Ile Pro Gly Ser Arg 275 280 285 Leu Leu Phe Leu Asp Glu Pro Tyr Gly His Asp Ala Phe Leu Ile Asp 290 295 300 Thr Glu Thr Val Ser Arg Met Val Cys Glu Phe Lys Arg Gln Leu Ile 305 310 315 320 Val Asp Asn 9 1149 DNA Caulobacter crescentus CDS (1)..(1146) RCO00727 9 atg gct gcg ctc gat ccg atc acg ccc gcc ggc ggg gga acc tgg cgg 48 Met Ala Ala Leu Asp Pro Ile Thr Pro Ala Gly Gly Gly Thr Trp Arg 1 5 10 15 ttt cct gcg aat gaa cct ctg cgg ctg gac tcc gga ggc gtc atc gaa 96 Phe Pro Ala Asn Glu Pro Leu Arg Leu Asp Ser Gly Gly Val Ile Glu 20 25 30 ggt ctg gaa atc gcc tac cag acc tac ggc cag ctg aac gcg gac aag 144 Gly Leu Glu Ile Ala Tyr Gln Thr Tyr Gly Gln Leu Asn Ala Asp Lys 35 40 45 tcc aac gcc gtc ctg atc tgc cac gcc ctg acg ggc gac cag cat gtg 192 Ser Asn Ala Val Leu Ile Cys His Ala Leu Thr Gly Asp Gln His Val 50 55 60 gcc tcg ccc cac ccc acc acc ggc aag ccc ggc tgg tgg caa cgc ctt 240 Ala Ser Pro His Pro Thr Thr Gly Lys Pro Gly Trp Trp Gln Arg Leu 65 70 75 80 gtt ggt ccc ggt aag ccg ctg gat ccc gcg cgg cac ttc atc atc tgc 288 Val Gly Pro Gly Lys Pro Leu Asp Pro Ala Arg His Phe Ile Ile Cys 85 90 95 tcg aac gtg atc ggc ggc tgc atg ggc tcg acg ggc ccg gcc tcg atc 336 Ser Asn Val Ile Gly Gly Cys Met Gly Ser Thr Gly Pro Ala Ser Ile 100 105 110 aat ccg gcc acg ggc aag acc tat ggc ctg tcg ttc cca gtc atc acc 384 Asn Pro Ala Thr Gly Lys Thr Tyr Gly Leu Ser Phe Pro Val Ile Thr 115 120 125 atc gcc gat atg gtg cgg gcc cag gcc atg ctg gtc tct gcg ctc ggg 432 Ile Ala Asp Met Val Arg Ala Gln Ala Met Leu Val Ser Ala Leu Gly 130 135 140 gtc gag acc ctg ttc gcc gtc gtc ggc ggc tcg atg ggc ggc atg cag 480 Val Glu Thr Leu Phe Ala Val Val Gly Gly Ser Met Gly Gly Met Gln 145 150 155 160 gtc cag caa tgg gcc gtg gac tat ccc gag cgg atg ttc agc gcc gtg 528 Val Gln Gln Trp Ala Val Asp Tyr Pro Glu Arg Met Phe Ser Ala Val 165 170 175 gtg ctg gcc tcg gcc tcg cgc cac tcg gcc cag aac atc gcg ttc cac 576 Val Leu Ala Ser Ala Ser Arg His Ser Ala Gln Asn Ile Ala Phe His 180 185 190 gag gtg ggc cgc cag gcg atc atg gcc gat ccc gac tgg cgc ggc ggc 624 Glu Val Gly Arg Gln Ala Ile Met Ala Asp Pro Asp Trp Arg Gly Gly 195 200 205 gcc tat gcc gag cac ggc gtg cgg ccc gag aag ggc ctg gcc gtg gcg 672 Ala Tyr Ala Glu His Gly Val Arg Pro Glu Lys Gly Leu Ala Val Ala 210 215 220 cgg atg gcc gcg cac atc acc tat ctg tcc gag ccc gcc ctg cag cgg 720 Arg Met Ala Ala His Ile Thr Tyr Leu Ser Glu Pro Ala Leu Gln Arg 225 230 235 240 aag ttc ggc cgc gag cta cag cgc gac ggc ctc tcc tgg ggc ttt gac 768 Lys Phe Gly Arg Glu Leu Gln Arg Asp Gly Leu Ser Trp Gly Phe Asp 245 250 255 gcc gac ttc cag gtc gag agc tat cta cgc cac cag ggg tcc agc ttc 816 Ala Asp Phe Gln Val Glu Ser Tyr Leu Arg His Gln Gly Ser Ser Phe 260 265 270 gtc gac cgg ttc gac gcc aac agc tat ctc tac atc acc cgg gcc atg 864 Val Asp Arg Phe Asp Ala Asn Ser Tyr Leu Tyr Ile Thr Arg Ala Met 275 280 285 gac tat ttc gac atc gcc gcc agc cat ggc ggg gtg ctg gcc aag gcg 912 Asp Tyr Phe Asp Ile Ala Ala Ser His Gly Gly Val Leu Ala Lys Ala 290 295 300 ttc acc cga gcg cgg aat gtg cgc ttc tgc gtg ctg agc ttc tcc agc 960 Phe Thr Arg Ala Arg Asn Val Arg Phe Cys Val Leu Ser Phe Ser Ser 305 310 315 320 gac tgg ctc tat ccg acc gcc gag aac cgc cac ctg gtc cgc gcc ctg 1008 Asp Trp Leu Tyr Pro Thr Ala Glu Asn Arg His Leu Val Arg Ala Leu 325 330 335 acc gcc gcc ggg gcc cgc gcg gcc ttc gcc gag atc gag agc gac aag 1056 Thr Ala Ala Gly Ala Arg Ala Ala Phe Ala Glu Ile Glu Ser Asp Lys 340 345 350 ggc cat gac gcc ttc ctg ctg gac gag ccg gtg atg gac gcc gcg ctg 1104 Gly His Asp Ala Phe Leu Leu Asp Glu Pro Val Met Asp Ala Ala Leu 355 360 365 gaa ggc ttc ctg gcc tcg gcc gaa cgc gat cgg ggg ctg gtt 1146 Glu Gly Phe Leu Ala Ser Ala Glu Arg Asp Arg Gly Leu Val 370 375 380 tga 1149 10 382 PRT Caulobacter crescentus 10 Met Ala Ala Leu Asp Pro Ile Thr Pro Ala Gly Gly Gly Thr Trp Arg 1 5 10 15 Phe Pro Ala Asn Glu Pro Leu Arg Leu Asp Ser Gly Gly Val Ile Glu 20 25 30 Gly Leu Glu Ile Ala Tyr Gln Thr Tyr Gly Gln Leu Asn Ala Asp Lys 35 40 45 Ser Asn Ala Val Leu Ile Cys His Ala Leu Thr Gly Asp Gln His Val 50 55 60 Ala Ser Pro His Pro Thr Thr Gly Lys Pro Gly Trp Trp Gln Arg Leu 65 70 75 80 Val Gly Pro Gly Lys Pro Leu Asp Pro Ala Arg His Phe Ile Ile Cys 85 90 95 Ser Asn Val Ile Gly Gly Cys Met Gly Ser Thr Gly Pro Ala Ser Ile 100 105 110 Asn Pro Ala Thr Gly Lys Thr Tyr Gly Leu Ser Phe Pro Val Ile Thr 115 120 125 Ile Ala Asp Met Val Arg Ala Gln Ala Met Leu Val Ser Ala Leu Gly 130 135 140 Val Glu Thr Leu Phe Ala Val Val Gly Gly Ser Met Gly Gly Met Gln 145 150 155 160 Val Gln Gln Trp Ala Val Asp Tyr Pro Glu Arg Met Phe Ser Ala Val 165 170 175 Val Leu Ala Ser Ala Ser Arg His Ser Ala Gln Asn Ile Ala Phe His 180 185 190 Glu Val Gly Arg Gln Ala Ile Met Ala Asp Pro Asp Trp Arg Gly Gly 195 200 205 Ala Tyr Ala Glu His Gly Val Arg Pro Glu Lys Gly Leu Ala Val Ala 210 215 220 Arg Met Ala Ala His Ile Thr Tyr Leu Ser Glu Pro Ala Leu Gln Arg 225 230 235 240 Lys Phe Gly Arg Glu Leu Gln Arg Asp Gly Leu Ser Trp Gly Phe Asp 245 250 255 Ala Asp Phe Gln Val Glu Ser Tyr Leu Arg His Gln Gly Ser Ser Phe 260 265 270 Val Asp Arg Phe Asp Ala Asn Ser Tyr Leu Tyr Ile Thr Arg Ala Met 275 280 285 Asp Tyr Phe Asp Ile Ala Ala Ser His Gly Gly Val Leu Ala Lys Ala 290 295 300 Phe Thr Arg Ala Arg Asn Val Arg Phe Cys Val Leu Ser Phe Ser Ser 305 310 315 320 Asp Trp Leu Tyr Pro Thr Ala Glu Asn Arg His Leu Val Arg Ala Leu 325 330 335 Thr Ala Ala Gly Ala Arg Ala Ala Phe Ala Glu Ile Glu Ser Asp Lys 340 345 350 Gly His Asp Ala Phe Leu Leu Asp Glu Pro Val Met Asp Ala Ala Leu 355 360 365 Glu Gly Phe Leu Ala Ser Ala Glu Arg Asp Arg Gly Leu Val 370

375 380 11 1140 DNA Neisseria gonorrhoeae CDS (1)..(1137) RNG00132 11 atg agt caa aat acc tcg gtg ggc att gta acg ccc caa aaa att ccg 48 Met Ser Gln Asn Thr Ser Val Gly Ile Val Thr Pro Gln Lys Ile Pro 1 5 10 15 ttt gaa atg ccg ctg gtt ttg gaa aac ggt aaa act ttg ccg cgt ttc 96 Phe Glu Met Pro Leu Val Leu Glu Asn Gly Lys Thr Leu Pro Arg Phe 20 25 30 gat ctg atg att gaa acc tac ggc gag ctg aat gct gaa aaa aac aat 144 Asp Leu Met Ile Glu Thr Tyr Gly Glu Leu Asn Ala Glu Lys Asn Asn 35 40 45 gcg gtt tta atc tgc cac gcg ctg tcg ggc aac cat cac gtt gcg ggc 192 Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val Ala Gly 50 55 60 agg cat tcg gcg gag gat aaa tat acg ggc tgg tgg gac aat atg gtc 240 Arg His Ser Ala Glu Asp Lys Tyr Thr Gly Trp Trp Asp Asn Met Val 65 70 75 80 ggt ccc gga aaa ccg att gat acg gaa cgt ttt ttc gtg gtc ggg ttg 288 Gly Pro Gly Lys Pro Ile Asp Thr Glu Arg Phe Phe Val Val Gly Leu 85 90 95 aac aat ctg ggc ggc tgc gac ggc agc agc ggg cct ttg tcg atc aat 336 Asn Asn Leu Gly Gly Cys Asp Gly Ser Ser Gly Pro Leu Ser Ile Asn 100 105 110 cct gaa acg ggc agg gaa tac ggc gcg gat ttt ccg atg gtt acg gtg 384 Pro Glu Thr Gly Arg Glu Tyr Gly Ala Asp Phe Pro Met Val Thr Val 115 120 125 aag gac tgg gta aaa tca caa gcc gcg ctt gcc gat tat ctc ggc atc 432 Lys Asp Trp Val Lys Ser Gln Ala Ala Leu Ala Asp Tyr Leu Gly Ile 130 135 140 gaa caa tgg gcg gcg gtt gtc ggc ggc agc ttg ggc ggc atg cag gct 480 Glu Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly Met Gln Ala 145 150 155 160 ttg cag tgg gcg att tcc tat ccc gaa cgt gtg cgc cac gcc ttg gtg 528 Leu Gln Trp Ala Ile Ser Tyr Pro Glu Arg Val Arg His Ala Leu Val 165 170 175 att gcg tct gcg ccg aaa ctg tcc gcg caa aat atc gcg ttt aat gat 576 Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala Phe Asn Asp 180 185 190 gta gca cgt cag gcg att ttg acc gac ccc gat ttc aat gaa gga cat 624 Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe Asn Glu Gly His 195 200 205 tac cgc agc cac aac acc gtt ccc gcg cgc ggt ttg cgg att gcc cgt 672 Tyr Arg Ser His Asn Thr Val Pro Ala Arg Gly Leu Arg Ile Ala Arg 210 215 220 atg atg gga cac att acg tat ctt gcc gaa gac ggt ttg ggc aaa aaa 720 Met Met Gly His Ile Thr Tyr Leu Ala Glu Asp Gly Leu Gly Lys Lys 225 230 235 240 ttc gga cgc gat ttg cgt tcc aac ggc tat caa tac ggc tat agc gtt 768 Phe Gly Arg Asp Leu Arg Ser Asn Gly Tyr Gln Tyr Gly Tyr Ser Val 245 250 255 gaa ttt gaa gta gaa tcc tat ctc cgc tat caa ggc gac aaa ttc gtc 816 Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys Phe Val 260 265 270 ggg cgg ttt gat gct aat aca tat ttg ctg atg acc aaa gct ttg gac 864 Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala Leu Asp 275 280 285 tat ttc gat ccg gcg gcg gat ttc ggc aac agc ctg acc cgc gcc gtg 912 Tyr Phe Asp Pro Ala Ala Asp Phe Gly Asn Ser Leu Thr Arg Ala Val 290 295 300 cag gat gtg cag gca aaa ttc ttt gtc gcc agc ttc agc acc gac tgg 960 Gln Asp Val Gln Ala Lys Phe Phe Val Ala Ser Phe Ser Thr Asp Trp 305 310 315 320 cgt ttc gcg ccc gaa cgt tcg cac gaa ctg gtc aag gca ctg att gcc 1008 Arg Phe Ala Pro Glu Arg Ser His Glu Leu Val Lys Ala Leu Ile Ala 325 330 335 gcc caa aaa tcc gtg cag tat atc gaa gtc aag tcc gca cac ggg cac 1056 Ala Gln Lys Ser Val Gln Tyr Ile Glu Val Lys Ser Ala His Gly His 340 345 350 gat gcc ttt tta atg gaa gac gaa gcc tat atg cgc gcc gta acg gct 1104 Asp Ala Phe Leu Met Glu Asp Glu Ala Tyr Met Arg Ala Val Thr Ala 355 360 365 tat atg aac aat gtt gac aag gat tgc cga tta tga 1140 Tyr Met Asn Asn Val Asp Lys Asp Cys Arg Leu 370 375 12 379 PRT Neisseria gonorrhoeae 12 Met Ser Gln Asn Thr Ser Val Gly Ile Val Thr Pro Gln Lys Ile Pro 1 5 10 15 Phe Glu Met Pro Leu Val Leu Glu Asn Gly Lys Thr Leu Pro Arg Phe 20 25 30 Asp Leu Met Ile Glu Thr Tyr Gly Glu Leu Asn Ala Glu Lys Asn Asn 35 40 45 Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val Ala Gly 50 55 60 Arg His Ser Ala Glu Asp Lys Tyr Thr Gly Trp Trp Asp Asn Met Val 65 70 75 80 Gly Pro Gly Lys Pro Ile Asp Thr Glu Arg Phe Phe Val Val Gly Leu 85 90 95 Asn Asn Leu Gly Gly Cys Asp Gly Ser Ser Gly Pro Leu Ser Ile Asn 100 105 110 Pro Glu Thr Gly Arg Glu Tyr Gly Ala Asp Phe Pro Met Val Thr Val 115 120 125 Lys Asp Trp Val Lys Ser Gln Ala Ala Leu Ala Asp Tyr Leu Gly Ile 130 135 140 Glu Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly Met Gln Ala 145 150 155 160 Leu Gln Trp Ala Ile Ser Tyr Pro Glu Arg Val Arg His Ala Leu Val 165 170 175 Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala Phe Asn Asp 180 185 190 Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe Asn Glu Gly His 195 200 205 Tyr Arg Ser His Asn Thr Val Pro Ala Arg Gly Leu Arg Ile Ala Arg 210 215 220 Met Met Gly His Ile Thr Tyr Leu Ala Glu Asp Gly Leu Gly Lys Lys 225 230 235 240 Phe Gly Arg Asp Leu Arg Ser Asn Gly Tyr Gln Tyr Gly Tyr Ser Val 245 250 255 Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys Phe Val 260 265 270 Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala Leu Asp 275 280 285 Tyr Phe Asp Pro Ala Ala Asp Phe Gly Asn Ser Leu Thr Arg Ala Val 290 295 300 Gln Asp Val Gln Ala Lys Phe Phe Val Ala Ser Phe Ser Thr Asp Trp 305 310 315 320 Arg Phe Ala Pro Glu Arg Ser His Glu Leu Val Lys Ala Leu Ile Ala 325 330 335 Ala Gln Lys Ser Val Gln Tyr Ile Glu Val Lys Ser Ala His Gly His 340 345 350 Asp Ala Phe Leu Met Glu Asp Glu Ala Tyr Met Arg Ala Val Thr Ala 355 360 365 Tyr Met Asn Asn Val Asp Lys Asp Cys Arg Leu 370 375 13 1140 DNA Neisseria meningitidis ser. A CDS (1)..(1137) RNM00815 13 atg agt caa aat gcc tcg gtg ggc att gta acg ccc caa aaa att ccg 48 Met Ser Gln Asn Ala Ser Val Gly Ile Val Thr Pro Gln Lys Ile Pro 1 5 10 15 ttt gaa atg ccg ctg gtt ttg gaa aac ggt aaa act ttg ccg cgt ttc 96 Phe Glu Met Pro Leu Val Leu Glu Asn Gly Lys Thr Leu Pro Arg Phe 20 25 30 gat ctg atg att gaa acc tac ggc gag ctg aat gcc gaa aaa aat aat 144 Asp Leu Met Ile Glu Thr Tyr Gly Glu Leu Asn Ala Glu Lys Asn Asn 35 40 45 gcg gtt tta atc tgt cat gcg ctg tca ggc aac cat cat gtt gcg ggc 192 Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val Ala Gly 50 55 60 agg cat tcg gcg gag gat aaa tat acg ggc tgg tgg gac aat atg gta 240 Arg His Ser Ala Glu Asp Lys Tyr Thr Gly Trp Trp Asp Asn Met Val 65 70 75 80 gga ccc ggc aaa ccg att gat aca gaa cgt ttt ttc gtg gtc ggt ttg 288 Gly Pro Gly Lys Pro Ile Asp Thr Glu Arg Phe Phe Val Val Gly Leu 85 90 95 aac aat ctg ggc ggc tgc gac ggc agc agc gga cct ttg tcg atc aat 336 Asn Asn Leu Gly Gly Cys Asp Gly Ser Ser Gly Pro Leu Ser Ile Asn 100 105 110 cct gaa acg ggc agg gaa tac ggc gcg gat ttt ccg gtg gtt acg gtg 384 Pro Glu Thr Gly Arg Glu Tyr Gly Ala Asp Phe Pro Val Val Thr Val 115 120 125 aag gac tgg gta aaa tcc caa gcc gcg ctt acc gat tat ctc ggc atc 432 Lys Asp Trp Val Lys Ser Gln Ala Ala Leu Thr Asp Tyr Leu Gly Ile 130 135 140 ggg caa tgg gcg gcg gtt gtc ggc ggc agc ttg ggc ggt atg cag gct 480 Gly Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly Met Gln Ala 145 150 155 160 ttg cag tgg acg att tcc tat ccc gag cgc gtg cgc cat gcc tta gtg 528 Leu Gln Trp Thr Ile Ser Tyr Pro Glu Arg Val Arg His Ala Leu Val 165 170 175 att gcg tcc gcg ccg aaa ctg tcc acg caa aat atc gcg ttt aat gat 576 Ile Ala Ser Ala Pro Lys Leu Ser Thr Gln Asn Ile Ala Phe Asn Asp 180 185 190 gta gca cgt cag gcg att ttg acc gat ccc gat ttc aac gaa gga cat 624 Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe Asn Glu Gly His 195 200 205 tac cgc agc cgc aac acc gtt ccc gct cgg ggc ttg cgg att gcc cgc 672 Tyr Arg Ser Arg Asn Thr Val Pro Ala Arg Gly Leu Arg Ile Ala Arg 210 215 220 atg atg ggg cac atc acc tat ctt gcc gaa gac ggt ttg ggc aaa aaa 720 Met Met Gly His Ile Thr Tyr Leu Ala Glu Asp Gly Leu Gly Lys Lys 225 230 235 240 ttc gga cgc gat ttg cgt tcc aac ggc tat caa tac ggc tat ggc gtt 768 Phe Gly Arg Asp Leu Arg Ser Asn Gly Tyr Gln Tyr Gly Tyr Gly Val 245 250 255 gaa ttt gaa gta gaa tcc tat ctg cgc tat caa ggc gat aaa ttc gtc 816 Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys Phe Val 260 265 270 ggg cgg ttt gat gcc aac acc tat ttg ctg atg acc aag gct ttg gac 864 Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala Leu Asp 275 280 285 tat ttc gat ccg gcg gcg gat ttc ggc aac agc ctg acc cgc gcc gtg 912 Tyr Phe Asp Pro Ala Ala Asp Phe Gly Asn Ser Leu Thr Arg Ala Val 290 295 300 cag gat gtt cag gca aaa ttc ttt gtc gcc agc ttc agc acc gat tgg 960 Gln Asp Val Gln Ala Lys Phe Phe Val Ala Ser Phe Ser Thr Asp Trp 305 310 315 320 cgt ttc gcg ccc gaa cgt tcg cac gaa ctg gtc aag gcc ctg att gcc 1008 Arg Phe Ala Pro Glu Arg Ser His Glu Leu Val Lys Ala Leu Ile Ala 325 330 335 gcc caa aaa tcc gtg cag tat atc gaa gtc aaa tcc gca cac ggg cac 1056 Ala Gln Lys Ser Val Gln Tyr Ile Glu Val Lys Ser Ala His Gly His 340 345 350 gat gcc ttt tta atg gaa gac gaa gcc tat atg cgt gcg gtc gcc gcc 1104 Asp Ala Phe Leu Met Glu Asp Glu Ala Tyr Met Arg Ala Val Ala Ala 355 360 365 tat atg aac aac gtt tat aag gaa tgt cag caa tga 1140 Tyr Met Asn Asn Val Tyr Lys Glu Cys Gln Gln 370 375 14 379 PRT Neisseria meningitidis ser. A 14 Met Ser Gln Asn Ala Ser Val Gly Ile Val Thr Pro Gln Lys Ile Pro 1 5 10 15 Phe Glu Met Pro Leu Val Leu Glu Asn Gly Lys Thr Leu Pro Arg Phe 20 25 30 Asp Leu Met Ile Glu Thr Tyr Gly Glu Leu Asn Ala Glu Lys Asn Asn 35 40 45 Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val Ala Gly 50 55 60 Arg His Ser Ala Glu Asp Lys Tyr Thr Gly Trp Trp Asp Asn Met Val 65 70 75 80 Gly Pro Gly Lys Pro Ile Asp Thr Glu Arg Phe Phe Val Val Gly Leu 85 90 95 Asn Asn Leu Gly Gly Cys Asp Gly Ser Ser Gly Pro Leu Ser Ile Asn 100 105 110 Pro Glu Thr Gly Arg Glu Tyr Gly Ala Asp Phe Pro Val Val Thr Val 115 120 125 Lys Asp Trp Val Lys Ser Gln Ala Ala Leu Thr Asp Tyr Leu Gly Ile 130 135 140 Gly Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly Met Gln Ala 145 150 155 160 Leu Gln Trp Thr Ile Ser Tyr Pro Glu Arg Val Arg His Ala Leu Val 165 170 175 Ile Ala Ser Ala Pro Lys Leu Ser Thr Gln Asn Ile Ala Phe Asn Asp 180 185 190 Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe Asn Glu Gly His 195 200 205 Tyr Arg Ser Arg Asn Thr Val Pro Ala Arg Gly Leu Arg Ile Ala Arg 210 215 220 Met Met Gly His Ile Thr Tyr Leu Ala Glu Asp Gly Leu Gly Lys Lys 225 230 235 240 Phe Gly Arg Asp Leu Arg Ser Asn Gly Tyr Gln Tyr Gly Tyr Gly Val 245 250 255 Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys Phe Val 260 265 270 Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala Leu Asp 275 280 285 Tyr Phe Asp Pro Ala Ala Asp Phe Gly Asn Ser Leu Thr Arg Ala Val 290 295 300 Gln Asp Val Gln Ala Lys Phe Phe Val Ala Ser Phe Ser Thr Asp Trp 305 310 315 320 Arg Phe Ala Pro Glu Arg Ser His Glu Leu Val Lys Ala Leu Ile Ala 325 330 335 Ala Gln Lys Ser Val Gln Tyr Ile Glu Val Lys Ser Ala His Gly His 340 345 350 Asp Ala Phe Leu Met Glu Asp Glu Ala Tyr Met Arg Ala Val Ala Ala 355 360 365 Tyr Met Asn Asn Val Tyr Lys Glu Cys Gln Gln 370 375 15 1140 DNA Pseudomonas fluorescens CDS (1)..(1137) RPU01633 15 atg cca gct gcc ttt ccc ccc gat tct gtt ggt ctg gtg acg ccg caa 48 Met Pro Ala Ala Phe Pro Pro Asp Ser Val Gly Leu Val Thr Pro Gln 1 5 10 15 acg gcg cac ttc agc gaa ccg ctg gcc ctg gcc tgc ggc cgt tcg ctg 96 Thr Ala His Phe Ser Glu Pro Leu Ala Leu Ala Cys Gly Arg Ser Leu 20 25 30 gcc gat tat gac ctg atc tac gaa acc tac ggc acg ctg aac gcg caa 144 Ala Asp Tyr Asp Leu Ile Tyr Glu Thr Tyr Gly Thr Leu Asn Ala Gln 35 40 45 gcg agc aac gcc gtg ctg atc tgc cac gcc ttg tcc ggc cac cac cat 192 Ala Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 gct gcg ggt tat cac agc gtc gac gac cgc aag ccc ggt tgg tgg gac 240 Ala Ala Gly Tyr His Ser Val Asp Asp Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 agc tgc atc ggc ccc ggc aaa ccg atc gac acc aac aag ttc ttc gtg 288 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Asn Lys Phe Phe Val 85 90 95 gtc agc ctg aac aac ctc ggc ggt tgc aat ggt tct acc ggc ccg agc 336 Val Ser Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Thr Gly Pro Ser 100 105 110 agc ctc aat ccg gaa acc ggc aag ccg ttc ggc gcc gac ttc ccg gtg 384 Ser Leu Asn Pro Glu Thr Gly Lys Pro Phe Gly Ala Asp Phe Pro Val 115 120 125 ctg acc gtg gaa gac tgg gtg cac agc cag gca cgc ctg gcc gac ctg 432 Leu Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Leu 130 135 140 ctc ggc atc ggc cag tgg gcg gcg gtg atc ggc ggc agc ctg ggc ggc 480 Leu Gly Ile Gly Gln Trp Ala Ala Val Ile Gly Gly Ser Leu Gly Gly 145 150 155 160 atg cag gcg ctg caa tgg acc atc acc tat ccg gat cgc gtt cgc cac 528 Met Gln Ala Leu Gln Trp Thr Ile Thr Tyr Pro Asp Arg Val Arg His 165 170 175 tgc ctg gcc atc gcc tcg gcc ccc aag ctg tcg gcg cag aac atc gcc 576 Cys Leu Ala Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 ttc aac gaa gtg gcg cgc cag gcg atc ctc act gac ccg gaa ttc cac 624 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Glu Phe His 195 200 205 ggc ggc tcg ttc cag gaa cac ggc gtg atc ccc aag cgc ggc ctg atg 672 Gly Gly Ser Phe Gln Glu His Gly Val Ile Pro Lys Arg Gly Leu Met 210 215 220 ctg gcg cgg atg gtg ggg cac atc acc tac ctg tcc gac gac tcc atg 720 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met 225 230 235 240 ggt gag aaa ttc ggc cgt ggc ctg aag agc gaa aag ctc aac tac gac 768 Gly Glu Lys Phe Gly Arg Gly Leu Lys Ser Glu Lys Leu Asn Tyr Asp 245 250 255 ttc cac agc gtc gag ttc cag gtc gaa agc tac ctg cgc tat cag ggc 816 Phe His Ser Val Glu Phe Gln Val Glu Ser Tyr Leu Arg Tyr Gln

Gly 260 265 270 gaa gag ttc tcc ggg cgc ttc gat gcc aac acc tat ctg ttg atg acc 864 Glu Glu Phe Ser Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr 275 280 285 aag gcg ctg gac tac ttc gat ccg gcg gcg aac ttc aac gat aac ctg 912 Lys Ala Leu Asp Tyr Phe Asp Pro Ala Ala Asn Phe Asn Asp Asn Leu 290 295 300 gcg aaa acc ttc gaa ggt gca aaa gcc aag ttc tgc gtg atg tcg ttc 960 Ala Lys Thr Phe Glu Gly Ala Lys Ala Lys Phe Cys Val Met Ser Phe 305 310 315 320 acc acc gac tgg cgc ttc tcc ccg gcc cgc tcg cga gaa ctg gtg gat 1008 Thr Thr Asp Trp Arg Phe Ser Pro Ala Arg Ser Arg Glu Leu Val Asp 325 330 335 gcg ctg atg gcg gcg cgc aaa gac gtc agc tac ctg gaa atc gac gcg 1056 Ala Leu Met Ala Ala Arg Lys Asp Val Ser Tyr Leu Glu Ile Asp Ala 340 345 350 ccc cag ggc cac gac gcc ttc ctg att ccg atc ccg cgc tac ttg cag 1104 Pro Gln Gly His Asp Ala Phe Leu Ile Pro Ile Pro Arg Tyr Leu Gln 355 360 365 gcg ttc ggc aat tac atg aac cgc att acg ttg tga 1140 Ala Phe Gly Asn Tyr Met Asn Arg Ile Thr Leu 370 375 16 379 PRT Pseudomonas fluorescens 16 Met Pro Ala Ala Phe Pro Pro Asp Ser Val Gly Leu Val Thr Pro Gln 1 5 10 15 Thr Ala His Phe Ser Glu Pro Leu Ala Leu Ala Cys Gly Arg Ser Leu 20 25 30 Ala Asp Tyr Asp Leu Ile Tyr Glu Thr Tyr Gly Thr Leu Asn Ala Gln 35 40 45 Ala Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 Ala Ala Gly Tyr His Ser Val Asp Asp Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Asn Lys Phe Phe Val 85 90 95 Val Ser Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Thr Gly Pro Ser 100 105 110 Ser Leu Asn Pro Glu Thr Gly Lys Pro Phe Gly Ala Asp Phe Pro Val 115 120 125 Leu Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Leu 130 135 140 Leu Gly Ile Gly Gln Trp Ala Ala Val Ile Gly Gly Ser Leu Gly Gly 145 150 155 160 Met Gln Ala Leu Gln Trp Thr Ile Thr Tyr Pro Asp Arg Val Arg His 165 170 175 Cys Leu Ala Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Glu Phe His 195 200 205 Gly Gly Ser Phe Gln Glu His Gly Val Ile Pro Lys Arg Gly Leu Met 210 215 220 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met 225 230 235 240 Gly Glu Lys Phe Gly Arg Gly Leu Lys Ser Glu Lys Leu Asn Tyr Asp 245 250 255 Phe His Ser Val Glu Phe Gln Val Glu Ser Tyr Leu Arg Tyr Gln Gly 260 265 270 Glu Glu Phe Ser Gly Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr 275 280 285 Lys Ala Leu Asp Tyr Phe Asp Pro Ala Ala Asn Phe Asn Asp Asn Leu 290 295 300 Ala Lys Thr Phe Glu Gly Ala Lys Ala Lys Phe Cys Val Met Ser Phe 305 310 315 320 Thr Thr Asp Trp Arg Phe Ser Pro Ala Arg Ser Arg Glu Leu Val Asp 325 330 335 Ala Leu Met Ala Ala Arg Lys Asp Val Ser Tyr Leu Glu Ile Asp Ala 340 345 350 Pro Gln Gly His Asp Ala Phe Leu Ile Pro Ile Pro Arg Tyr Leu Gln 355 360 365 Ala Phe Gly Asn Tyr Met Asn Arg Ile Thr Leu 370 375 17 1140 DNA Pseudomonas aeruginosa CDS (1)..(1137) RPA04460 17 atg ccc aca gtc ttc ccc gac gac tcc gtc ggt ctg gtc tcc ccc cag 48 Met Pro Thr Val Phe Pro Asp Asp Ser Val Gly Leu Val Ser Pro Gln 1 5 10 15 acg ctg cac ttc aac gaa ccg ctc gag ctg acc agc ggc aag tcc ctg 96 Thr Leu His Phe Asn Glu Pro Leu Glu Leu Thr Ser Gly Lys Ser Leu 20 25 30 gcc gag tac gac ctg gtg atc gaa acc tac ggc gag ctg aat gcc acg 144 Ala Glu Tyr Asp Leu Val Ile Glu Thr Tyr Gly Glu Leu Asn Ala Thr 35 40 45 cag agc aac gcg gtg ctg atc tgc cac gcc ctc tcc ggc cac cac cac 192 Gln Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 gcc gcc ggc tac cac agc gtc gac gag cgc aag ccg ggc tgg tgg gac 240 Ala Ala Gly Tyr His Ser Val Asp Glu Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 agc tgc atc ggt ccg ggc aag ccg atc gac acc cgc aag ttc ttc gtc 288 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Arg Lys Phe Phe Val 85 90 95 gtc gcc ctc aac aac ctc ggc ggt tgc aac gga tcc agc ggc ccc gcc 336 Val Ala Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Ser Gly Pro Ala 100 105 110 agc atc aat ccg gcg acc ggc aag gtc tac ggc gcg gac ttc ccg atg 384 Ser Ile Asn Pro Ala Thr Gly Lys Val Tyr Gly Ala Asp Phe Pro Met 115 120 125 gtt acg gtg gaa gac tgg gtg cat agc cag gcg cgc ctg gca gac cgc 432 Val Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Arg 130 135 140 ctc ggc atc cgc cag tgg gcc gcg gtg gtc ggc ggc agc ctc ggc ggc 480 Leu Gly Ile Arg Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly 145 150 155 160 atg cag gcg ctg caa tgg acc atc agc tat ccc gag cgc gtc cgt cac 528 Met Gln Ala Leu Gln Trp Thr Ile Ser Tyr Pro Glu Arg Val Arg His 165 170 175 tgc ctg tgc atc gcc agc gcg ccg aag ctg tcg gcg cag aac atc gcc 576 Cys Leu Cys Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 ttc aac gaa gtc gcc cgg cag gcg att ctt tcc gac cct gag ttc ctc 624 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Ser Asp Pro Glu Phe Leu 195 200 205 ggc ggc tac ttc cag gag cag ggc gtg att ccc aag cgc ggc ctc aag 672 Gly Gly Tyr Phe Gln Glu Gln Gly Val Ile Pro Lys Arg Gly Leu Lys 210 215 220 ctg gcg cgg atg gtc ggc cat atc acc tac ctg tcc gac gac gcc atg 720 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ala Met 225 230 235 240 ggc gcc aag ttc ggc cgt gta ctg aag acc gag aag ctc aac tac gac 768 Gly Ala Lys Phe Gly Arg Val Leu Lys Thr Glu Lys Leu Asn Tyr Asp 245 250 255 ctg cac agc gtc gag ttc cag gtc gag agt tac ctg cgc tac cag ggc 816 Leu His Ser Val Glu Phe Gln Val Glu Ser Tyr Leu Arg Tyr Gln Gly 260 265 270 gag gag ttc tcc acc cgc ttc gac gcc aat acc tac ctg ctg atg acc 864 Glu Glu Phe Ser Thr Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr 275 280 285 aag gcg ctg gac tac ttc gac ccc gcc gcc gcc cac ggc gac gac ctg 912 Lys Ala Leu Asp Tyr Phe Asp Pro Ala Ala Ala His Gly Asp Asp Leu 290 295 300 gtg cgc acc ctg gag ggc gtc gag gcg gac ttc tgc ctg atg tcc ttc 960 Val Arg Thr Leu Glu Gly Val Glu Ala Asp Phe Cys Leu Met Ser Phe 305 310 315 320 acc acc gac tgg cgt ttc tcg ccg gcc cgc tcg cgg gaa atc gtc gac 1008 Thr Thr Asp Trp Arg Phe Ser Pro Ala Arg Ser Arg Glu Ile Val Asp 325 330 335 gcc ctg atc gcg gcg aaa aag aac gtc agc tac ctg gag atc gac gcc 1056 Ala Leu Ile Ala Ala Lys Lys Asn Val Ser Tyr Leu Glu Ile Asp Ala 340 345 350 ccg caa ggc cac gac gcc ttc ctc atg ccg atc ccc cgg tac ctg caa 1104 Pro Gln Gly His Asp Ala Phe Leu Met Pro Ile Pro Arg Tyr Leu Gln 355 360 365 gcc ttc agc ggt tac atg aac cgc atc agc gtg tga 1140 Ala Phe Ser Gly Tyr Met Asn Arg Ile Ser Val 370 375 18 379 PRT Pseudomonas aeruginosa 18 Met Pro Thr Val Phe Pro Asp Asp Ser Val Gly Leu Val Ser Pro Gln 1 5 10 15 Thr Leu His Phe Asn Glu Pro Leu Glu Leu Thr Ser Gly Lys Ser Leu 20 25 30 Ala Glu Tyr Asp Leu Val Ile Glu Thr Tyr Gly Glu Leu Asn Ala Thr 35 40 45 Gln Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 Ala Ala Gly Tyr His Ser Val Asp Glu Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Arg Lys Phe Phe Val 85 90 95 Val Ala Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Ser Gly Pro Ala 100 105 110 Ser Ile Asn Pro Ala Thr Gly Lys Val Tyr Gly Ala Asp Phe Pro Met 115 120 125 Val Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Arg 130 135 140 Leu Gly Ile Arg Gln Trp Ala Ala Val Val Gly Gly Ser Leu Gly Gly 145 150 155 160 Met Gln Ala Leu Gln Trp Thr Ile Ser Tyr Pro Glu Arg Val Arg His 165 170 175 Cys Leu Cys Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Ser Asp Pro Glu Phe Leu 195 200 205 Gly Gly Tyr Phe Gln Glu Gln Gly Val Ile Pro Lys Arg Gly Leu Lys 210 215 220 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ala Met 225 230 235 240 Gly Ala Lys Phe Gly Arg Val Leu Lys Thr Glu Lys Leu Asn Tyr Asp 245 250 255 Leu His Ser Val Glu Phe Gln Val Glu Ser Tyr Leu Arg Tyr Gln Gly 260 265 270 Glu Glu Phe Ser Thr Arg Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr 275 280 285 Lys Ala Leu Asp Tyr Phe Asp Pro Ala Ala Ala His Gly Asp Asp Leu 290 295 300 Val Arg Thr Leu Glu Gly Val Glu Ala Asp Phe Cys Leu Met Ser Phe 305 310 315 320 Thr Thr Asp Trp Arg Phe Ser Pro Ala Arg Ser Arg Glu Ile Val Asp 325 330 335 Ala Leu Ile Ala Ala Lys Lys Asn Val Ser Tyr Leu Glu Ile Asp Ala 340 345 350 Pro Gln Gly His Asp Ala Phe Leu Met Pro Ile Pro Arg Tyr Leu Gln 355 360 365 Ala Phe Ser Gly Tyr Met Asn Arg Ile Ser Val 370 375 19 1146 DNA Burkholderia cepacia CDS (1)..(1143) RBU12675 19 atg gaa tcg atc ggt atc gtc gct ccc caa aaa atg cat ttc acc gag 48 Met Glu Ser Ile Gly Ile Val Ala Pro Gln Lys Met His Phe Thr Glu 1 5 10 15 ccg ctg ccg ttg cag aac ggc agt tcg ctc gcc ggt tac gac ctg atg 96 Pro Leu Pro Leu Gln Asn Gly Ser Ser Leu Ala Gly Tyr Asp Leu Met 20 25 30 gtc gag acc tac ggc acg ctc aac gcc gcg cgt agc aac gcg gtg ctg 144 Val Glu Thr Tyr Gly Thr Leu Asn Ala Ala Arg Ser Asn Ala Val Leu 35 40 45 gtg tgc cac gcg ctc aac gcg tcg cac cac gtg gcg ggc gtg tat gcc 192 Val Cys His Ala Leu Asn Ala Ser His His Val Ala Gly Val Tyr Ala 50 55 60 gac aac ccc agg gac atc ggc tgg tgg gac aac atg gtc ggc ccg ggc 240 Asp Asn Pro Arg Asp Ile Gly Trp Trp Asp Asn Met Val Gly Pro Gly 65 70 75 80 aag ccg ctc gac act gac aag ttc ttc gtg atc ggc gtg aac aac ctc 288 Lys Pro Leu Asp Thr Asp Lys Phe Phe Val Ile Gly Val Asn Asn Leu 85 90 95 gga tcg tgc ttc ggc tcg act ggg ccg atg agc atc gat ccg tct acc 336 Gly Ser Cys Phe Gly Ser Thr Gly Pro Met Ser Ile Asp Pro Ser Thr 100 105 110 ggc aat ccg tac ggc gcg acg ttt ccc gtc gtg acg gtg gaa gac tgg 384 Gly Asn Pro Tyr Gly Ala Thr Phe Pro Val Val Thr Val Glu Asp Trp 115 120 125 gtc aac gcc cag gcg cgc gtc gcg gat caa ttc ggc atc acg cgc ttt 432 Val Asn Ala Gln Ala Arg Val Ala Asp Gln Phe Gly Ile Thr Arg Phe 130 135 140 gcg gcg gtg atg ggc ggc agc ctc ggc ggc atg cag gcg ctc gcg tgg 480 Ala Ala Val Met Gly Gly Ser Leu Gly Gly Met Gln Ala Leu Ala Trp 145 150 155 160 agc atg atg tat ccg gag cgc gtc gct cac tgc atc gtg gtc gcg tcc 528 Ser Met Met Tyr Pro Glu Arg Val Ala His Cys Ile Val Val Ala Ser 165 170 175 aca ccc aag ctg tcg gcg cag aac atc gcg ttc aac gag gtt gcg cgc 576 Thr Pro Lys Leu Ser Ala Gln Asn Ile Ala Phe Asn Glu Val Ala Arg 180 185 190 tcg gcg atc ctg tcg gac ccg gac ttc cac ggc ggc aac tac tac gcg 624 Ser Ala Ile Leu Ser Asp Pro Asp Phe His Gly Gly Asn Tyr Tyr Ala 195 200 205 cac aac gtt aag ccg aag cgc ggc ctg cgc gtc gcg cgc atg atc ggc 672 His Asn Val Lys Pro Lys Arg Gly Leu Arg Val Ala Arg Met Ile Gly 210 215 220 cac atc acg tat ctg tcg gac gac gac atg gcc gag aaa ttc ggc cgc 720 His Ile Thr Tyr Leu Ser Asp Asp Asp Met Ala Glu Lys Phe Gly Arg 225 230 235 240 tcg ctg cgg cgc gcg gaa ggc gcg ctg gac gcg tac aac ttc aac ttc 768 Ser Leu Arg Arg Ala Glu Gly Ala Leu Asp Ala Tyr Asn Phe Asn Phe 245 250 255 gac gtg gag ttc gag gtg gag tcg tac ctg cgc tac cag ggc gac aag 816 Asp Val Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys 260 265 270 ttc gcc gac tac ttc gac gcg aat acg tat ctg ctg atc acc cgc gcg 864 Phe Ala Asp Tyr Phe Asp Ala Asn Thr Tyr Leu Leu Ile Thr Arg Ala 275 280 285 ctc gac tac ttc gat ccg gcc aag gcc ttc gcc ggc gac ctg acg gcc 912 Leu Asp Tyr Phe Asp Pro Ala Lys Ala Phe Ala Gly Asp Leu Thr Ala 290 295 300 gcg gtc gcg cac acc acg gcg aaa tat ctg atc gcc agc ttc acg acc 960 Ala Val Ala His Thr Thr Ala Lys Tyr Leu Ile Ala Ser Phe Thr Thr 305 310 315 320 gac tgg cgc ttc gcg ccg gcc cgc tcg cgt gaa ctg gtg aag gcg ctg 1008 Asp Trp Arg Phe Ala Pro Ala Arg Ser Arg Glu Leu Val Lys Ala Leu 325 330 335 ctc gat cac aag cgc acg gtc acc tac gcg gaa atc gac gcg ccg cac 1056 Leu Asp His Lys Arg Thr Val Thr Tyr Ala Glu Ile Asp Ala Pro His 340 345 350 ggc cac gac gcc ttc ctg ctc gac gac gcg cgc tat cac aac ctg atg 1104 Gly His Asp Ala Phe Leu Leu Asp Asp Ala Arg Tyr His Asn Leu Met 355 360 365 cgc gct tac tac gaa cgt att gcg aac gag gtg aac gca tga 1146 Arg Ala Tyr Tyr Glu Arg Ile Ala Asn Glu Val Asn Ala 370 375 380 20 381 PRT Burkholderia cepacia 20 Met Glu Ser Ile Gly Ile Val Ala Pro Gln Lys Met His Phe Thr Glu 1 5 10 15 Pro Leu Pro Leu Gln Asn Gly Ser Ser Leu Ala Gly Tyr Asp Leu Met 20 25 30 Val Glu Thr Tyr Gly Thr Leu Asn Ala Ala Arg Ser Asn Ala Val Leu 35 40 45 Val Cys His Ala Leu Asn Ala Ser His His Val Ala Gly Val Tyr Ala 50 55 60 Asp Asn Pro Arg Asp Ile Gly Trp Trp Asp Asn Met Val Gly Pro Gly 65 70 75 80 Lys Pro Leu Asp Thr Asp Lys Phe Phe Val Ile Gly Val Asn Asn Leu 85 90 95 Gly Ser Cys Phe Gly Ser Thr Gly Pro Met Ser Ile Asp Pro Ser Thr 100 105 110 Gly Asn Pro Tyr Gly Ala Thr Phe Pro Val Val Thr Val Glu Asp Trp 115 120 125 Val Asn Ala Gln Ala Arg Val Ala Asp Gln Phe Gly Ile Thr Arg Phe 130 135 140 Ala Ala Val Met Gly Gly Ser Leu Gly Gly Met Gln Ala Leu Ala Trp 145 150 155 160 Ser Met Met Tyr Pro Glu Arg Val Ala His Cys Ile Val Val Ala Ser 165 170 175 Thr Pro Lys Leu Ser Ala Gln Asn Ile Ala Phe Asn Glu Val Ala Arg 180 185 190 Ser Ala Ile Leu Ser Asp Pro Asp Phe His Gly Gly Asn Tyr Tyr Ala 195 200 205 His Asn Val Lys Pro Lys Arg Gly Leu Arg Val Ala Arg Met Ile Gly 210 215 220 His Ile Thr Tyr Leu Ser Asp Asp Asp Met Ala Glu Lys Phe Gly Arg 225 230 235 240 Ser Leu Arg Arg Ala Glu Gly Ala Leu Asp Ala Tyr Asn Phe Asn Phe

245 250 255 Asp Val Glu Phe Glu Val Glu Ser Tyr Leu Arg Tyr Gln Gly Asp Lys 260 265 270 Phe Ala Asp Tyr Phe Asp Ala Asn Thr Tyr Leu Leu Ile Thr Arg Ala 275 280 285 Leu Asp Tyr Phe Asp Pro Ala Lys Ala Phe Ala Gly Asp Leu Thr Ala 290 295 300 Ala Val Ala His Thr Thr Ala Lys Tyr Leu Ile Ala Ser Phe Thr Thr 305 310 315 320 Asp Trp Arg Phe Ala Pro Ala Arg Ser Arg Glu Leu Val Lys Ala Leu 325 330 335 Leu Asp His Lys Arg Thr Val Thr Tyr Ala Glu Ile Asp Ala Pro His 340 345 350 Gly His Asp Ala Phe Leu Leu Asp Asp Ala Arg Tyr His Asn Leu Met 355 360 365 Arg Ala Tyr Tyr Glu Arg Ile Ala Asn Glu Val Asn Ala 370 375 380 21 1134 DNA Nitrosomonas europaea CDS (1)..(1131) RNE02005 21 atg tcc aca caa gat tct gat tcg atc ggc atc gta tcg gca cga cgc 48 Met Ser Thr Gln Asp Ser Asp Ser Ile Gly Ile Val Ser Ala Arg Arg 1 5 10 15 gcc cat ttc gac acc ccg ctc agc ctg aaa agc gga gct gta ctg gac 96 Ala His Phe Asp Thr Pro Leu Ser Leu Lys Ser Gly Ala Val Leu Asp 20 25 30 agc tac gag ctc gtc tat gaa acc tat ggg gag ctg aat gca gac cga 144 Ser Tyr Glu Leu Val Tyr Glu Thr Tyr Gly Glu Leu Asn Ala Asp Arg 35 40 45 tcc aat gca gtg ctg atc tgc cat gct tta tcc ggc aac cac cat gtt 192 Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val 50 55 60 gcc ggt gtt tat gca gat aac ccc aag aat acc gga tgg tgg aac aac 240 Ala Gly Val Tyr Ala Asp Asn Pro Lys Asn Thr Gly Trp Trp Asn Asn 65 70 75 80 atg atc ggt ccg ggc aaa ccg gtc gat acc cga aaa ttc ttt gtc atc 288 Met Ile Gly Pro Gly Lys Pro Val Asp Thr Arg Lys Phe Phe Val Ile 85 90 95 ggt atc aat aat ctc ggg ggt tgc cat ggc tcc acc ggg ccc atc agc 336 Gly Ile Asn Asn Leu Gly Gly Cys His Gly Ser Thr Gly Pro Ile Ser 100 105 110 atc aac gac aag acc ggt aaa cgc ttc ggc ccg gat ttt ccg ctg gta 384 Ile Asn Asp Lys Thr Gly Lys Arg Phe Gly Pro Asp Phe Pro Leu Val 115 120 125 acg aca gct gac tgg gca aaa acc tat gtc cgt ttc gcc gat cag ttc 432 Thr Thr Ala Asp Trp Ala Lys Thr Tyr Val Arg Phe Ala Asp Gln Phe 130 135 140 agc atc gac tgt ttt gcc gcc gtc atc ggt ggc agt ctg ggc ggg atg 480 Ser Ile Asp Cys Phe Ala Ala Val Ile Gly Gly Ser Leu Gly Gly Met 145 150 155 160 tcg gcc atg caa ctg gcg ctc gat gca ccg gaa aga gtt cgt cat gcc 528 Ser Ala Met Gln Leu Ala Leu Asp Ala Pro Glu Arg Val Arg His Ala 165 170 175 ata gtg gtt gca gca tcg gcc agg ctg aca gca cag aac atc gct ttc 576 Ile Val Val Ala Ala Ser Ala Arg Leu Thr Ala Gln Asn Ile Ala Phe 180 185 190 aat gat gtc gcg cgt cag gcg att ctg acc gac cct gat ttt cac gac 624 Asn Asp Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe His Asp 195 200 205 ggc gac tat tat tcc cat ggc acc cac ccg cgc aga ggt tta cgc ctt 672 Gly Asp Tyr Tyr Ser His Gly Thr His Pro Arg Arg Gly Leu Arg Leu 210 215 220 gcc cgc atg ctt ggc cac atc acc tac ctg tcg gac gac tcc atg gcc 720 Ala Arg Met Leu Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met Ala 225 230 235 240 agc aaa ttc ggc cgt gag tta cgt aac ggc tcg ctt gct ttc aat tat 768 Ser Lys Phe Gly Arg Glu Leu Arg Asn Gly Ser Leu Ala Phe Asn Tyr 245 250 255 gat gtg gaa ttc cag atc gaa tcc tat ctg cac cat cag ggc gac aaa 816 Asp Val Glu Phe Gln Ile Glu Ser Tyr Leu His His Gln Gly Asp Lys 260 265 270 ttt gcc gac ctg ttc gac gca aac act tat ctg ctg atg acg aag gcg 864 Phe Ala Asp Leu Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala 275 280 285 ctc gat tat ttc gat ccg gcc cag gat tac gat ggc aac ctg agt gca 912 Leu Asp Tyr Phe Asp Pro Ala Gln Asp Tyr Asp Gly Asn Leu Ser Ala 290 295 300 gcc ttt gcc cgt gca caa gcg gat ttt ctg gta ctt tcc ttt act tcc 960 Ala Phe Ala Arg Ala Gln Ala Asp Phe Leu Val Leu Ser Phe Thr Ser 305 310 315 320 gac tgg cgt ttt tcc ccg gag cgt tcg cgc gat atc gtc aag gca ctg 1008 Asp Trp Arg Phe Ser Pro Glu Arg Ser Arg Asp Ile Val Lys Ala Leu 325 330 335 ctc gac aac aaa ctg aat gtc agt tat gcg gaa att ccc tcc tcg tac 1056 Leu Asp Asn Lys Leu Asn Val Ser Tyr Ala Glu Ile Pro Ser Ser Tyr 340 345 350 gga cat gat tcc ttt ctc atg cag gac gac tac tat cac cag ttg ata 1104 Gly His Asp Ser Phe Leu Met Gln Asp Asp Tyr Tyr His Gln Leu Ile 355 360 365 cgt gct tac atg aac aat atc gct ctc tag 1134 Arg Ala Tyr Met Asn Asn Ile Ala Leu 370 375 22 377 PRT Nitrosomonas europaea 22 Met Ser Thr Gln Asp Ser Asp Ser Ile Gly Ile Val Ser Ala Arg Arg 1 5 10 15 Ala His Phe Asp Thr Pro Leu Ser Leu Lys Ser Gly Ala Val Leu Asp 20 25 30 Ser Tyr Glu Leu Val Tyr Glu Thr Tyr Gly Glu Leu Asn Ala Asp Arg 35 40 45 Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly Asn His His Val 50 55 60 Ala Gly Val Tyr Ala Asp Asn Pro Lys Asn Thr Gly Trp Trp Asn Asn 65 70 75 80 Met Ile Gly Pro Gly Lys Pro Val Asp Thr Arg Lys Phe Phe Val Ile 85 90 95 Gly Ile Asn Asn Leu Gly Gly Cys His Gly Ser Thr Gly Pro Ile Ser 100 105 110 Ile Asn Asp Lys Thr Gly Lys Arg Phe Gly Pro Asp Phe Pro Leu Val 115 120 125 Thr Thr Ala Asp Trp Ala Lys Thr Tyr Val Arg Phe Ala Asp Gln Phe 130 135 140 Ser Ile Asp Cys Phe Ala Ala Val Ile Gly Gly Ser Leu Gly Gly Met 145 150 155 160 Ser Ala Met Gln Leu Ala Leu Asp Ala Pro Glu Arg Val Arg His Ala 165 170 175 Ile Val Val Ala Ala Ser Ala Arg Leu Thr Ala Gln Asn Ile Ala Phe 180 185 190 Asn Asp Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Asp Phe His Asp 195 200 205 Gly Asp Tyr Tyr Ser His Gly Thr His Pro Arg Arg Gly Leu Arg Leu 210 215 220 Ala Arg Met Leu Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met Ala 225 230 235 240 Ser Lys Phe Gly Arg Glu Leu Arg Asn Gly Ser Leu Ala Phe Asn Tyr 245 250 255 Asp Val Glu Phe Gln Ile Glu Ser Tyr Leu His His Gln Gly Asp Lys 260 265 270 Phe Ala Asp Leu Phe Asp Ala Asn Thr Tyr Leu Leu Met Thr Lys Ala 275 280 285 Leu Asp Tyr Phe Asp Pro Ala Gln Asp Tyr Asp Gly Asn Leu Ser Ala 290 295 300 Ala Phe Ala Arg Ala Gln Ala Asp Phe Leu Val Leu Ser Phe Thr Ser 305 310 315 320 Asp Trp Arg Phe Ser Pro Glu Arg Ser Arg Asp Ile Val Lys Ala Leu 325 330 335 Leu Asp Asn Lys Leu Asn Val Ser Tyr Ala Glu Ile Pro Ser Ser Tyr 340 345 350 Gly His Asp Ser Phe Leu Met Gln Asp Asp Tyr Tyr His Gln Leu Ile 355 360 365 Arg Ala Tyr Met Asn Asn Ile Ala Leu 370 375 23 1077 DNA Haemophilus influenzae CDS (1)..(1074) RHI02681 23 atg tct gtg caa aat gta gtg ctt ttt gac aca cag cct tta act ctg 48 Met Ser Val Gln Asn Val Val Leu Phe Asp Thr Gln Pro Leu Thr Leu 1 5 10 15 atg ctt ggc ggc aaa ctt tcc cat att aat gtc gcg tat caa act tat 96 Met Leu Gly Gly Lys Leu Ser His Ile Asn Val Ala Tyr Gln Thr Tyr 20 25 30 ggc acg ctc aat gcc gaa aaa aat aat gcg gta tta att tgc cac gct 144 Gly Thr Leu Asn Ala Glu Lys Asn Asn Ala Val Leu Ile Cys His Ala 35 40 45 ttg act ggt gat gct gag cct tat ttc gat gat ggt cga gat ggc tgg 192 Leu Thr Gly Asp Ala Glu Pro Tyr Phe Asp Asp Gly Arg Asp Gly Trp 50 55 60 tgg cag aat ttt atg gga gca ggt tta gca ttg gat acg gat cgt tat 240 Trp Gln Asn Phe Met Gly Ala Gly Leu Ala Leu Asp Thr Asp Arg Tyr 65 70 75 80 ttt ttt att agc tcg aac gta tta ggt ggt tgc aag gga aca act ggg 288 Phe Phe Ile Ser Ser Asn Val Leu Gly Gly Cys Lys Gly Thr Thr Gly 85 90 95 cct tca tca att aat ccg caa acg ggt aaa cct tat ggc agc caa ttt 336 Pro Ser Ser Ile Asn Pro Gln Thr Gly Lys Pro Tyr Gly Ser Gln Phe 100 105 110 cct aat att gtt gtg caa gat att gtt aaa gta caa aaa gcc ttg ctt 384 Pro Asn Ile Val Val Gln Asp Ile Val Lys Val Gln Lys Ala Leu Leu 115 120 125 gat cat ctt ggt att agc cat tta aaa gcc att att ggt gga tct ttt 432 Asp His Leu Gly Ile Ser His Leu Lys Ala Ile Ile Gly Gly Ser Phe 130 135 140 ggc ggc atg caa gcg aat caa tgg gcg att gat tat cct gat ttt atg 480 Gly Gly Met Gln Ala Asn Gln Trp Ala Ile Asp Tyr Pro Asp Phe Met 145 150 155 160 gat aat atc gtg aat ctt tgc tca tcc att tat ttt agt gct gaa gcc 528 Asp Asn Ile Val Asn Leu Cys Ser Ser Ile Tyr Phe Ser Ala Glu Ala 165 170 175 ata ggt ttt aat cac gta atg cgt caa gcg gtc att aat gat ccc aat 576 Ile Gly Phe Asn His Val Met Arg Gln Ala Val Ile Asn Asp Pro Asn 180 185 190 ttt aac ggc ggc gat tat tat gag ggt aca ccg cca gat caa ggg tta 624 Phe Asn Gly Gly Asp Tyr Tyr Glu Gly Thr Pro Pro Asp Gln Gly Leu 195 200 205 tct att gca cgt atg cta ggt atg ctg act tac cgc acc gat tta caa 672 Ser Ile Ala Arg Met Leu Gly Met Leu Thr Tyr Arg Thr Asp Leu Gln 210 215 220 ctt gcg aaa gcc ttt ggg cgt gcc aca aaa tca gat ggc agc ttt tgg 720 Leu Ala Lys Ala Phe Gly Arg Ala Thr Lys Ser Asp Gly Ser Phe Trp 225 230 235 240 ggc gat tac ttt caa gtg gaa tcc tat ctt tct tac caa ggc aaa aaa 768 Gly Asp Tyr Phe Gln Val Glu Ser Tyr Leu Ser Tyr Gln Gly Lys Lys 245 250 255 ttc tta gaa cgt ttt gat gcc aat agt tat ttg cat ttg tta cgt gcg 816 Phe Leu Glu Arg Phe Asp Ala Asn Ser Tyr Leu His Leu Leu Arg Ala 260 265 270 ttg gat atg tat gat cca agt ttg ggg tat gac aat gtt aaa gag gca 864 Leu Asp Met Tyr Asp Pro Ser Leu Gly Tyr Asp Asn Val Lys Glu Ala 275 280 285 tta tca cgt att aaa gca cgc tat acg ttg gtt tct gtg aca acg gat 912 Leu Ser Arg Ile Lys Ala Arg Tyr Thr Leu Val Ser Val Thr Thr Asp 290 295 300 caa ctt ttt aaa ccc att gat ctt tat aaa agt aaa cag ctt tta gag 960 Gln Leu Phe Lys Pro Ile Asp Leu Tyr Lys Ser Lys Gln Leu Leu Glu 305 310 315 320 caa agt gga gtc gat cta cat ttt tat gaa ttc cca tca gat tac gga 1008 Gln Ser Gly Val Asp Leu His Phe Tyr Glu Phe Pro Ser Asp Tyr Gly 325 330 335 cac gat gcg ttt tta gtg gat tat gat cag ttt gaa aaa cga att cga 1056 His Asp Ala Phe Leu Val Asp Tyr Asp Gln Phe Glu Lys Arg Ile Arg 340 345 350 gat ggt ttg gca ggt aat taa 1077 Asp Gly Leu Ala Gly Asn 355 24 358 PRT Haemophilus influenzae 24 Met Ser Val Gln Asn Val Val Leu Phe Asp Thr Gln Pro Leu Thr Leu 1 5 10 15 Met Leu Gly Gly Lys Leu Ser His Ile Asn Val Ala Tyr Gln Thr Tyr 20 25 30 Gly Thr Leu Asn Ala Glu Lys Asn Asn Ala Val Leu Ile Cys His Ala 35 40 45 Leu Thr Gly Asp Ala Glu Pro Tyr Phe Asp Asp Gly Arg Asp Gly Trp 50 55 60 Trp Gln Asn Phe Met Gly Ala Gly Leu Ala Leu Asp Thr Asp Arg Tyr 65 70 75 80 Phe Phe Ile Ser Ser Asn Val Leu Gly Gly Cys Lys Gly Thr Thr Gly 85 90 95 Pro Ser Ser Ile Asn Pro Gln Thr Gly Lys Pro Tyr Gly Ser Gln Phe 100 105 110 Pro Asn Ile Val Val Gln Asp Ile Val Lys Val Gln Lys Ala Leu Leu 115 120 125 Asp His Leu Gly Ile Ser His Leu Lys Ala Ile Ile Gly Gly Ser Phe 130 135 140 Gly Gly Met Gln Ala Asn Gln Trp Ala Ile Asp Tyr Pro Asp Phe Met 145 150 155 160 Asp Asn Ile Val Asn Leu Cys Ser Ser Ile Tyr Phe Ser Ala Glu Ala 165 170 175 Ile Gly Phe Asn His Val Met Arg Gln Ala Val Ile Asn Asp Pro Asn 180 185 190 Phe Asn Gly Gly Asp Tyr Tyr Glu Gly Thr Pro Pro Asp Gln Gly Leu 195 200 205 Ser Ile Ala Arg Met Leu Gly Met Leu Thr Tyr Arg Thr Asp Leu Gln 210 215 220 Leu Ala Lys Ala Phe Gly Arg Ala Thr Lys Ser Asp Gly Ser Phe Trp 225 230 235 240 Gly Asp Tyr Phe Gln Val Glu Ser Tyr Leu Ser Tyr Gln Gly Lys Lys 245 250 255 Phe Leu Glu Arg Phe Asp Ala Asn Ser Tyr Leu His Leu Leu Arg Ala 260 265 270 Leu Asp Met Tyr Asp Pro Ser Leu Gly Tyr Asp Asn Val Lys Glu Ala 275 280 285 Leu Ser Arg Ile Lys Ala Arg Tyr Thr Leu Val Ser Val Thr Thr Asp 290 295 300 Gln Leu Phe Lys Pro Ile Asp Leu Tyr Lys Ser Lys Gln Leu Leu Glu 305 310 315 320 Gln Ser Gly Val Asp Leu His Phe Tyr Glu Phe Pro Ser Asp Tyr Gly 325 330 335 His Asp Ala Phe Leu Val Asp Tyr Asp Gln Phe Glu Lys Arg Ile Arg 340 345 350 Asp Gly Leu Ala Gly Asn 355 25 1296 DNA Halobacterium sp CDS (1)..(1293) ETX_HALN1 25 atg ggc cac gat cac gga ctc cac acc aac agt gta cac gcc ggc cag 48 Met Gly His Asp His Gly Leu His Thr Asn Ser Val His Ala Gly Gln 1 5 10 15 cgc gtc gac ccg gcc acg ggc gct cgc gcg ccg cca ctc tac cag acc 96 Arg Val Asp Pro Ala Thr Gly Ala Arg Ala Pro Pro Leu Tyr Gln Thr 20 25 30 acg tcg tac gcc ttc gag gac agc gcc gat gcc gcc ggc cag ttc gcc 144 Thr Ser Tyr Ala Phe Glu Asp Ser Ala Asp Ala Ala Gly Gln Phe Ala 35 40 45 ctt gag cgg gac ggc tac atc tac tcg cgg ctg atg aac ccc acc gtg 192 Leu Glu Arg Asp Gly Tyr Ile Tyr Ser Arg Leu Met Asn Pro Thr Val 50 55 60 gag acc ctc cag gac cgc ctc gcc gcc ctc gaa ggc ggc gtc ggc gcg 240 Glu Thr Leu Gln Asp Arg Leu Ala Ala Leu Glu Gly Gly Val Gly Ala 65 70 75 80 gtc gcc acc gcg tcc gga atg gcc gcc ctg gac ctc gcg acg ttc ctg 288 Val Ala Thr Ala Ser Gly Met Ala Ala Leu Asp Leu Ala Thr Phe Leu 85 90 95 ctg gca cgc gcc ggc gac tcc gtc gtc gcc gcc agc gac ctc tac ggc 336 Leu Ala Arg Ala Gly Asp Ser Val Val Ala Ala Ser Asp Leu Tyr Gly 100 105 110 ggc acc gtg acg tac ctc acg cac agc gcc cag cgc cgc ggc gtc gac 384 Gly Thr Val Thr Tyr Leu Thr His Ser Ala Gln Arg Arg Gly Val Asp 115 120 125 acg acg ttc gtg gac gtc ctc gac tac gac gcc tac gcc gac gcc atc 432 Thr Thr Phe Val Asp Val Leu Asp Tyr Asp Ala Tyr Ala Asp Ala Ile 130 135 140 gac gcc gac acc gcc tac gtg ctc gtc gaa acc gtc ggc aac ccc agc 480 Asp Ala Asp Thr Ala Tyr Val Leu Val Glu Thr Val Gly Asn Pro Ser 145 150 155 160 ctg atc acg ccc gac ctc gaa cgc atc gcc gac atc gcc cac gac aac 528 Leu Ile Thr Pro Asp Leu Glu Arg Ile Ala Asp Ile Ala His Asp Asn 165 170 175 ggc gtt ccc ctg ctg gtg gac aac acg ttc gcg acc ccc gcg ctg gca 576 Gly Val Pro Leu Leu Val Asp Asn Thr Phe Ala Thr Pro Ala Leu Ala 180 185 190 acc ccg atc gac cac ggt gcc gac atc gtc tgg cac tcc acc acc aaa 624 Thr Pro Ile Asp His Gly Ala Asp Ile Val Trp His Ser Thr Thr Lys 195 200 205 tgg atc cac ggt gcc ggc acc acc gtc ggc ggc gcg ctc gtc gac gcc 672 Trp Ile His Gly Ala Gly Thr Thr Val Gly Gly Ala Leu Val Asp Ala 210 215

220 ggc agc ttc gac tgg gac gcc cac gcc gcc gac tac ccc gag atc gcc 720 Gly Ser Phe Asp Trp Asp Ala His Ala Ala Asp Tyr Pro Glu Ile Ala 225 230 235 240 cag gaa aac ccc gcc tac cac ggc gtg acc ttc acc gat cgc ttc ggg 768 Gln Glu Asn Pro Ala Tyr His Gly Val Thr Phe Thr Asp Arg Phe Gly 245 250 255 gac gcc gcg ttc acg tac gcc gca atc gcc cgc ggg ctg cgc gat ctg 816 Asp Ala Ala Phe Thr Tyr Ala Ala Ile Ala Arg Gly Leu Arg Asp Leu 260 265 270 ggc aac cag cag tcg ccg ttc gac gcc tgg cag acc ctc cag aag ctc 864 Gly Asn Gln Gln Ser Pro Phe Asp Ala Trp Gln Thr Leu Gln Lys Leu 275 280 285 gaa acg ctc ccg ctg cgc atg caa caa cac tgc cgg aac gcc cag ctc 912 Glu Thr Leu Pro Leu Arg Met Gln Gln His Cys Arg Asn Ala Gln Leu 290 295 300 gtc gcc gaa cac ctc cgg gac cac ccc aac gtg tcg tgg gtc aac tac 960 Val Ala Glu His Leu Arg Asp His Pro Asn Val Ser Trp Val Asn Tyr 305 310 315 320 ccc ggg ctg gcc gac cac gac acc cac gac aac gca acc acc tac ctc 1008 Pro Gly Leu Ala Asp His Asp Thr His Asp Asn Ala Thr Thr Tyr Leu 325 330 335 gat tcg ggc tac gga ggc atg ctc acg ttc ggc gtc gag gac ggc tac 1056 Asp Ser Gly Tyr Gly Gly Met Leu Thr Phe Gly Val Glu Asp Gly Tyr 340 345 350 gag gcc gcc caa tcg gtc acc gag gag acc acg ctt gcc agc ctg ctg 1104 Glu Ala Ala Gln Ser Val Thr Glu Glu Thr Thr Leu Ala Ser Leu Leu 355 360 365 gcg aac gtc ggc gac gcc aaa acg ctc gtg atc cac ccc gcc tcc acc 1152 Ala Asn Val Gly Asp Ala Lys Thr Leu Val Ile His Pro Ala Ser Thr 370 375 380 acc cac cag cag ctc acc ccc gaa gcc cag cgc gcc ggc ggt gtg cgc 1200 Thr His Gln Gln Leu Thr Pro Glu Ala Gln Arg Ala Gly Gly Val Arg 385 390 395 400 ccc gag atg gtg cgg gtg tcg gtc ggc atc gag gac ccc gcc gac atc 1248 Pro Glu Met Val Arg Val Ser Val Gly Ile Glu Asp Pro Ala Asp Ile 405 410 415 gtc gcg gac ctc gaa acc gcc atc gag gcc gcg gtc ggg tcg gcg 1293 Val Ala Asp Leu Glu Thr Ala Ile Glu Ala Ala Val Gly Ser Ala 420 425 430 tag 1296 26 431 PRT Halobacterium sp 26 Met Gly His Asp His Gly Leu His Thr Asn Ser Val His Ala Gly Gln 1 5 10 15 Arg Val Asp Pro Ala Thr Gly Ala Arg Ala Pro Pro Leu Tyr Gln Thr 20 25 30 Thr Ser Tyr Ala Phe Glu Asp Ser Ala Asp Ala Ala Gly Gln Phe Ala 35 40 45 Leu Glu Arg Asp Gly Tyr Ile Tyr Ser Arg Leu Met Asn Pro Thr Val 50 55 60 Glu Thr Leu Gln Asp Arg Leu Ala Ala Leu Glu Gly Gly Val Gly Ala 65 70 75 80 Val Ala Thr Ala Ser Gly Met Ala Ala Leu Asp Leu Ala Thr Phe Leu 85 90 95 Leu Ala Arg Ala Gly Asp Ser Val Val Ala Ala Ser Asp Leu Tyr Gly 100 105 110 Gly Thr Val Thr Tyr Leu Thr His Ser Ala Gln Arg Arg Gly Val Asp 115 120 125 Thr Thr Phe Val Asp Val Leu Asp Tyr Asp Ala Tyr Ala Asp Ala Ile 130 135 140 Asp Ala Asp Thr Ala Tyr Val Leu Val Glu Thr Val Gly Asn Pro Ser 145 150 155 160 Leu Ile Thr Pro Asp Leu Glu Arg Ile Ala Asp Ile Ala His Asp Asn 165 170 175 Gly Val Pro Leu Leu Val Asp Asn Thr Phe Ala Thr Pro Ala Leu Ala 180 185 190 Thr Pro Ile Asp His Gly Ala Asp Ile Val Trp His Ser Thr Thr Lys 195 200 205 Trp Ile His Gly Ala Gly Thr Thr Val Gly Gly Ala Leu Val Asp Ala 210 215 220 Gly Ser Phe Asp Trp Asp Ala His Ala Ala Asp Tyr Pro Glu Ile Ala 225 230 235 240 Gln Glu Asn Pro Ala Tyr His Gly Val Thr Phe Thr Asp Arg Phe Gly 245 250 255 Asp Ala Ala Phe Thr Tyr Ala Ala Ile Ala Arg Gly Leu Arg Asp Leu 260 265 270 Gly Asn Gln Gln Ser Pro Phe Asp Ala Trp Gln Thr Leu Gln Lys Leu 275 280 285 Glu Thr Leu Pro Leu Arg Met Gln Gln His Cys Arg Asn Ala Gln Leu 290 295 300 Val Ala Glu His Leu Arg Asp His Pro Asn Val Ser Trp Val Asn Tyr 305 310 315 320 Pro Gly Leu Ala Asp His Asp Thr His Asp Asn Ala Thr Thr Tyr Leu 325 330 335 Asp Ser Gly Tyr Gly Gly Met Leu Thr Phe Gly Val Glu Asp Gly Tyr 340 345 350 Glu Ala Ala Gln Ser Val Thr Glu Glu Thr Thr Leu Ala Ser Leu Leu 355 360 365 Ala Asn Val Gly Asp Ala Lys Thr Leu Val Ile His Pro Ala Ser Thr 370 375 380 Thr His Gln Gln Leu Thr Pro Glu Ala Gln Arg Ala Gly Gly Val Arg 385 390 395 400 Pro Glu Met Val Arg Val Ser Val Gly Ile Glu Asp Pro Ala Asp Ile 405 410 415 Val Ala Asp Leu Glu Thr Ala Ile Glu Ala Ala Val Gly Ser Ala 420 425 430 27 1143 DNA Thermus thermophilus CDS (1)..(1140) RTT00268 27 atg agc gag atc gcc ctc gag gcc tgg ggg gag cac gag gcc ctc ctc 48 Met Ser Glu Ile Ala Leu Glu Ala Trp Gly Glu His Glu Ala Leu Leu 1 5 10 15 ctc aag ccc ccc cgc tcc ccc ctc tcc atc ccc ccg ccc aag ccc cgc 96 Leu Lys Pro Pro Arg Ser Pro Leu Ser Ile Pro Pro Pro Lys Pro Arg 20 25 30 acc gcc gtc ctc ttc ccc agg cgg gag ggg ttc tac acg gag ctc ggg 144 Thr Ala Val Leu Phe Pro Arg Arg Glu Gly Phe Tyr Thr Glu Leu Gly 35 40 45 ggg tac ctc ccc gag gtg cgc ctc cgc ttt gag acc tac ggg acc ctc 192 Gly Tyr Leu Pro Glu Val Arg Leu Arg Phe Glu Thr Tyr Gly Thr Leu 50 55 60 tcc cgc agg cgg gat aac gcc gtc ctc gtc ttc cac gcc ctc acg ggg 240 Ser Arg Arg Arg Asp Asn Ala Val Leu Val Phe His Ala Leu Thr Gly 65 70 75 80 agc gcc cac ctg gcg ggg acc tac gac gag gaa acc ttt aga agc ctc 288 Ser Ala His Leu Ala Gly Thr Tyr Asp Glu Glu Thr Phe Arg Ser Leu 85 90 95 tcc ccc ctg gag cag gcc ttc ggc cgg gaa ggg tgg tgg gac agc ctg 336 Ser Pro Leu Glu Gln Ala Phe Gly Arg Glu Gly Trp Trp Asp Ser Leu 100 105 110 gtg ggg ccc ggg cgg atc ctg gac ccc gcc ctc tac tac gtg gtc tcc 384 Val Gly Pro Gly Arg Ile Leu Asp Pro Ala Leu Tyr Tyr Val Val Ser 115 120 125 gcc aac cac ctg gga agc tgc tac ggc tcc acc ggc ccc ctc tcc cta 432 Ala Asn His Leu Gly Ser Cys Tyr Gly Ser Thr Gly Pro Leu Ser Leu 130 135 140 gac ccc cac acg ggc cgc ccc tac ggg agg gac ttc cct ccc ctt acc 480 Asp Pro His Thr Gly Arg Pro Tyr Gly Arg Asp Phe Pro Pro Leu Thr 145 150 155 160 atc cgc gac ctg gcc cgg gcc cag gcg agg ctt ctg gac cat ctg ggg 528 Ile Arg Asp Leu Ala Arg Ala Gln Ala Arg Leu Leu Asp His Leu Gly 165 170 175 gtg gag aag gcc atc gtc atc ggg ggg agc ctc ggg ggg atg gtg gcc 576 Val Glu Lys Ala Ile Val Ile Gly Gly Ser Leu Gly Gly Met Val Ala 180 185 190 ctg gag ttc gcc ctc atg tac ccg gag agg gtg aag aag ctc gtg gtc 624 Leu Glu Phe Ala Leu Met Tyr Pro Glu Arg Val Lys Lys Leu Val Val 195 200 205 ctg gcg gcc ccc gca cgg cac ggc ccc tgg gcc cgg gcc ttc aac cac 672 Leu Ala Ala Pro Ala Arg His Gly Pro Trp Ala Arg Ala Phe Asn His 210 215 220 ctc tcc cgc cag gcc atc ctc caa gac ccc gag tac cag aag ggc aac 720 Leu Ser Arg Gln Ala Ile Leu Gln Asp Pro Glu Tyr Gln Lys Gly Asn 225 230 235 240 cct gcc ccc aag ggc atg gcc ctc gcc cgg gga atc gcc atg atg agc 768 Pro Ala Pro Lys Gly Met Ala Leu Ala Arg Gly Ile Ala Met Met Ser 245 250 255 tac cgg gcc ccc gag ggg ttt gag gcc cgc tgg ggc gcg gag ccc gag 816 Tyr Arg Ala Pro Glu Gly Phe Glu Ala Arg Trp Gly Ala Glu Pro Glu 260 265 270 ctc ggg gaa atc cac ctg gac tac cag ggg gag aag ttc ctc cgg cgc 864 Leu Gly Glu Ile His Leu Asp Tyr Gln Gly Glu Lys Phe Leu Arg Arg 275 280 285 ttc cac gcc gag agc tac ctc gtc ctc tcc cgg gcc atg gac aac cac 912 Phe His Ala Glu Ser Tyr Leu Val Leu Ser Arg Ala Met Asp Asn His 290 295 300 gac gtg ggc cgg ggc cgg ggc ggg gtg gag gag gcc ctg aag cgc ctc 960 Asp Val Gly Arg Gly Arg Gly Gly Val Glu Glu Ala Leu Lys Arg Leu 305 310 315 320 agg gcc atc ccc tcc ctc ttc gtg ggc att gac acc gac ctc ctc tac 1008 Arg Ala Ile Pro Ser Leu Phe Val Gly Ile Asp Thr Asp Leu Leu Tyr 325 330 335 ccc gcc tgg gag gtg agg cag gcg gcc aag gcg gcg ggg gcc cgc tac 1056 Pro Ala Trp Glu Val Arg Gln Ala Ala Lys Ala Ala Gly Ala Arg Tyr 340 345 350 cgg gag atc aaa agc ccc cac ggg cac gac gcc ttc ctc ata gag acc 1104 Arg Glu Ile Lys Ser Pro His Gly His Asp Ala Phe Leu Ile Glu Thr 355 360 365 gac cag gtg gag gag atc ctg gac gcc ttc ctc ccg tag 1143 Asp Gln Val Glu Glu Ile Leu Asp Ala Phe Leu Pro 370 375 380 28 380 PRT Thermus thermophilus 28 Met Ser Glu Ile Ala Leu Glu Ala Trp Gly Glu His Glu Ala Leu Leu 1 5 10 15 Leu Lys Pro Pro Arg Ser Pro Leu Ser Ile Pro Pro Pro Lys Pro Arg 20 25 30 Thr Ala Val Leu Phe Pro Arg Arg Glu Gly Phe Tyr Thr Glu Leu Gly 35 40 45 Gly Tyr Leu Pro Glu Val Arg Leu Arg Phe Glu Thr Tyr Gly Thr Leu 50 55 60 Ser Arg Arg Arg Asp Asn Ala Val Leu Val Phe His Ala Leu Thr Gly 65 70 75 80 Ser Ala His Leu Ala Gly Thr Tyr Asp Glu Glu Thr Phe Arg Ser Leu 85 90 95 Ser Pro Leu Glu Gln Ala Phe Gly Arg Glu Gly Trp Trp Asp Ser Leu 100 105 110 Val Gly Pro Gly Arg Ile Leu Asp Pro Ala Leu Tyr Tyr Val Val Ser 115 120 125 Ala Asn His Leu Gly Ser Cys Tyr Gly Ser Thr Gly Pro Leu Ser Leu 130 135 140 Asp Pro His Thr Gly Arg Pro Tyr Gly Arg Asp Phe Pro Pro Leu Thr 145 150 155 160 Ile Arg Asp Leu Ala Arg Ala Gln Ala Arg Leu Leu Asp His Leu Gly 165 170 175 Val Glu Lys Ala Ile Val Ile Gly Gly Ser Leu Gly Gly Met Val Ala 180 185 190 Leu Glu Phe Ala Leu Met Tyr Pro Glu Arg Val Lys Lys Leu Val Val 195 200 205 Leu Ala Ala Pro Ala Arg His Gly Pro Trp Ala Arg Ala Phe Asn His 210 215 220 Leu Ser Arg Gln Ala Ile Leu Gln Asp Pro Glu Tyr Gln Lys Gly Asn 225 230 235 240 Pro Ala Pro Lys Gly Met Ala Leu Ala Arg Gly Ile Ala Met Met Ser 245 250 255 Tyr Arg Ala Pro Glu Gly Phe Glu Ala Arg Trp Gly Ala Glu Pro Glu 260 265 270 Leu Gly Glu Ile His Leu Asp Tyr Gln Gly Glu Lys Phe Leu Arg Arg 275 280 285 Phe His Ala Glu Ser Tyr Leu Val Leu Ser Arg Ala Met Asp Asn His 290 295 300 Asp Val Gly Arg Gly Arg Gly Gly Val Glu Glu Ala Leu Lys Arg Leu 305 310 315 320 Arg Ala Ile Pro Ser Leu Phe Val Gly Ile Asp Thr Asp Leu Leu Tyr 325 330 335 Pro Ala Trp Glu Val Arg Gln Ala Ala Lys Ala Ala Gly Ala Arg Tyr 340 345 350 Arg Glu Ile Lys Ser Pro His Gly His Asp Ala Phe Leu Ile Glu Thr 355 360 365 Asp Gln Val Glu Glu Ile Leu Asp Ala Phe Leu Pro 370 375 380 29 1005 DNA Deinococcus radiodurans CDS (1)..(1002) RDR01287 29 gtg acc gcc gtg ctc gcg ggc cac gcc tct gcc ctg ctg ctg acc gaa 48 Val Thr Ala Val Leu Ala Gly His Ala Ser Ala Leu Leu Leu Thr Glu 1 5 10 15 gaa ccc gac tgt tcg ggg ccg cag acg gtc gtt ctc ttc cgg cgt gag 96 Glu Pro Asp Cys Ser Gly Pro Gln Thr Val Val Leu Phe Arg Arg Glu 20 25 30 ccg ctg ctg ctc gac tgc gga cgg gcg ctg agc gac gtg cgg gtg gcc 144 Pro Leu Leu Leu Asp Cys Gly Arg Ala Leu Ser Asp Val Arg Val Ala 35 40 45 ttt cac acc tac ggc acg ccg cgc gcc gac gcc acg ctg gtg ctg cac 192 Phe His Thr Tyr Gly Thr Pro Arg Ala Asp Ala Thr Leu Val Leu His 50 55 60 gcc ctg acc ggc gac agc gcg gtg cac gag tgg tgg ccc gac ttt ctg 240 Ala Leu Thr Gly Asp Ser Ala Val His Glu Trp Trp Pro Asp Phe Leu 65 70 75 80 ggc gcg ggc cgg cca ctg gac ccg gca gac gac tac gtg gtg tgc gcc 288 Gly Ala Gly Arg Pro Leu Asp Pro Ala Asp Asp Tyr Val Val Cys Ala 85 90 95 aac gtc ctc ggc ggg tgc gcc ggc acg acg agc gcc gct gaa ctc gcc 336 Asn Val Leu Gly Gly Cys Ala Gly Thr Thr Ser Ala Ala Glu Leu Ala 100 105 110 gcc acc tgt tcc gga ccg gtg ccg ctc agc ctg cgc gac atg gcc cgg 384 Ala Thr Cys Ser Gly Pro Val Pro Leu Ser Leu Arg Asp Met Ala Arg 115 120 125 gtg ggg cgc gcc ctg ctg gat tct ctc ggc gtg cga cgg gtg cgg gtc 432 Val Gly Arg Ala Leu Leu Asp Ser Leu Gly Val Arg Arg Val Arg Val 130 135 140 atc ggc gcg agc atg ggc ggg atg ctc gcc tac gcc tgg ctg ctg gag 480 Ile Gly Ala Ser Met Gly Gly Met Leu Ala Tyr Ala Trp Leu Leu Glu 145 150 155 160 tgc ccc gac ctg gtg gaa aag gcc gtg att ata gga gcc ccg gcg cgg 528 Cys Pro Asp Leu Val Glu Lys Ala Val Ile Ile Gly Ala Pro Ala Arg 165 170 175 cac tcg ccc tgg gct att gga ctg aac acg gcg gcc cgc agc gcc att 576 His Ser Pro Trp Ala Ile Gly Leu Asn Thr Ala Ala Arg Ser Ala Ile 180 185 190 gcc ctc gct ccc ggc ggc gag ggg ctg aag gtg gcg cgc cag att gcc 624 Ala Leu Ala Pro Gly Gly Glu Gly Leu Lys Val Ala Arg Gln Ile Ala 195 200 205 atg ctc agt tac cgc agc ccc gaa agc cta agc cgc acg cag gcg ggg 672 Met Leu Ser Tyr Arg Ser Pro Glu Ser Leu Ser Arg Thr Gln Ala Gly 210 215 220 cag cgc gtg ccg ggg gtg ccc gcc gtt acg tct tac ctg cac tac caa 720 Gln Arg Val Pro Gly Val Pro Ala Val Thr Ser Tyr Leu His Tyr Gln 225 230 235 240 ggc gaa aaa ctc gcc gcc cgc ttc gac gag cag acc tac tgc gcc ctc 768 Gly Glu Lys Leu Ala Ala Arg Phe Asp Glu Gln Thr Tyr Cys Ala Leu 245 250 255 acc tgg gcg atg gac gcc ttt cag ccg agc agc gcc gac ctc aaa gcg 816 Thr Trp Ala Met Asp Ala Phe Gln Pro Ser Ser Ala Asp Leu Lys Ala 260 265 270 gtg cgc gcg ccg gta ctc gtc gtc ggc atc tcc agc gat ctg ctc tac 864 Val Arg Ala Pro Val Leu Val Val Gly Ile Ser Ser Asp Leu Leu Tyr 275 280 285 ccc gcc gcc gag gtc cgc gcc tgc gcc gcc gag ctt ccc cac gcc gac 912 Pro Ala Ala Glu Val Arg Ala Cys Ala Ala Glu Leu Pro His Ala Asp 290 295 300 tac tgg gaa ctg ggc agc att cac ggc cac gac gcc ttt ttg atg gac 960 Tyr Trp Glu Leu Gly Ser Ile His Gly His Asp Ala Phe Leu Met Asp 305 310 315 320 cca cag gac ttg ccg gag cgg gtg ggg gcg ttt ctc agg agt 1002 Pro Gln Asp Leu Pro Glu Arg Val Gly Ala Phe Leu Arg Ser 325 330 tga 1005 30 334 PRT Deinococcus radiodurans 30 Val Thr Ala Val Leu Ala Gly His Ala Ser Ala Leu Leu Leu Thr Glu 1 5 10 15 Glu Pro Asp Cys Ser Gly Pro Gln Thr Val Val Leu Phe Arg Arg Glu 20 25 30 Pro Leu Leu Leu Asp Cys Gly Arg Ala Leu Ser Asp Val Arg Val Ala 35 40 45 Phe His Thr Tyr Gly Thr Pro Arg Ala Asp Ala Thr Leu Val Leu His 50 55 60 Ala Leu Thr Gly Asp Ser Ala Val His Glu Trp Trp Pro Asp Phe Leu 65 70 75 80 Gly Ala Gly Arg Pro Leu Asp Pro Ala Asp Asp Tyr Val Val Cys Ala 85 90

95 Asn Val Leu Gly Gly Cys Ala Gly Thr Thr Ser Ala Ala Glu Leu Ala 100 105 110 Ala Thr Cys Ser Gly Pro Val Pro Leu Ser Leu Arg Asp Met Ala Arg 115 120 125 Val Gly Arg Ala Leu Leu Asp Ser Leu Gly Val Arg Arg Val Arg Val 130 135 140 Ile Gly Ala Ser Met Gly Gly Met Leu Ala Tyr Ala Trp Leu Leu Glu 145 150 155 160 Cys Pro Asp Leu Val Glu Lys Ala Val Ile Ile Gly Ala Pro Ala Arg 165 170 175 His Ser Pro Trp Ala Ile Gly Leu Asn Thr Ala Ala Arg Ser Ala Ile 180 185 190 Ala Leu Ala Pro Gly Gly Glu Gly Leu Lys Val Ala Arg Gln Ile Ala 195 200 205 Met Leu Ser Tyr Arg Ser Pro Glu Ser Leu Ser Arg Thr Gln Ala Gly 210 215 220 Gln Arg Val Pro Gly Val Pro Ala Val Thr Ser Tyr Leu His Tyr Gln 225 230 235 240 Gly Glu Lys Leu Ala Ala Arg Phe Asp Glu Gln Thr Tyr Cys Ala Leu 245 250 255 Thr Trp Ala Met Asp Ala Phe Gln Pro Ser Ser Ala Asp Leu Lys Ala 260 265 270 Val Arg Ala Pro Val Leu Val Val Gly Ile Ser Ser Asp Leu Leu Tyr 275 280 285 Pro Ala Ala Glu Val Arg Ala Cys Ala Ala Glu Leu Pro His Ala Asp 290 295 300 Tyr Trp Glu Leu Gly Ser Ile His Gly His Asp Ala Phe Leu Met Asp 305 310 315 320 Pro Gln Asp Leu Pro Glu Arg Val Gly Ala Phe Leu Arg Ser 325 330 31 1461 DNA Saccharomyces cerevisiae CDS (1)..(1458) RSC08123 31 atg tcg cat act tta aaa tcg aaa acg ctc caa gag ctg gac att gag 48 Met Ser His Thr Leu Lys Ser Lys Thr Leu Gln Glu Leu Asp Ile Glu 1 5 10 15 gag att aag gaa act aac cca ttg ctc aaa cta gtt caa ggg cag agg 96 Glu Ile Lys Glu Thr Asn Pro Leu Leu Lys Leu Val Gln Gly Gln Arg 20 25 30 att gtt caa gtt ccg gaa cta gtg ctt gag tct ggc gtg gtc ata aat 144 Ile Val Gln Val Pro Glu Leu Val Leu Glu Ser Gly Val Val Ile Asn 35 40 45 aat ttc cct att gct tat aag acg tgg ggt aca ctg aat gaa gct ggt 192 Asn Phe Pro Ile Ala Tyr Lys Thr Trp Gly Thr Leu Asn Glu Ala Gly 50 55 60 gat aat gtt ctg gta att tgt cat gcc ttg act ggg tcc gca gat gtt 240 Asp Asn Val Leu Val Ile Cys His Ala Leu Thr Gly Ser Ala Asp Val 65 70 75 80 gct gac tgg tgg ggc cct ctt ctg ggt aac gac tta gca ttc gac cca 288 Ala Asp Trp Trp Gly Pro Leu Leu Gly Asn Asp Leu Ala Phe Asp Pro 85 90 95 tca agg ttt ttt atc ata tgt tta aac tct atg ggc tct cca tat ggg 336 Ser Arg Phe Phe Ile Ile Cys Leu Asn Ser Met Gly Ser Pro Tyr Gly 100 105 110 tct ttt tcg cca tta acg ata aat gag gag acg ggc gtt aga tat gga 384 Ser Phe Ser Pro Leu Thr Ile Asn Glu Glu Thr Gly Val Arg Tyr Gly 115 120 125 ccc gaa ttc cca tta tgt act gtg cgc gat gac gtt aga gct cac aga 432 Pro Glu Phe Pro Leu Cys Thr Val Arg Asp Asp Val Arg Ala His Arg 130 135 140 att gtt ctg gat tct ctg gga gta aag tca ata gcc tgt gtt att ggt 480 Ile Val Leu Asp Ser Leu Gly Val Lys Ser Ile Ala Cys Val Ile Gly 145 150 155 160 ggc tct atg ggg ggg atg ctg agt ttg gaa tgg gct gcc atg tat ggt 528 Gly Ser Met Gly Gly Met Leu Ser Leu Glu Trp Ala Ala Met Tyr Gly 165 170 175 aag gaa tat gtg aag aat atg gtt gct ctg gcg aca tca gca aga cat 576 Lys Glu Tyr Val Lys Asn Met Val Ala Leu Ala Thr Ser Ala Arg His 180 185 190 tct gcc tgg tgc ata tcg tgg tct gag gct caa aga caa tcg att tac 624 Ser Ala Trp Cys Ile Ser Trp Ser Glu Ala Gln Arg Gln Ser Ile Tyr 195 200 205 tca gat ccc aac tac ttg gac ggg tac tat ccg gta gag gag caa cct 672 Ser Asp Pro Asn Tyr Leu Asp Gly Tyr Tyr Pro Val Glu Glu Gln Pro 210 215 220 gtg gcc gga cta tcg gct gca cgt atg tct gca ttg ttg acg tac agg 720 Val Ala Gly Leu Ser Ala Ala Arg Met Ser Ala Leu Leu Thr Tyr Arg 225 230 235 240 aca aga aac agt ttc gag aac aaa ttc tcc aga aga tct cct tca ata 768 Thr Arg Asn Ser Phe Glu Asn Lys Phe Ser Arg Arg Ser Pro Ser Ile 245 250 255 gca caa caa caa aaa gct caa agg gag gag aca cgc aaa cca tct act 816 Ala Gln Gln Gln Lys Ala Gln Arg Glu Glu Thr Arg Lys Pro Ser Thr 260 265 270 gtc agc gaa cac tcc cta caa atc cac aat gat ggg tat aaa aca aaa 864 Val Ser Glu His Ser Leu Gln Ile His Asn Asp Gly Tyr Lys Thr Lys 275 280 285 gcc agc act gcc atc gct ggc att tct ggg caa aaa ggt caa agc gtg 912 Ala Ser Thr Ala Ile Ala Gly Ile Ser Gly Gln Lys Gly Gln Ser Val 290 295 300 gtg tcc acc gca tct tct tcg gat tca ttg aat tct tca aca tcg atg 960 Val Ser Thr Ala Ser Ser Ser Asp Ser Leu Asn Ser Ser Thr Ser Met 305 310 315 320 act tcg gta agt tct gta acg ggt gaa gtg aag gac ata aag cct gcg 1008 Thr Ser Val Ser Ser Val Thr Gly Glu Val Lys Asp Ile Lys Pro Ala 325 330 335 cag acg tat ttt tct gca caa agt tac ttg agg tac cag ggc aca aag 1056 Gln Thr Tyr Phe Ser Ala Gln Ser Tyr Leu Arg Tyr Gln Gly Thr Lys 340 345 350 ttc atc aat agg ttc gac gcc aat tgt tac att gcc atc aca cgt aaa 1104 Phe Ile Asn Arg Phe Asp Ala Asn Cys Tyr Ile Ala Ile Thr Arg Lys 355 360 365 ctg gat acg cac gat ttg gca aga gac aga gta gat gac atc act gag 1152 Leu Asp Thr His Asp Leu Ala Arg Asp Arg Val Asp Asp Ile Thr Glu 370 375 380 gtc ctt tct acc atc caa caa cca tcc ctg atc atc ggt atc caa tct 1200 Val Leu Ser Thr Ile Gln Gln Pro Ser Leu Ile Ile Gly Ile Gln Ser 385 390 395 400 gat gga ctg ttc aca tat tca gaa caa gaa ttt ttg gct gag cac ata 1248 Asp Gly Leu Phe Thr Tyr Ser Glu Gln Glu Phe Leu Ala Glu His Ile 405 410 415 ccg aag tcg caa tta gaa aaa att gaa tct ccc gaa ggc cac gat gcc 1296 Pro Lys Ser Gln Leu Glu Lys Ile Glu Ser Pro Glu Gly His Asp Ala 420 425 430 ttc cta ttg gag ttt aag ctg ata aac aaa ctg ata gta caa ttt tta 1344 Phe Leu Leu Glu Phe Lys Leu Ile Asn Lys Leu Ile Val Gln Phe Leu 435 440 445 aaa acc aac tgc aag gcc att acc gat gcc gct cca aga gct tgg gga 1392 Lys Thr Asn Cys Lys Ala Ile Thr Asp Ala Ala Pro Arg Ala Trp Gly 450 455 460 ggt gac gtt ggt aac gat gaa acg aag acg tct gtc ttt ggt gag gcc 1440 Gly Asp Val Gly Asn Asp Glu Thr Lys Thr Ser Val Phe Gly Glu Ala 465 470 475 480 gaa gaa gtt acc aac tgg tag 1461 Glu Glu Val Thr Asn Trp 485 32 486 PRT Saccharomyces cerevisiae 32 Met Ser His Thr Leu Lys Ser Lys Thr Leu Gln Glu Leu Asp Ile Glu 1 5 10 15 Glu Ile Lys Glu Thr Asn Pro Leu Leu Lys Leu Val Gln Gly Gln Arg 20 25 30 Ile Val Gln Val Pro Glu Leu Val Leu Glu Ser Gly Val Val Ile Asn 35 40 45 Asn Phe Pro Ile Ala Tyr Lys Thr Trp Gly Thr Leu Asn Glu Ala Gly 50 55 60 Asp Asn Val Leu Val Ile Cys His Ala Leu Thr Gly Ser Ala Asp Val 65 70 75 80 Ala Asp Trp Trp Gly Pro Leu Leu Gly Asn Asp Leu Ala Phe Asp Pro 85 90 95 Ser Arg Phe Phe Ile Ile Cys Leu Asn Ser Met Gly Ser Pro Tyr Gly 100 105 110 Ser Phe Ser Pro Leu Thr Ile Asn Glu Glu Thr Gly Val Arg Tyr Gly 115 120 125 Pro Glu Phe Pro Leu Cys Thr Val Arg Asp Asp Val Arg Ala His Arg 130 135 140 Ile Val Leu Asp Ser Leu Gly Val Lys Ser Ile Ala Cys Val Ile Gly 145 150 155 160 Gly Ser Met Gly Gly Met Leu Ser Leu Glu Trp Ala Ala Met Tyr Gly 165 170 175 Lys Glu Tyr Val Lys Asn Met Val Ala Leu Ala Thr Ser Ala Arg His 180 185 190 Ser Ala Trp Cys Ile Ser Trp Ser Glu Ala Gln Arg Gln Ser Ile Tyr 195 200 205 Ser Asp Pro Asn Tyr Leu Asp Gly Tyr Tyr Pro Val Glu Glu Gln Pro 210 215 220 Val Ala Gly Leu Ser Ala Ala Arg Met Ser Ala Leu Leu Thr Tyr Arg 225 230 235 240 Thr Arg Asn Ser Phe Glu Asn Lys Phe Ser Arg Arg Ser Pro Ser Ile 245 250 255 Ala Gln Gln Gln Lys Ala Gln Arg Glu Glu Thr Arg Lys Pro Ser Thr 260 265 270 Val Ser Glu His Ser Leu Gln Ile His Asn Asp Gly Tyr Lys Thr Lys 275 280 285 Ala Ser Thr Ala Ile Ala Gly Ile Ser Gly Gln Lys Gly Gln Ser Val 290 295 300 Val Ser Thr Ala Ser Ser Ser Asp Ser Leu Asn Ser Ser Thr Ser Met 305 310 315 320 Thr Ser Val Ser Ser Val Thr Gly Glu Val Lys Asp Ile Lys Pro Ala 325 330 335 Gln Thr Tyr Phe Ser Ala Gln Ser Tyr Leu Arg Tyr Gln Gly Thr Lys 340 345 350 Phe Ile Asn Arg Phe Asp Ala Asn Cys Tyr Ile Ala Ile Thr Arg Lys 355 360 365 Leu Asp Thr His Asp Leu Ala Arg Asp Arg Val Asp Asp Ile Thr Glu 370 375 380 Val Leu Ser Thr Ile Gln Gln Pro Ser Leu Ile Ile Gly Ile Gln Ser 385 390 395 400 Asp Gly Leu Phe Thr Tyr Ser Glu Gln Glu Phe Leu Ala Glu His Ile 405 410 415 Pro Lys Ser Gln Leu Glu Lys Ile Glu Ser Pro Glu Gly His Asp Ala 420 425 430 Phe Leu Leu Glu Phe Lys Leu Ile Asn Lys Leu Ile Val Gln Phe Leu 435 440 445 Lys Thr Asn Cys Lys Ala Ile Thr Asp Ala Ala Pro Arg Ala Trp Gly 450 455 460 Gly Asp Val Gly Asn Asp Glu Thr Lys Thr Ser Val Phe Gly Glu Ala 465 470 475 480 Glu Glu Val Thr Asn Trp 485 33 1470 DNA Schizosaccharomyces pombe CDS (1)..(1467) RSO01936 33 atg gaa tct caa tct ccg att gaa tca att gtc ttt act gac tcc tgt 48 Met Glu Ser Gln Ser Pro Ile Glu Ser Ile Val Phe Thr Asp Ser Cys 1 5 10 15 cat ccg tct cag caa gaa aat aaa ttt gtt cag ctt att tca gat caa 96 His Pro Ser Gln Gln Glu Asn Lys Phe Val Gln Leu Ile Ser Asp Gln 20 25 30 aaa att gca att gtt ccc aaa ttt acg ttg gag tgt ggc gac atc ctt 144 Lys Ile Ala Ile Val Pro Lys Phe Thr Leu Glu Cys Gly Asp Ile Leu 35 40 45 tac gat gtt ccc gtt gcc ttc aag act tgg ggt act ttg aat aaa gaa 192 Tyr Asp Val Pro Val Ala Phe Lys Thr Trp Gly Thr Leu Asn Lys Glu 50 55 60 gga aac aat tgt ctt ctt ctt tgt cat gct tta agt ggt tct gct gat 240 Gly Asn Asn Cys Leu Leu Leu Cys His Ala Leu Ser Gly Ser Ala Asp 65 70 75 80 gct gga gat tgg tgg ggt cct tta ctc ggt cct ggt cgt gcg ttt gat 288 Ala Gly Asp Trp Trp Gly Pro Leu Leu Gly Pro Gly Arg Ala Phe Asp 85 90 95 cca tca cat ttc ttt atc gta tgc ctt aat tct ctt ggt agc cca tac 336 Pro Ser His Phe Phe Ile Val Cys Leu Asn Ser Leu Gly Ser Pro Tyr 100 105 110 gga agc gcc tct cct gtt aca tgg aac gct gag act cat agt gtt tat 384 Gly Ser Ala Ser Pro Val Thr Trp Asn Ala Glu Thr His Ser Val Tyr 115 120 125 ggg cca gaa ttt cct tta gca acc ata cgt gat gat gta aac atc cat 432 Gly Pro Glu Phe Pro Leu Ala Thr Ile Arg Asp Asp Val Asn Ile His 130 135 140 aaa ctt att tta caa aga ttg ggt gta aag caa att gct atg gca gta 480 Lys Leu Ile Leu Gln Arg Leu Gly Val Lys Gln Ile Ala Met Ala Val 145 150 155 160 ggt ggc tcc atg ggt ggt atg ctg gtt ttg gag tgg gca ttt gat aag 528 Gly Gly Ser Met Gly Gly Met Leu Val Leu Glu Trp Ala Phe Asp Lys 165 170 175 gaa ttt gtg cga tca att gtt ccc att tct acc tct ctt cgt cat tcc 576 Glu Phe Val Arg Ser Ile Val Pro Ile Ser Thr Ser Leu Arg His Ser 180 185 190 gcg tgg tgc att agc tgg tct gaa gcg caa cgc cag agt ata tat tct 624 Ala Trp Cys Ile Ser Trp Ser Glu Ala Gln Arg Gln Ser Ile Tyr Ser 195 200 205 gac cct aag ttt aat gat gga tac tac ggc ata gac gat cag cct gta 672 Asp Pro Lys Phe Asn Asp Gly Tyr Tyr Gly Ile Asp Asp Gln Pro Val 210 215 220 agt ggc ctt gga gct gct cgt atg tct gcc ttg ttg aca tat cgc tcc 720 Ser Gly Leu Gly Ala Ala Arg Met Ser Ala Leu Leu Thr Tyr Arg Ser 225 230 235 240 aaa tgt tct ttc gaa cgt cgc ttt gcc cgt act gtt cct gat gcg tct 768 Lys Cys Ser Phe Glu Arg Arg Phe Ala Arg Thr Val Pro Asp Ala Ser 245 250 255 cgt cac ccc tat cca gat cgt tta ccc act cct ctc acg ccc agt aat 816 Arg His Pro Tyr Pro Asp Arg Leu Pro Thr Pro Leu Thr Pro Ser Asn 260 265 270 gca cat tgg gtc gtt cac aac gaa gga aac cgt aat cgc cgt gaa cga 864 Ala His Trp Val Val His Asn Glu Gly Asn Arg Asn Arg Arg Glu Arg 275 280 285 cct tgt cga tcc aat gga tca tca cct act tct gaa agt gct tta aat 912 Pro Cys Arg Ser Asn Gly Ser Ser Pro Thr Ser Glu Ser Ala Leu Asn 290 295 300 tcc ccc gcc tct tct gtc tcg tct tta cct tct tta ggt gcc tct cag 960 Ser Pro Ala Ser Ser Val Ser Ser Leu Pro Ser Leu Gly Ala Ser Gln 305 310 315 320 act aca gac agt tct tcc ctt aac cag agt tcg tta tta aga cgt cct 1008 Thr Thr Asp Ser Ser Ser Leu Asn Gln Ser Ser Leu Leu Arg Arg Pro 325 330 335 gct aat act tac ttc tct gcg caa tcg tat tta cgt tac caa gcg aag 1056 Ala Asn Thr Tyr Phe Ser Ala Gln Ser Tyr Leu Arg Tyr Gln Ala Lys 340 345 350 aag ttt gta agt cgc ttt gat gct aat tgt tac att tcg att act aaa 1104 Lys Phe Val Ser Arg Phe Asp Ala Asn Cys Tyr Ile Ser Ile Thr Lys 355 360 365 aag ttg gac acc cat gat att act cgt gga cgc ggt tca gac tct cct 1152 Lys Leu Asp Thr His Asp Ile Thr Arg Gly Arg Gly Ser Asp Ser Pro 370 375 380 aag gaa gtc atg aag gat ttg tct tta ccc gta ctc gta ctc ggt att 1200 Lys Glu Val Met Lys Asp Leu Ser Leu Pro Val Leu Val Leu Gly Ile 385 390 395 400 gaa agc gat ggt ctt ttc aca ttt gac gaa caa gtt gaa att gcc aaa 1248 Glu Ser Asp Gly Leu Phe Thr Phe Asp Glu Gln Val Glu Ile Ala Lys 405 410 415 tct ttt ccc aat gct acc ttg gaa aaa att att tcg gcc gaa ggc cac 1296 Ser Phe Pro Asn Ala Thr Leu Glu Lys Ile Ile Ser Ala Glu Gly His 420 425 430 gac ggt ttt ttg ctt gag ttt act caa gta aac tca cat att caa aaa 1344 Asp Gly Phe Leu Leu Glu Phe Thr Gln Val Asn Ser His Ile Gln Lys 435 440 445 ttc caa aag gaa cat tta att gat atc atg tct caa act aat tcc ttt 1392 Phe Gln Lys Glu His Leu Ile Asp Ile Met Ser Gln Thr Asn Ser Phe 450 455 460 gag cga ctt gat tcc caa gtt aat gat acc aac cgc gaa agc gtt ttt 1440 Glu Arg Leu Asp Ser Gln Val Asn Asp Thr Asn Arg Glu Ser Val Phe 465 470 475 480 gga gaa atg gaa gac ata acc tcc tgg taa 1470 Gly Glu Met Glu Asp Ile Thr Ser Trp 485 34 489 PRT Schizosaccharomyces pombe 34 Met Glu Ser Gln Ser Pro Ile Glu Ser Ile Val Phe Thr Asp Ser Cys 1 5 10 15 His Pro Ser Gln Gln Glu Asn Lys Phe Val Gln Leu Ile Ser Asp Gln 20 25 30 Lys Ile Ala Ile Val Pro Lys Phe Thr Leu Glu Cys Gly Asp Ile Leu 35 40 45 Tyr Asp Val Pro Val Ala Phe Lys Thr Trp Gly Thr Leu Asn Lys Glu 50 55 60 Gly Asn Asn Cys Leu Leu Leu Cys His Ala Leu Ser Gly Ser Ala Asp 65 70 75 80 Ala Gly Asp Trp Trp Gly Pro Leu Leu Gly Pro Gly Arg Ala Phe Asp 85 90 95 Pro Ser His Phe Phe Ile Val Cys Leu Asn Ser Leu Gly Ser Pro Tyr 100 105 110 Gly Ser Ala Ser Pro Val Thr Trp Asn Ala Glu

Thr His Ser Val Tyr 115 120 125 Gly Pro Glu Phe Pro Leu Ala Thr Ile Arg Asp Asp Val Asn Ile His 130 135 140 Lys Leu Ile Leu Gln Arg Leu Gly Val Lys Gln Ile Ala Met Ala Val 145 150 155 160 Gly Gly Ser Met Gly Gly Met Leu Val Leu Glu Trp Ala Phe Asp Lys 165 170 175 Glu Phe Val Arg Ser Ile Val Pro Ile Ser Thr Ser Leu Arg His Ser 180 185 190 Ala Trp Cys Ile Ser Trp Ser Glu Ala Gln Arg Gln Ser Ile Tyr Ser 195 200 205 Asp Pro Lys Phe Asn Asp Gly Tyr Tyr Gly Ile Asp Asp Gln Pro Val 210 215 220 Ser Gly Leu Gly Ala Ala Arg Met Ser Ala Leu Leu Thr Tyr Arg Ser 225 230 235 240 Lys Cys Ser Phe Glu Arg Arg Phe Ala Arg Thr Val Pro Asp Ala Ser 245 250 255 Arg His Pro Tyr Pro Asp Arg Leu Pro Thr Pro Leu Thr Pro Ser Asn 260 265 270 Ala His Trp Val Val His Asn Glu Gly Asn Arg Asn Arg Arg Glu Arg 275 280 285 Pro Cys Arg Ser Asn Gly Ser Ser Pro Thr Ser Glu Ser Ala Leu Asn 290 295 300 Ser Pro Ala Ser Ser Val Ser Ser Leu Pro Ser Leu Gly Ala Ser Gln 305 310 315 320 Thr Thr Asp Ser Ser Ser Leu Asn Gln Ser Ser Leu Leu Arg Arg Pro 325 330 335 Ala Asn Thr Tyr Phe Ser Ala Gln Ser Tyr Leu Arg Tyr Gln Ala Lys 340 345 350 Lys Phe Val Ser Arg Phe Asp Ala Asn Cys Tyr Ile Ser Ile Thr Lys 355 360 365 Lys Leu Asp Thr His Asp Ile Thr Arg Gly Arg Gly Ser Asp Ser Pro 370 375 380 Lys Glu Val Met Lys Asp Leu Ser Leu Pro Val Leu Val Leu Gly Ile 385 390 395 400 Glu Ser Asp Gly Leu Phe Thr Phe Asp Glu Gln Val Glu Ile Ala Lys 405 410 415 Ser Phe Pro Asn Ala Thr Leu Glu Lys Ile Ile Ser Ala Glu Gly His 420 425 430 Asp Gly Phe Leu Leu Glu Phe Thr Gln Val Asn Ser His Ile Gln Lys 435 440 445 Phe Gln Lys Glu His Leu Ile Asp Ile Met Ser Gln Thr Asn Ser Phe 450 455 460 Glu Arg Leu Asp Ser Gln Val Asn Asp Thr Asn Arg Glu Ser Val Phe 465 470 475 480 Gly Glu Met Glu Asp Ile Thr Ser Trp 485 35 1113 DNA Xylella almond CDS (1)..(1110) RXFX01562 35 atg acc gaa ttt atc cct ccg ggc agc cta ttc cat gcg ctc tcc tct 48 Met Thr Glu Phe Ile Pro Pro Gly Ser Leu Phe His Ala Leu Ser Ser 1 5 10 15 cca ttt gcg atg aag cgt ggc gga caa ctc cac cac gcc cgc atc gct 96 Pro Phe Ala Met Lys Arg Gly Gly Gln Leu His His Ala Arg Ile Ala 20 25 30 tac gaa aca tgg ggc cgc ctc aat gcc agc gcc acc aat gcc att ctg 144 Tyr Glu Thr Trp Gly Arg Leu Asn Ala Ser Ala Thr Asn Ala Ile Leu 35 40 45 atc atg cct ggc tta tca ccc aat gca cat gcc gca cac cat gac agc 192 Ile Met Pro Gly Leu Ser Pro Asn Ala His Ala Ala His His Asp Ser 50 55 60 aat gct gag cca ggc tgg tgg gag tca atg cta ggt cca ggc aaa ccc 240 Asn Ala Glu Pro Gly Trp Trp Glu Ser Met Leu Gly Pro Gly Lys Pro 65 70 75 80 atc gac aca gac cgt tgg ttc gtg atc tgt gtc aac tca ctt ggt agc 288 Ile Asp Thr Asp Arg Trp Phe Val Ile Cys Val Asn Ser Leu Gly Ser 85 90 95 tgc aaa gga tcg act ggc cct gca tcg tac aac ccc atc acg cag gcc 336 Cys Lys Gly Ser Thr Gly Pro Ala Ser Tyr Asn Pro Ile Thr Gln Ala 100 105 110 atg tat cgt ttg gac ttt cca gca ctg tca atc gaa gac ggg gcc aac 384 Met Tyr Arg Leu Asp Phe Pro Ala Leu Ser Ile Glu Asp Gly Ala Asn 115 120 125 tcc gca att gaa gtg gta cat gca ctg ggc atc aag caa ctt gcc agc 432 Ser Ala Ile Glu Val Val His Ala Leu Gly Ile Lys Gln Leu Ala Ser 130 135 140 ctg atc ggc aat tca atg ggc ggc atg acg gca ctg gcc atc ctg ctg 480 Leu Ile Gly Asn Ser Met Gly Gly Met Thr Ala Leu Ala Ile Leu Leu 145 150 155 160 tta cat cca gat ata gcc cgc agc cac atc aac atc tca ggc agc gcg 528 Leu His Pro Asp Ile Ala Arg Ser His Ile Asn Ile Ser Gly Ser Ala 165 170 175 cag gca tta ccg ttt tcc atc gcc att cgc tcg cta caa cgc gag gcg 576 Gln Ala Leu Pro Phe Ser Ile Ala Ile Arg Ser Leu Gln Arg Glu Ala 180 185 190 atc cgc ctg gac ccc cat tgg agg cag gga gac tac gac gac acc cac 624 Ile Arg Leu Asp Pro His Trp Arg Gln Gly Asp Tyr Asp Asp Thr His 195 200 205 tac ccg gaa tcg ggg cta cgc atc gca cgc aaa ctt ggg gtg atc acc 672 Tyr Pro Glu Ser Gly Leu Arg Ile Ala Arg Lys Leu Gly Val Ile Thr 210 215 220 tac cgc tcc gcg ctg gaa tgg gac ggg cgt ttt ggc cgg gta cgc ttg 720 Tyr Arg Ser Ala Leu Glu Trp Asp Gly Arg Phe Gly Arg Val Arg Leu 225 230 235 240 gat tcg gac caa acc aac gac aca cca ttc gga ctg gaa ttc caa att 768 Asp Ser Asp Gln Thr Asn Asp Thr Pro Phe Gly Leu Glu Phe Gln Ile 245 250 255 gaa aac tac ttg gaa agc cat gca cac cgc ttc gtg cac acc ttc gac 816 Glu Asn Tyr Leu Glu Ser His Ala His Arg Phe Val His Thr Phe Asp 260 265 270 cca aac tgc tac ctg tac ctg agc cgc tcc atg gac tgg ttc gac gtg 864 Pro Asn Cys Tyr Leu Tyr Leu Ser Arg Ser Met Asp Trp Phe Asp Val 275 280 285 gcc gag tac gcc aat gga gac att ctt gcc ggg ctg gcc agg atc cga 912 Ala Glu Tyr Ala Asn Gly Asp Ile Leu Ala Gly Leu Ala Arg Ile Arg 290 295 300 atc caa cgc gca ctc gcc atc ggt agc cat acc gac atc ctc ttt cca 960 Ile Gln Arg Ala Leu Ala Ile Gly Ser His Thr Asp Ile Leu Phe Pro 305 310 315 320 ata caa cag caa caa caa att gcc gaa ggg cta cgc cgt ggc ggt aca 1008 Ile Gln Gln Gln Gln Gln Ile Ala Glu Gly Leu Arg Arg Gly Gly Thr 325 330 335 cac gcc acc ttc ctg ggc ctt gac tca ccg cag ggg cat gat gcg ttc 1056 His Ala Thr Phe Leu Gly Leu Asp Ser Pro Gln Gly His Asp Ala Phe 340 345 350 ctt gtg gat atc gca aga ttt ggc cct cca gtg aag gaa ttt ctg gac 1104 Leu Val Asp Ile Ala Arg Phe Gly Pro Pro Val Lys Glu Phe Leu Asp 355 360 365 gaa ctg tga 1113 Glu Leu 370 36 370 PRT Xylella almond 36 Met Thr Glu Phe Ile Pro Pro Gly Ser Leu Phe His Ala Leu Ser Ser 1 5 10 15 Pro Phe Ala Met Lys Arg Gly Gly Gln Leu His His Ala Arg Ile Ala 20 25 30 Tyr Glu Thr Trp Gly Arg Leu Asn Ala Ser Ala Thr Asn Ala Ile Leu 35 40 45 Ile Met Pro Gly Leu Ser Pro Asn Ala His Ala Ala His His Asp Ser 50 55 60 Asn Ala Glu Pro Gly Trp Trp Glu Ser Met Leu Gly Pro Gly Lys Pro 65 70 75 80 Ile Asp Thr Asp Arg Trp Phe Val Ile Cys Val Asn Ser Leu Gly Ser 85 90 95 Cys Lys Gly Ser Thr Gly Pro Ala Ser Tyr Asn Pro Ile Thr Gln Ala 100 105 110 Met Tyr Arg Leu Asp Phe Pro Ala Leu Ser Ile Glu Asp Gly Ala Asn 115 120 125 Ser Ala Ile Glu Val Val His Ala Leu Gly Ile Lys Gln Leu Ala Ser 130 135 140 Leu Ile Gly Asn Ser Met Gly Gly Met Thr Ala Leu Ala Ile Leu Leu 145 150 155 160 Leu His Pro Asp Ile Ala Arg Ser His Ile Asn Ile Ser Gly Ser Ala 165 170 175 Gln Ala Leu Pro Phe Ser Ile Ala Ile Arg Ser Leu Gln Arg Glu Ala 180 185 190 Ile Arg Leu Asp Pro His Trp Arg Gln Gly Asp Tyr Asp Asp Thr His 195 200 205 Tyr Pro Glu Ser Gly Leu Arg Ile Ala Arg Lys Leu Gly Val Ile Thr 210 215 220 Tyr Arg Ser Ala Leu Glu Trp Asp Gly Arg Phe Gly Arg Val Arg Leu 225 230 235 240 Asp Ser Asp Gln Thr Asn Asp Thr Pro Phe Gly Leu Glu Phe Gln Ile 245 250 255 Glu Asn Tyr Leu Glu Ser His Ala His Arg Phe Val His Thr Phe Asp 260 265 270 Pro Asn Cys Tyr Leu Tyr Leu Ser Arg Ser Met Asp Trp Phe Asp Val 275 280 285 Ala Glu Tyr Ala Asn Gly Asp Ile Leu Ala Gly Leu Ala Arg Ile Arg 290 295 300 Ile Gln Arg Ala Leu Ala Ile Gly Ser His Thr Asp Ile Leu Phe Pro 305 310 315 320 Ile Gln Gln Gln Gln Gln Ile Ala Glu Gly Leu Arg Arg Gly Gly Thr 325 330 335 His Ala Thr Phe Leu Gly Leu Asp Ser Pro Gln Gly His Asp Ala Phe 340 345 350 Leu Val Asp Ile Ala Arg Phe Gly Pro Pro Val Lys Glu Phe Leu Asp 355 360 365 Glu Leu 370 37 1113 DNA Xylella oleander CDS (1)..(1110) RXFY01729 37 atg acc gaa ttt atc cct ccg ggc agc cta ttc cat gcg ctc tcc tct 48 Met Thr Glu Phe Ile Pro Pro Gly Ser Leu Phe His Ala Leu Ser Ser 1 5 10 15 cca ttt gcg atg aag cgt ggc gga caa ctc cac cac gcc cgc atc gct 96 Pro Phe Ala Met Lys Arg Gly Gly Gln Leu His His Ala Arg Ile Ala 20 25 30 tac gaa aca tgg ggc cgc ctc aat gcc agc gcc acc aat gcc att ctg 144 Tyr Glu Thr Trp Gly Arg Leu Asn Ala Ser Ala Thr Asn Ala Ile Leu 35 40 45 atc atg cct ggc tta tca ccc aat gca cat gcc gca cac cat gac agc 192 Ile Met Pro Gly Leu Ser Pro Asn Ala His Ala Ala His His Asp Ser 50 55 60 aat gct gag cca ggc tgg tgg gag tca atg cta ggt cca ggc aaa ccc 240 Asn Ala Glu Pro Gly Trp Trp Glu Ser Met Leu Gly Pro Gly Lys Pro 65 70 75 80 atc gac aca gac cgt tgg ttc gtg atc tgt gtc aac tca ctt ggt agc 288 Ile Asp Thr Asp Arg Trp Phe Val Ile Cys Val Asn Ser Leu Gly Ser 85 90 95 tgc aaa gga tcg act ggc cct gca tcg tac aac ccc atc acg cag gcc 336 Cys Lys Gly Ser Thr Gly Pro Ala Ser Tyr Asn Pro Ile Thr Gln Ala 100 105 110 atg tat cgt ttg gac ttt cca gca ctg tca atc gaa gac ggg gcc aac 384 Met Tyr Arg Leu Asp Phe Pro Ala Leu Ser Ile Glu Asp Gly Ala Asn 115 120 125 gcc gca att gaa gtg gta cat gca ctg ggc atc aag caa ctt gcc agc 432 Ala Ala Ile Glu Val Val His Ala Leu Gly Ile Lys Gln Leu Ala Ser 130 135 140 ctg atc ggc aat tca atg ggg ggc atg acg aca ctg gcc atc ctg ctg 480 Leu Ile Gly Asn Ser Met Gly Gly Met Thr Thr Leu Ala Ile Leu Leu 145 150 155 160 tta cat cca gat att gcc cgc agc cac atc aac atc tca ggc agc gcg 528 Leu His Pro Asp Ile Ala Arg Ser His Ile Asn Ile Ser Gly Ser Ala 165 170 175 cag gca tta ccg ttt tcc atc gcc att cgc tcg cta caa cgc gag gcg 576 Gln Ala Leu Pro Phe Ser Ile Ala Ile Arg Ser Leu Gln Arg Glu Ala 180 185 190 atc cgc ctg gac ccc cat tgg aag cag gga gac tac gac gac acc cac 624 Ile Arg Leu Asp Pro His Trp Lys Gln Gly Asp Tyr Asp Asp Thr His 195 200 205 tac ccg gaa tcg ggg cta cgc atc gca cgc aaa ctc ggg gtg atc acc 672 Tyr Pro Glu Ser Gly Leu Arg Ile Ala Arg Lys Leu Gly Val Ile Thr 210 215 220 tac cgc tcc gcg ctg gaa tgg gac ggg cgt ttt ggc cgg gta cgc ttg 720 Tyr Arg Ser Ala Leu Glu Trp Asp Gly Arg Phe Gly Arg Val Arg Leu 225 230 235 240 gat tcg gac caa acc aac gac aca cca ttc gga ctg gaa ttc caa att 768 Asp Ser Asp Gln Thr Asn Asp Thr Pro Phe Gly Leu Glu Phe Gln Ile 245 250 255 gaa aac tac ttg gaa agc cat gca cac cgc ttc gtg cac acc ttc gac 816 Glu Asn Tyr Leu Glu Ser His Ala His Arg Phe Val His Thr Phe Asp 260 265 270 cca aac tgc tac ctg tac ctg agc cgc tcc atg gac tgg ttc gac gtg 864 Pro Asn Cys Tyr Leu Tyr Leu Ser Arg Ser Met Asp Trp Phe Asp Val 275 280 285 gcc gag tac gcc aat gga gac att ctt gcc ggg ctg gcc agg atc cga 912 Ala Glu Tyr Ala Asn Gly Asp Ile Leu Ala Gly Leu Ala Arg Ile Arg 290 295 300 atc caa cgc gca ctt gcc atc ggt agc cat acc gac atc ctc ttt cca 960 Ile Gln Arg Ala Leu Ala Ile Gly Ser His Thr Asp Ile Leu Phe Pro 305 310 315 320 ata caa cag caa caa caa att gcc gaa ggg cta cgc cgt ggc ggt aca 1008 Ile Gln Gln Gln Gln Gln Ile Ala Glu Gly Leu Arg Arg Gly Gly Thr 325 330 335 cac gcc acc ttc ctg ggc ctt gac tca ccg cag gga cat gat gcg ttc 1056 His Ala Thr Phe Leu Gly Leu Asp Ser Pro Gln Gly His Asp Ala Phe 340 345 350 ctt gtg gat atc gca gga ttt ggc cct cca gtg aag gaa ttt ctg ggc 1104 Leu Val Asp Ile Ala Gly Phe Gly Pro Pro Val Lys Glu Phe Leu Gly 355 360 365 gaa ctg tga 1113 Glu Leu 370 38 370 PRT Xylella oleander 38 Met Thr Glu Phe Ile Pro Pro Gly Ser Leu Phe His Ala Leu Ser Ser 1 5 10 15 Pro Phe Ala Met Lys Arg Gly Gly Gln Leu His His Ala Arg Ile Ala 20 25 30 Tyr Glu Thr Trp Gly Arg Leu Asn Ala Ser Ala Thr Asn Ala Ile Leu 35 40 45 Ile Met Pro Gly Leu Ser Pro Asn Ala His Ala Ala His His Asp Ser 50 55 60 Asn Ala Glu Pro Gly Trp Trp Glu Ser Met Leu Gly Pro Gly Lys Pro 65 70 75 80 Ile Asp Thr Asp Arg Trp Phe Val Ile Cys Val Asn Ser Leu Gly Ser 85 90 95 Cys Lys Gly Ser Thr Gly Pro Ala Ser Tyr Asn Pro Ile Thr Gln Ala 100 105 110 Met Tyr Arg Leu Asp Phe Pro Ala Leu Ser Ile Glu Asp Gly Ala Asn 115 120 125 Ala Ala Ile Glu Val Val His Ala Leu Gly Ile Lys Gln Leu Ala Ser 130 135 140 Leu Ile Gly Asn Ser Met Gly Gly Met Thr Thr Leu Ala Ile Leu Leu 145 150 155 160 Leu His Pro Asp Ile Ala Arg Ser His Ile Asn Ile Ser Gly Ser Ala 165 170 175 Gln Ala Leu Pro Phe Ser Ile Ala Ile Arg Ser Leu Gln Arg Glu Ala 180 185 190 Ile Arg Leu Asp Pro His Trp Lys Gln Gly Asp Tyr Asp Asp Thr His 195 200 205 Tyr Pro Glu Ser Gly Leu Arg Ile Ala Arg Lys Leu Gly Val Ile Thr 210 215 220 Tyr Arg Ser Ala Leu Glu Trp Asp Gly Arg Phe Gly Arg Val Arg Leu 225 230 235 240 Asp Ser Asp Gln Thr Asn Asp Thr Pro Phe Gly Leu Glu Phe Gln Ile 245 250 255 Glu Asn Tyr Leu Glu Ser His Ala His Arg Phe Val His Thr Phe Asp 260 265 270 Pro Asn Cys Tyr Leu Tyr Leu Ser Arg Ser Met Asp Trp Phe Asp Val 275 280 285 Ala Glu Tyr Ala Asn Gly Asp Ile Leu Ala Gly Leu Ala Arg Ile Arg 290 295 300 Ile Gln Arg Ala Leu Ala Ile Gly Ser His Thr Asp Ile Leu Phe Pro 305 310 315 320 Ile Gln Gln Gln Gln Gln Ile Ala Glu Gly Leu Arg Arg Gly Gly Thr 325 330 335 His Ala Thr Phe Leu Gly Leu Asp Ser Pro Gln Gly His Asp Ala Phe 340 345 350 Leu Val Asp Ile Ala Gly Phe Gly Pro Pro Val Lys Glu Phe Leu Gly 355 360 365 Glu Leu 370 39 1578 DNA Emericella nidulans CDS (1)..(1575) REN00010 39 atg agt ccg ctg aac ggc gtc gct cgt tcc ttt ccg cgg ccc ttc cag 48 Met Ser Pro Leu Asn Gly Val Ala Arg Ser Phe Pro Arg Pro Phe Gln 1 5 10 15 gcc gtg acc agg cgg cct ttt cga gtt gtc cag ccg gcc atc gcc tgt 96 Ala Val Thr Arg Arg Pro Phe Arg Val Val Gln Pro Ala Ile Ala Cys 20 25 30 ccg tcc aac agc cgg tcg ttt aac cat tct cga tca tta cga tca acg 144 Pro Ser Asn Ser Arg Ser Phe Asn His Ser Arg Ser Leu Arg Ser Thr 35 40 45 ggg tct cag tcc ccc gct cca tcc cca cgc gac tcc tcg aat ccc gcg 192 Gly Ser Gln Ser Pro Ala Pro Ser Pro Arg Asp Ser Ser Asn Pro Ala 50 55

60 ctg tcc ttc cct tgc ctc gac gcc cag gag gcc aag tcc gct ctt ctt 240 Leu Ser Phe Pro Cys Leu Asp Ala Gln Glu Ala Lys Ser Ala Leu Leu 65 70 75 80 tcc gcg cga tct ctt ggt tca ggc cct gaa ccc tcc tat acc gcc ggc 288 Ser Ala Arg Ser Leu Gly Ser Gly Pro Glu Pro Ser Tyr Thr Ala Gly 85 90 95 cac cac gaa cga ttc cat tcc gac gaa ccg ctg ctc ctt gat tgg ggc 336 His His Glu Arg Phe His Ser Asp Glu Pro Leu Leu Leu Asp Trp Gly 100 105 110 ggt ttg ctt cca gaa ttt gat atc gca tat gag aca tgg ggc cag ctg 384 Gly Leu Leu Pro Glu Phe Asp Ile Ala Tyr Glu Thr Trp Gly Gln Leu 115 120 125 aac gag aag aag gat aat gtc att ctg ctg cat acc ggt ctg tct gca 432 Asn Glu Lys Lys Asp Asn Val Ile Leu Leu His Thr Gly Leu Ser Ala 130 135 140 tct agc cat gcg cac agc acc gaa gcg aac ccg aag ccc ggc tgg tgg 480 Ser Ser His Ala His Ser Thr Glu Ala Asn Pro Lys Pro Gly Trp Trp 145 150 155 160 gag aaa ttc ata ggt cct ggg aag acg cta gat acg gac aag tac ttt 528 Glu Lys Phe Ile Gly Pro Gly Lys Thr Leu Asp Thr Asp Lys Tyr Phe 165 170 175 gtg atc tgc acc aat gtc ctt gga ggg tgc tac ggt agc acg ggg ccc 576 Val Ile Cys Thr Asn Val Leu Gly Gly Cys Tyr Gly Ser Thr Gly Pro 180 185 190 tcg acg gtg gac ccg tcg gat ggg aag aag tat gct acg cgg ttt ccc 624 Ser Thr Val Asp Pro Ser Asp Gly Lys Lys Tyr Ala Thr Arg Phe Pro 195 200 205 atc ctg aca att gaa gat atg gtg cga gcg cag ttc cgc ctt ttg gac 672 Ile Leu Thr Ile Glu Asp Met Val Arg Ala Gln Phe Arg Leu Leu Asp 210 215 220 cat ctt ggg gtt cgg aaa ctc tac gcg tcc gtc ggc tcc agc atg ggt 720 His Leu Gly Val Arg Lys Leu Tyr Ala Ser Val Gly Ser Ser Met Gly 225 230 235 240 ggt atg cag agt ctt gca gcc ggt gtt ctg ttc cca gag cga gtg ggc 768 Gly Met Gln Ser Leu Ala Ala Gly Val Leu Phe Pro Glu Arg Val Gly 245 250 255 aag att gtg tcg att agc ggt tgt gct cga agc cat ccg tac agc att 816 Lys Ile Val Ser Ile Ser Gly Cys Ala Arg Ser His Pro Tyr Ser Ile 260 265 270 gct atg cgc cat acc cag cgg cag gtg ttg atg atg gat cca aat tgg 864 Ala Met Arg His Thr Gln Arg Gln Val Leu Met Met Asp Pro Asn Trp 275 280 285 gct cga ggt ttc tac tac gat tcg atc cca cct cat tca ggc atg aag 912 Ala Arg Gly Phe Tyr Tyr Asp Ser Ile Pro Pro His Ser Gly Met Lys 290 295 300 ctc gct cgc gag att gcc acc gtc acg tac cgc agc gga cca gaa tgg 960 Leu Ala Arg Glu Ile Ala Thr Val Thr Tyr Arg Ser Gly Pro Glu Trp 305 310 315 320 gag aaa cgc ttt ggt cgg aaa cgg gct gat ccg agc aaa cag cct gcg 1008 Glu Lys Arg Phe Gly Arg Lys Arg Ala Asp Pro Ser Lys Gln Pro Ala 325 330 335 ctt tgc ccc gac ttt ctc atc gag acg tat ctc gac cac gcc ggt gaa 1056 Leu Cys Pro Asp Phe Leu Ile Glu Thr Tyr Leu Asp His Ala Gly Glu 340 345 350 aaa ttc tgc ttg gaa tac gat gcc aac agc ctg ctc tac atc tcc aag 1104 Lys Phe Cys Leu Glu Tyr Asp Ala Asn Ser Leu Leu Tyr Ile Ser Lys 355 360 365 gcg atg gat ctg ttt gac cta ggg ttg act cag caa ctc gcg acg aag 1152 Ala Met Asp Leu Phe Asp Leu Gly Leu Thr Gln Gln Leu Ala Thr Lys 370 375 380 aag cag agg gcg gag gcc cag gcg aag att agc agc gga aca aac act 1200 Lys Gln Arg Ala Glu Ala Gln Ala Lys Ile Ser Ser Gly Thr Asn Thr 385 390 395 400 gtc aat gat gcg tcg tgc agc ctt aca ctt cct gaa cag cca tac cag 1248 Val Asn Asp Ala Ser Cys Ser Leu Thr Leu Pro Glu Gln Pro Tyr Gln 405 410 415 gag cag cca tct gcc tcg aca tcc gcc gag cag tct gct tcc gct tca 1296 Glu Gln Pro Ser Ala Ser Thr Ser Ala Glu Gln Ser Ala Ser Ala Ser 420 425 430 gag acc ggg tcg gct ccg aac gat ctt gtt gcc ggg ctt gcg ccg ctg 1344 Glu Thr Gly Ser Ala Pro Asn Asp Leu Val Ala Gly Leu Ala Pro Leu 435 440 445 aaa gac cat cag gtg ctg gta atc gga gtc gca agc gac att ctc ttc 1392 Lys Asp His Gln Val Leu Val Ile Gly Val Ala Ser Asp Ile Leu Phe 450 455 460 ccg gcg tgg caa cag cgc gag atc gcg gag act ctg att caa gca ggg 1440 Pro Ala Trp Gln Gln Arg Glu Ile Ala Glu Thr Leu Ile Gln Ala Gly 465 470 475 480 aac aag acc gtg gag cat att gag ctg ggc aac gac gtg tct ctc ttt 1488 Asn Lys Thr Val Glu His Ile Glu Leu Gly Asn Asp Val Ser Leu Phe 485 490 495 ggt cat gac aca ttc ctc ctt gat gtc aga acg tcg gag gcg cag ttc 1536 Gly His Asp Thr Phe Leu Leu Asp Val Arg Thr Ser Glu Ala Gln Phe 500 505 510 gca agt tcc gta cta gtc ggc tcg cac ata att gta caa tag 1578 Ala Ser Ser Val Leu Val Gly Ser His Ile Ile Val Gln 515 520 525 40 525 PRT Emericella nidulans 40 Met Ser Pro Leu Asn Gly Val Ala Arg Ser Phe Pro Arg Pro Phe Gln 1 5 10 15 Ala Val Thr Arg Arg Pro Phe Arg Val Val Gln Pro Ala Ile Ala Cys 20 25 30 Pro Ser Asn Ser Arg Ser Phe Asn His Ser Arg Ser Leu Arg Ser Thr 35 40 45 Gly Ser Gln Ser Pro Ala Pro Ser Pro Arg Asp Ser Ser Asn Pro Ala 50 55 60 Leu Ser Phe Pro Cys Leu Asp Ala Gln Glu Ala Lys Ser Ala Leu Leu 65 70 75 80 Ser Ala Arg Ser Leu Gly Ser Gly Pro Glu Pro Ser Tyr Thr Ala Gly 85 90 95 His His Glu Arg Phe His Ser Asp Glu Pro Leu Leu Leu Asp Trp Gly 100 105 110 Gly Leu Leu Pro Glu Phe Asp Ile Ala Tyr Glu Thr Trp Gly Gln Leu 115 120 125 Asn Glu Lys Lys Asp Asn Val Ile Leu Leu His Thr Gly Leu Ser Ala 130 135 140 Ser Ser His Ala His Ser Thr Glu Ala Asn Pro Lys Pro Gly Trp Trp 145 150 155 160 Glu Lys Phe Ile Gly Pro Gly Lys Thr Leu Asp Thr Asp Lys Tyr Phe 165 170 175 Val Ile Cys Thr Asn Val Leu Gly Gly Cys Tyr Gly Ser Thr Gly Pro 180 185 190 Ser Thr Val Asp Pro Ser Asp Gly Lys Lys Tyr Ala Thr Arg Phe Pro 195 200 205 Ile Leu Thr Ile Glu Asp Met Val Arg Ala Gln Phe Arg Leu Leu Asp 210 215 220 His Leu Gly Val Arg Lys Leu Tyr Ala Ser Val Gly Ser Ser Met Gly 225 230 235 240 Gly Met Gln Ser Leu Ala Ala Gly Val Leu Phe Pro Glu Arg Val Gly 245 250 255 Lys Ile Val Ser Ile Ser Gly Cys Ala Arg Ser His Pro Tyr Ser Ile 260 265 270 Ala Met Arg His Thr Gln Arg Gln Val Leu Met Met Asp Pro Asn Trp 275 280 285 Ala Arg Gly Phe Tyr Tyr Asp Ser Ile Pro Pro His Ser Gly Met Lys 290 295 300 Leu Ala Arg Glu Ile Ala Thr Val Thr Tyr Arg Ser Gly Pro Glu Trp 305 310 315 320 Glu Lys Arg Phe Gly Arg Lys Arg Ala Asp Pro Ser Lys Gln Pro Ala 325 330 335 Leu Cys Pro Asp Phe Leu Ile Glu Thr Tyr Leu Asp His Ala Gly Glu 340 345 350 Lys Phe Cys Leu Glu Tyr Asp Ala Asn Ser Leu Leu Tyr Ile Ser Lys 355 360 365 Ala Met Asp Leu Phe Asp Leu Gly Leu Thr Gln Gln Leu Ala Thr Lys 370 375 380 Lys Gln Arg Ala Glu Ala Gln Ala Lys Ile Ser Ser Gly Thr Asn Thr 385 390 395 400 Val Asn Asp Ala Ser Cys Ser Leu Thr Leu Pro Glu Gln Pro Tyr Gln 405 410 415 Glu Gln Pro Ser Ala Ser Thr Ser Ala Glu Gln Ser Ala Ser Ala Ser 420 425 430 Glu Thr Gly Ser Ala Pro Asn Asp Leu Val Ala Gly Leu Ala Pro Leu 435 440 445 Lys Asp His Gln Val Leu Val Ile Gly Val Ala Ser Asp Ile Leu Phe 450 455 460 Pro Ala Trp Gln Gln Arg Glu Ile Ala Glu Thr Leu Ile Gln Ala Gly 465 470 475 480 Asn Lys Thr Val Glu His Ile Glu Leu Gly Asn Asp Val Ser Leu Phe 485 490 495 Gly His Asp Thr Phe Leu Leu Asp Val Arg Thr Ser Glu Ala Gln Phe 500 505 510 Ala Ser Ser Val Leu Val Gly Ser His Ile Ile Val Gln 515 520 525 41 1170 DNA Mesorhizobium loti CDS (1)..(1167) NP_104621 41 atg gcc gct ctg cgc gca gga aag acc aac aac gag gcc gac cag ccg 48 Met Ala Ala Leu Arg Ala Gly Lys Thr Asn Asn Glu Ala Asp Gln Pro 1 5 10 15 tcg agc ccg gtg ttg cgc ttc ggg gcg gac aag ccg ctc aag ctc gac 96 Ser Ser Pro Val Leu Arg Phe Gly Ala Asp Lys Pro Leu Lys Leu Asp 20 25 30 gcc ggc acg ctt ttg tcg ccg ttc cag atc gcc tat cag acc tac ggc 144 Ala Gly Thr Leu Leu Ser Pro Phe Gln Ile Ala Tyr Gln Thr Tyr Gly 35 40 45 acg ctg aac gat gcc cgc tcc aat gcc atc ctc gtc tgc cat gcg ctg 192 Thr Leu Asn Asp Ala Arg Ser Asn Ala Ile Leu Val Cys His Ala Leu 50 55 60 acc ggc gac cag cat gtc gcc aac acc aat ccg gtg acc ggc aag ccg 240 Thr Gly Asp Gln His Val Ala Asn Thr Asn Pro Val Thr Gly Lys Pro 65 70 75 80 gga tgg tgg gaa gtg ctg atc ggc ccc ggc agg atc atc gac acc aac 288 Gly Trp Trp Glu Val Leu Ile Gly Pro Gly Arg Ile Ile Asp Thr Asn 85 90 95 cgt ttc ttc gtc atc tgc tcc aac gtc atc ggc ggt tgt ctg ggc tcc 336 Arg Phe Phe Val Ile Cys Ser Asn Val Ile Gly Gly Cys Leu Gly Ser 100 105 110 acc ggc ccg gcc tcg acc aac ccc gcc acc ggc aag ccc tac ggg ctc 384 Thr Gly Pro Ala Ser Thr Asn Pro Ala Thr Gly Lys Pro Tyr Gly Leu 115 120 125 gac ctg ccg gtc atc acc atc cgc gat atg gtg cgc gcg cag cag atg 432 Asp Leu Pro Val Ile Thr Ile Arg Asp Met Val Arg Ala Gln Gln Met 130 135 140 ctg atc gat cat ttc ggc atc gag aaa ctg ttc tgc gtg ctc ggc ggc 480 Leu Ile Asp His Phe Gly Ile Glu Lys Leu Phe Cys Val Leu Gly Gly 145 150 155 160 tcg atg ggc gga atg cag gtg ctg gaa tgg gcg tcg agc tac ccc gag 528 Ser Met Gly Gly Met Gln Val Leu Glu Trp Ala Ser Ser Tyr Pro Glu 165 170 175 cgc gtc ttt tcg gca ctg ccg atc gcc acc ggc gcg cgc cat tcc tcg 576 Arg Val Phe Ser Ala Leu Pro Ile Ala Thr Gly Ala Arg His Ser Ser 180 185 190 cag aac atc gcc ttc cac gag gtc ggc cgg cag gct gtc atg gcc gat 624 Gln Asn Ile Ala Phe His Glu Val Gly Arg Gln Ala Val Met Ala Asp 195 200 205 ccg gac tgg cac ggc ggc aaa tat ttc gaa aac ggc aaa cgc ccg gaa 672 Pro Asp Trp His Gly Gly Lys Tyr Phe Glu Asn Gly Lys Arg Pro Glu 210 215 220 aag ggc ctg gcg gta gcg cgc atg gcc gcc cac ata acc tat ctg tcg 720 Lys Gly Leu Ala Val Ala Arg Met Ala Ala His Ile Thr Tyr Leu Ser 225 230 235 240 gaa gcc gcc ctg cac cgg aaa ttc ggc cgc aat ctg cag gat cgc gag 768 Glu Ala Ala Leu His Arg Lys Phe Gly Arg Asn Leu Gln Asp Arg Glu 245 250 255 gcg ctg acc ttc ggc ttc gac gcc gac ttc cag atc gaa agc tat ctg 816 Ala Leu Thr Phe Gly Phe Asp Ala Asp Phe Gln Ile Glu Ser Tyr Leu 260 265 270 cgc cac caa ggc atg acc ttc gtc gac cgc ttc gac gcc aat tcc tat 864 Arg His Gln Gly Met Thr Phe Val Asp Arg Phe Asp Ala Asn Ser Tyr 275 280 285 ctc tac atg acg cgg tcg atg gac tat ttc gac ctc gcc gcc gat cat 912 Leu Tyr Met Thr Arg Ser Met Asp Tyr Phe Asp Leu Ala Ala Asp His 290 295 300 ggc ggg cgg ctg gcg gat gcc ttt gcc ggc acc aaa acc cgc ttc tgc 960 Gly Gly Arg Leu Ala Asp Ala Phe Ala Gly Thr Lys Thr Arg Phe Cys 305 310 315 320 ctg gtg tcc ttc acc tcg gat tgg ttg ttt ccg acc gaa gag agc cgc 1008 Leu Val Ser Phe Thr Ser Asp Trp Leu Phe Pro Thr Glu Glu Ser Arg 325 330 335 tcg atc gtg cac gcg ctc aac gcc gcc ggc gcg tcc gtg tcc ttc gtc 1056 Ser Ile Val His Ala Leu Asn Ala Ala Gly Ala Ser Val Ser Phe Val 340 345 350 gaa atc gag acc gac cgc ggc cac gat gcc ttc ctg ctc gac gag ccg 1104 Glu Ile Glu Thr Asp Arg Gly His Asp Ala Phe Leu Leu Asp Glu Pro 355 360 365 gaa ctg ttc gcc gcc atc aac ggc ttc atc ggc tcc gcg gcg cgg gcg 1152 Glu Leu Phe Ala Ala Ile Asn Gly Phe Ile Gly Ser Ala Ala Arg Ala 370 375 380 aga ggg cta agc gca tga 1170 Arg Gly Leu Ser Ala 385 42 389 PRT Mesorhizobium loti 42 Met Ala Ala Leu Arg Ala Gly Lys Thr Asn Asn Glu Ala Asp Gln Pro 1 5 10 15 Ser Ser Pro Val Leu Arg Phe Gly Ala Asp Lys Pro Leu Lys Leu Asp 20 25 30 Ala Gly Thr Leu Leu Ser Pro Phe Gln Ile Ala Tyr Gln Thr Tyr Gly 35 40 45 Thr Leu Asn Asp Ala Arg Ser Asn Ala Ile Leu Val Cys His Ala Leu 50 55 60 Thr Gly Asp Gln His Val Ala Asn Thr Asn Pro Val Thr Gly Lys Pro 65 70 75 80 Gly Trp Trp Glu Val Leu Ile Gly Pro Gly Arg Ile Ile Asp Thr Asn 85 90 95 Arg Phe Phe Val Ile Cys Ser Asn Val Ile Gly Gly Cys Leu Gly Ser 100 105 110 Thr Gly Pro Ala Ser Thr Asn Pro Ala Thr Gly Lys Pro Tyr Gly Leu 115 120 125 Asp Leu Pro Val Ile Thr Ile Arg Asp Met Val Arg Ala Gln Gln Met 130 135 140 Leu Ile Asp His Phe Gly Ile Glu Lys Leu Phe Cys Val Leu Gly Gly 145 150 155 160 Ser Met Gly Gly Met Gln Val Leu Glu Trp Ala Ser Ser Tyr Pro Glu 165 170 175 Arg Val Phe Ser Ala Leu Pro Ile Ala Thr Gly Ala Arg His Ser Ser 180 185 190 Gln Asn Ile Ala Phe His Glu Val Gly Arg Gln Ala Val Met Ala Asp 195 200 205 Pro Asp Trp His Gly Gly Lys Tyr Phe Glu Asn Gly Lys Arg Pro Glu 210 215 220 Lys Gly Leu Ala Val Ala Arg Met Ala Ala His Ile Thr Tyr Leu Ser 225 230 235 240 Glu Ala Ala Leu His Arg Lys Phe Gly Arg Asn Leu Gln Asp Arg Glu 245 250 255 Ala Leu Thr Phe Gly Phe Asp Ala Asp Phe Gln Ile Glu Ser Tyr Leu 260 265 270 Arg His Gln Gly Met Thr Phe Val Asp Arg Phe Asp Ala Asn Ser Tyr 275 280 285 Leu Tyr Met Thr Arg Ser Met Asp Tyr Phe Asp Leu Ala Ala Asp His 290 295 300 Gly Gly Arg Leu Ala Asp Ala Phe Ala Gly Thr Lys Thr Arg Phe Cys 305 310 315 320 Leu Val Ser Phe Thr Ser Asp Trp Leu Phe Pro Thr Glu Glu Ser Arg 325 330 335 Ser Ile Val His Ala Leu Asn Ala Ala Gly Ala Ser Val Ser Phe Val 340 345 350 Glu Ile Glu Thr Asp Arg Gly His Asp Ala Phe Leu Leu Asp Glu Pro 355 360 365 Glu Leu Phe Ala Ala Ile Asn Gly Phe Ile Gly Ser Ala Ala Arg Ala 370 375 380 Arg Gly Leu Ser Ala 385 43 1155 DNA acremonium crysogenum CDS (1)..(1152) P39058 43 tgt cgc ctc aga tcg cca atc gct tcg agg ctt cgc tag atg ccc aag 48 Cys Arg Leu Arg Ser Pro Ile Ala Ser Arg Leu Arg Xaa Met Pro Lys 1 5 10 15 aca tag cca gaa tat cgc tct tca cac tgg aat ctg gcg tca tcc ttc 96 Thr Xaa Pro Glu Tyr Arg Ser Ser His Trp Asn Leu Ala Ser Ser Phe 20 25 30 gcg atg tac ccg tgg cat aca aat cgt ggg gtc gca tga atg tct caa 144 Ala Met Tyr Pro Trp His Thr Asn Arg Gly Val Ala Xaa Met Ser Gln 35 40 45 ggg ata act gcg tca tcg tct gcc aca cct tga cga gca gcg ccc atg 192 Gly Ile Thr Ala Ser Ser Ser Ala Thr Pro Xaa Arg Ala Ala Pro Met 50 55 60 tca cct cgt ggt ggc cca cac tgt ttg gcc aag gca ggg ctt tcg ata 240 Ser Pro Arg Gly Gly Pro His Cys Leu Ala Lys Ala Gly Leu Ser Ile 65 70 75 80 cct ctc gct act tca tca tct gcc taa

att atc tcg gga gcc cct ttg 288 Pro Leu Ala Thr Ser Ser Ser Ala Xaa Ile Ile Ser Gly Ala Pro Leu 85 90 95 gga gtg ctg gac cat gtt cac cgg acc ccg atg cag aag gcc agc gcc 336 Gly Val Leu Asp His Val His Arg Thr Pro Met Gln Lys Ala Ser Ala 100 105 110 cgt acg ggg cca agt ttc ctc gca cga cga ttc gag atg atg ttc gta 384 Arg Thr Gly Pro Ser Phe Leu Ala Arg Arg Phe Glu Met Met Phe Val 115 120 125 ttc atc gcc agg tgc tcg aca ggt tag gcg tca ggc aaa ttg ctg ccg 432 Phe Ile Ala Arg Cys Ser Thr Gly Xaa Ala Ser Gly Lys Leu Leu Pro 130 135 140 tag tcg gcg cat cca tgg gtg gaa tgc aca ctc tgg aat ggg cct tct 480 Xaa Ser Ala His Pro Trp Val Glu Cys Thr Leu Trp Asn Gly Pro Ser 145 150 155 160 ttg gtc ccg agt acg tgc gaa aga ttg tgc cca tcg cga cat cat gcc 528 Leu Val Pro Ser Thr Cys Glu Arg Leu Cys Pro Ser Arg His His Ala 165 170 175 gtc aga gcg gct ggt gcg cag ctt ggt tcg aga cac aga ggc agt gca 576 Val Arg Ala Ala Gly Ala Gln Leu Gly Ser Arg His Arg Gly Ser Ala 180 185 190 tct atg atg acc cca agt acc tgg acg ggg agt acg acg tag acg acc 624 Ser Met Met Thr Pro Ser Thr Trp Thr Gly Ser Thr Thr Xaa Thr Thr 195 200 205 agc ctg tcc ggg ggc tcg aaa cag cgc gca aga ttg cga atc tca cgt 672 Ser Leu Ser Gly Gly Ser Lys Gln Arg Ala Arg Leu Arg Ile Ser Arg 210 215 220 aca aga gca aac ctg cga tgg acg agc gct tcc ata tgg ctc cag gag 720 Thr Arg Ala Asn Leu Arg Trp Thr Ser Ala Ser Ile Trp Leu Gln Glu 225 230 235 240 tcc aag ccg gcc gga ata tca gca gcc agg atg cga aga agg aaa tca 768 Ser Lys Pro Ala Gly Ile Ser Ala Ala Arg Met Arg Arg Arg Lys Ser 245 250 255 acg gca cag aca gcg gca aca gcc acc gtg ctg gcc agc cca ttg aag 816 Thr Ala Gln Thr Ala Ala Thr Ala Thr Val Leu Ala Ser Pro Leu Lys 260 265 270 ccg tat ctt cct atc tcc ggt acc agg ccc aga agt ttg ccg cga gct 864 Pro Tyr Leu Pro Ile Ser Gly Thr Arg Pro Arg Ser Leu Pro Arg Ala 275 280 285 tcg acg cca act gct aca tcg cca tga cac tca agt tcg aca ccc acg 912 Ser Thr Pro Thr Ala Thr Ser Pro Xaa His Ser Ser Ser Thr Pro Thr 290 295 300 aca tca gca gag gcc ggg cag gat caa tcc cgg agg ctc tgg caa tga 960 Thr Ser Ala Glu Ala Gly Gln Asp Gln Ser Arg Arg Leu Trp Gln Xaa 305 310 315 320 tta cac aac cag cgt tga tca ttt gcg cca ggt cag acg gtc tgt act 1008 Leu His Asn Gln Arg Xaa Ser Phe Ala Pro Gly Gln Thr Val Cys Thr 325 330 335 cgt ttg acg agc acg ttg aga tgg ggc gca gta tcc caa aca gtc gtc 1056 Arg Leu Thr Ser Thr Leu Arg Trp Gly Ala Val Ser Gln Thr Val Val 340 345 350 ttt gcg tgg tgg aca cga atg agg gtc atg act tct ttg taa tgg aag 1104 Phe Ala Trp Trp Thr Arg Met Arg Val Met Thr Ser Leu Xaa Trp Lys 355 360 365 cgg aca agg tta atg atg ccg tca gag gat tcc tcg atc agt cat taa 1152 Arg Thr Arg Leu Met Met Pro Ser Glu Asp Ser Ser Ile Ser His Xaa 370 375 380 tgt 1155 44 384 PRT acremonium crysogenum unsure 13 .. 13 All occurrences of Xaa indicate any amino acid unsure 18 .. 18 All occurrences of Xaa indicate any amino acid unsure 45 .. 45 All occurrences of Xaa indicate any amino acid unsure 59 .. 59 All occurrences of Xaa indicate any amino acid unsure 89 .. 89 All occurrences of Xaa indicate any amino acid unsure 137 .. 137 All occurrences of Xaa indicate any amino acid unsure 145 .. 145 All occurrences of Xaa indicate any amino acid unsure 206 .. 206 All occurrences of Xaa indicate any amino acid unsure 297 .. 297 All occurrences of Xaa indicate any amino acid unsure 320 .. 320 All occurrences of Xaa indicate any amino acid unsure 326 .. 326 All occurrences of Xaa indicate any amino acid unsure 366 .. 366 All occurrences of Xaa indicate any amino acid unsure 384 .. 384 All occurrences of Xaa indicate any amino acid 44 Cys Arg Leu Arg Ser Pro Ile Ala Ser Arg Leu Arg Xaa Met Pro Lys 1 5 10 15 Thr Xaa Pro Glu Tyr Arg Ser Ser His Trp Asn Leu Ala Ser Ser Phe 20 25 30 Ala Met Tyr Pro Trp His Thr Asn Arg Gly Val Ala Xaa Met Ser Gln 35 40 45 Gly Ile Thr Ala Ser Ser Ser Ala Thr Pro Xaa Arg Ala Ala Pro Met 50 55 60 Ser Pro Arg Gly Gly Pro His Cys Leu Ala Lys Ala Gly Leu Ser Ile 65 70 75 80 Pro Leu Ala Thr Ser Ser Ser Ala Xaa Ile Ile Ser Gly Ala Pro Leu 85 90 95 Gly Val Leu Asp His Val His Arg Thr Pro Met Gln Lys Ala Ser Ala 100 105 110 Arg Thr Gly Pro Ser Phe Leu Ala Arg Arg Phe Glu Met Met Phe Val 115 120 125 Phe Ile Ala Arg Cys Ser Thr Gly Xaa Ala Ser Gly Lys Leu Leu Pro 130 135 140 Xaa Ser Ala His Pro Trp Val Glu Cys Thr Leu Trp Asn Gly Pro Ser 145 150 155 160 Leu Val Pro Ser Thr Cys Glu Arg Leu Cys Pro Ser Arg His His Ala 165 170 175 Val Arg Ala Ala Gly Ala Gln Leu Gly Ser Arg His Arg Gly Ser Ala 180 185 190 Ser Met Met Thr Pro Ser Thr Trp Thr Gly Ser Thr Thr Xaa Thr Thr 195 200 205 Ser Leu Ser Gly Gly Ser Lys Gln Arg Ala Arg Leu Arg Ile Ser Arg 210 215 220 Thr Arg Ala Asn Leu Arg Trp Thr Ser Ala Ser Ile Trp Leu Gln Glu 225 230 235 240 Ser Lys Pro Ala Gly Ile Ser Ala Ala Arg Met Arg Arg Arg Lys Ser 245 250 255 Thr Ala Gln Thr Ala Ala Thr Ala Thr Val Leu Ala Ser Pro Leu Lys 260 265 270 Pro Tyr Leu Pro Ile Ser Gly Thr Arg Pro Arg Ser Leu Pro Arg Ala 275 280 285 Ser Thr Pro Thr Ala Thr Ser Pro Xaa His Ser Ser Ser Thr Pro Thr 290 295 300 Thr Ser Ala Glu Ala Gly Gln Asp Gln Ser Arg Arg Leu Trp Gln Xaa 305 310 315 320 Leu His Asn Gln Arg Xaa Ser Phe Ala Pro Gly Gln Thr Val Cys Thr 325 330 335 Arg Leu Thr Ser Thr Leu Arg Trp Gly Ala Val Ser Gln Thr Val Val 340 345 350 Phe Ala Trp Trp Thr Arg Met Arg Val Met Thr Ser Leu Xaa Trp Lys 355 360 365 Arg Thr Arg Leu Met Met Pro Ser Glu Asp Ser Ser Ile Ser His Xaa 370 375 380 45 1077 DNA Pseudomonas putida CDS (1)..(1074) AAK49778 45 atg tca act gtc ttt ccc gaa gat tcc gtc ggt ctg gta gta cgg caa 48 Met Ser Thr Val Phe Pro Glu Asp Ser Val Gly Leu Val Val Arg Gln 1 5 10 15 acc tcc cgg ttc gat gaa ccg ctg gca ctg gcc tgt ggc cgt tca ctg 96 Thr Ser Arg Phe Asp Glu Pro Leu Ala Leu Ala Cys Gly Arg Ser Leu 20 25 30 gcc agt tac gaa ctg gtc tac gag acc tat ggc acc ctg aac gcc agc 144 Ala Ser Tyr Glu Leu Val Tyr Glu Thr Tyr Gly Thr Leu Asn Ala Ser 35 40 45 gcg agc aac gcc gtg ctg atc tgc cat gcc ctg tcc ggc cac cac cat 192 Ala Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 gcc gct ggc tac cat gcc gcc acc gac cgc aag ccg ggc tgg tgg gac 240 Ala Ala Gly Tyr His Ala Ala Thr Asp Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 agc tgc atc ggc ccc gga aaa ccg atc gat acc aac cgc ttc ttc gtg 288 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Asn Arg Phe Phe Val 85 90 95 gtc agc ctg aac aac ctc ggc ggc tgc aac ggc agc acc ggc ccc agc 336 Val Ser Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Thr Gly Pro Ser 100 105 110 agt gtc aac cca gcc acc ggt aaa ccc tat ggc gcc gag ttc ccg gta 384 Ser Val Asn Pro Ala Thr Gly Lys Pro Tyr Gly Ala Glu Phe Pro Val 115 120 125 ttg acc gtg gaa gac tgg gtg cac agc cag gca cgg ctg gcc gac cgc 432 Leu Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Arg 130 135 140 ctg ggc atc cag cag tgg gca gct atc gtc ggc ggt agc ctg ggt ggc 480 Leu Gly Ile Gln Gln Trp Ala Ala Ile Val Gly Gly Ser Leu Gly Gly 145 150 155 160 atg cag gcg ctg caa tgg acg atg acc tac ccc gag cgc gta cgc cac 528 Met Gln Ala Leu Gln Trp Thr Met Thr Tyr Pro Glu Arg Val Arg His 165 170 175 tgc gtc gac att gcc tcg gcc ccc aag ctg tcg gcg cag aac atc gcc 576 Cys Val Asp Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 ttc aac gag gtg gcg cgt cag gcc att ctt acc gac cct gag tac cgc 624 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Glu Tyr Arg 195 200 205 aga ggc tcg ttt cca gga cca ggt gtg atc ccc aag cgc ggc ctg atg 672 Arg Gly Ser Phe Pro Gly Pro Gly Val Ile Pro Lys Arg Gly Leu Met 210 215 220 ctg gca cgg atg gtc ggc cac att acc tat ctg tcc gat gat tcg atg 720 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met 225 230 235 240 ggt gaa aaa ttc ggc cga gag ctg aaa gcg aca agc tca act acg act 768 Gly Glu Lys Phe Gly Arg Glu Leu Lys Ala Thr Ser Ser Thr Thr Thr 245 250 255 tcc aca gcg tcg agt tcc agg tcg aaa gct acc tgc gct atc agg gcg 816 Ser Thr Ala Ser Ser Ser Arg Ser Lys Ala Thr Cys Ala Ile Arg Ala 260 265 270 agg agt ttt ccg gcc gtt tcg acg cca aca cct acc ttg atg acc aag 864 Arg Ser Phe Pro Ala Val Ser Thr Pro Thr Pro Thr Leu Met Thr Lys 275 280 285 gca ctg gac tat ttc gac ccg gcc gcc acg cac ggt ggt gat ctg gcc 912 Ala Leu Asp Tyr Phe Asp Pro Ala Ala Thr His Gly Gly Asp Leu Ala 290 295 300 gcc acc ctg gcc cac gtc acg gcg gac tac tgc atc tgt cgt tca cca 960 Ala Thr Leu Ala His Val Thr Ala Asp Tyr Cys Ile Cys Arg Ser Pro 305 310 315 320 ccg act gcg ctt ctc tcc ggc ccg ttc gcg cga gat cgt cga cgc gct 1008 Pro Thr Ala Leu Leu Ser Gly Pro Phe Ala Arg Asp Arg Arg Arg Ala 325 330 335 gat ggc cgc gcg caa gaa cgt ctg cta cct gga gat cga ttc gcc cta 1056 Asp Gly Arg Ala Gln Glu Arg Leu Leu Pro Gly Asp Arg Phe Ala Leu 340 345 350 cgg gca cga tgc att tcc tga 1077 Arg Ala Arg Cys Ile Ser 355 46 358 PRT Pseudomonas putida 46 Met Ser Thr Val Phe Pro Glu Asp Ser Val Gly Leu Val Val Arg Gln 1 5 10 15 Thr Ser Arg Phe Asp Glu Pro Leu Ala Leu Ala Cys Gly Arg Ser Leu 20 25 30 Ala Ser Tyr Glu Leu Val Tyr Glu Thr Tyr Gly Thr Leu Asn Ala Ser 35 40 45 Ala Ser Asn Ala Val Leu Ile Cys His Ala Leu Ser Gly His His His 50 55 60 Ala Ala Gly Tyr His Ala Ala Thr Asp Arg Lys Pro Gly Trp Trp Asp 65 70 75 80 Ser Cys Ile Gly Pro Gly Lys Pro Ile Asp Thr Asn Arg Phe Phe Val 85 90 95 Val Ser Leu Asn Asn Leu Gly Gly Cys Asn Gly Ser Thr Gly Pro Ser 100 105 110 Ser Val Asn Pro Ala Thr Gly Lys Pro Tyr Gly Ala Glu Phe Pro Val 115 120 125 Leu Thr Val Glu Asp Trp Val His Ser Gln Ala Arg Leu Ala Asp Arg 130 135 140 Leu Gly Ile Gln Gln Trp Ala Ala Ile Val Gly Gly Ser Leu Gly Gly 145 150 155 160 Met Gln Ala Leu Gln Trp Thr Met Thr Tyr Pro Glu Arg Val Arg His 165 170 175 Cys Val Asp Ile Ala Ser Ala Pro Lys Leu Ser Ala Gln Asn Ile Ala 180 185 190 Phe Asn Glu Val Ala Arg Gln Ala Ile Leu Thr Asp Pro Glu Tyr Arg 195 200 205 Arg Gly Ser Phe Pro Gly Pro Gly Val Ile Pro Lys Arg Gly Leu Met 210 215 220 Leu Ala Arg Met Val Gly His Ile Thr Tyr Leu Ser Asp Asp Ser Met 225 230 235 240 Gly Glu Lys Phe Gly Arg Glu Leu Lys Ala Thr Ser Ser Thr Thr Thr 245 250 255 Ser Thr Ala Ser Ser Ser Arg Ser Lys Ala Thr Cys Ala Ile Arg Ala 260 265 270 Arg Ser Phe Pro Ala Val Ser Thr Pro Thr Pro Thr Leu Met Thr Lys 275 280 285 Ala Leu Asp Tyr Phe Asp Pro Ala Ala Thr His Gly Gly Asp Leu Ala 290 295 300 Ala Thr Leu Ala His Val Thr Ala Asp Tyr Cys Ile Cys Arg Ser Pro 305 310 315 320 Pro Thr Ala Leu Leu Ser Gly Pro Phe Ala Arg Asp Arg Arg Arg Ala 325 330 335 Asp Gly Arg Ala Gln Glu Arg Leu Leu Pro Gly Asp Arg Phe Ala Leu 340 345 350 Arg Ala Arg Cys Ile Ser 355 47 52 DNA Artificial sequence Description of the artificial sequencePCR primer 47 cccgggatcc gctagcggcg cgccggccgg cccggtgtga aataccgcac ag 52 48 53 DNA Artificial sequence Description of the artificial sequencePCR primer 48 tctagactcg agcggccgcg gccggccttt aaattgaaga cgaaagggcc tcg 53 49 47 DNA Artificial sequence Description of the artificial sequencePCR primer 49 gagatctaga cccggggatc cgctagcggg ctgctaaagg aagcgga 47 50 38 DNA Artificial sequence Description of the artificial sequencePCR primer 50 gagaggcgcg ccgctagcgt gggcgaagaa ctccagca 38 51 34 DNA Artificial sequence Description of the artificial sequencePCR primer 51 gagagggcgg ccgcgcaaag tcccgcttcg tgaa 34 52 34 DNA Artificial sequence Description of the artificial sequencePCR primer 52 gagagggcgg ccgctcaagt cggtcaagcc acgc 34 53 140 DNA Artificial sequence Description of the artificial sequencePCR primer 53 tcgaatttaa atctcgagag gcctgacgtc gggcccggta ccacgcgtca tatgactagt 60 tcggacctag ggatatcgtc gacatcgatg ctcttctgcg ttaattaaca attgggatcc 120 tctagacccg ggatttaaat 140 54 140 DNA Artificial sequence Description of the artificial sequencePCR primer 54 gatcatttaa atcccgggtc tagaggatcc caattgttaa ttaacgcaga agagcatcga 60 tgtcgacgat atccctaggt ccgaactagt catatgacgc gtggtaccgg gcccgacgtc 120 aggcctctcg agatttaaat 140 55 33 DNA Artificial sequence Description of the artificial sequencePCR primer 55 gagagcggcc gccgatcctt tttaacccat cac 33 56 32 DNA Artificial sequence Description of the artificial sequencePCR primer 56 aggagcggcc gccatcggca ttttcttttg cg 32 57 5091 DNA Artificial sequence Description of the artificial sequenceplas mid 57 gccgcgactg ccttcgcgaa gccttgcccc gcggaaattt cctccaccga gttcgtgcac 60 acccctatgc caagcttctt tcaccctaaa ttcgagagat tggattctta ccgtggaaat 120 tcttcgcaaa aatcgtcccc tgatcgccct tgcgacgttg gcgtcggtgc cgctggttgc 180 gcttggcttg accgacttga tcagcggccg ctcgatttaa atctcgagag gcctgacgtc 240 gggcccggta ccacgcgtca tatgactagt tcggacctag ggatatcgtc gacatcgatg 300 ctcttctgcg ttaattaaca attgggatcc tctagacccg ggatttaaat cgctagcggg 360 ctgctaaagg aagcggaaca cgtagaaagc cagtccgcag aaacggtgct gaccccggat 420 gaatgtcagc tactgggcta tctggacaag ggaaaacgca agcgcaaaga gaaagcaggt 480 agcttgcagt gggcttacat ggcgatagct agactgggcg gttttatgga cagcaagcga 540 accggaattg ccagctgggg cgccctctgg taaggttggg aagccctgca aagtaaactg 600 gatggctttc ttgccgccaa ggatctgatg gcgcagggga tcaagatctg atcaagagac 660 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc 720 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc 780 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc 840 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg 900 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt 960 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc 1020 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga 1080 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga 1140 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct 1200 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc 1260 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt 1320 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat

attgctgaag agcttggcgg 1380 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat 1440 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc 1500 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa 1560 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 1620 ctcatgctgg agttcttcgc ccacgctagc ggcgcgccgg ccggcccggt gtgaaatacc 1680 gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga 1740 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 1800 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 1860 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 1920 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 1980 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 2040 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 2100 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 2160 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 2220 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 2280 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 2340 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 2400 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 2460 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 2520 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 2580 cttcacctag atccttttaa aggccggccg cggccgcgca aagtcccgct tcgtgaaaat 2640 tttcgtgccg cgtgattttc cgccaaaaac tttaacgaac gttcgttata atggtgtcat 2700 gaccttcacg acgaagtact aaaattggcc cgaatcatca gctatggatc tctctgatgt 2760 cgcgctggag tccgacgcgc tcgatgctgc cgtcgattta aaaacggtga tcggattttt 2820 ccgagctctc gatacgacgg acgcgccagc atcacgagac tgggccagtg ccgcgagcga 2880 cctagaaact ctcgtggcgg atcttgagga gctggctgac gagctgcgtg ctcggccagc 2940 gccaggagga cgcacagtag tggaggatgc aatcagttgc gcctactgcg gtggcctgat 3000 tcctccccgg cctgacccgc gaggacggcg cgcaaaatat tgctcagatg cgtgtcgtgc 3060 cgcagccagc cgcgagcgcg ccaacaaacg ccacgccgag gagctggagg cggctaggtc 3120 gcaaatggcg ctggaagtgc gtcccccgag cgaaattttg gccatggtcg tcacagagct 3180 ggaagcggca gcgagaatta tcgcgatcgt ggcggtgccc gcaggcatga caaacatcgt 3240 aaatgccgcg tttcgtgtgc cgtggccgcc caggacgtgt cagcgccgcc accacctgca 3300 ccgaatcggc agcagcgtcg cgcgtcgaaa aagcgcacag gcggcaagaa gcgataagct 3360 gcacgaatac ctgaaaaatg ttgaacgccc cgtgagcggt aactcacagg gcgtcggcta 3420 acccccagtc caaacctggg agaaagcgct caaaaatgac tctagcggat tcacgagaca 3480 ttgacacacc ggcctggaaa ttttccgctg atctgttcga cacccatccc gagctcgcgc 3540 tgcgatcacg tggctggacg agcgaagacc gccgcgaatt cctcgctcac ctgggcagag 3600 aaaatttcca gggcagcaag acccgcgact tcgccagcgc ttggatcaaa gacccggaca 3660 cggagaaaca cagccgaagt tataccgagt tggttcaaaa tcgcttgccc ggtgccagta 3720 tgttgctctg acgcacgcgc agcacgcagc cgtgcttgtc ctggacattg atgtgccgag 3780 ccaccaggcc ggcgggaaaa tcgagcacgt aaaccccgag gtctacgcga ttttggagcg 3840 ctgggcacgc ctggaaaaag cgccagcttg gatcggcgtg aatccactga gcgggaaatg 3900 ccagctcatc tggctcattg atccggtgta tgccgcagca ggcatgagca gcccgaatat 3960 gcgcctgctg gctgcaacga ccgaggaaat gacccgcgtt ttcggcgctg accaggcttt 4020 ttcacatagg ctgagccgtg gccactgcac tctccgacga tcccagccgt accgctggca 4080 tgcccagcac aatcgcgtgg atcgcctagc tgatcttatg gaggttgctc gcatgatctc 4140 aggcacagaa aaacctaaaa aacgctatga gcaggagttt tctagcggac gggcacgtat 4200 cgaagcggca agaaaagcca ctgcggaagc aaaagcactt gccacgcttg aagcaagcct 4260 gccgagcgcc gctgaagcgt ctggagagct gatcgacggc gtccgtgtcc tctggactgc 4320 tccagggcgt gccgcccgtg atgagacggc ttttcgccac gctttgactg tgggatacca 4380 gttaaaagcg gctggtgagc gcctaaaaga caccaagggt catcgagcct acgagcgtgc 4440 ctacaccgtc gctcaggcgg tcggaggagg ccgtgagcct gatctgccgc cggactgtga 4500 ccgccagacg gattggccgc gacgtgtgcg cggctacgtc gctaaaggcc agccagtcgt 4560 ccctgctcgt cagacagaga cgcagagcca gccgaggcga aaagctctgg ccactatggg 4620 aagacgtggc ggtaaaaagg ccgcagaacg ctggaaagac ccaaacagtg agtacgcccg 4680 agcacagcga gaaaaactag ctaagtccag tcaacgacaa gctaggaaag ctaaaggaaa 4740 tcgcttgacc attgcaggtt ggtttatgac tgttgaggga gagactggct cgtggccgac 4800 aatcaatgaa gctatgtctg aatttagcgt gtcacgtcag accgtgaata gagcacttaa 4860 ggtctgcggg cattgaactt ccacgaggac gccgaaagct tcccagtaaa tgtgccatct 4920 cgtaggcaga aaacggttcc cccgtagggt ctctctcttg gcctcctttc taggtcgggc 4980 tgattgctct tgaagctctc taggggggct cacaccatag gcagataacg ttccccaccg 5040 gctcgcctcg taagcgcaca aggactgctc ccaaagatct tcaaagccac t 5091 58 4323 DNA Artificial sequence Description of the artificial sequenceplasmid 58 tctctcagcg tatggttgtc gcctgagctg tagttgcctt catcgatgaa ctgctgtaca 60 ttttgatacg tttttccgtc accgtcaaag attgatttat aatcctctac accgttgatg 120 ttcaaagagc tgtctgatgc tgatacgtta acttgtgcag ttgtcagtgt ttgtttgccg 180 taatgtttac cggagaaatc agtgtagaat aaacggattt ttccgtcaga tgtaaatgtg 240 gctgaacctg accattcttg tgtttggtct tttaggatag aatcatttgc atcgaatttg 300 tcgctgtctt taaagacgcg gccagcgttt ttccagctgt caatagaagt ttcgccgact 360 ttttgataga acatgtaaat cgatgtgtca tccgcatttt taggatctcc ggctaatgca 420 aagacgatgt ggtagccgtg atagtttgcg acagtgccgt cagcgttttg taatggccag 480 ctgtcccaaa cgtccaggcc ttttgcagaa gagatatttt taattgtgga cgaatcaaat 540 tcagaaactt gatatttttc atttttttgc tgttcaggga tttgcagcat atcatggcgt 600 gtaatatggg aaatgccgta tgtttcctta tatggctttt ggttcgtttc tttcgcaaac 660 gcttgagttg cgcctcctgc cagcagtgcg gtagtaaagg ttaatactgt tgcttgtttt 720 gcaaactttt tgatgttcat cgttcatgtc tcctttttta tgtactgtgt tagcggtctg 780 cttcttccag ccctcctgtt tgaagatggc aagttagtta cgcacaataa aaaaagacct 840 aaaatatgta aggggtgacg ccaaagtata cactttgccc tttacacatt ttaggtcttg 900 cctgctttat cagtaacaaa cccgcgcgat ttacttttcg acctcattct attagactct 960 cgtttggatt gcaactggtc tattttcctc ttttgtttga tagaaaatca taaaaggatt 1020 tgcagactac gggcctaaag aactaaaaaa tctatctgtt tcttttcatt ctctgtattt 1080 tttatagttt ctgttgcatg ggcataaagt tgccttttta atcacaattc agaaaatatc 1140 ataatatctc atttcactaa ataatagtga acggcaggta tatgtgatgg gttaaaaagg 1200 atcggcggcc gctcgattta aatctcgaga ggcctgacgt cgggcccggt accacgcgtc 1260 atatgactag ttcggaccta gggatatcgt cgacatcgat gctcttctgc gttaattaac 1320 aattgggatc ctctagaccc gggatttaaa tcgctagcgg gctgctaaag gaagcggaac 1380 acgtagaaag ccagtccgca gaaacggtgc tgaccccgga tgaatgtcag ctactgggct 1440 atctggacaa gggaaaacgc aagcgcaaag agaaagcagg tagcttgcag tgggcttaca 1500 tggcgatagc tagactgggc ggttttatgg acagcaagcg aaccggaatt gccagctggg 1560 gcgccctctg gtaaggttgg gaagccctgc aaagtaaact ggatggcttt cttgccgcca 1620 aggatctgat ggcgcagggg atcaagatct gatcaagaga caggatgagg atcgtttcgc 1680 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc 1740 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca 1800 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg 1860 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg 1920 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag 1980 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg 2040 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc 2100 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa 2160 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac 2220 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat 2280 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac 2340 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc 2400 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt 2460 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc 2520 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg 2580 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg 2640 cccacgctag cggcgcgccg gccggcccgg tgtgaaatac cgcacagatg cgtaaggaga 2700 aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt 2760 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca 2820 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa 2880 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat 2940 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc 3000 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc 3060 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt 3120 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac 3180 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg 3240 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca 3300 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc 3360 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa 3420 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 3480 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac 3540 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta 3600 aaggccggcc gcggccgcca tcggcatttt cttttgcgtt tttatttgtt aactgttaat 3660 tgtccttgtt caaggatgct gtctttgaca acagatgttt tcttgccttt gatgttcagc 3720 aggaagctcg gcgcaaacgt tgattgtttg tctgcgtaga atcctctgtt tgtcatatag 3780 cttgtaatca cgacattgtt tcctttcgct tgaggtacag cgaagtgtga gtaagtaaag 3840 gttacatcgt taggatcaag atccattttt aacacaaggc cagttttgtt cagcggcttg 3900 tatgggccag ttaaagaatt agaaacataa ccaagcatgt aaatatcgtt agacgtaatg 3960 ccgtcaatcg tcatttttga tccgcgggag tcagtgaaca ggtaccattt gccgttcatt 4020 ttaaagacgt tcgcgcgttc aatttcatct gttactgtgt tagatgcaat cagcggtttc 4080 atcacttttt tcagtgtgta atcatcgttt agctcaatca taccgagagc gccgtttgct 4140 aactcagccg tgcgtttttt atcgctttgc agaagttttt gactttcttg acggaagaat 4200 gatgtgcttt tgccatagta tgctttgtta aataaagatt cttcgccttg gtagccatct 4260 tcagttccag tgtttgcttc aaatactaag tatttgtggc ctttatcttc tacgtagtga 4320 gga 4323 59 35 DNA Artificial sequence Description of the artificial sequencePCR Primer 59 gagagagaga cgcgtcccag tggctgagac gcatc 35 60 34 DNA Artificial sequence Description of the artificial sequencePCR Primer 60 ctctctctgt cgacgaattc aatcttacgg cctg 34 61 5860 DNA Artificial sequence Description of the artificial sequenceplasmid 61 cccggtacca cgcgtcccag tggctgagac gcatccgcta aagccccagg aaccctgtgc 60 agaaagaaaa cactcctctg gctaggtaga cacagtttat aaaggtagag ttgagcgggt 120 aactgtcagc acgtagatcg aaaggtgcac aaaggtggcc ctggtcgtac agaaatatgg 180 cggttcctcg cttgagagtg cggaacgcat tagaaacgtc gctgaacgga tcgttgccac 240 caagaaggct ggaaatgatg tcgtggttgt ctgctccgca atgggagaca ccacggatga 300 acttctagaa cttgcagcgg cagtgaatcc cgttccgcca gctcgtgaaa tggatatgct 360 cctgactgct ggtgagcgta tttctaacgc tctcgtcgcc atggctattg agtcccttgg 420 cgcagaagcc caatctttca cgggctctca ggctggtgtg ctcaccaccg agcgccacgg 480 aaacgcacgc attgttgatg tcactccagg tcgtgtgcgt gaagcactcg atgagggcaa 540 gatctgcatt gttgctggtt tccagggtgt taataaagaa acccgcgatg tcaccacgtt 600 gggtcgtggt ggttctgaca ccactgcagt tgcgttggca gctgctttga acgctgatgt 660 gtgtgagatt tactcggacg ttgacggtgt gtataccgct gacccgcgca tcgttcctaa 720 tgcacagaag ctggaaaagc tcagcttcga agaaatgctg gaacttgctg ctgttggctc 780 caagattttg gtgctgcgca gtgttgaata cgctcgtgca ttcaatgtgc cacttcgcgt 840 acgctcgtct tatagtaatg atcccggcac tttgattgcc ggctctatgg aggatattcc 900 tgtggaagaa gcagtcctta ccggtgtcgc aaccgacaag tccgaagcca aagtaaccgt 960 tctgggtatt tccgataagc caggcgaggc tgcgaaggtt ttccgtgcgt tggctgatgc 1020 agaaatcaac attgacatgg ttctgcagaa cgtctcttct gtagaagacg gcaccaccga 1080 catcaccttc acctgccctc gttccgacgg ccgccgcgcg atggagatct tgaagaagct 1140 tcaggttcag ggcaactgga ccaatgtgct ttacgacgac caggtcggca aagtctccct 1200 cgtgggtgct ggcatgaagt ctcacccagg tgttaccgca gagttcatgg aagctctgcg 1260 cgatgtcaac gtgaacatcg aattgatttc cacctctgag attcgtattt ccgtgctgat 1320 ccgtgaagat gatctggatg ctgctgcacg tgcattgcat gagcagttcc agctgggcgg 1380 cgaagacgaa gccgtcgttt atgcaggcac cggacgctaa agttttaaag gagtagtttt 1440 acaatgacca ccatcgcagt tgttggtgca accggccagg tcggccaggt tatgcgcacc 1500 cttttggaag agcgcaattt cccagctgac actgttcgtt tctttgcttc cccacgttcc 1560 gcaggccgta agattgaatt cgtcgacatc gatgctcttc tgcgttaatt aacaattggg 1620 atcctctaga cccgggattt aaatcgctag cgggctgcta aaggaagcgg aacacgtaga 1680 aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga 1740 caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat 1800 agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct 1860 ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct 1920 gatggcgcag gggatcaaga tctgatcaag agacaggatg aggatcgttt cgcatgattg 1980 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg 2040 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg 2100 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg 2160 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg 2220 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc 2280 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc 2340 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc 2400 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc 2460 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg 2520 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct 2580 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt 2640 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc 2700 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt 2760 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc 2820 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg 2880 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccacgc 2940 tagcggcgcg ccggccggcc cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc 3000 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 3060 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata 3120 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 3180 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 3240 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 3300 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 3360 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 3420 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 3480 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 3540 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 3600 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc 3660 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 3720 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 3780 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 3840 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaaggccg 3900 gccgcggccg ccatcggcat tttcttttgc gtttttattt gttaactgtt aattgtcctt 3960 gttcaaggat gctgtctttg acaacagatg ttttcttgcc tttgatgttc agcaggaagc 4020 tcggcgcaaa cgttgattgt ttgtctgcgt agaatcctct gtttgtcata tagcttgtaa 4080 tcacgacatt gtttcctttc gcttgaggta cagcgaagtg tgagtaagta aaggttacat 4140 cgttaggatc aagatccatt tttaacacaa ggccagtttt gttcagcggc ttgtatgggc 4200 cagttaaaga attagaaaca taaccaagca tgtaaatatc gttagacgta atgccgtcaa 4260 tcgtcatttt tgatccgcgg gagtcagtga acaggtacca tttgccgttc attttaaaga 4320 cgttcgcgcg ttcaatttca tctgttactg tgttagatgc aatcagcggt ttcatcactt 4380 ttttcagtgt gtaatcatcg tttagctcaa tcataccgag agcgccgttt gctaactcag 4440 ccgtgcgttt tttatcgctt tgcagaagtt tttgactttc ttgacggaag aatgatgtgc 4500 ttttgccata gtatgctttg ttaaataaag attcttcgcc ttggtagcca tcttcagttc 4560 cagtgtttgc ttcaaatact aagtatttgt ggcctttatc ttctacgtag tgaggatctc 4620 tcagcgtatg gttgtcgcct gagctgtagt tgccttcatc gatgaactgc tgtacatttt 4680 gatacgtttt tccgtcaccg tcaaagattg atttataatc ctctacaccg ttgatgttca 4740 aagagctgtc tgatgctgat acgttaactt gtgcagttgt cagtgtttgt ttgccgtaat 4800 gtttaccgga gaaatcagtg tagaataaac ggatttttcc gtcagatgta aatgtggctg 4860 aacctgacca ttcttgtgtt tggtctttta ggatagaatc atttgcatcg aatttgtcgc 4920 tgtctttaaa gacgcggcca gcgtttttcc agctgtcaat agaagtttcg ccgacttttt 4980 gatagaacat gtaaatcgat gtgtcatccg catttttagg atctccggct aatgcaaaga 5040 cgatgtggta gccgtgatag tttgcgacag tgccgtcagc gttttgtaat ggccagctgt 5100 cccaaacgtc caggcctttt gcagaagaga tatttttaat tgtggacgaa tcaaattcag 5160 aaacttgata tttttcattt ttttgctgtt cagggatttg cagcatatca tggcgtgtaa 5220 tatgggaaat gccgtatgtt tccttatatg gcttttggtt cgtttctttc gcaaacgctt 5280 gagttgcgcc tcctgccagc agtgcggtag taaaggttaa tactgttgct tgttttgcaa 5340 actttttgat gttcatcgtt catgtctcct tttttatgta ctgtgttagc ggtctgcttc 5400 ttccagccct cctgtttgaa gatggcaagt tagttacgca caataaaaaa agacctaaaa 5460 tatgtaaggg gtgacgccaa agtatacact ttgcccttta cacattttag gtcttgcctg 5520 ctttatcagt aacaaacccg cgcgatttac ttttcgacct cattctatta gactctcgtt 5580 tggattgcaa ctggtctatt ttcctctttt gtttgataga aaatcataaa aggatttgca 5640 gactacgggc ctaaagaact aaaaaatcta tctgtttctt ttcattctct gtatttttta 5700 tagtttctgt tgcatgggca taaagttgcc tttttaatca caattcagaa aatatcataa 5760 tatctcattt cactaaataa tagtgaacgg caggtatatg tgatgggtta aaaaggatcg 5820 gcggccgctc gatttaaatc tcgagaggcc tgacgtcggg 5860 62 38 DNA Artificial sequence Description of the artificial sequencePCR Primer 62 cggcaccacc gacatcatct tcacctgccc tcgttccg 38 63 38 DNA Artificial sequence Description of the artificial sequencePCR Primer 63 cggaacgagg gcaggtgaag atgatgtcgg tggtgccg 38 64 1266 DNA LysC mutant CDS (1)..(1266) 64 gtg gcc ctg gtc gta cag aaa tat ggc ggt tcc tcg ctt gag agt gcg 48 Val Ala Leu Val Val Gln Lys Tyr Gly Gly Ser Ser Leu Glu Ser Ala 1 5 10 15 gaa cgc att aga aac gtc gct gaa cgg atc gtt gcc acc aag aag gct 96 Glu Arg Ile Arg Asn Val Ala Glu Arg Ile Val Ala Thr Lys Lys Ala 20 25 30

gga aat gat gtc gtg gtt gtc tgc tcc gca atg gga gac acc acg gat 144 Gly Asn Asp Val Val Val Val Cys Ser Ala Met Gly Asp Thr Thr Asp 35 40 45 gaa ctt cta gaa ctt gca gcg gca gtg aat ccc gtt ccg cca gct cgt 192 Glu Leu Leu Glu Leu Ala Ala Ala Val Asn Pro Val Pro Pro Ala Arg 50 55 60 gaa atg gat atg ctc ctg act gct ggt gag cgt att tct aac gct ctc 240 Glu Met Asp Met Leu Leu Thr Ala Gly Glu Arg Ile Ser Asn Ala Leu 65 70 75 80 gtc gcc atg gct att gag tcc ctt ggc gca gaa gcc caa tct ttc acg 288 Val Ala Met Ala Ile Glu Ser Leu Gly Ala Glu Ala Gln Ser Phe Thr 85 90 95 ggc tct cag gct ggt gtg ctc acc acc gag cgc cac gga aac gca cgc 336 Gly Ser Gln Ala Gly Val Leu Thr Thr Glu Arg His Gly Asn Ala Arg 100 105 110 att gtt gat gtc act cca ggt cgt gtg cgt gaa gca ctc gat gag ggc 384 Ile Val Asp Val Thr Pro Gly Arg Val Arg Glu Ala Leu Asp Glu Gly 115 120 125 aag atc tgc att gtt gct ggt ttc cag ggt gtt aat aaa gaa acc cgc 432 Lys Ile Cys Ile Val Ala Gly Phe Gln Gly Val Asn Lys Glu Thr Arg 130 135 140 gat gtc acc acg ttg ggt cgt ggt ggt tct gac acc act gca gtt gcg 480 Asp Val Thr Thr Leu Gly Arg Gly Gly Ser Asp Thr Thr Ala Val Ala 145 150 155 160 ttg gca gct gct ttg aac gct gat gtg tgt gag att tac tcg gac gtt 528 Leu Ala Ala Ala Leu Asn Ala Asp Val Cys Glu Ile Tyr Ser Asp Val 165 170 175 gac ggt gtg tat acc gct gac ccg cgc atc gtt cct aat gca cag aag 576 Asp Gly Val Tyr Thr Ala Asp Pro Arg Ile Val Pro Asn Ala Gln Lys 180 185 190 ctg gaa aag ctc agc ttc gaa gaa atg ctg gaa ctt gct gct gtt ggc 624 Leu Glu Lys Leu Ser Phe Glu Glu Met Leu Glu Leu Ala Ala Val Gly 195 200 205 tcc aag att ttg gtg ctg cgc agt gtt gaa tac gct cgt gca ttc aat 672 Ser Lys Ile Leu Val Leu Arg Ser Val Glu Tyr Ala Arg Ala Phe Asn 210 215 220 gtg cca ctt cgc gta cgc tcg tct tat agt aat gat ccc ggc act ttg 720 Val Pro Leu Arg Val Arg Ser Ser Tyr Ser Asn Asp Pro Gly Thr Leu 225 230 235 240 att gcc ggc tct atg gag gat att cct gtg gaa gaa gca gtc ctt acc 768 Ile Ala Gly Ser Met Glu Asp Ile Pro Val Glu Glu Ala Val Leu Thr 245 250 255 ggt gtc gca acc gac aag tcc gaa gcc aaa gta acc gtt ctg ggt att 816 Gly Val Ala Thr Asp Lys Ser Glu Ala Lys Val Thr Val Leu Gly Ile 260 265 270 tcc gat aag cca ggc gag gct gcg aag gtt ttc cgt gcg ttg gct gat 864 Ser Asp Lys Pro Gly Glu Ala Ala Lys Val Phe Arg Ala Leu Ala Asp 275 280 285 gca gaa atc aac att gac atg gtt ctg cag aac gtc tct tct gta gaa 912 Ala Glu Ile Asn Ile Asp Met Val Leu Gln Asn Val Ser Ser Val Glu 290 295 300 gac ggc acc acc gac atc atc ttc acc tgc cct cgt tcc gac ggc cgc 960 Asp Gly Thr Thr Asp Ile Ile Phe Thr Cys Pro Arg Ser Asp Gly Arg 305 310 315 320 cgc gcg atg gag atc ttg aag aag ctt cag gtt cag ggc aac tgg acc 1008 Arg Ala Met Glu Ile Leu Lys Lys Leu Gln Val Gln Gly Asn Trp Thr 325 330 335 aat gtg ctt tac gac gac cag gtc ggc aaa gtc tcc ctc gtg ggt gct 1056 Asn Val Leu Tyr Asp Asp Gln Val Gly Lys Val Ser Leu Val Gly Ala 340 345 350 ggc atg aag tct cac cca ggt gtt acc gca gag ttc atg gaa gct ctg 1104 Gly Met Lys Ser His Pro Gly Val Thr Ala Glu Phe Met Glu Ala Leu 355 360 365 cgc gat gtc aac gtg aac atc gaa ttg att tcc acc tct gag att cgt 1152 Arg Asp Val Asn Val Asn Ile Glu Leu Ile Ser Thr Ser Glu Ile Arg 370 375 380 att tcc gtg ctg atc cgt gaa gat gat ctg gat gct gct gca cgt gca 1200 Ile Ser Val Leu Ile Arg Glu Asp Asp Leu Asp Ala Ala Ala Arg Ala 385 390 395 400 ttg cat gag cag ttc cag ctg ggc ggc gaa gac gaa gcc gtc gtt tat 1248 Leu His Glu Gln Phe Gln Leu Gly Gly Glu Asp Glu Ala Val Val Tyr 405 410 415 gca ggc acc gga cgc taa 1266 Ala Gly Thr Gly Arg 420 65 421 PRT LysC mutant 65 Val Ala Leu Val Val Gln Lys Tyr Gly Gly Ser Ser Leu Glu Ser Ala 1 5 10 15 Glu Arg Ile Arg Asn Val Ala Glu Arg Ile Val Ala Thr Lys Lys Ala 20 25 30 Gly Asn Asp Val Val Val Val Cys Ser Ala Met Gly Asp Thr Thr Asp 35 40 45 Glu Leu Leu Glu Leu Ala Ala Ala Val Asn Pro Val Pro Pro Ala Arg 50 55 60 Glu Met Asp Met Leu Leu Thr Ala Gly Glu Arg Ile Ser Asn Ala Leu 65 70 75 80 Val Ala Met Ala Ile Glu Ser Leu Gly Ala Glu Ala Gln Ser Phe Thr 85 90 95 Gly Ser Gln Ala Gly Val Leu Thr Thr Glu Arg His Gly Asn Ala Arg 100 105 110 Ile Val Asp Val Thr Pro Gly Arg Val Arg Glu Ala Leu Asp Glu Gly 115 120 125 Lys Ile Cys Ile Val Ala Gly Phe Gln Gly Val Asn Lys Glu Thr Arg 130 135 140 Asp Val Thr Thr Leu Gly Arg Gly Gly Ser Asp Thr Thr Ala Val Ala 145 150 155 160 Leu Ala Ala Ala Leu Asn Ala Asp Val Cys Glu Ile Tyr Ser Asp Val 165 170 175 Asp Gly Val Tyr Thr Ala Asp Pro Arg Ile Val Pro Asn Ala Gln Lys 180 185 190 Leu Glu Lys Leu Ser Phe Glu Glu Met Leu Glu Leu Ala Ala Val Gly 195 200 205 Ser Lys Ile Leu Val Leu Arg Ser Val Glu Tyr Ala Arg Ala Phe Asn 210 215 220 Val Pro Leu Arg Val Arg Ser Ser Tyr Ser Asn Asp Pro Gly Thr Leu 225 230 235 240 Ile Ala Gly Ser Met Glu Asp Ile Pro Val Glu Glu Ala Val Leu Thr 245 250 255 Gly Val Ala Thr Asp Lys Ser Glu Ala Lys Val Thr Val Leu Gly Ile 260 265 270 Ser Asp Lys Pro Gly Glu Ala Ala Lys Val Phe Arg Ala Leu Ala Asp 275 280 285 Ala Glu Ile Asn Ile Asp Met Val Leu Gln Asn Val Ser Ser Val Glu 290 295 300 Asp Gly Thr Thr Asp Ile Ile Phe Thr Cys Pro Arg Ser Asp Gly Arg 305 310 315 320 Arg Ala Met Glu Ile Leu Lys Lys Leu Gln Val Gln Gly Asn Trp Thr 325 330 335 Asn Val Leu Tyr Asp Asp Gln Val Gly Lys Val Ser Leu Val Gly Ala 340 345 350 Gly Met Lys Ser His Pro Gly Val Thr Ala Glu Phe Met Glu Ala Leu 355 360 365 Arg Asp Val Asn Val Asn Ile Glu Leu Ile Ser Thr Ser Glu Ile Arg 370 375 380 Ile Ser Val Leu Ile Arg Glu Asp Asp Leu Asp Ala Ala Ala Arg Ala 385 390 395 400 Leu His Glu Gln Phe Gln Leu Gly Gly Glu Asp Glu Ala Val Val Tyr 405 410 415 Ala Gly Thr Gly Arg 420 66 5860 DNA Artificial sequence Description of the artificial sequenceplasmid 66 cccggtacca cgcgtcccag tggctgagac gcatccgcta aagccccagg aaccctgtgc 60 agaaagaaaa cactcctctg gctaggtaga cacagtttat aaaggtagag ttgagcgggt 120 aactgtcagc acgtagatcg aaaggtgcac aaaggtggcc ctggtcgtac agaaatatgg 180 cggttcctcg cttgagagtg cggaacgcat tagaaacgtc gctgaacgga tcgttgccac 240 caagaaggct ggaaatgatg tcgtggttgt ctgctccgca atgggagaca ccacggatga 300 acttctagaa cttgcagcgg cagtgaatcc cgttccgcca gctcgtgaaa tggatatgct 360 cctgactgct ggtgagcgta tttctaacgc tctcgtcgcc atggctattg agtcccttgg 420 cgcagaagcc caatctttca cgggctctca ggctggtgtg ctcaccaccg agcgccacgg 480 aaacgcacgc attgttgatg tcactccagg tcgtgtgcgt gaagcactcg atgagggcaa 540 gatctgcatt gttgctggtt tccagggtgt taataaagaa acccgcgatg tcaccacgtt 600 gggtcgtggt ggttctgaca ccactgcagt tgcgttggca gctgctttga acgctgatgt 660 gtgtgagatt tactcggacg ttgacggtgt gtataccgct gacccgcgca tcgttcctaa 720 tgcacagaag ctggaaaagc tcagcttcga agaaatgctg gaacttgctg ctgttggctc 780 caagattttg gtgctgcgca gtgttgaata cgctcgtgca ttcaatgtgc cacttcgcgt 840 acgctcgtct tatagtaatg atcccggcac tttgattgcc ggctctatgg aggatattcc 900 tgtggaagaa gcagtcctta ccggtgtcgc aaccgacaag tccgaagcca aagtaaccgt 960 tctgggtatt tccgataagc caggcgaggc tgcgaaggtt ttccgtgcgt tggctgatgc 1020 agaaatcaac attgacatgg ttctgcagaa cgtctcttct gtagaagacg gcaccaccga 1080 catcatcttc acctgccctc gttccgacgg ccgccgcgcg atggagatct tgaagaagct 1140 tcaggttcag ggcaactgga ccaatgtgct ttacgacgac caggtcggca aagtctccct 1200 cgtgggtgct ggcatgaagt ctcacccagg tgttaccgca gagttcatgg aagctctgcg 1260 cgatgtcaac gtgaacatcg aattgatttc cacctctgag attcgtattt ccgtgctgat 1320 ccgtgaagat gatctggatg ctgctgcacg tgcattgcat gagcagttcc agctgggcgg 1380 cgaagacgaa gccgtcgttt atgcaggcac cggacgctaa agttttaaag gagtagtttt 1440 acaatgacca ccatcgcagt tgttggtgca accggccagg tcggccaggt tatgcgcacc 1500 cttttggaag agcgcaattt cccagctgac actgttcgtt tctttgcttc cccacgttcc 1560 gcaggccgta agattgaatt cgtcgacatc gatgctcttc tgcgttaatt aacaattggg 1620 atcctctaga cccgggattt aaatcgctag cgggctgcta aaggaagcgg aacacgtaga 1680 aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga 1740 caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat 1800 agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct 1860 ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct 1920 gatggcgcag gggatcaaga tctgatcaag agacaggatg aggatcgttt cgcatgattg 1980 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg 2040 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg 2100 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg 2160 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg 2220 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc 2280 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc 2340 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc 2400 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc 2460 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg 2520 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct 2580 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt 2640 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc 2700 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt 2760 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc 2820 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg 2880 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccacgc 2940 tagcggcgcg ccggccggcc cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc 3000 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 3060 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata 3120 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 3180 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 3240 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 3300 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 3360 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 3420 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 3480 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 3540 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 3600 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc 3660 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 3720 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 3780 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 3840 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaaggccg 3900 gccgcggccg ccatcggcat tttcttttgc gtttttattt gttaactgtt aattgtcctt 3960 gttcaaggat gctgtctttg acaacagatg ttttcttgcc tttgatgttc agcaggaagc 4020 tcggcgcaaa cgttgattgt ttgtctgcgt agaatcctct gtttgtcata tagcttgtaa 4080 tcacgacatt gtttcctttc gcttgaggta cagcgaagtg tgagtaagta aaggttacat 4140 cgttaggatc aagatccatt tttaacacaa ggccagtttt gttcagcggc ttgtatgggc 4200 cagttaaaga attagaaaca taaccaagca tgtaaatatc gttagacgta atgccgtcaa 4260 tcgtcatttt tgatccgcgg gagtcagtga acaggtacca tttgccgttc attttaaaga 4320 cgttcgcgcg ttcaatttca tctgttactg tgttagatgc aatcagcggt ttcatcactt 4380 ttttcagtgt gtaatcatcg tttagctcaa tcataccgag agcgccgttt gctaactcag 4440 ccgtgcgttt tttatcgctt tgcagaagtt tttgactttc ttgacggaag aatgatgtgc 4500 ttttgccata gtatgctttg ttaaataaag attcttcgcc ttggtagcca tcttcagttc 4560 cagtgtttgc ttcaaatact aagtatttgt ggcctttatc ttctacgtag tgaggatctc 4620 tcagcgtatg gttgtcgcct gagctgtagt tgccttcatc gatgaactgc tgtacatttt 4680 gatacgtttt tccgtcaccg tcaaagattg atttataatc ctctacaccg ttgatgttca 4740 aagagctgtc tgatgctgat acgttaactt gtgcagttgt cagtgtttgt ttgccgtaat 4800 gtttaccgga gaaatcagtg tagaataaac ggatttttcc gtcagatgta aatgtggctg 4860 aacctgacca ttcttgtgtt tggtctttta ggatagaatc atttgcatcg aatttgtcgc 4920 tgtctttaaa gacgcggcca gcgtttttcc agctgtcaat agaagtttcg ccgacttttt 4980 gatagaacat gtaaatcgat gtgtcatccg catttttagg atctccggct aatgcaaaga 5040 cgatgtggta gccgtgatag tttgcgacag tgccgtcagc gttttgtaat ggccagctgt 5100 cccaaacgtc caggcctttt gcagaagaga tatttttaat tgtggacgaa tcaaattcag 5160 aaacttgata tttttcattt ttttgctgtt cagggatttg cagcatatca tggcgtgtaa 5220 tatgggaaat gccgtatgtt tccttatatg gcttttggtt cgtttctttc gcaaacgctt 5280 gagttgcgcc tcctgccagc agtgcggtag taaaggttaa tactgttgct tgttttgcaa 5340 actttttgat gttcatcgtt catgtctcct tttttatgta ctgtgttagc ggtctgcttc 5400 ttccagccct cctgtttgaa gatggcaagt tagttacgca caataaaaaa agacctaaaa 5460 tatgtaaggg gtgacgccaa agtatacact ttgcccttta cacattttag gtcttgcctg 5520 ctttatcagt aacaaacccg cgcgatttac ttttcgacct cattctatta gactctcgtt 5580 tggattgcaa ctggtctatt ttcctctttt gtttgataga aaatcataaa aggatttgca 5640 gactacgggc ctaaagaact aaaaaatcta tctgtttctt ttcattctct gtatttttta 5700 tagtttctgt tgcatgggca taaagttgcc tttttaatca caattcagaa aatatcataa 5760 tatctcattt cactaaataa tagtgaacgg caggtatatg tgatgggtta aaaaggatcg 5820 gcggccgctc gatttaaatc tcgagaggcc tgacgtcggg 5860 67 29 DNA Artificial sequence Description of the artificial sequencePCR primer 67 gagactcgag gttggctggt catcatagg 29 68 34 DNA Artificial sequence Description of the artificial sequencePCR primer 68 gaagagagca tatgtcagcg ctctagtttg gttc 34 69 6472 DNA Artificial sequence Description of the artificial sequenceplasmid 69 tcgaggttgg ctggtcatca taggaatcaa cctggccact ttatggtggg caccaccgtc 60 gcaaacaaca tatcttgcag caggcgtgtc gattctttcc gccatcattg tttggtttct 120 tcccggcgca cacccgctat ggaatcgccg tcgcattgct tcacgcaaac aacagtccac 180 cggtagacgt cgacaagccc ccaaacgatc aagccaccct caaacggcgg aatttagcca 240 acaacaatag actagacaga gctgtccatg tagcatgaac tcgattatca actgccacga 300 gaggtcgggg tcatgctcac caccacaggg acgctcacgc accaaaaaat cggagacttt 360 tacaccgaag ccggagcgac gcttcacgac gtaaccatcg cctaccaagc atggggccac 420 tacaccggca ccaatctcat cgttctcgaa catgccctga ccggcgactc taacgctatt 480 tcatggtggg acggactgat tggccctggc aaagcactcg acaccaaccg ctactgcatc 540 ctatgcacca acgtgctcgg aggatgcaaa ggatccaccg gaccgagcag tccacaccca 600 gacggaaaac catggggatc cagatttcca gccctttcaa tccgtgacct tgtcaatgcc 660 gaaaaacaac ttttcgacca cctcggcatc aataaaattc acgcaatcat cggcggatcc 720 atgggaggcg cacgcaccct cgaatgggct gcactccacc cacacatgat gacgactgga 780 ttcgtcatag cagtctcagc acgcgcaagc gcttggcaaa tcggtattca aactgcacaa 840 atcagcgcca tagaactcga cccccactgg aacggcggcg attactacag cggtcacgca 900 ccatgggaag gaatcgccgc cgctcgccgg atcgcccacc tcacctatcg cggcgaacta 960 gaaatagacg aacgattcgg cacttccgca caacacggtg aaaacccact cggccccttc 1020 cgagatccac atcaacgttt tgcggtcacg agctacctcc aacaccaagg catcaaactc 1080 gctcaacgat tcgatgcagg tagttacgtc gtgcttaccg aagccctcaa tcgtcatgac 1140 atcggacgcg gccgaggcgg actcaacaaa gccctcagcg caatcacagt ccccatcatg 1200 attgctggcg ttgataccga tattctctac ccctatcacc agcaagaaca cctatcacga 1260 aatctaggca acctactcgc tatggcaaaa atcagctcac cagtaggcca cgacgctttc 1320 ctcacagaat tccgacaaat ggagcgaatc ctaagacatt tcatggagct ttcggaagga 1380 atcgacgatt ccttccgaac caaactagag cgctgacata tgactagttc ggacctaggg 1440 atatcgtcga catcgatgct cttctgcgtt aattaacaat tgggatcctc tagacccggg 1500 atttaaatcg ctagcgggct gctaaaggaa gcggaacacg tagaaagcca gtccgcagaa 1560 acggtgctga ccccggatga atgtcagcta ctgggctatc tggacaaggg aaaacgcaag 1620 cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg cgatagctag actgggcggt 1680 tttatggaca gcaagcgaac cggaattgcc agctggggcg ccctctggta aggttgggaa 1740 gccctgcaaa gtaaactgga tggctttctt gccgccaagg atctgatggc gcaggggatc 1800 aagatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca 1860 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac 1920 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt 1980 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc 2040 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg 2100 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc 2160 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc 2220 ggctacctgc ccattcgacc accaagcgaa

acatcgcatc gagcgagcac gtactcggat 2280 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc 2340 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca 2400 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga 2460 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat 2520 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc 2580 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact 2640 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc 2700 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg 2760 atcctccagc gcggggatct catgctggag ttcttcgccc acgctagcgg cgcgccggcc 2820 ggcccggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc 2880 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc 2940 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat 3000 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt 3060 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg 3120 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 3180 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 3240 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 3300 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 3360 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 3420 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 3480 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt 3540 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 3600 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 3660 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 3720 gagattatca aaaaggatct tcacctagat ccttttaaag gccggccgcg gccgcgcaaa 3780 gtcccgcttc gtgaaaattt tcgtgccgcg tgattttccg ccaaaaactt taacgaacgt 3840 tcgttataat ggtgtcatga ccttcacgac gaagtactaa aattggcccg aatcatcagc 3900 tatggatctc tctgatgtcg cgctggagtc cgacgcgctc gatgctgccg tcgatttaaa 3960 aacggtgatc ggatttttcc gagctctcga tacgacggac gcgccagcat cacgagactg 4020 ggccagtgcc gcgagcgacc tagaaactct cgtggcggat cttgaggagc tggctgacga 4080 gctgcgtgct cggccagcgc caggaggacg cacagtagtg gaggatgcaa tcagttgcgc 4140 ctactgcggt ggcctgattc ctccccggcc tgacccgcga ggacggcgcg caaaatattg 4200 ctcagatgcg tgtcgtgccg cagccagccg cgagcgcgcc aacaaacgcc acgccgagga 4260 gctggaggcg gctaggtcgc aaatggcgct ggaagtgcgt cccccgagcg aaattttggc 4320 catggtcgtc acagagctgg aagcggcagc gagaattatc gcgatcgtgg cggtgcccgc 4380 aggcatgaca aacatcgtaa atgccgcgtt tcgtgtgccg tggccgccca ggacgtgtca 4440 gcgccgccac cacctgcacc gaatcggcag cagcgtcgcg cgtcgaaaaa gcgcacaggc 4500 ggcaagaagc gataagctgc acgaatacct gaaaaatgtt gaacgccccg tgagcggtaa 4560 ctcacagggc gtcggctaac ccccagtcca aacctgggag aaagcgctca aaaatgactc 4620 tagcggattc acgagacatt gacacaccgg cctggaaatt ttccgctgat ctgttcgaca 4680 cccatcccga gctcgcgctg cgatcacgtg gctggacgag cgaagaccgc cgcgaattcc 4740 tcgctcacct gggcagagaa aatttccagg gcagcaagac ccgcgacttc gccagcgctt 4800 ggatcaaaga cccggacacg gagaaacaca gccgaagtta taccgagttg gttcaaaatc 4860 gcttgcccgg tgccagtatg ttgctctgac gcacgcgcag cacgcagccg tgcttgtcct 4920 ggacattgat gtgccgagcc accaggccgg cgggaaaatc gagcacgtaa accccgaggt 4980 ctacgcgatt ttggagcgct gggcacgcct ggaaaaagcg ccagcttgga tcggcgtgaa 5040 tccactgagc gggaaatgcc agctcatctg gctcattgat ccggtgtatg ccgcagcagg 5100 catgagcagc ccgaatatgc gcctgctggc tgcaacgacc gaggaaatga cccgcgtttt 5160 cggcgctgac caggcttttt cacataggct gagccgtggc cactgcactc tccgacgatc 5220 ccagccgtac cgctggcatg cccagcacaa tcgcgtggat cgcctagctg atcttatgga 5280 ggttgctcgc atgatctcag gcacagaaaa acctaaaaaa cgctatgagc aggagttttc 5340 tagcggacgg gcacgtatcg aagcggcaag aaaagccact gcggaagcaa aagcacttgc 5400 cacgcttgaa gcaagcctgc cgagcgccgc tgaagcgtct ggagagctga tcgacggcgt 5460 ccgtgtcctc tggactgctc cagggcgtgc cgcccgtgat gagacggctt ttcgccacgc 5520 tttgactgtg ggataccagt taaaagcggc tggtgagcgc ctaaaagaca ccaagggtca 5580 tcgagcctac gagcgtgcct acaccgtcgc tcaggcggtc ggaggaggcc gtgagcctga 5640 tctgccgccg gactgtgacc gccagacgga ttggccgcga cgtgtgcgcg gctacgtcgc 5700 taaaggccag ccagtcgtcc ctgctcgtca gacagagacg cagagccagc cgaggcgaaa 5760 agctctggcc actatgggaa gacgtggcgg taaaaaggcc gcagaacgct ggaaagaccc 5820 aaacagtgag tacgcccgag cacagcgaga aaaactagct aagtccagtc aacgacaagc 5880 taggaaagct aaaggaaatc gcttgaccat tgcaggttgg tttatgactg ttgagggaga 5940 gactggctcg tggccgacaa tcaatgaagc tatgtctgaa tttagcgtgt cacgtcagac 6000 cgtgaataga gcacttaagg tctgcgggca ttgaacttcc acgaggacgc cgaaagcttc 6060 ccagtaaatg tgccatctcg taggcagaaa acggttcccc cgtagggtct ctctcttggc 6120 ctcctttcta ggtcgggctg attgctcttg aagctctcta ggggggctca caccataggc 6180 agataacgtt ccccaccggc tcgcctcgta agcgcacaag gactgctccc aaagatcttc 6240 aaagccactg ccgcgactgc cttcgcgaag ccttgccccg cggaaatttc ctccaccgag 6300 ttcgtgcaca cccctatgcc aagcttcttt caccctaaat tcgagagatt ggattcttac 6360 cgtggaaatt cttcgcaaaa atcgtcccct gatcgccctt gcgacgttgg cgtcggtgcc 6420 gctggttgcg cttggcttga ccgacttgat cagcggccgc tcgatttaaa tc 6472

Claims

1. A method for the fermentative production of L-methionine, which comprises the following steps: a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, the coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes for a protein with homoserine O-acetyltransferase (metA) activity, wherein said heterologous nucleotide sequence comprises a nucleotide sequence encoding a metA protein having an amino acid sequence with 95% homology or more to the sequence as set forth in SEQ ID NO: 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, or 46; b) concentrating L-methionine in the medium or in the bacterial cells; and c) isolating L-methionine.

2. The method of claim 1, wherein the metA-encoding sequence is derived from any of the following organisms: TABLE-US-00014 Mycobacterium leprae ATCC 43910 Mycobacterium tuberculosis CDC1551 ATCC 25584 Chlorobium tepidum ATCC 49652 Pseudomonas aeruginosa ATCC 17933 Caulobacter crescentus ATCC 19089 Neisseria gonorrhoeae ATCC 53420 Neisseria meningitidis ATCC 53414 Pseudomonas fluorescens ATCC 13525 Burkholderia cepacia ATCC 25416 Nitrosomonas europaea ATCC 19718 Haemophilus influenzae ATCC 51907 Halobacterium sp NRC1 ATCC 33170 Thermus thermophilus ATCC 27634 Deinococcus radiodurans ATCC 13939 Saccharomyces cerevisiae ATCC 10751 Schizosaccharomyces pombe ATCC 24969 Xylella fastidiosa ATCC 35881 Emericella nidulans ATCC 36104 Mesorhizobium loti ATCC 35173 Acremonium crysogenum ATCC 11550 Pseudomonas putida ATCC 47054 Staphylococcus aureus ATCC 35556

3. The method of claim 1, wherein the meta-encoding sequence comprises a coding sequence as set forth in SEQ ID NO: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, or 45.

4. The method of claim 1, wherein the meta-encoding sequence encodes for a protein with meta activity, said protein comprising an amino acid sequence as set forth in SEQ ID NO: 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44 or 46.

5. The method of claim 1, wherein the meta encoding sequence is a DNA or RNA which can be replicated in coryneform bacteria or is stably integrated into the chromosome.

6. The method of claim 5, wherein a) a bacteria strain transformed with a plasmid vector carrying at least one copy of the meta encoding sequence under the control of regulatory sequences is used, or b) a strain in which the meta encoding sequence has been integrated into the bacteria chromosome is used.

7. The method of claim 1, wherein the meta encoding sequence is overexpressed.

8. The method of claim 1, wherein bacteria are fermented in which additionally at least one further gene of the biosynthetic pathway of L-methionine has been overexpressed or mutated such that its activity is not influenced by metabolic metabolites.

9. The method of claim 1, wherein bacteria are fermented in which at least one metabolic pathway, which reduces the production of L-methionine, is at least partially switched off.

10. The method of claim 1, wherein coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among a) a lysC gene derived from a coryneform bacterium, which encodes an aspartate kinase, b) a glyceraldehyde-3-phosphate dehydrogenase-encoding gene gap, c) a 3-phosphoglycerate kinase-encoding gene pgk, d) a pyruvate carboxylase-encoding gene pyc, e) a triose phosphate isomerase-encoding gene tpi, f) a methylene tetrahydrofolate reductase-encoding gene metF, g) a cystathionine gamma-synthase-encoding gene metB, h) a cystathionine gamma-lyase-encoding gene metC, i) a serine hydroxymethyltransferase-encoding gene glyA, j) a O-acetylhomoserine sulfhydrylase-encoding gene metY, k) a vitamin B12-dependent methionine synthase-encoding gene metH, l) a phosphoserine aminotransferase-encoding gene serC, m) a phosphoserine phosphatase-encoding gene serB, n) a serine acetyltransferase-encoding gene cysE, and o) a gene hom, which encodes a homoserine dehydrogenase, is overexpressed or mutated in such a way that the activity of the corresponding proteins is influenced by metabolic metabolites to a smaller extent, if at all, compared to nonmutated proteins.

11. The method of claim 1, wherein coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among a) a homoserine kinase-encoding gene thrB, b) a threonine dehydratase-encoding gene ilvA, c) a threonine synthase-encoding gene thrC, d) a meso-diaminopimelate D-dehydrogenase-encoding gene ddh, e) a phosphoenolpyruvate carboxykinase-encoding gene pck, f) a glucose-6-phosphate 6-isomerase-encoding gene pgi, g) a pyruvate oxidase-encoding gene poxB, h) a dihydrodipicolinate synthase-encoding gene dapA, i) a dihydrodipicolinate reductase-encoding gene dapB; and j) a diaminopicolinate decarboxylase-encoding gene, is attenuated by changing the rate of expression or by introducing a specific mutation.

12. The method of claim 1, wherein the coryneform bacterium is of the species Corynebacterium glutamicum.

13. A method for producing an L-methionine-containing animal feed additive from a fermentation medium, comprising a) fermenting in a fermentation medium cells of a coryneform bacterium for producing L-methionine, the coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes for a protein with homoserine O-acetyltransferase (meta) activity, wherein said heterologous nucleotide sequence comprises a nucleotide sequence encoding a meta protein having an amino acid sequence with 95% homology or more to the sequence as set forth in SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, or 46; b) removing water from the L-methionine-containing fermentation medium and/or removing from 0 to 100% by weight of biomass formed during fermentation; and c) drying the medium to obtain an L-methionine-containing animal feed additive in a powder or a granule form.

14. A method for the production of L-methionine, which comprises the following steps: a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, said coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes a protein with homoserine O-acetyltransferase (meta) activity, wherein the heterologous nucleotide sequence comprises a nucleotide sequence having 95% homology or more to the sequence as set forth in SEQ ID NO: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, or 45; b) concentrating L-methionine in the medium or in the bacterial cells; and c) isolating L-methionine.

15. The method of claim 14, wherein the meta-encoding nucleotide sequence comprises a nucleotide sequence as set forth in SEQ ID NO: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, or 45.

16. The method of claim 14, wherein the meta-encoding nucleotide sequence encodes a protein with meta activity, said protein comprising an amino acid sequence as set forth in SEQ ID NO: 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44 or 46.

17. The method of claim 14, wherein the metA encoding sequence is a DNA or RNA which can be replicated in coryneform bacteria or is stably integrated into the chromosome.

18. The method of claim 14, wherein a) a bacteria strain transformed with a plasmid vector carrying at least one copy of the metA encoding sequence under the control of regulatory sequences is used, or b) a strain in which the meta encoding sequence has been integrated into the bacteria chromosome is used.

19. The method of claim 14, wherein the meta encoding sequence is overexpressed.

20. The method of claim 14, wherein bacteria are fermented in which additionally at least one further gene of the biosynthetic pathway of L-methionine has been overexpressed or mutated such that its activity is not influenced by metabolic metabolites.

21. The method of claim 14, wherein the coryneform bacterium is of the species Corynebacterium glutamicum.

22. A method for producing an L-methionine-containing animal feed additive from a fermentation medium, comprising a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, said coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes a protein with homoserine O-acetyltransferase (meta) activity, wherein the heterologous nucleotide sequence comprises a nucleotide sequence having 95% homology or more to the sequence as set forth in SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, or 45; b) removing water from the L-methionine-containing fermentation medium and/or removing from 0 to 100% by weight of biomass formed during fermentation; and c) drying the medium to obtain an L-methionine-containing animal feed additive in a powder or a granule form.

23. A method for the fermentative production of L-methionine, which comprises the following steps: a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, the coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes for a protein with homoserine O-acetyltransferase (meta) activity, wherein said heterologous nucleotide sequence comprises a nucleotide sequence encoding a metA protein having an amino acid sequence with 95% homology or more to the sequence as set forth in SEQ ID NO: 2; b) concentrating L-methionine in the medium or in the bacterial cells; and c) isolating L-methionine; wherein coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among a) a gene lysC derived from a coryneform bacterium, which encodes an aspartate kinase, a) a glyceraldehyde-3-phosphate dehydrogenase-encoding gene gap, b) a 3-phosphoglycerate kinase-encoding gene pgk, c) a pyruvate carboxylase-encoding gene pyc, d) a triose phosphate isomerase-encoding gene tpi, e) a methylene tetrahydrofolate reductase-encoding gene metF, f) a cystathionine gamma-synthase-encoding gene metB, g) a cystathionine gamma-lyase-encoding gene metC, h) a serine hydroxymethyltransferase-encoding gene glyA, i) a O-acetylhomoserine sulfhydrylase-encoding gene metY, j) a vitamin B12-dependent methionine synthase-encoding gene metH, k) a phosphoserine aminotransferase-encoding gene serC, l) a phosphoserine phosphatase-encoding gene serB, m) a serine acetyltransferase-encoding gene cysE, and n) a gene hom, which encodes a homoserine dehydrogenase, is overexpressed or mutated in such a way that the activity of the corresponding proteins is influenced by metabolic metabolites to a smaller extent, if at all, compared to nonmutated proteins.

24. A method for the fermentative production of L-methionine, which comprises the following steps: a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, the coryneform bacteria expressing at least one heterologous nucleotide sequence which encodes for a protein with homoserine O-acetyltransferase (metA) activity, wherein said heterologous nucleotide sequence comprises a nucleotide sequence encoding a metA protein having an amino acid sequence with 95% homology or more to the sequence as set forth in SEQ ID NO: 2; b) concentrating L-methionine in the medium or in the bacterial cells; and c) isolating L-methionine; wherein coryneform bacteria are fermented in which, at the same time, at least one of the genes selected from among a) a homoserine kinase-encoding gene thrB, b) a threonine dehydratase-encoding gene ilvA, c) a threonine synthase-encoding gene thrC, d) a meso-diaminopimelate D-dehydrogenase-encoding gene ddh, e) a phosphoenolpyruvate carboxykinase-encoding gene pck, f) a glucose-6-phosphate 6-isomerase-encoding gene pgi, g) a pyruvate oxidase-encoding gene poxB, h) a dihydrodipicolinate synthase-encoding gene dapA, i) a dihydrodipicolinate reductase-encoding gene dapB; and j) a diaminopicolinate decarboxylase-encoding gene, is attenuated by changing the rate of expression or by introducing a specific mutation.