KR20070077628A - Fusaricidin synthetase and gene thereof - Google Patents

Abstract

A fusaricidin synthetase isolated from Paenibacillus polymyxa E681 strain is provided to increase the productivity of the fusaricidin and be usefully used for preparing a novel antibiotic. The polypeptide concerned with the synthesis of fusaricidin is described as SEQ ID : NO. 2, wherein the fusaricidin is fusaricidin A, fusaricidin B, fusaricidin C, fusaricidin D, LI-F03, LI-F04, LI-F05, LI-F07, or LI-F08. The method for preparing the fusaricidin or a derivative thereof comprises the steps of: (a) introducing a gene coding the polypeptide into an expression vector; (b) introducing the expression vector into a host cell to transform it; (c) culturing the transformant; and (d) isolating and purifying the fusaricidin or the derivative thereof from the culture material obtained from the step(c).

Description

Fusaricidin synthetase and gene etc}

1 is a diagram showing the structure of fuzacidin isolated from the strains of Panibacillus polymixsa E681;

FIG. 2 is a diagram showing the domain structure of the fuzzy cydin biosynthetic enzyme gene found from the genome of the F. genus E681 strain;

A: A (adenylation) domain;

C: C (condensation) domain;

E: E (epimeration) domain;

T: T (thiolation) domain and

TE: TE (termination) domain,

FIG. 3 is a diagram showing the structure of fuzacidin predicted from the domain structure of the fuzacidine biosynthetic enzyme gene found from the genome of the F. genus E681 strain.

The present invention relates to a fuzacidin biosynthetic enzyme isolated from a genus of Paenibacillus , a Gram-positive bacterium, and a gene encoding the same. Specifically, the present invention relates to a penibacilli polymyxa E681 ( Paenibacillus). polymyxa E681) relates to a fuzacidine biosynthetic enzyme isolated from a strain, a gene encoding the same, a method for producing fuzacidine or a derivative thereof using the gene.

Non-ribosomal peptide synthetase (abbreviated as 'NRPS') consists of one or more open reading frames (ORFs) that make up the NRPS complex, and each NRPS or NRPS subunit contains one or more modules. do. A module is defined as a catalytic unit that connects one building block (eg, one amino acid) to a growing peptide chain. The order and specificity of the biosynthetic module forming the NRPS protein template governs the sequence and structure of the final peptide product. Thus, NRPS processes not related to ribosomal-mediated RNA translation according to the genetic code can produce peptides that exhibit vast structural diversity compared to peptides translated from RNA templates by ribosomes. These include the formation of linkages of D- and L-amino acids with hydroxy acids, mutations in the main peptide chain forming linear, cyclic or branched cyclic structures, and oxidation, acylation, glycosylation, N-methylation and heterocycle ring formation. Additional structural modifications included.

Fuzisidine biosynthetic enzyme, one of NRPS, sequentially binds each amino acid monomer constituting fuzacidin, and induces modification of amino acids if necessary to complete chain of whole amino acids and form ring structure to synthesize peptide antibiotic do. Each module of NRPS consists of at least A, C and T domains, where the A domain (adenylation domain) selects and activates amino acid monomers, the C domain (condensation domain) catalyzes the formation of peptide bonds, and the T domain (thiolation domain). , Also called PCP), is responsible for rotating the phosphopantheteine group to deliver amino acid monomers to the peptide chain under synthesis.

Recently, the tertiary structure of the A domain, which recognizes phenylalanine in the Gramicidin biosynthesis gene, has been identified, revealing that eight amino acid residues are involved in the site of binding to specific amino acids (Conti E. et al., EMBO J). 16: 4174-4183, 1997). Comparing the amino acid sequence of the A domain with the amino acid sequence of the known A domain, it was confirmed that the A domain that recognizes the same amino acid has very high homology with the 8 amino acid residues, and thus the 8 amino acids Analysis of the residues reveals which amino acids a particular A domain recognizes and binds (Challis GL et al., Chem . Biol . 7: 211-224. 2000).

In addition to these core domains, there are E domains and TE domains, which serve to convert L-amino acids to D-amino acids, which are generally driven by specific amino acid motifs or features. Is characterized.

Modular structures can be used to design new enzymes with new specificities by varying the number and location of modules at the DNA level by genetic engineering manipulations and in vivo recombination. For example, residues that exchange domains derived from heterologous NRPS using recombinant techniques (Schneider et al., Mol . Gen. Genet. , 257: 308-318, 1998) or form substrate binding pockets of the A domain. By designing new substrate specificities in the module by changing the chemistry (Cane et al., Chem . Biol . Vol . 6: 319-325, 1999).

Fuzacidin is an antibiotic isolated from the genus of Panibacillus, consisting of a ring structure of six amino acid residues, and containing guanidino-3-hydroxypentadecanoic acid (15-guanidino-3-hydroxypentadecanoic acid). (See Figure 3). So far, fujarycidin LI-F03, LI-F04, LI-F05, LI-F07 and LI-F08 have been reported from Penicillus polymix (Kurusu K, Ohba K, Arai T and Fukushima K. J. Antibiotics 40: 1506-1514, 1987), fuzisidine A, B, C and D (Kajimura Y and Kaneda M. J. Antibiotics 49: 129-135, 1996; Kajimura Y and Kaneda M. J. Antibiotics 50: 220-228, 1997). The amino acid chains of fuzacidin are not encoded by genes and synthesized by ribosomes like ordinary polypeptides, but by enzymes called non-ribosomal peptide synthetase (NRPS) (Marahiel MA, Stachelhaus T and ... Mootz HD Chem Rev 97 : 2651-2673, 1997; Doekel S and Marahiel MA Metab Eng 6:... 64-77, 2001).

Fuzacidin is a Fusarium of Fusarium oxysporum), Aspergillus and this (Aspergillus niger), Aspergillus duck material (Aspergillus oryzae), and has excellent fungicidal activity against phytopathogenic fungi, such as penny room Solarium somi (Penicillum thomii) Din when especially purpurea spot B is Candida albicans (Candida albicans ) and Saccharomyces cerevisiae also have bactericidal activity. Fuzacidin also has excellent bactericidal activity against Gram-positive bacteria, including Staphylococcus aureus (Kajimura Y and Kaneda M. J. Antibiotics 49: 129-135, 1996; Kajimura Y and Kaneda M. J). Antibiotics 50: 220-228, 1997). Meanwhile, antifungal activity has been reported against Leptosphaeria maculans , which causes black root rot of canola (Beatty PH and Jensen SE. Can. J. Microbiol . 48: 159-169, 2002).

As described above, since fuzisidine has been reported to have excellent bactericidal activity against pathogenic gram-positive bacteria and phytopathogenic fungi, it is considered to be commercially viable and productivity improvement of fuzacidine is required. Examples of introducing antibiotic biosynthesis genes into host bacteria that are easy to industrially cultivate as a method of increasing antibiotic production have been reported (Eppelmann K, Doekel S and Marahiel MA. J. Biol . Chem . 276: 34824-34831, 2001; Pfeifer BA and Khosla C. Microbiol. Mol . Biol . Rev. 65: 106-118, 2001) There have been reports of increased productivity by replacing the promoter of the antibiotic biosynthesis gene with a strong one (Tsuge K, Akiyama T and Shoda M. J.). Bacteriol . 183: 6265-6273, 2001). However, in the case of fuzacidin, no biosynthetic gene is known so far, and no attempt was made to increase the productivity of the method described above. On the other hand, through reconstitution of modules or domains of antibiotic biosynthetic genes (Mootz HD, Schwarzer D and Marahiel MA. Proc . Natl . Acad . Sci . USA 97: 5848-5853, 2000; Ferra FD, Rodriguez F, Tortora O. Tosi C and Grandi G. J. Biol . Chem . 272: 25304-25309, 1997) by substituting specific amino acids in the domain (Eppelmann K, Stachelhaus T and Marahiel MA. Biochemistry 41: 9718-9726, 2000) The development of is also being tried in this way, it is possible to develop antibiotics with good or new activity. However, this attempt could not be made because the fuzacidine biosynthesis gene is unknown.

Therefore, securing fuzisidine biosynthesis genes can be used to increase fuzasidine productivity or to develop new antibiotics based on fuzasidine.

Accordingly, the present inventors have identified that the strain produces fuzacidin through the isolation and purification and characterization of fuzacidin in the F. E681 strain of Panicivacillus polymixsa E681, gene encoding NRPS through genome-wide sequencing analysis Was discovered and separated, and the present invention was completed by confirming that the gene is fuzacidine biosynthesis gene through domain analysis.

An object of the present invention is the Paenibacillus poly68 E Pa. polymyxa E681) to provide a fuzacidine biosynthesis enzyme isolated from a strain, a gene encoding the same, a method for producing fuzacidine or a derivative thereof using the gene.

The present invention provides a polypeptide involved in fuzacidine synthesis.

The present invention also provides a gene encoding the polypeptide.

The present invention also provides a recombinant vector comprising the gene.

The present invention also provides a host cell transformed with the vector.

In addition, the present invention

1) introducing into the expression vector a gene encoding a polypeptide involved in the fuzacidine synthesis;

2) transforming the expression vector into which the gene of step 1) is introduced into a host cell; And

3) culturing the transformant of step 2) and

4) It provides a method for producing fujarycidin or derivatives thereof comprising the step of separating and purifying fuzacidin or derivatives thereof from the culture of step 3).

Hereinafter, the present invention will be described in detail.

The present invention provides a polypeptide involved in fuzacidin synthesis and a gene encoding the same.

Polypeptides involved in the fuzisidine synthesis of the invention are set forth in SEQ ID NO: 2, and variants thereof, ie polypeptides in which one or more modules, domains and / or amino acids have been added, deleted, substituted but exhibit functional equivalence. Belongs to the category. In addition, the genes encoding the polypeptides and variants thereof all fall within the scope of the present invention, and are preferably set forth as SEQ ID NO: 1.

The fuzacidin is L-Thr (threonine), D-Val (valine) is bonded to the amino group of GHPD (15-guanidino-3-hydroxypentadecanoic acid), and then L-Val or L-Tyr (tyrosine) is Is fuzisidine, which is bonded to D-allo-Thr, D-Asn or D-Gln and finally D-Ala (alanine) or D-Val is formed to form a polyketidic ring (see FIG. 1). . Fuzzysidine produced from the Phenibacillus polymyxa strain of the present invention is fuzzysidine A, fuzzysidine B, fuzzysidine C, fuzzysidine D and LI-F03, LI-F04, LI- F07 and LI-F08, but are not necessarily limited thereto.

We completely decoded the genome sequence of the F. genus E681 strain using a whole-genome shotgun sequencing strategy. As a result, the genome of the F. genus E681 strain has a total length of about 5.4 Mbps. As a single circular chromosome structure was confirmed. Fusucidine biosynthesis genes were also identified from the genome.

From the genomic sequences of the E681 strain, Critica (Badger JH and Olsen GJ, Mol . Biol. Evol . 16: 512, 1999), glimmer (Delcher AL et al., Nucleic Acids Res. 27: 4636, 1999) and zcurve (Guo F.-B. et al., Nucleic Acids Res. 31: 1780, 2003) and more than 4,800 protein coding genes were identified and translated into amino acid sequences to estimate the function of each gene product. Then searched in published protein sequence database (Altschul SF et al., Nucleic Acids Res. 25: 3389-3402, 1997), followed by domain and protein family analysis (Bateman A. et al., Nucleic Acids Res . 32 ( Database issue): D138-141, 2004; Haft DH et al., Nucleic Acids Res. 31: 371-373, 2003), motif and pattern search (Hulo N. et al., Nucleic Acids Res. 32 (Database issue) : D134-137, 2004) and location predictive analysis of proteins (Gardy JL et al., Nucleic Acids Res. 31: 3613-3617, 2003).

As a result of the gene sequence search, the NRPS gene encoding the biosynthesis of at least four different antibiotics was found.

The active amino acid table associated with the substrate specificity of each A domain is summarized by Challis et al. (Challis GL et al., Chem . Biol . 7: 211-224, 2000) . As a result of the comparative analysis, it was confirmed that one of these genes was a gene encoding fuzacidine biosynthesis enzyme (see FIG. 2).

Polypeptides involved in the fuzacidine synthesis of the invention comprise one or more modules, each module comprising two or more domains selected from the group consisting of A, C, T, E or TE domains. Do.

Specifically, the polypeptide is the following six modules,

First module: C (condensation) -A (adenylation) -T (thiolation) domain;

Second module: C-A-T-E (epimeration) domain;

Third module: C-A-T domain;

Fourth and fifth modules: C-A-T-E domain; And

Sixth module: C-A-T-TE (termination) domain (see Figure 2).

In the case of constructing fuzacidine biosynthesis by the above module, each A domain recognized amino acids such as Thr, Leu / Ile / Val, Tyr, Thr, Asn, and Val as described in FIG. 2. However, as shown in Example 1, fuzacidin A and B were isolated from E681 strain, and fuzacidin A was L-Thr, D-Val, L-Val, D-allo-Thr, D-Asn, D -Ala and the like and fuzacidin B includes L-Thr, D-Val, L-Val, D-allo-Thr, D-Gln, D-Ala and the like. On the other hand, when the entire genome sequence of the E681 strain was analyzed and analyzed, it was found that only one enzyme gene was involved in fuzacidin synthesis. Therefore, the third A domain recognizes Tyr or Val, the fifth A domain recognizes Asn or Gln, and the sixth A domain recognizes Val or Ala. Fuzacidin structure predicted by this analysis is L-Thr (threonine), D-Val (caline) is combined with amino group of 15-guanidino-3-hydroxypentadecanoic acid (GHPD), and then L-Tyr ( tyrosine) or L-Val bound, and D-allo-Thr, D-Asn or D-Glu was finally found after binding to D-Val or D-Ala, resulting in a polyketidic peptide ring. (See Figure 3).

This fact indicates that the gene of the present invention is a biosynthetic enzyme that produces fuzacidin and also synthesizes various kinds of fuzacidin (see FIG. 3).

The genes of the invention can be designed using genetic recombination techniques to change the number and position of modules or domains at the DNA level to design new enzymes with distinct specificities. For example, by exchanging domains derived from heterologous NRPS (Schneider et al., Mol . Gen. Genet. , 257: 308-318, 1998), or by changing residues that form substrate binding pockets of the A domain Design a new substrate specificity within (Cane and Walsh, Chem . Biol . 6: 319-325, 1999), or in addition, one or more modules, domains or amino acids are added, substituted or deleted or D- and L-amino acids and hydroxy Additional structural modifications, including linkage of acids, mutations in peptide chains and oxidation, acylation, glycosylation, N-methylation and heterocycle ring formation.

Therefore, it is possible to develop fuzacidine derivatives or new antibiotics by the above method using the gene of the present invention.

The present invention also provides a recombinant vector comprising the gene and a host cell transformed therewith.

The gene encoding fuzisidine synthetase of the present invention can be cloned into a suitable vector such as BAC, plasmid, fosmid, etc., and the vector is introduced into a suitable host cell capable of producing fuzzy cydin antibiotic. Can be.

Preferred host cells include the F. bacilli, E. coli , Bacillus subtilis , etc., and the introduction of recombinant vectors can also be introduced through known techniques, ie, heat shock, electroshock, etc. In addition to the above expression host, it will be apparent to those skilled in the art that various strains can be easily used depending on the vector depending on the purpose of expression.

In addition, the present invention

1) introducing into a expression vector a gene encoding a polypeptide involved in fuzacidine synthesis;

2) transforming the expression vector into which the gene of step 1) is introduced into a host cell; And

3) culturing the transformant of step 2) and

4) It provides a method for producing fujarycidin or derivatives thereof comprising the step of separating and purifying fuzacidin or derivatives thereof from the culture of step 3).

Hereinafter, preferred examples are provided to aid in understanding the present invention.

However, the following examples are merely provided to more easily understand the present invention, and the contents of the present invention are not limited by the examples.

< Example  1> Penny Bacillus From Polymix Fuzacidin  Separation and Characterization

<1-1> Penny Bacillus Polymix  Strain culture

Using the media used by Paulus and Gray (Paulus H and Gray E. J. Biol . Chem . 239: 865-871, 1964), the Panicacillus polymyx E681 strain (KCTC 8801P) was subjected to aerobic conditions at 25 ° C, 180 ° C. After incubation for 3 days at rpm, the supernatant was obtained after centrifugation (7,000 rpm, 10 min).

<1-2> by LC / MS analysis system Fuzacidin  Confirm

The components of the supernatant were analyzed using an LC / MS analysis system. For separation and purification of the supernatant, the culture medium was centrifuged at 8,000 rpm for 20 minutes to remove the cells, and the obtained culture supernatant was extracted with butanol and concentrated under reduced pressure using a rotary evaporator to remove butanol, followed by chloroform: methanol (4: 1 to 2: 2). It developed on the silica gel column which has 1) as a solvent, and obtained the active fraction. The reaction mixture was concentrated under reduced pressure, and then separated on Sephadex LH-20 using methanol as a solvent, followed by analysis on high pressure liquid chromatography (HPLC). 8 mg of cydin B was obtained.

The cultured supernatant was analyzed on LC / MS (Thermo electron Co., USA) under the condition of 0.2 ml / min using acetonitrile and water to which 0.1% formic acid was added. It was confirmed that the (M + H) ⁺ ion peak had values of 883, 897, and 911, and had molecular weights such as fuzacidin A, fuzacidin B, and LI-F05b, respectively. LI-F05b is a series compound with a fusicidin-like structure with differences in amino acid residues (Kurusu K, Ohba K, Arai T and Fukushima K. J. Antibiotics 40: 1506-1514, 1987).

< Example  2> Fuzacidin  Base Sequence Determination of Biosynthetic Genes

Using the whole-genome shotgun sequencing strategy for the whole genome, the genome sequence of the F. genus E681 strain was completely decoded and the fuzacidin biosynthesis gene was identified.

<2-1> Library Production

Incubating the strains of Panibacillus polymyxa E681 by the method of Example 1 Genome Analysis, A laboratory manual Vol. Chromosomal DNA was isolated and disrupted according to the method described in III Cloning systems (CSHL Press, Cold Spring Harbor, NY, USA) to produce a pelletized library for sequencing.

The high molecular weight chromosome DNA was crushed with a VCX-500 sonicator (Sonics, Newtown, CT, USA) six times with 19% intensity and pulse on / off time of 0.3 / 3 seconds. DNA fragments inserted into the library were recovered using 2 kb, 5 kb, 8 kb and 10 kb, and were used for pUC18, pUC19, pUC118 or pBCKS (Stratagene, La Jolla, CA, USA), pTrueBlue (Genomics One ( Laval, Quebec, Canada) vector was used, and at the same time, fosmid and BAC libraries of about 40 kb and ~ 100 kb in size were prepared to form contig skeleton structures. Utilized.

The phosmid library was prepared using a phosmid library production kit (CopyControl ^™ fosmid library production kit, Epicentre Biotechnologies, Madison, WI, USA), and the BAC library was prepared by introducing chromosomal DNA cut with Hind III into pIndigo 536 vector. (Peterson DG et al., J. Agric . Genomics , 5, 2000; www.ncgr.org/research/jag; Luo M. et al., Genome 44: 154-62, 2001).

E. coli reaction for the plasmid library Electroporation was introduced into DH10B and then plated on LB agar plate medium containing X-gal / IPTG / Amp (Ampicillin). White recombinant colonies were inoculated into 96 deep-well plates containing LB (Amp) liquid medium and shaken for 48 hours at 250 rpm in an incubator at 37 ° C to recover cells, followed by plasmid DNA was isolated and purified.

<2-2> Sequence Analysis

DNA sequencing reactions were performed using a BigDye ^™ terminator cycle sequencing kit (Applied Biosystems, CA, USA), and the reactions were analyzed by ABI 3700 and 3730 DNA analyzers (Applied Biosystems, Foster City, CA, USA). Sequencing result file processing was used phred / phrap / consed program (http://www.phrap.org). All result files were phreded to assign nucleotide sequences, and only good quality results were taken to mask vector sequences. Phrap was used for sequence merging and consed for contig identification, editing, and primer design.

About 61,700 sequence fragments (corresponding to multiples of 6.7) were obtained from the plasmid and phosmid / BAC ends, and sequence consolidation resulted in more than 800 contiguous sequences. .

First, clones connecting the contigs due to the sequences at both ends were searched, and then a base sequence was determined by designing and synthesizing a primer capable of reading a portion of the sequence gap. The large portion of the sequence gap larger than 15 kb was subjected to limited shot sequencing by selecting phosphmids linking the portions. Misaligned sequences by repetitive sequences such as rRNA genes or translocation enzyme genes were identified and corrected in the consed program. To remove the physical gap, primers were designed at the end of each contig and recombinant PCR was performed, or RT-PCR was performed to secure chromosomal sequences of unknown regions. After removing all gaps to obtain a complete circular chromosome sequence, the Phred value was checked and the region was less accurate, and the region was re-analyzed by PCR amplification. The final target value of accuracy was set to> 99.99% (<1 bp base error per 10 kb).

Finally, the genome of the F. E681 strain of the F. nifolia polymix was composed of a single circular chromosome structure (% G + C: 45.8) with a total sequence of about 5.4 Mbps.

<2-3> Protein Prediction Analysis from Genes

From this genome, Critica (Badger JH and Olsen GJ, Mol . Biol . Evol . 16: 512, 1999), glimmer (Delcher AL et al., Nucleic Acids Res . 27: 4636, 1999) and zcurve (Guo F.-B et al., Nucleic Acids Res . 31: 1780, 2003), and more than 4,800 protein-coding genes were identified and translated into amino acid sequences to estimate the function of each gene product. A blastp search was performed on the sequence database (Altschul SF, et al., Nucleic Acids Res. 25: 3389-3402, 1997). Target databases include COG (Tatusov RL et al., BMC Bioinformatics . 4:41, 2003), UniProt Knowledgebase (Bairoch A. et al., Nucleic Acids Res. 33 (Database issue): D154-159, 2005), NCBI -NR (ftp://ftp.ncbi.nih.gov/blast/db/nr.tar.gz), KEGG-Genes (Kanehisa M. et al., Nucleic Acids Res. 32 (Database issue): D277-280 , 2004).

Domain and protein family analyzes include Pfam (Bateman A. et al., Nucleic Acids Res . 32 (Database issue): D138-141, 2004) and TIGRFAMs (Haft DH, et al., Nucleic Acids Res. 31: 371-373 , 2003), and Prosite database (Hulo N. et al., Nucleic Acids Res . 32 (Database issue): D134-137, 2004) was used for motif and pattern search.

Psort-B was used to predict the position of the protein (Gardy JL et al., Nucleic Acids Res ., 31: 3613-3617, 2003). Proteins were named hierarchically in consideration of the reliability of the search results. In the search for UniProt, a protein without an E value of less than 10 ^-5 was named a hypothetical protein.

Gene sequencing revealed four NRPS genes encoding at least four different antibiotic synthesis, and the substrate specificity of the A domain of the gene was determined by Challis et al. (Challis GL et al., Chem . Biol . 7: 211- 224, 2000) and compared with the substrate specificity-related active amino acid table summarized as a result of the gene was confirmed that one of the genes coding for fuzisidine biosynthesis enzyme (see Figure 2).

< Example  3> Fuzacidin  From the nucleotide sequence of the biosynthetic gene Fuzacidin  Structure prediction

NRPS is a high molecular weight protein that selectively binds to and activates certain amino acids and collects them sequentially. Recognition and activation of amino acids takes place in the A domain. Recently, the tertiary structure of the A domain, which recognizes phenylalanine in the Gramicidin biosynthesis gene, has been identified, suggesting that eight amino acid residues are involved in binding to specific amino acids (Conti E, Stachelhaus T, Marahiel MA and Brick). P. EMBO J. 16: 4174-4183, 1997). The amino acid sequence of the A domain was compared with the amino acid sequence of the known A domain. As a result, the A domain that recognized the same amino acid showed that the eight amino acid residues had very high homology. Thus, analyzing the eight amino acid residues, it is possible to know which amino acids the specific A domain recognizes and binds (Challis GL, Ravel J and Townsend CA. Chem . Biol . 7: 211-224, 2000).

As a result of analyzing the fuzacidine biosynthesis gene of the present invention in comparison with the specific activity related amino acid table summarized by Challis et al., Each A domain is represented as Thr, Leu / Ile / Val, Tyr, Thr, Amino acids such as Asn and Val were recognized. Through this analysis, the structure of fuzacidin is as shown in FIG. 3. However, as shown in Example 1, fuzacidin A and B were isolated from E681 strain, and fuzacidin A was L-Thr, D-Val, L-Val, D-allo-Thr, D-Asn, D -Ala and the like and fuzacidin B includes L-Thr, D-Val, L-Val, D-allo-Thr, D-Gln, D-Ala and the like. On the other hand, as a result of decoding the entire genome sequence of the E681 strain and analyzed, there is only one gene involved in fuzacidine synthesis. Therefore, the third A domain recognizes Tyr or Val, the fifth A domain recognizes Asn or Gln, and the sixth A domain recognizes Val or Ala. Therefore, it can be seen that the fuzacidine biosynthesis gene of the E681 strain can synthesize various fuzacidins.

As shown in the above example, it was confirmed that the F. bacilli strain E681 produced Fusacidin, and the entire genome was analyzed to confirm the fuzacidin biosynthesis gene. The present invention was completed by confirming the synthesis of zarycidine.

As described above, the inventors have confirmed that the strain produces fuzacidin through the purification and characterization of fuzacidin in the strain of Penicillus polymixsa E681, the genome sequence analysis and domain analysis It was confirmed that the gene is fuzacidine biosynthesis gene through. Thus, the fuzacidine biosynthesis enzyme of the present invention can be usefully used to increase the productivity of fuzacidine and to prepare new antibiotics.

<110> Korea research institute of bioscience and biotechnology <120> Fusaricidin synthetase and gene <130> 5p-12-22 <160> 2 <170> KopatentIn 1.71 <210> 1 <211> 23727 <212> DNA <213> DNA sequence <400> 1 atgaatgaca tgcagttata tgatttaaca aatgcgcaga agcgtatatg gtataccgaa 60 ttactctacc cagatacgtc agtgtcacag ctttccggta cagctaaaat gaaggggcac 120 atcaatattg ctgcctttat gcagtccatt aatttgatta tcaaacagta tgatgcgttc 180 cgcatccgta ttacctcagt ggatggattg cctcagcagt acgtcgttcc ttatgaagag 240 agacagttgg agtgcctgga tcttagccac tatgaaagtg tatctgaggt ggaagcctta 300 cttgagcaac acaaaagcaa acctttgccc ctgctggatt ctgagctctt ccagttttta 360 attgtgaaga ttagcgagga agagtattgg attaatatca agatgcacca tattatttct 420 gacggaatat caatggtggt ctatggcaat aagctgacag aattttacat ggagttaatt 480 caaggaaatg aaccgcagct gggcgaagat tgctcgtata ttcaatatat ttcagatgag 540 aatgcatatg aactttctga cagataccaa aaggataagg catactggct ggataaattt 600 tctgatttgc ctgagcttac gggttggaag tcatataacc cgttatcttt aagcacccac 660 gccgtccggg agcattttac cgtaccagaa gtgctatatc acgagctgca agcattttgc 720 caacagaata ggatttcttt gttccagttc ttcatgggtg cgatgtatat ctacatacac 780 aaaatgacga atcagccgga tgtggtgatt ggaacttcgt tcgctaaccg ggggaacaaa 840 aaagagaagc aaaagatagg tatgttcgtc agcaccgctg ctgccagaac atacgtcaaa 900 aaggatatgg atgtgttgag cttcctgcag gatgtagcca gagatcagat gtcagtcctg 960 cggcatcaga agtatccata caatcagtta attcaggatc ttagagaaat gcatggtaac 1020 aaggatattc agcggctttt tggcgtttca atggaatatc gtcttatcaa ttgggttgat 1080 ttggatgatg tgcgtatttt gacagattat gatttctgcg gggacgaagt gaacgatttc 1140 gtgcttcata tcgtagagat cctggatgaa ggcgaactag tactggatgt cgattatcgg 1200 acagagctgt ttgaacgcag tgaagttaag gacatggttt cccagttgct tacgatcgcc 1260 gagcagatca ttcattcacc tcagctttct attgcagagg taaacttatt gggtgaacca 1320 gaagagcagt ccattttggc tctttcggaa ggtgctgcag tcgattatcc acgtgagaag 1380 accatccatg gcttattcga ggaacaagcc gagcgcacgc cagatcacgt agccgttcag 1440 atggacgagc agagcattac atacctagct ctaaacgagc aggctaacca gcttgcgaga 1500 tatttgcgct ccgagggagt aggagcagat acgctcgtag ggattatggc tgaccgttcc 1560 ttggagatgg tcattgggat gttggccatt ttgaaagcag gtggtgccta tgtaccgatt 1620 gatcctgatt atcccgagga acgtatccat tatatgctgg aggattcagg ggtccgtctg 1680 ttgctcaccc aaagtcatct atgggagagc accacttttg acggaaagct tgtgagtttg 1740 gacgaagcta caacgtatac aggagatgct tccaatctgg agagtatttc gggaccaagc 1800 catctagcct atgttatcta cacgtcgggt acgaccggca agccgaaggg cacgctgatt 1860 gaacacaaaa atgtagttcg actgctcttt aacgataaaa atctatttga tttcagctct 1920 caggatacgt ggacgctatt ccattcgttc tgcttcgatt tctccgtttg ggagatgtat 1980 ggagcgcttc tttacggagg gaaattggtg attgttccat ctctcacagc caagagccca 2040 gcagctttct tggagttgtt gaaagacaac caagtcacca ttttaaatca gacgccgacg 2100 tatttttatc aggtgctaca ggaagagtta acgcactctt cgacagagct tggccttaga 2160 aaaatcattt ttggtggaga ggccctaagt ccatctcttc taagaaactg gcgggtcaag 2220 tatcctgatg tgcagctgat taatatgtac ggaattacgg aaacaacggt ccatgtcact 2280 tacaaggaaa tcacggaaca tgagattgaa gcggggaaaa gcaatattgg cagaacgatt 2340 cccacactta gcgcttatat tctcgatgag caaagacgcc ttcagcctgt tggggttcca 2400 ggagagctat acattgcggg agacggtctt gcccgtgggt atttgaatcg gccggatttg 2460 acgtctgaga aattcgttga gcatccgtat cgggcgggag agcggctgta ccgaactggg 2520 gatcttgctc gttggttgcc tgatggcaat attgaatatt tggggcggat cgaccatcag 2580 gtcaaaattc gcggctaccg aattgagctt ggcgaggtag aagcccaaat tctcaaggct 2640 ccgaacgtac gagaaacgat tgtcctcgca cgggaagacg aacagggcca aaaattgctg 2700 tgcgcctact atgtagcctc cagtgatctt tcgccggggg aattgcgggc tcagctggcg 2760 gcggaactcc ccgcttacat gattccttct tattttgtcc ggctggagca aatgccgctt 2820 acaccaaatg gaaaactgga tcgccgtgcg ttgccggctc ctgaaagcag cgtacaatcc 2880 ggcgaggctt atttggctcc gagaactgct gtggaagctc agatggtact catctggcaa 2940 gatatcttgg gcgttgctcg tgttggtgtc agagataatt tctttgaaat tggtggccac 3000 tctttgcggg caacagtgct cgtttcacgg attcacaaag aattgggatg tagcatttcg 3060 ctgcgtgagg tgttccagtc acctacggtc gaatccttgg cgcaacttgt gaaaaaacac 3120 attccgaccc tgtacgaatc cattccacag gcaacggaaa gcgaagctta cccagtgtcc 3180 tcagcgcaaa agcggttata cgtgctgaga cagatggacg ggggagagct cagctacaat 3240 atgccagggg tcttcacagt ggatggaccg ttggatcgca cgcggctgga gtctgcgttc 3300 caggcactga tccggcgtca tgaatccctg agaaccggct tttatatgca ggatggagag 3360 cttgttcagc gtgtgcatag gaatgtgccg ttcgcgttga actacacaga ggcttcggtg 3420 gaggagacgg atacgctcat tcacaacttt attcgtgcct ttgatctgag ccaggctcca 3480 ttactgcgtg ttagcttggt gaagctccag gaggagcgtc atctgttgct gtttgatatg 3540 catcacatta tttcagatgg ggtttctatt caaatattga tagaggaact tactcatttg 3600 tatcaaggag aacagctacc agagctgcac atccagtaca aggattatgc cgtatggcaa 3660 cgggaacagt cagagaacca atggcaagat cttgagaaat attggctgca atcctttgag 3720 ggagagttgc cagtattgga tttgcctaca gacttccagc gaccttcggt tcggagcttc 3780 gagggtagcc gaattgattt tacattggat gaatctggaa ataaggcgat acaagagctt 3840 gcatcccgta caggtactac actgtatatg gtattgctgg ccgcttattc ggtactactg 3900 cacaaatata caggacaaga ggacatcgtc gtaggttctc cagtagccgg aagaccgcag 3960 gctgagcttg agggcattat cggaatgttt gtcaacacac tggccttgcg cagctacccg 4020 acaggagata aaacctttca ggattatctt ctggaaatca aggaaacggc gctcaaggcg 4080 tttgagcatc aggattaccc ttttgaaaaa ttggtcgaaa agctgggcgt aggacgtgat 4140 gtcagccgca atccgctctt tgacacctta ttggtattac aaaataccga gcaggaagag 4200 caggaaatgg acggagtgca ctttactcct tacttgatgg acaccgtcac agccaaattt 4260 gatctgtccc tcaatgtaga ggagaagggg tcaaaattag cctttggcct cgagtatagt 4320 acggctttat atcggcgtga aagtgtagag cgatttgcaa cgcacttgct ccgggttctg 4380 cacgcagtct cggtcaatcc tcagttgcaa ctggccgaga tagaaatgat cacaccggag 4440 gagaaagtac aaatcgttga agtttttaac gcgacatcgg ctccttatcc aagggacaag 4500 accattcatg agctgttcgc ggaacaagcc aagcgcacac cggagcagac ggcgcttgta 4560 ttcggcgatg tccagctaac gtaccttgaa ttggaagaca aggcgagccg actggcccaa 4620 acactgcgtc gtttgggaac gttgagggag cagcctgtgg ccgtgatggg cggacgaagc 4680 atcgagatgg tcattggtat gctcgcggtg cttcaggcag gtggagccta tgtgccgatt 4740 gatcctgatt acccggaaga tcgggttcgt tatatgctta atgattccga cgccaagcta 4800 ttattggtgc aaaagggcga gcttataaat gtagactacg gtataccgat tgtcgatctt 4860 agcagtaaag aggcttatgc tgctgaacct gcccagccgg agacggctca aggatcgcag 4920 gggcttgctt atgtcatcta tacatcgggt acgacgggta gaccgaaggg cgttatggtt 4980 gaacaccgga acgtggtccg tctggtcaaa gagaccaact atgtggagct gaatgaatcc 5040 acacgaattt tgcagacagg agccgtggcc tttgatgctt ctacattcga aatatgggga 5100 gcgttgctta acggtgggca gctctatttt gtagagaatg acgacattct gattgctgat 5160 aggctcaaag cggctattgc caagtacggg attacaacat tgtggcttac ttcacccctt 5220 ttcaatcagc tttccctaca ggatgagtac ctgttcagag ggctcaaagc attattagtc 5280 ggcggtgacg tactgtctct atctcatatg aatcgtgtaa tcgaggctaa tcccgatctt 5340 atccctatca attgctatgg tccgacagag aatacgacct tctccaccac ctacaagatt 5400 ccaggtcgtg ccgaaggggg cgtgccgatt ggtcgtccaa ttagtaattc gaccgcttat 5460 gtggtcaatg gatcgctgca attacagcct attggtgctt ggggtgaact cattgtcggc 5520 ggtgaaggtg tagcacgcgg atatctcaat cgtcctgatc tcacagcaga gaaatttgtt 5580 cctagtcctg tgaaggacgg agaaccgtgc taccgaactg gggatttggt acgctggctt 5640 ccagatgggg atttggagtt taaaggaaga attgatgagc aggtcaaaat acgtggttac 5700 cgcatcgaac tccctgaaat cgaggcccaa ctggccaagg tggagtcagt aatcgacgcc 5760 gtagtggtcg ttcgcgcgga tgagcttgga gagaagcagc tttgcgctta ttatgtggcg 5820 gatcgtacgc tcacggcagg cgaagtacgc ctttccctat cgcaggtact tccgggctat 5880 atgattccat cctactttat ccagatggac cgtatgccat taacgtcaaa cggaaaagtg 5940 gaccgcaggt ctctgccagc tcctcaagta ggcgcgcata caggacggaa gtatacagct 6000 cctcgtacac cggctgaaga agctttggca tctgtctggc aaggggtgct gggtgccgaa 6060 caggtgggta tccatgacaa tttctttgaa ttgggcggag actccattaa ggccattcag 6120 gtgtcgtcac ggttactgca ggccggctat cggttagaga tgaagcagct gttcaaatcg 6180 ccaaccattg ctgagctagg cgctgaaata caaacggctg tgcatatggc tgaacaggga 6240 gttgtgcgtg gaacgactcg cttgactcca gtccaacagt ggttctttgg acggaagcag 6300 gcagagcctc atcacttcaa tcaagcggtt atgctgtatc gtgaacaagg gtttgaggaa 6360 aaggccttgc atcaggtgct aagaaaactc gctgagcatc atgacgccct tcgcatggtt 6420 ttccgtcaga cagagcatgg ctacgaggct tggaatcgtg atcttgaaga aggagagctg 6480 tatagcctgt tcaccgctga tttacggaat gaatccgatc cgactgcagc cattacatcg 6540 ctgtcggatg acattcagcg cagtatcaat ctggcagaag gtccgctgct gaagttagga 6600 cttttccatt gtcaggatgg agaccacctt ctaatcgtga tccaccattt ggtggtggac 6660 ggagtatcct ggcggatttt gttcgaggat attgcagcag gctatgagca ggtgattcag 6720 ggacaagcgc tgacattccc acagaagacg gattccttcc gtgactgggg cgacgccctt 6780 gctcgttatt cggaaggtcc tgaaatggag actcatcggg cgtattggag agagctggag 6840 gatcagccac tcgaacagtt gccgaaggat gaggctgtgg aaagccttct tttacaggat 6900 agcaaagtag taacagcaca atggactata gaagaaaccg accaattgtt gagaaaagcc 6960 catcgtgctt atcaaacaga gacgaatgat ctgctattga ctgctctggg catggcggta 7020 tccaaatggt ctggcatcgg aaaggttgct gtgaatctgg aaggacacgg tcgtgagccg 7080 attataccga atatcgacat cacccgtacc gtaggctggt ttacaagtca atttccggtg 7140 attttagact tggggaataa cccggaagtg gcctccttga tcaagtctgt gaaagagggg 7200 ctgcgccgaa ttccgaacaa aggtattggc tatgggttgc ttaaaacgat ggcaagtcag 7260 ttggatgaag gcagcttcag cttgcagcct gagatttctt ttaactatct gggacaattt 7320 gatcaggatt tgcaaggaag ctcgttgcag atttctcctt atccgaccgg aagcgcgcaa 7380 agtttgttgg aggaaccagc ctatacgcta gatatcaatg gcatggtgac ggacggagcc 7440 ctgactctga cgattactta taacggaaaa cagtataagt catctacgat ggaacagctc 7500 gctggatata ttgaagaaag cctgcgggag cttctccagc attgcgtaac ccaagaaaaa 7560 accgtattga caccaagcga tgtgctcgcg aagggtctaa gcattgccga tctggaggag 7620 ctttctaagc agaccagcca cataggcgat attgagaatg tatatagtct gacgccgatg 7680 cagaagggca tgctgttcca tgatatgttt gagccgcata caggtgcata ttttgagcag 7740 gctgcctttg actttaaggg tagctttgat ccgaccgcct tcggacacag tctggatgca 7800 gtggtggagc gtcatgctat cctgcgcacg aacttttaca gcggatgggg tagcgagcct 7860 ttgcaggttg tatttcggca cagaggcgct aaattggtgt acgaagacct gcgggagatg 7920 aatgcatcgc agcgcgaagc ttacctgaag acatttggtg ctaaggacaa agcactgggc 7980 ttcaacctag ctgaagacga gcttctccgt gtatcaattt tacaaacaga tgaagagagc 8040 ttccgtctct tatggagctt tcaccacatc gtcatggatg ggtggtgtgt tccgttaatt 8100 acgcaggagg tatttgaaca ctattttgcc ctcctggaag gaagagagcc gcagttggca 8160 gaggttcatc cgtacagtcg atatatcgaa tggctagaac agcaggatga agcagttgcg 8220 tccaactatt ggagccgata tctggccggt tacgagcagc agacgctttt acctcaagtt 8280 ggtggagcaa gtaagggaga aggctatata gcagaaaagc tgaattatcc tctcagcagg 8340 gaattgactg agcgccttga aaaggtggcc agggatgctc atgtcacgat gaatatattg 8400 ctgcagtccc tctggggcat tgcgcttcaa cgctataacg gtagccggga tgtcgtgtac 8460 ggaagtgtag tatcaggccg accagcagaa attcctggca ttgatcggat gatcggtttg 8520 ttcatcaata cgattcctgt tcgtgtgaag acagaggaga atctcccctt caccgtcctg 8580 atgaagcagc agcaggaaca atatatggct tctcatatgt atggcaccta cccgttgttt 8640 gagattcagg ctcagacgga tcagaagcag gatctaatct cccatattat ggtgtttgag 8700 aactatcctg tggaggagga ggtagagcgt ctgggtggtg gcgaggctgc ctttgagatt 8760 gaggaggcgg agcttcttga gcaaacgaat tacgatttta atttaattgt cctccctggc 8820 gaagagatga gattgctgtt ccagtacaat gcacttgttt atgacccagt gacaattggg 8880 caaatcaagg gccatctggt tcacctcatg gaacaaattg tagagaaccc tgctatttcc 8940 gtggatgctc tagaattaat cacgccgcag gagagagaac agattctgaa cgtatgggga 9000 aatacaaaag ccatttacga gcactataac acgttccacg ggctgttgga ggaacaggcg 9060 ggacgaacgc cggatgcggc tgccatttgg ttcgaggacg agagtctgac ctatgccgag 9120 ctcaatgcaa aagccaatgg actggcgaga aggctccgca ctcagggaat caagacggga 9180 gatctggtgg gactgattgc tgaacggtcg ctcgaaatga tcgttgggat ctacggcatt 9240 atgaaagccg ggggtgccta tgttccaatc gatccagagt atccgcaaga acgaatcagc 9300 tacatgcttg aagattccgg ggcgaagctg atccttacac aggcccatct cttggagcat 9360 ctcggatgga cggaaaatgt tttgctgctg gatgaatcat cgacctatga tgccgacacc 9420 tcgaatttgg aggatactgc tggcccggat gatctggctt acgtgatcta cacttcaggt 9480 acgacggggc agcctaaggg cgtattagtc gagcatcggg gactacccaa tctttcggac 9540 gtatatgggg cacacttcga agttacaccg caggatcgga tcgttcagtt tgcaagcctg 9600 tcgtttgatg cgtcggtttc ggaaatttta acggcgctga gccacggggg tgttctgtgc 9660 atcccttctg cacaagatat tttagatcat gccctgttcg agcagttcat gaacgataag 9720 gggattacgg tagcgacttt gccacccgct tacgctatcc accttgatcc agagcgtttg 9780 ccaacactgc ggtgcctgct aaccgctgga tcgaccgcat cgatcgagtt gatcgaagag 9840 tggaggaagc atgtacgtta ctctaatggc tatggcccaa cagaggactc cgtatgcacc 9900 accatctggt ctgtcccgga cagtgaggaa gcaacgaata ttgtatctat tggacgacct 9960 attgctaacc atagtgtgta catcttggat gaccatttta gattgcaacc tgtcggtgta 10020 gctggagagc tatgcatttc gagtatcggg ttagcacggg ggtatcataa tcggcctgag 10080 ttaatggatg agaagttcgt ggacaatccg tttgctccag gagaacgtat gtatcggacg 10140 ggtgacctgg ttcgctggtt accgaatgga aacatcgagt acttaggtcg aatagatcac 10200 caagtcaaaa tccgcggcta ccgtatcgag ctaggcgagg tagaagcaca aatgctcaga 10260 gtgccgtccg ttcaggaagt cgtagccatg gctgtagagg gcgatgacgg ctacaaagat 10320 ctagtcgctt atttcgtagc tgctcagaaa cttgaggtat ccgaacttcg tgccgtcctg 10380 tcggagatgt tacctggata tatgatccct tcccgcttca tacaactgga ggacatgctt 10440 ctgacgtcga acggtaaaat cgatcgaaaa gcgctgcagg gcgagcgtgg atgggcagcg 10500 gcttcgtctg aggctccaag gacacctttg gaaatccaat tagcagaaat atggcaagag 10560 gtgctgggtg tagagagcgc gggagtgaag gataattttt tccattttgg aggtcattca 10620 ctgcgtgcag ccctgctagt ctcacgaatt cgcaaggaaa tgaatcgcga gattagtctg 10680 agagcagtgt tcgagtctcc tactattgag ggattggctc gtgtcattga gggctataca 10740 ccgctgaatt tcgaagaaat tcctacagcg ggagcgagag agcattatcc attgtcctcg 10800 gcccaaaaac gactgtttat tctaagtcag ctggaaggtg gagagctgag ctacaatatg 10860 ccgggtatcc ttaccgttga gggagctttg gatcgggaac ggctagagca ggcattccgt 10920 cgtctgattc accgccatgg ttcgctgcgt actcgttttg tgactgtgaa cggtgaacct 10980 gtacagcagc tcctgacgga tgttccgttt actgtggaat atgcggagtt gagcgaggaa 11040 gaggcaggag ctacccttca gcagtttgtc cgtccttttg atttaggtgt agctccattg 11100 ctgcgggtcg gccttattcg aattgcacat gagcgccatt tactattgtt cgacatgcat 11160 catattgtct cagatggggt ttctatgaat attctcatag aagagtttct ccgcttctac 11220 caagaggagg acgtatttcc tgagctacag atccagtaca cagactatgc tgtatggcag 11280 caagagcagc tcggaagcga gcgtcttaag gcccaggaag cttactggct ggatgctttc 11340 cgcggaagct tgccagtgct ggatttgcca ggagatgaag ttcgtcctgc ggtgcgaagc 11400 tttgcgggcg atcgaatcga cttccaaata gattcttctc tgagtgcttc acttcaggag 11460 ctggctaccc gaacgggttc cactctgttc atggtactgc tggcagccta tacagcgctc 11520 ttgcacaagt acacaggtca ggaagatgtc attgtcggtt cacctgtggc aggaagatcc 11580 catgcgacac tcgaaggcct catcggtatg ttcgtcggca cagtggcact tcgtacttat 11640 ccagaaggag aaaagccttt cgaggcttat ctgcaggaag tgaaggaaac agcgctgcgg 11700 gcctatgaaa accaggatta cccgttcgag gagctggtag aaaagctgga gcttcagcgt 11760 gatttgagcc gtaatccgct atttgatacc atgtttgtcc tgcaaaatat cgagcaggga 11820 gaacaagaaa tagaaggatt gcgcttcact ccttacgata atgtacatcc ggctgccaag 11880 ttcgatctca cgctgaccgt gagtgaagta gacggggcat tgaactgcac gcttgagtac 11940 gcgactgcga tctacaagca agagactgct gagcggatgg caggccactt tgtacagctt 12000 attcgggaag ccatcgctaa tccggcactg ccgttgtcat cccttgatat cgtgacacct 12060 caggaaaaat caaggctgat gaaagcgccg gacgaagcca aggcagatta tcctcgtgac 12120 aagacgatcc atgcgctgtt cgaggagcag gccgcacgta ctccgaatgc agtggcagtc 12180 gtatgtgaag atgcagccct gtcctacagc gagctgaacg agcgggccaa tggacttgcc 12240 cgaacgctga gggaacgtgg tttgcaacca gacggtttgg ctggaatcat ggcggatcgt 12300 tcccttgaaa tggtggtcgg gattttagcc atcttgaagg caggcggagc ctatgtccct 12360 gtagaccctg aatatccaga ggaccgcatt cactttatgc ttgaggactc gggagccaag 12420 ctactgctga cacaagcgca tctggagcaa cgtgtctcct tcgctgggaa catcgtaagt 12480 ctggataaaa cagcttccta taaggaagat gtctcaaacc tgcagcctgc agccggacca 12540 aaccatcttg cctacgtcat ctacacatcg ggtacgacag gcaagcccaa gggaacgctg 12600 atcgagcata aaaatgtagt tcgcttgctc tttaatgata aaaatatgtt tgacttcgat 12660 gctcaggata cgtggacact gttccattca ttctgttttg acttctctgt atgggaaatg 12720 tacggagcat tgctgaacgg aggacggttg gtcatcgttc catcgcttac cgcgaagagt 12780 ccagatcgtt tcttgcaatt gcttaaggat cagaaggtca ccgttttgaa ccagacaccg 12840 acgtacttct atcagttgct acaggaagag cttggtcatc aggcggcaga actgagcctc 12900 cgtatgatta tcttcggtgg agaggcatta accccggccc tgctcaagga ctggagaacg 12960 aagtatccgc aagtacagct cattaacatg tacggcatta ccgaaacgac cgtgcatgta 13020 acctacaagg aaattacaga gttggaaatt gaacagggcc gcagcaatat cggcaccacg 13080 attccgacgc tgcgagcgta cattttggat gaacaacgcc gtccacagcc gattggcatt 13140 ccgggtgaac tctatgtggc gggcgtaggc ctggcgcgag gttatctgaa ccgaccggaa 13200 ttgacggaag agaagtttgt cgctcatccg tttgaagcgg gcgagcgcat gtaccgctcg 13260 ggtgacttgg cacgctggtt gccggatggc agcatggagt atttgggacg gattgaccat 13320 caggtaaaaa tccgtggtta ccgtatcgag ctgggcgaag tggaagcgaa gctgctccat 13380 gctccgtctg taagggaggc cgttgtgctc gcccgagagg atggaagtgg acaaaaagtg 13440 cttatcggct atttcactgc cgatcagatg ctgacggtag gcgagttgag aaaagccttg 13500 gctgccgaac tgcctgctta tatgattcca tcttacttta tgcaattgga acagatgcct 13560 ttgacgccaa atggcaagct ggatcgcaaa gcgcttccgg ctccggaggc caatgtgcag 13620 actggagcgg tttatgaacc gccaaggacg aaggctgagg aaactttggc ttccgtatgg 13680 caaggtgtgc tgggagcgca gcaggtcggc atccatgatc atttcttcga tctgggtggt 13740 gactctatca aggcgattca agtgtcctcg agattgttcc aagctggata taaattagag 13800 atgaaggatc tcttcaaata tccgacaatt gccgagctaa gcccgtatct tcaggcagct 13860 ggacgtacag cggaacaggg cgaaattaaa ggtgcagcag agttaatgcc aattcagcgt 13920 tggttctttg aacgccatac agcggagccg caccattata atcatgccgt catgctctat 13980 cggaaagacg gctttgatga ggctgcactt cggttgacaa tggaccaaat tgcgatccat 14040 catgacgcgc tgcgcatggt tttccggcct acagaagctg ggtatgcagc ttggaatcgg 14100 ggaacggacg aaggcgagct ctacacattg gacattgccg atgtgcggca ggtggaagac 14160 cagacagctg cggttcaggc acaagccgat gccattcagg caagctttga cttggaaggg 14220 ggcccactgt tcaagctagg cctgttccat tgtgacgatg gcgatcattt gttgattgtc 14280 attcatcacc tcttggttga cggcgtatcc tggcgcatcc tgtttgagga cattgcagcg 14340 gggtacgagc aggcattgaa tggacaagca atcatccttc cacaaaagac cgattcgtat 14400 cttgtatggt ctgagcaagc gacgaagtat gcagcagggc ctgctctgga caaggagcgt 14460 gcatactggc agctgattga ggaggcgatt ttggccccac tgccgaagga tgaggatcag 14520 aagccgggca ccattcggga tactgaatcg gttacggtaa cgtggtctgc gcaggaaact 14580 gacctgctgt tgcgacaagc gaaccgggcg tatcatacgg agacgaatga cttgctcttg 14640 actgctctgg gggcagccat tcagcgctgg acaggcatgg agcagatttt ggtcaatctt 14700 gaagggcatg gacgggaaat gatcgtacca gacctggata ttacccgtac cgtgggttgg 14760 ttcacaaccc agtatccggt tctgctgaac ctgcaaggca gacaggaagt atctgcgcga 14820 atcaagcgca tcaaggagaa tttgcgagag gttccgcaca aaggaatcgg ctacggtctt 14880 ctgaagtata tagcaccgga gaaaagtgtc ggattcggcg tagagccgga gatttccttc 14940 aattatcttg ggcagtttga tcaggatttg gaggggaatg cccttagcct atccacacat 15000 tcagtgggta aggcgctcag cgatctcaca cctcagcaat atgctctgga tgtgaatggc 15060 atgattgccg aaggccagct atcactgacg attacgtaca gcagcaggca gtatcgtaag 15120 gagacggtga gtcattttgc tgaattatta cagtcaagcc ttagcgaggt catcctgcat 15180 tgtgtggctc aggagcgttc gcagcttacg cctagtgatg ttctgttcca aggactgacc 15240 ctggagcagc ttgatcgact cacagcccaa acggcccata ttggagagat tgaggatgtg 15300 tacaagctga cggcaatgca gaagggaatg ctcttccaca gcttgctgga gccggactcc 15360 tcttcatact ttgagcaggc aacgtttgag ctacgtggca gcttcgatgt agatatcttc 15420 ttcgagagct tccaggcttt ggcgcaacga catgccatac tacgtaccgg cttttacaac 15480 aacattgctg atgtgccgct gcaagttgcc tttaagcaac ggttaatccc tctgcactac 15540 gtagatttgc gcgatgcatc tatgcaggaa caagaagccc gaatcaaggc ttatattgct 15600 gaagatatgg ttaaggggtt cagcttgtca gaagatccgc tgatgcgagt gaccgtcttg 15660 cagaaggatc aaagctgtct tgtattgtgg agcttccatc atattgtcat ggacggctgg 15720 tgtatcccga tcattacaca ggagctgttc gattattatt ctgccaagaa acagcaaata 15780 cagcctgtcc tgccgccagc tcagccctac agccgttata tagaatggct tgatgcacag 15840 gatgaacaag aggcttcaac gtattggagc caatatctcg aagattatga cgggaatact 15900 gtactgccgg aaggtaaaac gaaatctcaa gcgaaagaag cgggctatgt tctaaaagag 15960 catgttctcc atctgggtgc atcgttgacc ggtaaaatgg atgttgtcgc gaagcgcaat 16020 cacgtgaccg tcaacacact catgcagaca gcttggggtc tgattcttca acgttataat 16080 gccagctcgg atgtcgtttt cggcggcgtt gtgtcgggta gacccgctga gattgcaggg 16140 atcgaaaata tggtgggtct gttcattaat acagtaccta tccgtgtaca gtcatccaaa 16200 gacgaagcct ttgtaaaggt gatgaaacgt acacaggcac agtcattggc tggtcgtgcc 16260 tatgacacct atccactgta tgagattcag gggaaaacaa cccaaaagca ggacttgatt 16320 tctcatatta tgatctttga aaattacccg ctcgacgagc aggtggagca atcggggaat 16380 caaacggagg acaatctcga agtcgccaac ttcaccatgt ttgaacaaac caactatgac 16440 tttaacctgg ttgtaattcc gggcgaggac atcaaggtct gcattcgcta taatgcttcg 16500 gtttacgagc aagaaagcat tgcacgaatc ggaggacact tgttgcagat gctcgatcag 16560 gtggctgctc gtccgcaggc gacgatacag gaactggaga ttgtaacatc tgaggaacgg 16620 atgaatctgc ttgactgggg cggcaaggcc catgcttatc caagtgatca gggcctgcat 16680 accttgtttg aggaacaggt agtccgtacg ccggataaga tcgcggcaat aagcggtgac 16740 attcagatca catatcggga gctgaacgag caggcgaaca gactggcttc gaccttgata 16800 gcccaaggac tgcggagtga acaagtagtg ggtctgttgg cggatcggtc agtggagctg 16860 ctagttgcca ttatgggcgt actcaaagcg ggcggagcct atgtaccgat tgatcctgaa 16920 tatccgcagg agcggattca gtatattctg aaggactctg gcgctgagat tctgctcaca 16980 cagagccacc tgacgaagct agcgtccttt gagggaatgg ttatggaatt ggattcatca 17040 cacatctacg gaactggagt gaataatcct aatattcctg tgagaggcaa cgatctggtg 17100 tatctaatct atacttcggg tacaacagga aatccgaagg gaaccatgat taaccataaa 17160 ggaatcgtga actacatctg gtgggccaat aaagtctatt gtgcagggca accaacggac 17220 ttcccgctgt attcatccat ttcgtttgac ttgacgatga catcaatgtt tactccatta 17280 ataaatggag gaatagtacg gatttatgat ggtatagata aagcggaggt tgtacagcat 17340 attttgcgcg aaaatgcggt ggacattctc aagctgacgc cgactcatct cagtctgatt 17400 aaagatatgg ccattccagc ggaaagtcgc attcagcagc ttattgtggg tggagagaat 17460 ttgaccacgc atttgtcgaa aatcattaca gatctctttg gcggcaacat caaaatctac 17520 aatgaatacg gtccaaccga aacggttgtc ggctgcatga ttcacctgta cgatcctgcg 17580 aaggatacac gtgagtccgt accgattggg ttgccgtccg acaacatata cattcatatc 17640 ctggatgaac agcttcgtct tgtaccgtta ggtgtagagg gcgaaatgta catcgccgga 17700 gacggagtag cccgtggata cctgaaccgt ccggatctta ccgcagagaa attcattaga 17760 aatccgttcg ctgcggaagg aaatatgtat cgcactgggg atttggctcg tcgccttcct 17820 aatggagaca ttgagtacat tggacgcatt gaccatcaag ttaaaatacg gggctatcgt 17880 attgagcttg gtgagattga ggccaagctg ctggacattc cacttgtcga ggaagctctc 17940 gttgttgcgt gggcagatgc tcatgggcag aaatcgctgt gtgcttactt cgtagcggat 18000 cgcgaaatgt ctgtcagcga gctgagaaac gaactgtctg ccggactgcc tgcatatatg 18060 attccgtctt actttgtcca actggacgtg atgcctctga caccgaatgg taagctggat 18120 cgcaaggcac tgcctgaacc gaactcgggt ataaaggcgg gagcagactt taccgctccg 18180 cgcacggatg tggagaatat tctggcttca atctggcaag gtgtactcgg cgtgccgctt 18240 gtcggcatac atgataattt ctttgagctt ggaggtgact cgatcaaatc catccaagta 18300 tcttcaaggc ttctccaagc aggctacaag cttgaaatga aggatttgtt cggttatccg 18360 acaattgcag agctggcgca gcgcgttagt gtggtcagcc gaattgcgga ccaaagcgag 18420 gtacacggat cggtaagact ttcacccgct cagcacagat tcttcgatga acagtcgatg 18480 gatctgcatc actttaatca gtcggtcatg ttgtaccgac gggatggctt caataccgat 18540 gcgctcgctg aggttgttcg gaaaattgca gagcatcatg atgctttacg actggtgttc 18600 cgccaaggag agcagggatt ggaggcctgg aaccggagca tgggtgaggg tgaactctat 18660 agcctccaaa tccacgacct gcgggatgag acagacccgg cttcagcaat agaagcgggt 18720 gcggaagcca ttcagcgcag catctctctg gaggatggac ctctctttag actgggtctg 18780 ttccgctgtg cggaaggcga acatctgttg atcgttattc atcatctggc tgtggatggc 18840 gtatcctggc gtattctctt tgaggacctg caggaaggct acgagcaggc agcacgtgga 18900 gaagcggtca agtttccaca gaagacggat tcgtaccgtg catgggttga gggaatcaca 18960 caatttgcaa acagcccggc ggctgaacaa gaacgcagct attgggcaga ggtagaggga 19020 gatggctttg tccctcttcc caaagacaag gtagacggcg ctcttctcat caaagacagt 19080 gaggctgtca cggtgaggtg gtcaccagaa gagacagagc agttcctgaa agaagcgaac 19140 cgcacttaca atacggaggt caacgatctg ctcctgacgg ctttaggtat ggctgttcac 19200 gagtggacag gaatcgaacg tgtaggcatc cttctggagg gacatggacg ggagcctgtt 19260 gtgccggaac tggatattac tcgcacaata ggctggttta caagtcaata ccctgtcgcc 19320 cttgagatgg gaggggaatt ggagatcggc gccagaatca agcacgtcaa ggaaggcttg 19380 cgtcgtatcc cgaacaaagg tgtcggatat ggtattttga aatatttaag cgacggttcc 19440 gatgtctcct ccttctcggc tgaacctgag attaccttca actacttggg acagttcgac 19500 caggatcttg caggagggat gatggaagta tcgtcttact cagtaggacc tgaggtcagt 19560 gagcagatgg tgcagcatca ggcagtgaac attaatggac tgattgccga aggacagctt 19620 caactttcgg tcagctataa tcgtcatcag ctccacgggg agtccgtggc taagtttgtt 19680 ggcattctga agaaccgtct cagcgaagtc attggacatt gcgtaagtaa ggaaagaaca 19740 gaacttacac caagtgatgt actcctcaaa gatatcagct tagaaaagat tgaggagcta 19800 gaagagcaga cacggcatat cggcagtatt gaaaatatgt ataagctaac accgatgcaa 19860 aaaggaatgt tgttccacag cttgctggag cctcattcgg aagtctactt tgagcaggcc 19920 aaatttgaaa ttcagggagc attctatcct gaggatttca aacgcagctt aaaatatctg 19980 atgaaacggc atgccatatt gagaacgaat ttccatgccg ggtggggcga tttccctatt 20040 cagattgtgt tcaaagaaag agcgtgtgac ttcgtatacg aggatctgca cgagctggaa 20100 gccgatgaaa ttcaagcgcg tcttgcagct tatactgctc aggataaagc aagaggcttt 20160 aatcttgctg aagaagcgtt gctgcgtgtt gctattctac gtacagcaga agaggcttac 20220 catttgctgt ggagctctca tcacatcatt ttggatggct ggtgtatgcc gcttgtgctc 20280 caggaagtgt ttgagacata cggagctcta cgtgagcaaa gggaacctga gcttcctgca 20340 gctgtatcgt acagccagta tattcaatgg ttggagaagc aaggcgagga agaggcatcc 20400 tcttactgga gagggtacct ggaaggctac gagcagcaga cgaagctgcc acaagccatc 20460 acacaaccat cggcaaaagc agaagcctac gtgtcggaga agctggtatt cacgttggat 20520 gcggaattga ccgatcgcct ggaacaggtg gctaagcagc atcaggtgac gatgaatacg 20580 ttgatgcaag cagcctgggg aatcgtgttg cagcgctaca atagaagcca ggatgtcgtc 20640 ttcggaagtg tggtatcggg cagacctgcc gagattccgg gtatcgaaag catgatcggt 20700 ctcttcatta atacagttcc agttcgtgtt caggccgagg gaagcgattc gttctctcat 20760 gtgatgaaga gacagcagga attatacttg gcaggacatg cttatgattc ctatccgctc 20820 tatgagattc aggcacagag cgagcaaaag caagatttga tctctcatat tatggtattt 20880 gagaattatc cggtagaaga gcatctggaa gagaaaattg ccaatgatga ggctgaatac 20940 aaaattacgg acgttcagat gtttgaacag acgaattatg attttaacct cattgtgctg 21000 ccaggacgta gtctggagtt cttatatcgt tataatgctc gcgtctatga tcgggagagc 21060 gtggaacgga ttcaaggaca cttgacgaga attctgacaa gtgtatctgt acaacctgct 21120 atccgtattg acgagctgga gctgattacg ccagaagaga aatcacaaat tatagaggtg 21180 tggggagata cagcagcccc ttatccgcgt gagaagaccc ttcacggcat atttgaggaa 21240 aaggctgcgc tcacaccgga tcgtacagca cttatttacg gtgaaacggt gcttacctac 21300 ggagagcttc atcaacaggc caaccgcctg gcgcgtacgc tgcgtgttca aggggtcaga 21360 ccagatcagc cggtcggcat tatggtcgag cgttcgcttg agatgatcat tggtatccat 21420 gctattctaa aagcgggtgg cgcctatgta ccgattgatc cgggattccc tgaggatcgt 21 480 attcgtcaca tgctggagga ttcaggagcg aagcttctac tgacgaagaa ccatctcaaa 21540 gatcgtttcc cattcactgg cacgatcctg tcacttgacg atccgcaggc gtatcatgct 21600 gatgactcca atctggagcc agtcgcagga ccagagcatc tggcgtatat catttacacg 21660 tcaggttcga ctggcaagcc gaagggtgta atgattgagc atcactctgc tgtccatacg 21720 ctgagtcagt tggaagctga atatccgatg ttggcaggcg accgtttcct gctcaaaacg 21780 acattcacct ttgacttctc cgtgccggag ctgttctgct ggttctttgg acaggggacg 21840 ctcgtgatcc tgccgccggg cgtggacaaa gatccggtcg ccctgctgga ggctgtggat 21900 gcgaaccaaa ttacgcatct taatttggta ccttcgatgc tcagcgtgct cgttcaatat 21960 ttgaaagaaa gtagtaccca aggattcctt actttgaaat acctgtttgc ctgcggcgag 22020 acgctgcctg ccaaacttgt ggaagagtat tataaagtat ctccttacgc agtactggaa 22080 aacatctacg gtcctacgga agcagccgta tatgcgactc ggtatacaac gagccttgag 22140 actgccgctc taacgcatgt gccgatcggc aaaccgtacg ctaacgtcca agtatggatg 22200 atggacagcg cttctcaggt atcacctgtg ggggtaccgg gagaactctg cattgcgggc 22260 gaaggggtag cgagagggta cttcaatcag ccggacctga cggcagagaa gttcattcct 22320 catccgtaca agccgggaga aaggatttac cgaacaggcg atttggctcg gtgggtgcca 22380 gacgggaata ttgagtactt gggacggatc gatcaccagg taaaaatccg cggttaccgc 22440 attgagctgg gagaagtgga agcacaaatt ctgaaggtgc catcggtgca ggaagcggtt 22500 gttctagcac tggctgattc tactggaagt actcagcttt gtgcatactt tgtggccgaa 22560 gaggggcttg cagcgggcgt actacgcgag gcactggcca gcgagctgcc aagctacatg 22620 attccgactg ctttcgtaca gttggcacaa atgccgctga atccgaatgg aaaattggat 22680 cgcaaagcgc taccggcacc ggaaacactt ctgcggagca cagcggagta tatcgcgccg 22740 cgtacgcaga cagaagtaga gcttgctcag atttggtccg aggtgctcgg cgttcaggaa 22800 atcgggatca gggatcattt cttcgaactt gggggccatt ccctgaaagt attgggcttg 22860 atccaaagga tctcgtccgg tatgggcgtc cagctcccac tccaagtcgt gtttaatctg 22920 ccgactgtgg aagaaatggc gcatgaaatt ttcaagctgc aggcaacaac ttctgccgat 22980 gaacaagaaa tggaaattat ccacttccca gggaaaggaa tacttaaagt attttgcttc 23040 cctcctcggg taggtcactc tctggggtac tatgagatgg ccaaggagct ggatgggctt 23100 tgcgaggtgt acgggatgga atttatcggc gaccgtttcc agggtcaaga tatgctggat 23160 cgatacatcg atgccatcgt ggatattcaa gcagagggcc catatatatt cttgggatac 23220 tcacttggag gaaatctggc cttcgaggta gctaaagcca tggaaatccg aggtcaccat 23280 gttggagaca ttattatggt agatgctatg agaaagatgt ccaaggatga atcgacaccg 23340 gaggagcttg aagagattgt cgaaacggtg ctggacagca ttagggagca gtacaaagca 23400 ttcctggccg atccagccga cagggagcga gtcatggaca aaatgttggt ttactccacc 23460 taccgcgatg aacttattaa cgcaggtgaa gttcatgcaa atatccatgc cctgattgca 23520 gaggatgata gtgttggtcc agatacatca ttagataaat tgctatggca acaggcgaca 23580 cttggccaat acaaagaata cgaagtcatc ggaacgcatg atgtgctgct tgattccggt 23640 tttgttgggg aaaatgctaa agtactgaga cagatacttg gtaaaattgc agaaacctca 23700 tctaaaaaca agcctatttt gtcctaa 23727 <210> 2 <211> 7908 <212> PRT <213> amino acid sequence <400> 2 Met Asn Asp Met Gln Leu Tyr Asp Leu Thr Asn Ala Gln Lys Arg Ile   1 5 10 15 Trp Tyr Thr Glu Leu Leu Tyr Pro Asp Thr Ser Val Ser Gln Leu Ser              20 25 30 Gly Thr Ala Lys Met Lys Gly His Ile Asn Ile Ala Ala Phe Met Gln          35 40 45 Ser Ile Asn Leu Ile Ile Lys Gln Tyr Asp Ala Phe Arg Ile Arg Ile      50 55 60 Thr Ser Val Asp Gly Leu Pro Gln Gln Tyr Val Val Pro Tyr Glu Glu  65 70 75 80 Arg Gln Leu Glu Cys Leu Asp Leu Ser His Tyr Glu Ser Val Ser Glu                  85 90 95 Val Glu Ala Leu Leu Glu Gln His Lys Ser Lys Pro Leu Pro Leu Leu             100 105 110 Asp Ser Glu Leu Phe Gln Phe Leu Ile Val Lys Ile Ser Glu Glu Glu         115 120 125 Tyr Trp Ile Asn Ile Lys Met His His Ile Ile Ser Asp Gly Ile Ser     130 135 140 Met Val Val Tyr Gly Asn Lys Leu Thr Glu Phe Tyr Met Glu Leu Ile 145 150 155 160 Gln Gly Asn Glu Pro Gln Leu Gly Glu Asp Cys Ser Tyr Ile Gln Tyr                 165 170 175 Ile Ser Asp Glu Asn Ala Tyr Glu Leu Ser Asp Arg Tyr Gln Lys Asp             180 185 190 Lys Ala Tyr Trp Leu Asp Lys Phe Ser Asp Leu Pro Glu Leu Thr Gly         195 200 205 Trp Lys Ser Tyr Asn Pro Leu Ser Leu Ser Thr His Ala Val Arg Glu     210 215 220 His Phe Thr Val Pro Glu Val Leu Tyr His Glu Leu Gln Ala Phe Cys 225 230 235 240 Gln Gln Asn Arg Ile Ser Leu Phe Gln Phe Phe Met Gly Ala Met Tyr                 245 250 255 Ile Tyr Ile His Lys Met Thr Asn Gln Pro Asp Val Val Ile Gly Thr             260 265 270 Ser Phe Ala Asn Arg Gly Asn Lys Lys Glu Lys Gln Lys Ile Gly Met         275 280 285 Phe Val Ser Thr Ala Ala Ala Arg Thr Tyr Val Lys Lys Asp Met Asp     290 295 300 Val Leu Ser Phe Leu Gln Asp Val Ala Arg Asp Gln Met Ser Val Leu 305 310 315 320 Arg His Gln Lys Tyr Pro Tyr Asn Gln Leu Ile Gln Asp Leu Arg Glu                 325 330 335 Met His Gly Asn Lys Asp Ile Gln Arg Leu Phe Gly Val Ser Met Glu             340 345 350 Tyr Arg Leu Ile Asn Trp Val Asp Leu Asp Asp Val Arg Ile Leu Thr         355 360 365 Asp Tyr Asp Phe Cys Gly Asp Glu Val Asn Asp Phe Val Leu His Ile     370 375 380 Val Glu Ile Leu Asp Glu Gly Glu Leu Val Leu Asp Val Asp Tyr Arg 385 390 395 400 Thr Glu Leu Phe Glu Arg Ser Glu Val Lys Asp Met Val Ser Gln Leu                 405 410 415 Leu Thr Ile Ala Glu Gln Ile Ile His Ser Pro Gln Leu Ser Ile Ala             420 425 430 Glu Val Asn Leu Leu Gly Glu Pro Glu Glu Gln Ser Ile Leu Ala Leu         435 440 445 Ser Glu Gly Ala Ala Val Asp Tyr Pro Arg Glu Lys Thr Ile His Gly     450 455 460 Leu Phe Glu Glu Gln Ala Glu Arg Thr Pro Asp His Val Ala Val Gln 465 470 475 480 Met Asp Glu Gln Ser Ile Thr Tyr Leu Ala Leu Asn Glu Gln Ala Asn                 485 490 495 Gln Leu Ala Arg Tyr Leu Arg Ser Glu Gly Val Gly Ala Asp Thr Leu             500 505 510 Val Gly Ile Met Ala Asp Arg Ser Leu Glu Met Val Ile Gly Met Leu         515 520 525 Ala Ile Leu Lys Ala Gly Gly Ala Tyr Val Pro Ile Asp Pro Asp Tyr     530 535 540 Pro Glu Glu Arg Ile His Tyr Met Leu Glu Asp Ser Gly Val Arg Leu 545 550 555 560 Leu Leu Thr Gln Ser His Leu Trp Glu Ser Thr Thr Phe Asp Gly Lys                 565 570 575 Leu Val Ser Leu Asp Glu Ala Thr Thr Tyr Thr Gly Asp Ala Ser Asn             580 585 590 Leu Glu Ser Ile Ser Gly Pro Ser His Leu Ala Tyr Val Ile Tyr Thr         595 600 605 Ser Gly Thr Thr Gly Lys Pro Lys Gly Thr Leu Ile Glu His Lys Asn     610 615 620 Val Val Arg Leu Leu Phe Asn Asp Lys Asn Leu Phe Asp Phe Ser Ser 625 630 635 640 Gln Asp Thr Trp Thr Leu Phe His Ser Phe Cys Phe Asp Phe Ser Val                 645 650 655 Trp Glu Met Tyr Gly Ala Leu Leu Tyr Gly Gly Lys Leu Val Ile Val             660 665 670 Pro Ser Leu Thr Ala Lys Ser Pro Ala Ala Phe Leu Glu Leu Leu Lys         675 680 685 Asp Asn Gln Val Thr Ile Leu Asn Gln Thr Pro Thr Tyr Phe Tyr Gln     690 695 700 Val Leu Gln Glu Glu Leu Thr His Ser Ser Thr Glu Leu Gly Leu Arg 705 710 715 720 Lys Ile Ile Phe Gly Gly Glu Ala Leu Ser Pro Ser Leu Leu Arg Asn                 725 730 735 Trp Arg Val Lys Tyr Pro Asp Val Gln Leu Ile Asn Met Tyr Gly Ile             740 745 750 Thr Glu Thr Thr Val His Val Thr Tyr Lys Glu Ile Thr Glu His Glu         755 760 765 Ile Glu Ala Gly Lys Ser Asn Ile Gly Arg Thr Ile Pro Thr Leu Ser     770 775 780 Ala Tyr Ile Leu Asp Glu Gln Arg Arg Leu Gln Pro Val Gly Val Pro 785 790 795 800 Gly Glu Leu Tyr Ile Ala Gly Asp Gly Leu Ala Arg Gly Tyr Leu Asn                 805 810 815 Arg Pro Asp Leu Thr Ser Glu Lys Phe Val Glu His Pro Tyr Arg Ala             820 825 830 Gly Glu Arg Leu Tyr Arg Thr Gly Asp Leu Ala Arg Trp Leu Pro Asp         835 840 845 Gly Asn Ile Glu Tyr Leu Gly Arg Ile Asp His Gln Val Lys Ile Arg     850 855 860 Gly Tyr Arg Ile Glu Leu Gly Glu Val Glu Ala Gln Ile Leu Lys Ala 865 870 875 880 Pro Asn Val Arg Glu Thr Ile Val Leu Ala Arg Glu Asp Glu Gln Gly                 885 890 895 Gln Lys Leu Leu Cys Ala Tyr Tyr Val Ala Ser Ser Asp Leu Ser Pro             900 905 910 Gly Glu Leu Arg Ala Gln Leu Ala Ala Glu Leu Pro Ala Tyr Met Ile         915 920 925 Pro Ser Tyr Phe Val Arg Leu Glu Gln Met Pro Leu Thr Pro Asn Gly     930 935 940 Lys Leu Asp Arg Arg Ala Leu Pro Ala Pro Glu Ser Ser Val Gln Ser 945 950 955 960 Gly Glu Ala Tyr Leu Ala Pro Arg Thr Ala Val Glu Ala Gln Met Val                 965 970 975 Leu Ile Trp Gln Asp Ile Leu Gly Val Ala Arg Val Gly Val Arg Asp             980 985 990 Asn Phe Phe Glu Ile Gly Gly His Ser Leu Arg Ala Thr Val Leu Val         995 1000 1005 Ser Arg Ile His Lys Glu Leu Gly Cys Ser Ile Ser Leu Arg Glu Val    1010 1015 1020 Phe Gln Ser Pro Thr Val Glu Ser Leu Ala Gln Leu Val Lys Lys His 1025 1030 1035 1040 Ile Pro Thr Leu Tyr Glu Ser Ile Pro Gln Ala Thr Glu Ser Glu Ala                1045 1050 1055 Tyr Pro Val Ser Ser Ala Gln Lys Arg Leu Tyr Val Leu Arg Gln Met            1060 1065 1070 Asp Gly Gly Glu Leu Ser Tyr Asn Met Pro Gly Val Phe Thr Val Asp        1075 1080 1085 Gly Pro Leu Asp Arg Thr Arg Leu Glu Ser Ala Phe Gln Ala Leu Ile    1090 1095 1100 Arg Arg His Glu Ser Leu Arg Thr Gly Phe Tyr Met Gln Asp Gly Glu 1105 1110 1115 1120 Leu Val Gln Arg Val His Arg Asn Val Pro Phe Ala Leu Asn Tyr Thr                1125 1130 1135 Glu Ala Ser Val Glu Glu Thr Asp Thr Leu Ile His Asn Phe Ile Arg            1140 1145 1150 Ala Phe Asp Leu Ser Gln Ala Pro Leu Leu Arg Val Ser Leu Val Lys        1155 1160 1165 Leu Gln Glu Glu Arg His Leu Leu Leu Phe Asp Met His His Ile Ile    1170 1175 1180 Ser Asp Gly Val Ser Ile Gln Ile Leu Ile Glu Glu Leu Thr His Leu 1185 1190 1195 1200 Tyr Gln Gly Glu Gln Leu Pro Glu Leu His Ile Gln Tyr Lys Asp Tyr                1205 1210 1215 Ala Val Trp Gln Arg Glu Gln Ser Glu Asn Gln Trp Gln Asp Leu Glu            1220 1225 1230 Lys Tyr Trp Leu Gln Ser Phe Glu Gly Glu Leu Pro Val Leu Asp Leu        1235 1240 1245 Pro Thr Asp Phe Gln Arg Pro Ser Val Arg Ser Phe Glu Gly Ser Arg    1250 1255 1260 Ile Asp Phe Thr Leu Asp Glu Ser Gly Asn Lys Ala Ile Gln Glu Leu 1265 1270 1275 1280 Ala Ser Arg Thr Gly Thr Thr Leu Tyr Met Val Leu Leu Ala Ala Tyr                1285 1290 1295 Ser Val Leu Leu His Lys Tyr Thr Gly Gln Glu Asp Ile Val Val Gly            1300 1305 1310 Ser Pro Val Ala Gly Arg Pro Gln Ala Glu Leu Glu Gly Ile Ile Gly        1315 1320 1325 Met Phe Val Asn Thr Leu Ala Leu Arg Ser Tyr Pro Thr Gly Asp Lys    1330 1335 1340 Thr Phe Gln Asp Tyr Leu Leu Glu Ile Lys Glu Thr Ala Leu Lys Ala 1345 1350 1355 1360 Phe Glu His Gln Asp Tyr Pro Phe Glu Lys Leu Val Glu Lys Leu Gly                1365 1370 1375 Val Gly Arg Asp Val Ser Arg Asn Pro Leu Phe Asp Thr Leu Leu Val            1380 1385 1390 Leu Gln Asn Thr Glu Gln Glu Glu Gln Glu Met Asp Gly Val His Phe        1395 1400 1405 Thr Pro Tyr Leu Met Asp Thr Val Thr Ala Lys Phe Asp Leu Ser Leu    1410 1415 1420 Asn Val Glu Glu Lys Gly Ser Lys Leu Ala Phe Gly Leu Glu Tyr Ser 1425 1430 1435 1440 Thr Ala Leu Tyr Arg Arg Glu Ser Val Glu Arg Phe Ala Thr His Leu                1445 1450 1455 Leu Arg Val Leu His Ala Val Ser Val Asn Pro Gln Leu Gln Leu Ala            1460 1465 1470 Glu Ile Glu Met Ile Thr Pro Glu Glu Lys Val Gln Ile Val Glu Val        1475 1480 1485 Phe Asn Ala Thr Ser Ala Pro Tyr Pro Arg Asp Lys Thr Ile His Glu    1490 1495 1500 Leu Phe Ala Glu Gln Ala Lys Arg Thr Pro Glu Gln Thr Ala Leu Val 1505 1510 1515 1520 Phe Gly Asp Val Gln Leu Thr Tyr Leu Glu Leu Glu Asp Lys Ala Ser                1525 1530 1535 Arg Leu Ala Gln Thr Leu Arg Arg Leu Gly Thr Leu Arg Glu Gln Pro            1540 1545 1550 Val Ala Val Met Gly Gly Arg Ser Ile Glu Met Val Ile Gly Met Leu        1555 1560 1565 Ala Val Leu Gln Ala Gly Gly Ala Tyr Val Pro Ile Asp Pro Asp Tyr    1570 1575 1580 Pro Glu Asp Arg Val Arg Tyr Met Leu Asn Asp Ser Asp Ala Lys Leu 1585 1590 1595 1600 Leu Leu Val Gln Lys Gly Glu Leu Ile Asn Val Asp Tyr Gly Ile Pro                1605 1610 1615 Ile Val Asp Leu Ser Ser Lys Glu Ala Tyr Ala Ala Glu Pro Ala Gln            1620 1625 1630 Pro Glu Thr Ala Gln Gly Ser Gln Gly Leu Ala Tyr Val Ile Tyr Thr        1635 1640 1645 Ser Gly Thr Thr Gly Arg Pro Lys Gly Val Met Val Glu His Arg Asn    1650 1655 1660 Val Val Arg Leu Val Lys Glu Thr Asn Tyr Val Glu Leu Asn Glu Ser 1665 1670 1675 1680 Thr Arg Ile Leu Gln Thr Gly Ala Val Ala Phe Asp Ala Ser Thr Phe                1685 1690 1695 Glu Ile Trp Gly Ala Leu Leu Asn Gly Gly Gln Leu Tyr Phe Val Glu            1700 1705 1710 Asn Asp Asp Ile Leu Ile Ala Asp Arg Leu Lys Ala Ala Ile Ala Lys        1715 1720 1725 Tyr Gly Ile Thr Thr Leu Trp Leu Thr Ser Pro Leu Phe Asn Gln Leu    1730 1735 1740 Ser Leu Gln Asp Glu Tyr Leu Phe Arg Gly Leu Lys Ala Leu Leu Val 1745 1750 1755 1760 Gly Gly Asp Val Leu Ser Leu Ser His Met Asn Arg Val Ile Glu Ala                1765 1770 1775 Asn Pro Asp Leu Ile Pro Ile Asn Cys Tyr Gly Pro Thr Glu Asn Thr            1780 1785 1790 Thr Phe Ser Thr Thr Tyr Lys Ile Pro Gly Arg Ala Glu Gly Gly Val        1795 1800 1805 Pro Ile Gly Arg Pro Ile Ser Asn Ser Thr Ala Tyr Val Val Asn Gly    1810 1815 1820 Ser Leu Gln Leu Gln Pro Ile Gly Ala Trp Gly Glu Leu Ile Val Gly 1825 1830 1835 1840 Gly Glu Gly Val Ala Arg Gly Tyr Leu Asn Arg Pro Asp Leu Thr Ala                1845 1850 1855 Glu Lys Phe Val Pro Ser Pro Val Lys Asp Gly Glu Pro Cys Tyr Arg            1860 1865 1870 Thr Gly Asp Leu Val Arg Trp Leu Pro Asp Gly Asp Leu Glu Phe Lys        1875 1880 1885 Gly Arg Ile Asp Glu Gln Val Lys Ile Arg Gly Tyr Arg Ile Glu Leu    1890 1895 1900 Pro Glu Ile Glu Ala Gln Leu Ala Lys Val Glu Ser Val Ile Asp Ala 1905 1910 1915 1920 Val Val Val Val Arg Ala Asp Glu Leu Gly Glu Lys Gln Leu Cys Ala                1925 1930 1935 Tyr Tyr Val Ala Asp Arg Thr Leu Thr Ala Gly Glu Val Arg Leu Ser            1940 1945 1950 Leu Ser Gln Val Leu Pro Gly Tyr Met Ile Pro Ser Tyr Phe Ile Gln        1955 1960 1965 Met Asp Arg Met Pro Leu Thr Ser Asn Gly Lys Val Asp Arg Arg Ser    1970 1975 1980 Leu Pro Ala Pro Gln Val Gly Ala His Thr Gly Arg Lys Tyr Thr Ala 1985 1990 1995 2000 Pro Arg Thr Pro Ala Glu Glu Ala Leu Ala Ser Val Trp Gln Gly Val                2005 2010 2015 Leu Gly Ala Glu Gln Val Gly Ile His Asp Asn Phe Phe Glu Leu Gly            2020 2025 2030 Gly Asp Ser Ile Lys Ala Ile Gln Val Ser Ser Arg Leu Leu Gln Ala        2035 2040 2045 Gly Tyr Arg Leu Glu Met Lys Gln Leu Phe Lys Ser Pro Thr Ile Ala    2050 2055 2060 Glu Leu Gly Ala Glu Ile Gln Thr Ala Val His Met Ala Glu Gln Gly 2065 2070 2075 2080 Val Val Arg Gly Thr Thr Arg Leu Thr Pro Val Gln Gln Trp Phe Phe                2085 2090 2095 Gly Arg Lys Gln Ala Glu Pro His His Phe Asn Gln Ala Val Met Leu            2100 2105 2110 Tyr Arg Glu Gln Gly Phe Glu Glu Lys Ala Leu His Gln Val Leu Arg        2115 2120 2125 Lys Leu Ala Glu His His Asp Ala Leu Arg Met Val Phe Arg Gln Thr    2130 2135 2140 Glu His Gly Tyr Glu Ala Trp Asn Arg Asp Leu Glu Glu Gly Glu Leu 2145 2150 2155 2160 Tyr Ser Leu Phe Thr Ala Asp Leu Arg Asn Glu Ser Asp Pro Thr Ala                2165 2170 2175 Ala Ile Thr Ser Leu Ser Asp Asp Ile Gln Arg Ser Ile Asn Leu Ala            2180 2185 2190 Glu Gly Pro Leu Leu Lys Leu Gly Leu Phe His Cys Gln Asp Gly Asp        2195 2200 2205 His Leu Leu Ile Val Ile His His Leu Val Val Asp Gly Val Ser Trp    2210 2215 2220 Arg Ile Leu Phe Glu Asp Ile Ala Ala Gly Tyr Glu Gln Val Ile Gln 2225 2230 2235 2240 Gly Gln Ala Leu Thr Phe Pro Gln Lys Thr Asp Ser Phe Arg Asp Trp                2245 2250 2255 Gly Asp Ala Leu Ala Arg Tyr Ser Glu Gly Pro Glu Met Glu Thr His            2260 2265 2270 Arg Ala Tyr Trp Arg Glu Leu Glu Asp Gln Pro Leu Glu Gln Leu Pro        2275 2280 2285 Lys Asp Glu Ala Val Glu Ser Leu Leu Leu Gln Asp Ser Lys Val Val    2290 2295 2300 Thr Ala Gln Trp Thr Ile Glu Glu Thr Asp Gln Leu Leu Arg Lys Ala 2305 2310 2315 2320 His Arg Ala Tyr Gln Thr Glu Thr Asn Asp Leu Leu Leu Thr Ala Leu                2325 2330 2335 Gly Met Ala Val Ser Lys Trp Ser Gly Ile Gly Lys Val Ala Val Asn            2340 2345 2350 Leu Glu Gly His Gly Arg Glu Pro Ile Ile Pro Asn Ile Asp Ile Thr        2355 2360 2365 Arg Thr Val Gly Trp Phe Thr Ser Gln Phe Pro Val Ile Leu Asp Leu    2370 2375 2380 Gly Asn Asn Pro Glu Val Ala Ser Leu Ile Lys Ser Val Lys Glu Gly 2385 2390 2395 2400 Leu Arg Arg Ile Pro Asn Lys Gly Ile Gly Tyr Gly Leu Leu Lys Thr                2405 2410 2415 Met Ala Ser Gln Leu Asp Glu Gly Ser Phe Ser Leu Gln Pro Glu Ile            2420 2425 2430 Ser Phe Asn Tyr Leu Gly Gln Phe Asp Gln Asp Leu Gln Gly Ser Ser        2435 2440 2445 Leu Gln Ile Ser Pro Tyr Pro Thr Gly Ser Ala Gln Ser Leu Leu Glu    2450 2455 2460 Glu Pro Ala Tyr Thr Leu Asp Ile Asn Gly Met Val Thr Asp Gly Ala 2465 2470 2475 2480 Leu Thr Leu Thr Ile Thr Tyr Asn Gly Lys Gln Tyr Lys Ser Ser Thr                2485 2490 2495 Met Glu Gln Leu Ala Gly Tyr Ile Glu Glu Ser Leu Arg Glu Leu Leu            2500 2505 2510 Gln His Cys Val Thr Gln Glu Lys Thr Val Leu Thr Pro Ser Asp Val        2515 2520 2525 Leu Ala Lys Gly Leu Ser Ile Ala Asp Leu Glu Glu Leu Ser Lys Gln    2530 2535 2540 Thr Ser His Ile Gly Asp Ile Glu Asn Val Tyr Ser Leu Thr Pro Met 2545 2550 2555 2560 Gln Lys Gly Met Leu Phe His Asp Met Phe Glu Pro His Thr Gly Ala                2565 2570 2575 Tyr Phe Glu Gln Ala Ala Phe Asp Phe Lys Gly Ser Phe Asp Pro Thr            2580 2585 2590 Ala Phe Gly His Ser Leu Asp Ala Val Val Glu Arg His Ala Ile Leu        2595 2600 2605 Arg Thr Asn Phe Tyr Ser Gly Trp Gly Ser Glu Pro Leu Gln Val Val    2610 2615 2620 Phe Arg His Arg Gly Ala Lys Leu Val Tyr Glu Asp Leu Arg Glu Met 2625 2630 2635 2640 Asn Ala Ser Gln Arg Glu Ala Tyr Leu Lys Thr Phe Gly Ala Lys Asp                2645 2650 2655 Lys Ala Leu Gly Phe Asn Leu Ala Glu Asp Glu Leu Leu Arg Val Ser            2660 2665 2670 Ile Leu Gln Thr Asp Glu Glu Ser Phe Arg Leu Leu Trp Ser Phe His        2675 2680 2685 His Ile Val Met Asp Gly Trp Cys Val Pro Leu Ile Thr Gln Glu Val    2690 2695 2700 Phe Glu His Tyr Phe Ala Leu Leu Glu Gly Arg Glu Pro Gln Leu Ala 2705 2710 2715 2720 Glu Val His Pro Tyr Ser Arg Tyr Ile Glu Trp Leu Glu Gln Gln Asp                2725 2730 2735 Glu Ala Val Ala Ser Asn Tyr Trp Ser Arg Tyr Leu Ala Gly Tyr Glu            2740 2745 2750 Gln Gln Thr Leu Leu Pro Gln Val Gly Gly Ala Ser Lys Gly Glu Gly        2755 2760 2765 Tyr Ile Ala Glu Lys Leu Asn Tyr Pro Leu Ser Arg Glu Leu Thr Glu    2770 2775 2780 Arg Leu Glu Lys Val Ala Arg Asp Ala His Val Thr Met Asn Ile Leu 2785 2790 2795 2800 Leu Gln Ser Leu Trp Gly Ile Ala Leu Gln Arg Tyr Asn Gly Ser Arg                2805 2810 2815 Asp Val Val Tyr Gly Ser Val Val Ser Gly Arg Pro Ala Glu Ile Pro            2820 2825 2830 Gly Ile Asp Arg Met Ile Gly Leu Phe Ile Asn Thr Ile Pro Val Arg        2835 2840 2845 Val Lys Thr Glu Glu Asn Leu Pro Phe Thr Val Leu Met Lys Gln Gln    2850 2855 2860 Gln Glu Gln Tyr Met Ala Ser His Met Tyr Gly Thr Tyr Pro Leu Phe 2865 2870 2875 2880 Glu Ile Gln Ala Gln Thr Asp Gln Lys Gln Asp Leu Ile Ser His Ile                2885 2890 2895 Met Val Phe Glu Asn Tyr Pro Val Glu Glu Glu Val Glu Arg Leu Gly            2900 2905 2910 Gly Gly Glu Ala Ala Phe Glu Ile Glu Glu Ala Glu Leu Leu Glu Gln        2915 2920 2925 Thr Asn Tyr Asp Phe Asn Leu Ile Val Leu Pro Gly Glu Glu Met Arg    2930 2935 2940 Leu Leu Phe Gln Tyr Asn Ala Leu Val Tyr Asp Pro Val Thr Ile Gly 2945 2950 2955 2960 Gln Ile Lys Gly His Leu Val His Leu Met Glu Gln Ile Val Glu Asn                2965 2970 2975 Pro Ala Ile Ser Val Asp Ala Leu Glu Leu Ile Thr Pro Gln Glu Arg            2980 2985 2990 Glu Gln Ile Leu Asn Val Trp Gly Asn Thr Lys Ala Ile Tyr Glu His        2995 3000 3005 Tyr Asn Thr Phe His Gly Leu Leu Glu Glu Gln Ala Gly Arg Thr Pro    3010 3015 3020 Asp Ala Ala Ala Ile Trp Phe Glu Asp Glu Ser Leu Thr Tyr Ala Glu 3025 3030 3035 3040 Leu Asn Ala Lys Ala Asn Gly Leu Ala Arg Arg Leu Arg Thr Gln Gly                3045 3050 3055 Ile Lys Thr Gly Asp Leu Val Gly Leu Ile Ala Glu Arg Ser Leu Glu            3060 3065 3070 Met Ile Val Gly Ile Tyr Gly Ile Met Lys Ala Gly Gly Ala Tyr Val        3075 3080 3085 Pro Ile Asp Pro Glu Tyr Pro Gln Glu Arg Ile Ser Tyr Met Leu Glu    3090 3095 3100 Asp Ser Gly Ala Lys Leu Ile Leu Thr Gln Ala His Leu Leu Glu His 3105 3110 3115 3120 Leu Gly Trp Thr Glu Asn Val Leu Leu Leu Asp Glu Ser Ser Thr Tyr                3125 3130 3135 Asp Ala Asp Thr Ser Asn Leu Glu Asp Thr Ala Gly Pro Asp Asp Leu            3140 3145 3150 Ala Tyr Val Ile Tyr Thr Ser Gly Thr Thr Gly Gln Pro Lys Gly Val        3155 3160 3165 Leu Val Glu His Arg Gly Leu Pro Asn Leu Ser Asp Val Tyr Gly Ala    3170 3175 3180 His Phe Glu Val Thr Pro Gln Asp Arg Ile Val Gln Phe Ala Ser Leu 3185 3190 3195 3200 Ser Phe Asp Ala Ser Val Ser Glu Ile Leu Thr Ala Leu Ser His Gly                3205 3210 3215 Gly Val Leu Cys Ile Pro Ser Ala Gln Asp Ile Leu Asp His Ala Leu            3220 3225 3230 Phe Glu Gln Phe Met Asn Asp Lys Gly Ile Thr Val Ala Thr Leu Pro        3235 3240 3245 Pro Ala Tyr Ala Ile His Leu Asp Pro Glu Arg Leu Pro Thr Leu Arg    3250 3255 3260 Cys Leu Leu Thr Ala Gly Ser Thr Ala Ser Ile Glu Leu Ile Glu Glu 3265 3270 3275 3280 Trp Arg Lys His Val Arg Tyr Ser Asn Gly Tyr Gly Pro Thr Glu Asp                3285 3290 3295 Ser Val Cys Thr Thr Ile Trp Ser Val Pro Asp Ser Glu Glu Ala Thr            3300 3305 3310 Asn Ile Val Ser Ile Gly Arg Pro Ile Ala Asn His Ser Val Tyr Ile        3315 3320 3325 Leu Asp Asp His Phe Arg Leu Gln Pro Val Gly Val Ala Gly Glu Leu    3330 3335 3340 Cys Ile Ser Ser Ile Gly Leu Ala Arg Gly Tyr His Asn Arg Pro Glu 3345 3350 3355 3360 Leu Met Asp Glu Lys Phe Val Asp Asn Pro Phe Ala Pro Gly Glu Arg                3365 3370 3375 Met Tyr Arg Thr Gly Asp Leu Val Arg Trp Leu Pro Asn Gly Asn Ile            3380 3385 3390 Glu Tyr Leu Gly Arg Ile Asp His Gln Val Lys Ile Arg Gly Tyr Arg        3395 3400 3405 Ile Glu Leu Gly Glu Val Glu Ala Gln Met Leu Arg Val Pro Ser Val    3410 3415 3420 Gln Glu Val Val Ala Met Ala Val Glu Gly Asp Asp Gly Tyr Lys Asp 3425 3430 3435 3440 Leu Val Ala Tyr Phe Val Ala Ala Gln Lys Leu Glu Val Ser Glu Leu                3445 3450 3455 Arg Ala Val Leu Ser Glu Met Leu Pro Gly Tyr Met Ile Pro Ser Arg            3460 3465 3470 Phe Ile Gln Leu Glu Asp Met Leu Leu Thr Ser Asn Gly Lys Ile Asp        3475 3480 3485 Arg Lys Ala Leu Gln Gly Glu Arg Gly Trp Ala Ala Ala Ser Ser Glu    3490 3495 3500 Ala Pro Arg Thr Pro Leu Glu Ile Gln Leu Ala Glu Ile Trp Gln Glu 3505 3510 3515 3520 Val Leu Gly Val Glu Ser Ala Gly Val Lys Asp Asn Phe Phe His Phe                3525 3530 3535 Gly Gly His Ser Leu Arg Ala Ala Leu Leu Val Ser Arg Ile Arg Lys            3540 3545 3550 Glu Met Asn Arg Glu Ile Ser Leu Arg Ala Val Phe Glu Ser Pro Thr        3555 3560 3565 Ile Glu Gly Leu Ala Arg Val Ile Glu Gly Tyr Thr Pro Leu Asn Phe    3570 3575 3580 Glu Glu Ile Pro Thr Ala Gly Ala Arg Glu His Tyr Pro Leu Ser Ser 3585 3590 3595 3600 Ala Gln Lys Arg Leu Phe Ile Leu Ser Gln Leu Glu Gly Gly Glu Leu                3605 3610 3615 Ser Tyr Asn Met Pro Gly Ile Leu Thr Val Glu Gly Ala Leu Asp Arg            3620 3625 3630 Glu Arg Leu Glu Gln Ala Phe Arg Arg Leu Ile His Arg His Gly Ser        3635 3640 3645 Leu Arg Thr Arg Phe Val Thr Val Asn Gly Glu Pro Val Gln Gln Leu    3650 3655 3660 Leu Thr Asp Val Pro Phe Thr Val Glu Tyr Ala Glu Leu Ser Glu Glu 3665 3670 3675 3680 Glu Ala Gly Ala Thr Leu Gln Gln Phe Val Arg Pro Phe Asp Leu Gly                3685 3690 3695 Val Ala Pro Leu Leu Arg Val Gly Leu Ile Arg Ile Ala His Glu Arg            3700 3705 3710 His Leu Leu Leu Phe Asp Met His His Ile Val Ser Asp Gly Val Ser        3715 3720 3725 Met Asn Ile Leu Ile Glu Glu Phe Leu Arg Phe Tyr Gln Glu Glu Asp    3730 3735 3740 Val Phe Pro Glu Leu Gln Ile Gln Tyr Thr Asp Tyr Ala Val Trp Gln 3745 3750 3755 3760 Gln Glu Gln Leu Gly Ser Glu Arg Leu Lys Ala Gln Glu Ala Tyr Trp                3765 3770 3775 Leu Asp Ala Phe Arg Gly Ser Leu Pro Val Leu Asp Leu Pro Gly Asp            3780 3785 3790 Glu Val Arg Pro Ala Val Arg Ser Phe Ala Gly Asp Arg Ile Asp Phe        3795 3800 3805 Gln Ile Asp Ser Ser Leu Ser Ala Ser Leu Gln Glu Leu Ala Thr Arg    3810 3815 3820 Thr Gly Ser Thr Leu Phe Met Val Leu Leu Ala Ala Tyr Thr Ala Leu 3825 3830 3835 3840 Leu His Lys Tyr Thr Gly Gln Glu Asp Val Ile Val Gly Ser Pro Val                3845 3850 3855 Ala Gly Arg Ser His Ala Thr Leu Glu Gly Leu Ile Gly Met Phe Val            3860 3865 3870 Gly Thr Val Ala Leu Arg Thr Tyr Pro Glu Gly Glu Lys Pro Phe Glu        3875 3880 3885 Ala Tyr Leu Gln Glu Val Lys Glu Thr Ala Leu Arg Ala Tyr Glu Asn    3890 3895 3900 Gln Asp Tyr Pro Phe Glu Glu Leu Val Glu Lys Leu Glu Leu Gln Arg 3905 3910 3915 3920 Asp Leu Ser Arg Asn Pro Leu Phe Asp Thr Met Phe Val Leu Gln Asn                3925 3930 3935 Ile Glu Gln Gly Glu Gln Glu Ile Glu Gly Leu Arg Phe Thr Pro Tyr            3940 3945 3950 Asp Asn Val His Pro Ala Ala Lys Phe Asp Leu Thr Leu Thr Val Ser        3955 3960 3965 Glu Val Asp Gly Ala Leu Asn Cys Thr Leu Glu Tyr Ala Thr Ala Ile    3970 3975 3980 Tyr Lys Gln Glu Thr Ala Glu Arg Met Ala Gly His Phe Val Gln Leu 3985 3990 3995 4000 Ile Arg Glu Ala Ile Ala Asn Pro Ala Leu Pro Leu Ser Ser Leu Asp                4005 4010 4015 Ile Val Thr Pro Gln Glu Lys Ser Arg Leu Met Lys Ala Pro Asp Glu            4020 4025 4030 Ala Lys Ala Asp Tyr Pro Arg Asp Lys Thr Ile His Ala Leu Phe Glu        4035 4040 4045 Glu Gln Ala Ala Arg Thr Pro Asn Ala Val Ala Val Val Cys Glu Asp    4050 4055 4060 Ala Ala Leu Ser Tyr Ser Glu Leu Asn Glu Arg Ala Asn Gly Leu Ala 4065 4070 4075 4080 Arg Thr Leu Arg Glu Arg Gly Leu Gln Pro Asp Gly Leu Ala Gly Ile                4085 4090 4095 Met Ala Asp Arg Ser Leu Glu Met Val Val Gly Ile Leu Ala Ile Leu            4100 4105 4110 Lys Ala Gly Gly Ala Tyr Val Pro Val Asp Pro Glu Tyr Pro Glu Asp        4115 4120 4125 Arg Ile His Phe Met Leu Glu Asp Ser Gly Ala Lys Leu Leu Leu Thr    4130 4135 4140 Gln Ala His Leu Glu Gln Arg Val Ser Phe Ala Gly Asn Ile Val Ser 4145 4150 4155 4160 Leu Asp Lys Thr Ala Ser Tyr Lys Glu Asp Val Ser Asn Leu Gln Pro                4165 4170 4175 Ala Ala Gly Pro Asn His Leu Ala Tyr Val Ile Tyr Thr Ser Gly Thr            4180 4185 4190 Thr Gly Lys Pro Lys Gly Thr Leu Ile Glu His Lys Asn Val Val Arg        4195 4200 4205 Leu Leu Phe Asn Asp Lys Asn Met Phe Asp Phe Asp Ala Gln Asp Thr    4210 4215 4220 Trp Thr Leu Phe His Ser Phe Cys Phe Asp Phe Ser Val Trp Glu Met 4225 4230 4235 4240 Tyr Gly Ala Leu Leu Asn Gly Gly Arg Leu Val Ile Val Pro Ser Leu                4245 4250 4255 Thr Ala Lys Ser Pro Asp Arg Phe Leu Gln Leu Leu Lys Asp Gln Lys            4260 4265 4270 Val Thr Val Leu Asn Gln Thr Pro Thr Tyr Phe Tyr Gln Leu Leu Gln        4275 4280 4285 Glu Glu Leu Gly His Gln Ala Ala Glu Leu Ser Leu Arg Met Ile Ile    4290 4295 4300 Phe Gly Gly Glu Ala Leu Thr Pro Ala Leu Leu Lys Asp Trp Arg Thr 4305 4310 4315 4320 Lys Tyr Pro Gln Val Gln Leu Ile Asn Met Tyr Gly Ile Thr Glu Thr                4325 4330 4335 Thr Val His Val Thr Tyr Lys Glu Ile Thr Glu Leu Glu Ile Glu Gln            4340 4345 4350 Gly Arg Ser Asn Ile Gly Thr Thr Ile Pro Thr Leu Arg Ala Tyr Ile        4355 4360 4365 Leu Asp Glu Gln Arg Arg Pro Gln Pro Ile Gly Ile Pro Gly Glu Leu    4370 4375 4380 Tyr Val Ala Gly Val Gly Leu Ala Arg Gly Tyr Leu Asn Arg Pro Glu 4385 4390 4395 4400 Leu Thr Glu Glu Lys Phe Val Ala His Pro Phe Glu Ala Gly Glu Arg                4405 4410 4415 Met Tyr Arg Ser Gly Asp Leu Ala Arg Trp Leu Pro Asp Gly Ser Met            4420 4425 4430 Glu Tyr Leu Gly Arg Ile Asp His Gln Val Lys Ile Arg Gly Tyr Arg        4435 4440 4445 Ile Glu Leu Gly Glu Val Glu Ala Lys Leu Leu His Ala Pro Ser Val    4450 4455 4460 Arg Glu Ala Val Val Leu Ala Arg Glu Asp Gly Ser Gly Gln Lys Val 4465 4470 4475 4480 Leu Ile Gly Tyr Phe Thr Ala Asp Gln Met Leu Thr Val Gly Glu Leu                4485 4490 4495 Arg Lys Ala Leu Ala Ala Glu Leu Pro Ala Tyr Met Ile Pro Ser Tyr            4500 4505 4510 Phe Met Gln Leu Glu Gln Met Pro Leu Thr Pro Asn Gly Lys Leu Asp        4515 4520 4525 Arg Lys Ala Leu Pro Ala Pro Glu Ala Asn Val Gln Thr Gly Ala Val    4530 4535 4540 Tyr Glu Pro Pro Arg Thr Lys Ala Glu Glu Thr Leu Ala Ser Val Trp 4545 4550 4555 4560 Gln Gly Val Leu Gly Ala Gln Gln Val Gly Ile His Asp His Phe Phe                4565 4570 4575 Asp Leu Gly Gly Asp Ser Ile Lys Ala Ile Gln Val Ser Ser Arg Leu            4580 4585 4590 Phe Gln Ala Gly Tyr Lys Leu Glu Met Lys Asp Leu Phe Lys Tyr Pro        4595 4600 4605 Thr Ile Ala Glu Leu Ser Pro Tyr Leu Gln Ala Ala Gly Arg Thr Ala    4610 4615 4620 Glu Gln Gly Glu Ile Lys Gly Ala Ala Glu Leu Met Pro Ile Gln Arg 4625 4630 4635 4640 Trp Phe Phe Glu Arg His Thr Ala Glu Pro His His Tyr Asn His Ala                4645 4650 4655 Val Met Leu Tyr Arg Lys Asp Gly Phe Asp Glu Ala Ala Leu Arg Leu            4660 4665 4670 Thr Met Asp Gln Ile Ala Ile His His Asp Ala Leu Arg Met Val Phe        4675 4680 4685 Arg Pro Thr Glu Ala Gly Tyr Ala Ala Trp Asn Arg Gly Thr Asp Glu    4690 4695 4700 Gly Glu Leu Tyr Thr Leu Asp Ile Ala Asp Val Arg Gln Val Glu Asp 4705 4710 4715 4720 Gln Thr Ala Ala Val Gln Ala Gln Ala Asp Ala Ile Gln Ala Ser Phe                4725 4730 4735 Asp Leu Glu Gly Gly Pro Leu Phe Lys Leu Gly Leu Phe His Cys Asp            4740 4745 4750 Asp Gly Asp His Leu Leu Ile Val Ile His His Leu Leu Val Asp Gly        4755 4760 4765 Val Ser Trp Arg Ile Leu Phe Glu Asp Ile Ala Ala Gly Tyr Glu Gln    4770 4775 4780 Ala Leu Asn Gly Gln Ala Ile Ile Leu Pro Gln Lys Thr Asp Ser Tyr 4785 4790 4795 4800 Leu Val Trp Ser Glu Gln Ala Thr Lys Tyr Ala Ala Gly Pro Ala Leu                4805 4810 4815 Asp Lys Glu Arg Ala Tyr Trp Gln Leu Ile Glu Glu Ala Ile Leu Ala            4820 4825 4830 Pro Leu Pro Lys Asp Glu Asp Gln Lys Pro Gly Thr Ile Arg Asp Thr        4835 4840 4845 Glu Ser Val Thr Val Thr Trp Ser Ala Gln Glu Thr Asp Leu Leu Leu    4850 4855 4860 Arg Gln Ala Asn Arg Ala Tyr His Thr Glu Thr Asn Asp Leu Leu Leu 4865 4870 4875 4880 Thr Ala Leu Gly Ala Ala Ile Gln Arg Trp Thr Gly Met Glu Gln Ile                4885 4890 4895 Leu Val Asn Leu Glu Gly His Gly Arg Glu Met Ile Val Pro Asp Leu            4900 4905 4910 Asp Ile Thr Arg Thr Val Gly Trp Phe Thr Thr Gln Tyr Pro Val Leu        4915 4920 4925 Leu Asn Leu Gln Gly Arg Gln Glu Val Ser Ala Arg Ile Lys Arg Ile    4930 4935 4940 Lys Glu Asn Leu Arg Glu Val Pro His Lys Gly Ile Gly Tyr Gly Leu 4945 4950 4955 4960 Leu Lys Tyr Ile Ala Pro Glu Lys Ser Val Gly Phe Gly Val Glu Pro                4965 4970 4975 Glu Ile Ser Phe Asn Tyr Leu Gly Gln Phe Asp Gln Asp Leu Glu Gly            4980 4985 4990 Asn Ala Leu Ser Leu Ser Thr His Ser Val Gly Lys Ala Leu Ser Asp        4995 5000 5005 Leu Thr Pro Gln Gln Tyr Ala Leu Asp Val Asn Gly Met Ile Ala Glu    5010 5015 5020 Gly Gln Leu Ser Leu Thr Ile Thr Tyr Ser Ser Arg Gln Tyr Arg Lys 5025 5030 5035 5040 Glu Thr Val Ser His Phe Ala Glu Leu Leu Gln Ser Ser Leu Ser Glu                5045 5050 5055 Val Ile Leu His Cys Val Ala Gln Glu Arg Ser Gln Leu Thr Pro Ser            5060 5065 5070 Asp Val Leu Phe Gln Gly Leu Thr Leu Glu Gln Leu Asp Arg Leu Thr        5075 5080 5085 Ala Gln Thr Ala His Ile Gly Glu Ile Glu Asp Val Tyr Lys Leu Thr    5090 5095 5100 Ala Met Gln Lys Gly Met Leu Phe His Ser Leu Leu Glu Pro Asp Ser 5105 5110 5115 5120 Ser Ser Tyr Phe Glu Gln Ala Thr Phe Glu Leu Arg Gly Ser Phe Asp                5125 5130 5135 Val Asp Ile Phe Phe Glu Ser Phe Gln Ala Leu Ala Gln Arg His Ala            5140 5145 5150 Ile Leu Arg Thr Gly Phe Tyr Asn Asn Ile Ala Asp Val Pro Leu Gln        5155 5160 5165 Val Ala Phe Lys Gln Arg Leu Ile Pro Leu His Tyr Val Asp Leu Arg    5170 5175 5180 Asp Ala Ser Met Gln Glu Gln Glu Ala Arg Ile Lys Ala Tyr Ile Ala 5185 5190 5195 5200 Glu Asp Met Val Lys Gly Phe Ser Leu Ser Glu Asp Pro Leu Met Arg                5205 5210 5215 Val Thr Val Leu Gln Lys Asp Gln Ser Cys Leu Val Leu Trp Ser Phe            5220 5225 5230 His His Ile Val Met Asp Gly Trp Cys Ile Pro Ile Ile Thr Gln Glu        5235 5240 5245 Leu Phe Asp Tyr Tyr Ser Ala Lys Lys Gln Gln Ile Gln Pro Val Leu    5250 5255 5260 Pro Pro Ala Gln Pro Tyr Ser Arg Tyr Ile Glu Trp Leu Asp Ala Gln 5265 5270 5275 5280 Asp Glu Gln Glu Ala Ser Thr Tyr Trp Ser Gln Tyr Leu Glu Asp Tyr                5285 5290 5295 Asp Gly Asn Thr Val Leu Pro Glu Gly Lys Thr Lys Ser Gln Ala Lys            5300 5305 5310 Glu Ala Gly Tyr Val Leu Lys Glu His Val Leu His Leu Gly Ala Ser        5315 5320 5325 Leu Thr Gly Lys Met Asp Val Val Ala Lys Arg Asn His Val Thr Val    5330 5335 5340 Asn Thr Leu Met Gln Thr Ala Trp Gly Leu Ile Leu Gln Arg Tyr Asn 5345 5350 5355 5360 Ala Ser Ser Asp Val Val Phe Gly Gly Val Val Ser Gly Arg Pro Ala                5365 5370 5375 Glu Ile Ala Gly Ile Glu Asn Met Val Gly Leu Phe Ile Asn Thr Val            5380 5385 5390 Pro Ile Arg Val Gln Ser Ser Lys Asp Glu Ala Phe Val Lys Val Met        5395 5400 5405 Lys Arg Thr Gln Ala Gln Ser Leu Ala Gly Arg Ala Tyr Asp Thr Tyr    5410 5415 5420 Pro Leu Tyr Glu Ile Gln Gly Lys Thr Thr Gln Lys Gln Asp Leu Ile 5425 5430 5435 5440 Ser His Ile Met Ile Phe Glu Asn Tyr Pro Leu Asp Glu Gln Val Glu                5445 5450 5455 Gln Ser Gly Asn Gln Thr Glu Asp Asn Leu Glu Val Ala Asn Phe Thr            5460 5465 5470 Met Phe Glu Gln Thr Asn Tyr Asp Phe Asn Leu Val Val Ile Pro Gly        5475 5480 5485 Glu Asp Ile Lys Val Cys Ile Arg Tyr Asn Ala Ser Val Tyr Glu Gln    5490 5495 5500 Glu Ser Ile Ala Arg Ile Gly Gly His Leu Leu Gln Met Leu Asp Gln 5505 5510 5515 5520 Val Ala Ala Arg Pro Gln Ala Thr Ile Gln Glu Leu Glu Ile Val Thr                5525 5530 5535 Ser Glu Glu Arg Met Asn Leu Leu Asp Trp Gly Gly Lys Ala His Ala            5540 5545 5550 Tyr Pro Ser Asp Gln Gly Leu His Thr Leu Phe Glu Glu Gln Val Val        5555 5560 5565 Arg Thr Pro Asp Lys Ile Ala Ala Ile Ser Gly Asp Ile Gln Ile Thr    5570 5575 5580 Tyr Arg Glu Leu Asn Glu Gln Ala Asn Arg Leu Ala Ser Thr Leu Ile 5585 5590 5595 5600 Ala Gln Gly Leu Arg Ser Glu Gln Val Val Gly Leu Leu Ala Asp Arg                5605 5610 5615 Ser Val Glu Leu Leu Val Ala Ile Met Gly Val Leu Lys Ala Gly Gly            5620 5625 5630 Ala Tyr Val Pro Ile Asp Pro Glu Tyr Pro Gln Glu Arg Ile Gln Tyr        5635 5640 5645 Ile Leu Lys Asp Ser Gly Ala Glu Ile Leu Leu Thr Gln Ser His Leu    5650 5655 5660 Thr Lys Leu Ala Ser Phe Glu Gly Met Val Met Glu Leu Asp Ser Ser 5665 5670 5675 5680 His Ile Tyr Gly Thr Gly Val Asn Asn Pro Asn Ile Pro Val Arg Gly                5685 5690 5695 Asn Asp Leu Val Tyr Leu Ile Tyr Thr Ser Gly Thr Thr Gly Asn Pro            5700 5705 5710 Lys Gly Thr Met Ile Asn His Lys Gly Ile Val Asn Tyr Ile Trp Trp        5715 5720 5725 Ala Asn Lys Val Tyr Cys Ala Gly Gln Pro Thr Asp Phe Pro Leu Tyr    5730 5735 5740 Ser Ser Ile Ser Phe Asp Leu Thr Met Thr Ser Met Phe Thr Pro Leu 5745 5750 5755 5760 Ile Asn Gly Gly Ile Val Arg Ile Tyr Asp Gly Ile Asp Lys Ala Glu                5765 5770 5775 Val Val Gln His Ile Leu Arg Glu Asn Ala Val Asp Ile Leu Lys Leu            5780 5785 5790 Thr Pro Thr His Leu Ser Leu Ile Lys Asp Met Ala Ile Pro Ala Glu        5795 5800 5805 Ser Arg Ile Gln Gln Leu Ile Val Gly Gly Glu Asn Leu Thr Thr His    5810 5815 5820 Leu Ser Lys Ile Ile Thr Asp Leu Phe Gly Gly Asn Ile Lys Ile Tyr 5825 5830 5835 5840 Asn Glu Tyr Gly Pro Thr Glu Thr Val Val Gly Cys Met Ile His Leu                5845 5850 5855 Tyr Asp Pro Ala Lys Asp Thr Arg Glu Ser Val Pro Ile Gly Leu Pro            5860 5865 5870 Ser Asp Asn Ile Tyr Ile His Ile Leu Asp Glu Gln Leu Arg Leu Val        5875 5880 5885 Pro Leu Gly Val Glu Gly Glu Met Tyr Ile Ala Gly Asp Gly Val Ala    5890 5895 5900 Arg Gly Tyr Leu Asn Arg Pro Asp Leu Thr Ala Glu Lys Phe Ile Arg 5905 5910 5915 5920 Asn Pro Phe Ala Ala Glu Gly Asn Met Tyr Arg Thr Gly Asp Leu Ala                5925 5930 5935 Arg Arg Leu Pro Asn Gly Asp Ile Glu Tyr Ile Gly Arg Ile Asp His            5940 5945 5950 Gln Val Lys Ile Arg Gly Tyr Arg Ile Glu Leu Gly Glu Ile Glu Ala        5955 5960 5965 Lys Leu Leu Asp Ile Pro Leu Val Glu Glu Ala Leu Val Val Ala Trp    5970 5975 5980 Ala Asp Ala His Gly Gln Lys Ser Leu Cys Ala Tyr Phe Val Ala Asp 5985 5990 5995 6000 Arg Glu Met Ser Val Ser Glu Leu Arg Asn Glu Leu Ser Ala Gly Leu                6005 6010 6015 Pro Ala Tyr Met Ile Pro Ser Tyr Phe Val Gln Leu Asp Val Met Pro            6020 6025 6030 Leu Thr Pro Asn Gly Lys Leu Asp Arg Lys Ala Leu Pro Glu Pro Asn        6035 6040 6045 Ser Gly Ile Lys Ala Gly Ala Asp Phe Thr Ala Pro Arg Thr Asp Val    6050 6055 6060 Glu Asn Ile Leu Ala Ser Ile Trp Gln Gly Val Leu Gly Val Pro Leu 6065 6070 6075 6080 Val Gly Ile His Asp Asn Phe Phe Glu Leu Gly Gly Asp Ser Ile Lys                6085 6090 6095 Ser Ile Gln Val Ser Ser Arg Leu Leu Gln Ala Gly Tyr Lys Leu Glu            6100 6105 6110 Met Lys Asp Leu Phe Gly Tyr Pro Thr Ile Ala Glu Leu Ala Gln Arg        6115 6120 6125 Val Ser Val Val Ser Arg Ile Ala Asp Gln Ser Glu Val His Gly Ser    6130 6135 6140 Val Arg Leu Ser Pro Ala Gln His Arg Phe Phe Asp Glu Gln Ser Met 6145 6150 6155 6160 Asp Leu His His Phe Asn Gln Ser Val Met Leu Tyr Arg Arg Asp Gly                6165 6170 6175 Phe Asn Thr Asp Ala Leu Ala Glu Val Val Arg Lys Ile Ala Glu His            6180 6185 6190 His Asp Ala Leu Arg Leu Val Phe Arg Gln Gly Glu Gln Gly Leu Glu        6195 6200 6205 Ala Trp Asn Arg Ser Met Gly Glu Gly Glu Leu Tyr Ser Leu Gln Ile    6210 6215 6220 His Asp Leu Arg Asp Glu Thr Asp Pro Ala Ser Ala Ile Glu Ala Gly 6225 6230 6235 6240 Ala Glu Ala Ile Gln Arg Ser Ile Ser Leu Glu Asp Gly Pro Leu Phe                6245 6250 6255 Arg Leu Gly Leu Phe Arg Cys Ala Glu Gly Glu His Leu Leu Ile Val            6260 6265 6270 Ile His His Leu Ala Val Asp Gly Val Ser Trp Arg Ile Leu Phe Glu        6275 6280 6285 Asp Leu Gln Glu Gly Tyr Glu Gln Ala Ala Arg Gly Glu Ala Val Lys    6290 6295 6300 Phe Pro Gln Lys Thr Asp Ser Tyr Arg Ala Trp Val Glu Gly Ile Thr 6305 6310 6315 6320 Gln Phe Ala Asn Ser Pro Ala Ala Glu Gln Glu Arg Ser Tyr Trp Ala                6325 6330 6335 Glu Val Glu Gly Asp Gly Phe Val Pro Leu Pro Lys Asp Lys Val Asp            6340 6345 6350 Gly Ala Leu Leu Ile Lys Asp Ser Glu Ala Val Thr Val Arg Trp Ser        6355 6360 6365 Pro Glu Glu Thr Glu Gln Phe Leu Lys Glu Ala Asn Arg Thr Tyr Asn    6370 6375 6380 Thr Glu Val Asn Asp Leu Leu Leu Thr Ala Leu Gly Met Ala Val His 6385 6390 6395 6400 Glu Trp Thr Gly Ile Glu Arg Val Gly Ile Leu Leu Glu Gly His Gly                6405 6410 6415 Arg Glu Pro Val Val Pro Glu Leu Asp Ile Thr Arg Thr Ile Gly Trp            6420 6425 6430 Phe Thr Ser Gln Tyr Pro Val Ala Leu Glu Met Gly Gly Glu Leu Glu        6435 6440 6445 Ile Gly Ala Arg Ile Lys His Val Lys Glu Gly Leu Arg Arg Ile Pro    6450 6455 6460 Asn Lys Gly Val Gly Tyr Gly Ile Leu Lys Tyr Leu Ser Asp Gly Ser 6465 6470 6475 6480 Asp Val Ser Ser Phe Ser Ala Glu Pro Glu Ile Thr Phe Asn Tyr Leu                6485 6490 6495 Gly Gln Phe Asp Gln Asp Leu Ala Gly Gly Met Met Glu Val Ser Ser            6500 6505 6510 Tyr Ser Val Gly Pro Glu Val Ser Glu Gln Met Val Gln His Gln Ala        6515 6520 6525 Val Asn Ile Asn Gly Leu Ile Ala Glu Gly Gln Leu Gln Leu Ser Val    6530 6535 6540 Ser Tyr Asn Arg His Gln Leu His Gly Glu Ser Val Ala Lys Phe Val 6545 6550 6555 6560 Gly Ile Leu Lys Asn Arg Leu Ser Glu Val Ile Gly His Cys Val Ser                6565 6570 6575 Lys Glu Arg Thr Glu Leu Thr Pro Ser Asp Val Leu Leu Lys Asp Ile            6580 6585 6590 Ser Leu Glu Lys Ile Glu Glu Leu Glu Glu Gln Thr Arg His Ile Gly        6595 6600 6605 Ser Ile Glu Asn Met Tyr Lys Leu Thr Pro Met Gln Lys Gly Met Leu    6610 6615 6620 Phe His Ser Leu Leu Glu Pro His Ser Glu Val Tyr Phe Glu Gln Ala 6625 6630 6635 6640 Lys Phe Glu Ile Gln Gly Ala Phe Tyr Pro Glu Asp Phe Lys Arg Ser                6645 6650 6655 Leu Lys Tyr Leu Met Lys Arg His Ala Ile Leu Arg Thr Asn Phe His            6660 6665 6670 Ala Gly Trp Gly Asp Phe Pro Ile Gln Ile Val Phe Lys Glu Arg Ala        6675 6680 6685 Cys Asp Phe Val Tyr Glu Asp Leu His Glu Leu Glu Ala Asp Glu Ile    6690 6695 6700 Gln Ala Arg Leu Ala Ala Tyr Thr Ala Gln Asp Lys Ala Arg Gly Phe 6705 6710 6715 6720 Asn Leu Ala Glu Glu Ala Leu Leu Arg Val Ala Ile Leu Arg Thr Ala                6725 6730 6735 Glu Glu Ala Tyr His Leu Leu Trp Ser Ser His His Ile Ile Leu Asp            6740 6745 6750 Gly Trp Cys Met Pro Leu Val Leu Gln Glu Val Phe Glu Thr Tyr Gly        6755 6760 6765 Ala Leu Arg Glu Gln Arg Glu Pro Glu Leu Pro Ala Ala Val Ser Tyr    6770 6775 6780 Ser Gln Tyr Ile Gln Trp Leu Glu Lys Gln Gly Glu Glu Glu Ala Ser 6785 6790 6795 6800 Ser Tyr Trp Arg Gly Tyr Leu Glu Gly Tyr Glu Gln Gln Thr Lys Leu                6805 6810 6815 Pro Gln Ala Ile Thr Gln Pro Ser Ala Lys Ala Glu Ala Tyr Val Ser            6820 6825 6830 Glu Lys Leu Val Phe Thr Leu Asp Ala Glu Leu Thr Asp Arg Leu Glu        6835 6840 6845 Gln Val Ala Lys Gln His Gln Val Thr Met Asn Thr Leu Met Gln Ala    6850 6855 6860 Ala Trp Gly Ile Val Leu Gln Arg Tyr Asn Arg Ser Gln Asp Val Val 6865 6870 6875 6880 Phe Gly Ser Val Val Ser Gly Arg Pro Ala Glu Ile Pro Gly Ile Glu                6885 6890 6895 Ser Met Ile Gly Leu Phe Ile Asn Thr Val Pro Val Arg Val Gln Ala            6900 6905 6910 Glu Gly Ser Asp Ser Phe Ser His Val Met Lys Arg Gln Gln Glu Leu        6915 6920 6925 Tyr Leu Ala Gly His Ala Tyr Asp Ser Tyr Pro Leu Tyr Glu Ile Gln    6930 6935 6940 Ala Gln Ser Glu Gln Lys Gln Asp Leu Ile Ser His Ile Met Val Phe 6945 6950 6955 6960 Glu Asn Tyr Pro Val Glu Glu His Leu Glu Glu Lys Ile Ala Asn Asp                6965 6970 6975 Glu Ala Glu Tyr Lys Ile Thr Asp Val Gln Met Phe Glu Gln Thr Asn            6980 6985 6990 Tyr Asp Phe Asn Leu Ile Val Leu Pro Gly Arg Ser Leu Glu Phe Leu        6995 7000 7005 Tyr Arg Tyr Asn Ala Arg Val Tyr Asp Arg Glu Ser Val Glu Arg Ile    7010 7015 7020 Gln Gly His Leu Thr Arg Ile Leu Thr Ser Val Ser Val Gln Pro Ala 7025 7030 7035 7040 Ile Arg Ile Asp Glu Leu Glu Leu Ile Thr Pro Glu Glu Lys Ser Gln                7045 7050 7055 Ile Ile Glu Val Trp Gly Asp Thr Ala Ala Pro Tyr Pro Arg Glu Lys            7060 7065 7070 Thr Leu His Gly Ile Phe Glu Glu Lys Ala Ala Leu Thr Pro Asp Arg        7075 7080 7085 Thr Ala Leu Ile Tyr Gly Glu Thr Val Leu Thr Tyr Gly Glu Leu His    7090 7095 7100 Gln Gln Ala Asn Arg Leu Ala Arg Thr Leu Arg Val Gln Gly Val Arg 7105 7110 7115 7120 Pro Asp Gln Pro Val Gly Ile Met Val Glu Arg Ser Leu Glu Met Ile                7125 7130 7135 Ile Gly Ile His Ala Ile Leu Lys Ala Gly Gly Ala Tyr Val Pro Ile            7140 7145 7150 Asp Pro Gly Phe Pro Glu Asp Arg Ile Arg His Met Leu Glu Asp Ser        7155 7160 7165 Gly Ala Lys Leu Leu Leu Thr Lys Asn His Leu Lys Asp Arg Phe Pro    7170 7175 7180 Phe Thr Gly Thr Ile Leu Ser Leu Asp Asp Pro Gln Ala Tyr His Ala 7185 7190 7195 7200 Asp Asp Ser Asn Leu Glu Pro Val Ala Gly Pro Glu His Leu Ala Tyr                7205 7210 7215 Ile Ile Tyr Thr Ser Gly Ser Thr Gly Lys Pro Lys Gly Val Met Ile            7220 7225 7230 Glu His His Ser Ala Val His Thr Leu Ser Gln Leu Glu Ala Glu Tyr        7235 7240 7245 Pro Met Leu Ala Gly Asp Arg Phe Leu Leu Lys Thr Thr Phe Thr Phe    7250 7255 7260 Asp Phe Ser Val Pro Glu Leu Phe Cys Trp Phe Phe Gly Gln Gly Thr 7265 7270 7275 7280 Leu Val Ile Leu Pro Pro Gly Val Asp Lys Asp Pro Val Ala Leu Leu                7285 7290 7295 Glu Ala Val Asp Ala Asn Gln Ile Thr His Leu Asn Leu Val Pro Ser            7300 7305 7310 Met Leu Ser Val Leu Val Gln Tyr Leu Lys Glu Ser Ser Thr Gln Gly        7315 7320 7325 Phe Leu Thr Leu Lys Tyr Leu Phe Ala Cys Gly Glu Thr Leu Pro Ala    7330 7335 7340 Lys Leu Val Glu Glu Tyr Tyr Lys Val Ser Pro Tyr Ala Val Leu Glu 7345 7350 7355 7360 Asn Ile Tyr Gly Pro Thr Glu Ala Ala Val Tyr Ala Thr Arg Tyr Thr                7365 7370 7375 Thr Ser Leu Glu Thr Ala Ala Leu Thr His Val Pro Ile Gly Lys Pro            7380 7385 7390 Tyr Ala Asn Val Gln Val Trp Met Met Asp Ser Ala Ser Gln Val Ser        7395 7400 7405 Pro Val Gly Val Pro Gly Glu Leu Cys Ile Ala Gly Glu Gly Val Ala    7410 7415 7420 Arg Gly Tyr Phe Asn Gln Pro Asp Leu Thr Ala Glu Lys Phe Ile Pro 7425 7430 7435 7440 His Pro Tyr Lys Pro Gly Glu Arg Ile Tyr Arg Thr Gly Asp Leu Ala                7445 7450 7455 Arg Trp Val Pro Asp Gly Asn Ile Glu Tyr Leu Gly Arg Ile Asp His            7460 7465 7470 Gln Val Lys Ile Arg Gly Tyr Arg Ile Glu Leu Gly Glu Val Glu Ala        7475 7480 7485 Gln Ile Leu Lys Val Pro Ser Val Gln Glu Ala Val Val Leu Ala Leu    7490 7495 7500 Ala Asp Ser Thr Gly Ser Thr Gln Leu Cys Ala Tyr Phe Val Ala Glu 7505 7510 7515 7520 Glu Gly Leu Ala Ala Gly Val Leu Arg Glu Ala Leu Ala Ser Glu Leu                7525 7530 7535 Pro Ser Tyr Met Ile Pro Thr Ala Phe Val Gln Leu Ala Gln Met Pro            7540 7545 7550 Leu Asn Pro Asn Gly Lys Leu Asp Arg Lys Ala Leu Pro Ala Pro Glu        7555 7560 7565 Thr Leu Leu Arg Ser Thr Ala Glu Tyr Ile Ala Pro Arg Thr Gln Thr    7570 7575 7580 Glu Val Glu Leu Ala Gln Ile Trp Ser Glu Val Leu Gly Val Gln Glu 7585 7590 7595 7600 Ile Gly Ile Arg Asp His Phe Phe Glu Leu Gly Gly His Ser Leu Lys                7605 7610 7615 Val Leu Gly Leu Ile Gln Arg Ile Ser Ser Gly Met Gly Val Gln Leu            7620 7625 7630 Pro Leu Gln Val Val Phe Asn Leu Pro Thr Val Glu Glu Met Ala His        7635 7640 7645 Glu Ile Phe Lys Leu Gln Ala Thr Thr Ser Ser Ala Asp Glu Gln Glu Met    7650 7655 7660 Glu Ile Ile His Phe Pro Gly Lys Gly Ile Leu Lys Val Phe Cys Phe 7665 7670 7675 7680 Pro Pro Arg Val Gly His Ser Leu Gly Tyr Tyr Glu Met Ala Lys Glu                7685 7690 7695 Leu Asp Gly Leu Cys Glu Val Tyr Gly Met Glu Phe Ile Gly Asp Arg            7700 7705 7710 Phe Gln Gly Gln Asp Met Leu Asp Arg Tyr Ile Asp Ala Ile Val Asp        7715 7720 7725 Ile Gln Ala Glu Gly Pro Tyr Ile Phe Leu Gly Tyr Ser Leu Gly Gly    7730 7735 7740 Asn Leu Ala Phe Glu Val Ala Lys Ala Met Glu Ile Arg Gly His His 7745 7750 7755 7760 Val Gly Asp Ile Ile Met Val Asp Ala Met Arg Lys Met Ser Lys Asp                7765 7770 7775 Glu Ser Thr Pro Glu Glu Leu Glu Glu Ile Val Glu Thr Val Leu Asp            7780 7785 7790 Ser Ile Arg Glu Gln Tyr Lys Ala Phe Leu Ala Asp Pro Ala Asp Arg        7795 7800 7805 Glu Arg Val Met Asp Lys Met Leu Val Tyr Ser Thr Tyr Arg Asp Glu    7810 7815 7820 Leu Ile Asn Ala Gly Glu Val His Ala Asn Ile His Ala Leu Ile Ala 7825 7830 7835 7840 Glu Asp Asp Ser Val Gly Pro Asp Thr Ser Leu Asp Lys Leu Leu Trp                7845 7850 7855 Gln Gln Ala Thr Leu Gly Gln Tyr Lys Glu Tyr Glu Val Ile Gly Thr            7860 7865 7870 His Asp Val Leu Leu Asp Ser Gly Phe Val Gly Glu Asn Ala Lys Val        7875 7880 7885 Leu Arg Gln Ile Leu Gly Lys Ile Ala Glu Thr Ser Ser Lys Asn Lys    7890 7895 7900 Pro Ile Leu Ser 7905

Claims (10)

Polypeptides or variants thereof involved in fuzacidine synthesis. The polypeptide of claim 1 set forth in SEQ ID NO: 2. The method of claim 1, wherein the fusicidin is fuzacidin A, fuzacidin B, fuzacidin C, fuzacidin D and LI-F03, LI-F04, LI-F05, LI-F07 And LI-F08. The polypeptide of claim 1, wherein the polypeptide comprises one or more modules, wherein the module comprises two or more domains selected from the group consisting of A, C, T, E, TE domains. The method of claim 4, wherein the addition, substitution or deletion of one or more modules, domains or amino acids or the linking of D- and L-amino acids with hydroxy acids, mutations and oxidation, acylation, glycosylation, N- in the peptide chains. A polypeptide comprising additional structural modifications, including methylation and heterocycle ring formation. A gene encoding the polypeptide of claim 1. The gene of claim 6, wherein the gene is set forth in SEQ ID NO: 1. Recombinant vector comprising the gene of claim 6. A host cell transformed with the vector of claim 8. 1) introducing the gene of claim 6 into an expression vector; 2) transforming the expression vector into which the gene of step 1) is introduced into a host cell; And 3) culturing the transformant of step 2) and 4) A method for preparing fuzacidine or a derivative thereof, comprising separating and purifying fuzacidine or a derivative thereof from the culture of step 3).

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