Patent Publication Number: US-2003229916-A1

Title: Phosphoribosylaminoimidazole carboxylase

Description:
[0001] This application claims the benefit of U.S. Provisional Application No. 60/341,955, filed Dec. 19, 2001, the entire content of which is herein incorporated by reference. 
    
    
     
       FIELD OF INVENTION  
       [0002] The field of invention relates to plant molecular biology, and more specifically, to nucleic acid fragments encoding phosphoribosylamino-imidazole carboxylase in plants and seeds.  
       BACKGROUND OF INVENTION  
       [0003] Purine and pyrimidine nucleotides are produced in the cell by de novo biosynthetic pathways and by salvage pathways. Salvage pathways function to recover nucleotide bases released during the degradation of nucleic acids. Purines are components of DNA and RNA. Regulation of purine synthesis and metabolism within the cell is critical to functions of all cells. Most mutations affecting nucleotide biosynthetic enzymes are lethal, although certain redundancy and salvage pathways may moderate the deleterious effects of some of these mutations. The detailed pathway of purine biosynthesis was worked out in the 1950s. The origin of carbon atom 6 of the purine ring from carbon dioxide during the synthesis of the purine ring by chicken liver was attributed to phosphoribosylaminoimidazole carboxylase (AIR carboxylase) (Lukens, L. N. and Buchanan, J. M.,  J. Biol. Chem.  234(7):1799-1805 (1959)). AIR carboxylase is an enzyme that functions in purine metabolism converting 5-amino-1-ribosylimidazole 5′-phosphate (AIR) and carbon dioxide to 5-amino-1-ribosyl-4-imidazolecarboxylic acid 5′-phosphate (carboxy-AIR).  
       [0004] De novo synthesis of the purine, inosine monophosphate, is achieved via ten enzymatic reactions. In eukaryotic cells several of the steps are catalyzed by multifunctional proteins. However, the regulation of de novo purine biosynthesis in the nodules of tropical legumes (mothbean;  Vigna aconitifolia ) revealed that the AIR carboxylase (NCBI General Identification (GI) No. 1709930) and the next enzyme in the purine pathway (5-aminoimidazole-4-N-succinocarboxamide ribonucleotide (SAICAR) synthetase) are distinct proteins in mothbean, unlike in animals where both activities are associated with a single bifunctional polypeptide (Chapman, K. A.; Delauney, A. J.; Kim, J. H. and Verma, D. P.,  Plant Mol. Biol.  24(2)389-395 (1994)). The activities and some properties of AIR carboxylase and SAICAR synthetase were also examined from wheat seedlings ( Triticum aestivum ) and gel filtration (no sequence information) showed that in higher plants the two were also individual proteins (Dolgikh, E. A.; Dolgikh, V. V. and Domkin, V. D., Russ.  J. Plant Physiol.  43(1):10-16 (1996)).  
       SUMMARY OF INVENTION  
       [0005] The present invention includes isolated polynucleotides comprising a nucleotide sequence encoding a polypeptide having phosphoribosylaminoimidazole carboxylase activity wherein the amino acid sequence of the polypeptide and the amino acid sequence of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17 have preferably at least 80% sequence identity. It is also preferred that the identity be at least 85%, at least 90%, or at least 95%. The present invention also includes isolated polynucleotides comprising the complement of the nucleotide sequence. More specifically, the present invention includes isolated polynucleotides encoding the polypeptide sequence of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17 or nucleotide sequences comprising the nucleotide sequence of SEQ ID NO:1, 3, 5, 7, 9, 11 or 16.  
       [0006] In a first embodiment, the present invention includes an isolated polynucleotide comprising: (a) a nucleotide sequence encoding a polypeptide having phosphoribosylaminoimidazole carboxylase activity, wherein the polypeptide has an amino acid sequence of at least 80%, 85%, 90%, or 95% sequence identity, based on the Clustal V method of alignment, when compared to one of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17, or (b) a complement of the nucleotide sequence, wherein the complement and the nucleotide sequence consist of the same number of nucleotides and are 100% complementary. The polypeptide preferably comprises the amino acid sequence of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17. The nucleotide sequence preferably comprises the nucleotide sequence of SEQ ID NO:1, 3, 5, 7, 9, 11 or 16.  
       [0007] In a second embodiment, the present invention concerns a recombinant DNA construct comprising any of the isolated polynucleotides of the present invention operably linked to at least one regulatory sequence, and a cell, a plant, and a seed comprising the recombinant DNA construct.  
       [0008] In a third embodiment, the present invention includes a vector comprising any of the isolated polynucleotides of the present invention.  
       [0009] In a fourth embodiment, the present invention concerns a method for transforming a cell comprising transforming a cell with any of the isolated polynucleotides of the present invention. The cell transformed by this method is also included. Advantageously, the cell is eukaryotic, e.g., a yeast or plant cell, or prokaryotic, e.g., a bacterium.  
       [0010] In a fifth embodiment, the present invention includes a method for producing a transgenic plant comprising transforming a plant cell with any of the isolated polynucleotides of the present invention and regenerating a plant from the transformed plant cell, a transgenic plant produced by this method, and seed obtained from this transgenic plant.  
       [0011] In a sixth embodiment, the present invention includes an isolated polypeptide having phosphoribosylaminoimidazole carboxylase activity, wherein the polypeptide has an amino acid sequence of at least 80%, 85%, 90%, or 95% identity, based on the Clustal V method of alignment, when compared to one of SEQ ID NO:2, 4, 6, 8, 10,12 or 17. The polypeptide preferably comprises one of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17.  
       [0012] In a seventh embodiment, the present invention includes a method for isolating a polypeptide having phosphoribosylaminoimidazole carboxylase activity comprising isolating the polypeptide from a cell or culture medium of the cell, wherein the cell comprises a recombinant DNA construct comprising a polynucleotide of the present invention operably linked to at least one regulatory sequence.  
       [0013] In an eighth embodiment, this invention includes a method for selecting a transformed cell comprising: (a) transforming a host cell with the recombinant DNA construct or an expression cassette of the present invention; and (b) growing the transformed host cell, preferably a plant cell, under conditions that allow expression of the phosphoribosylaminoimidazole carboxylase polynucleotide in an amount sufficient to complement a null mutant in order to provide a positive selection means.  
       [0014] In a ninth embodiment, this invention includes a method of altering the level of expression of a phosphoribosylaminoimidazole carboxylase protein in a host cell comprising: (a) transforming a host cell with a recombinant DNA construct of the present invention; and (b) growing the transformed host cell under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of altered levels of the phosphoribosylaminoimidazole carboxylase protein in the transformed host cell.  
       [0015] A further embodiment of the instant invention is a method for evaluating at least one compound for its ability to inhibit the activity of a phosphoribosylaminoimidazole carboxylase, the method comprising the steps of: (a) introducing into a host cell a recombinant DNA construct comprising a nucleic acid fragment encoding a phosphoribosylaminoimidazole carboxylase polypeptide, operably linked to at least one regulatory sequence; (b) growing the host cell under conditions that are suitable for expression of the recombinant DNA construct wherein expression of the recombinant DNA construct results in production of phosphoribosylaminoimidazole carboxylase polypeptide in the host cell; (c) optionally purifying the phosphoribosylaminoimidazole carboxylase polypeptide expressed by recombinant DNA construct in the host cell; (d) treating the phosphoribosylaminoimidazole carboxylase polypeptide with a compound to be tested; (e) comparing the activity of the phosphoribosylaminoimidazole carboxylase polypeptide that has been treated with a test compound to the activity of an untreated phosphoribosylaminoimidazole carboxylase polypeptide, and (f) selecting compounds with potential for inhibitory activity. 
     
    
    
     BRIEF DESCRIPTION OF THE DRAWING AND SEQUENCE LISTINGS  
     [0016]FIGS. 1A and 1B depict the ten enzymatic reactions that are required for the de novo synthesis of the purine, inosine monophosphate (inosinate), from the precursor phosphoribosylpyrophosphate (PRPP). Phosphoribosylaminoimidazole (AIR) carboxylase is the sixth step and it converts 5-amino-1-ribosylimidazole 5′-phosphate (AIR) and carbon dioxide to 5-amino-1-ribosyl-4-imidazolecarboxylic acid 5′-phosphate (carboxy-AIR).  
     [0017]FIGS. 2A, 2B,  2 C and  2 D depict the amino acid sequence alignment of the phosphoribosylaminoimidazole carboxylases encoded by the following:  
     [0018] (a) nucleotide sequence derived from corn clone cpd1c.pk002.l23 (SEQ ID NO:2),  
     [0019] (b) nucleotide sequence derived from corn clone cho1c.pk003.f1 (SEQ ID NO:4),  
     [0020] (c) nucleotide sequence derived from corn clone cpi1c.pk011.g12 (SEQ ID NO:6),  
     [0021] (d) nucleotide sequence derived from rice clone rdi2c.pk010.p22 (SEQ ID NO:8),  
     [0022] (e) nucleotide sequence derived from brassica clone ebp1f.pk002.g18 (SEQ ID NO:1 7), (f) nucleotide sequence from  Vigna aconitifolia  (NCBI General Identification (GI) No. 1709930; SEQ ID NO:15), (g) nucleotide sequence from  Arabidopsis thaliana  (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) and  
     [0023] (h) nucleotide sequence from  Nicotiana tabacum  (NCBI General Identification (GI) No. 13173434; SEQ ID NO:14). Dashes are used by the program to maximize alignment of the sequences.  
     [0024]FIG. 3 depicts the amino acid alignment of the catalytic region of  Vigna aconitifolia  (NCBI General Identification (GI) No. 1709930; SEQ ID NO:15) (nucleotides 387 to 557) with the amino acid sequences encoded by the following:  
     [0025] (a) nucleotide sequence derived from corn clone cpd1c.pk002.l 23 (SEQ ID NO:2),  
     [0026] (b) nucleotide sequence derived from corn clone cho1c.pk003.f1 (SEQ ID NO:4),  
     [0027] (c) nucleotide sequence derived from corn clone cpi1c.pk011.g12 (SEQ ID NO:6),  
     [0028] (d) nucleotide sequence derived from rice clone rdi2c.pk010.p22 (SEQ ID NO:8),  
     [0029] (e) nucleotide sequence derived from brassica clone ebp1f.pk002.g18 (SEQ ID NO:17), (f) nucleotide sequence from  Arabidopsis thaliana  (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) and (g) nucleotide sequence from  Nicotiana tabacum  (NCBI General Identification (GI) No.13173434; SEQ ID NO:14). When all the amino acids match the residue of the Consensus the residue of the Consensus will show, otherwise a “.” will show. 
    
    
     [0030] Table 1 lists the polypeptides that are described herein, the designation of the cDNA clones that comprise the nucleic acid fragments encoding polypeptides representing all or a substantial portion of these polypeptides, and the corresponding identifier (SEQ ID NO:) as used in the attached Sequence Listing. Table 1 also identifies the cDNA clones as individual ESTs (“EST”), the sequences of the entire cDNA inserts comprising the indicated cDNA clones (“FIS”), contigs assembled from two or more ESTs (“Contig”), contigs assembled from an FIS and one or more ESTs (“Contig*”), or sequences encoding the entire or functional protein derived from an FIS, a contig, or an FIS and PCR fragment sequence (“CGS”). The sequence descriptions and Sequence Listing attached hereto comply with the rules governing nucleotide and/or amino acid sequence disclosures in patent applications as set forth in 37 C.F.R. §1.821-1.825.  
               TABLE 1                          Phosphoribosylaminoimidazole Carboxylase                         SEQ ID NO:                                 Plant   Clone Designation   Status   (Nucleotide)   (Amino Acid)                                         Corn   cpd1c.pk002.123:fis   CGS   1   2       Corn   cho1c.pk003.f1:fis   CGS   3   4       Corn   cpi1c.pk011.g12:fis   FIS   5   6       Rice   rdi2c.pk010.p22:fis   CGS   7   8       Soybean   sfl1.pk0118.d10:fis   FIS   9   10       Wheat   Contig of   FIS   11   12           wre1n.pk0004.g6:fis       Brassica   ebp1f.pk002.g18:fis   CGS   16   17                  
 
     [0031] SEQ ID NO:13 is the amino acid sequence of  Arabidopsis thaliana  (NCBI General Identification (GI) No. 7436526).  
     [0032] SEQ ID NO:14 is the amino acid sequence of  Nicotiana tabacum  (NCBI General Identification (GI) No. 13173434).  
     [0033] SEQ ID NO:15 is the amino acid sequence of  Vigna aconitifolia  (NCBI General Identification (GI) No. 1709930).  
     [0034] The Sequence Listing contains the one letter code for nucleotide sequence characters and the three letter codes for amino acids as defined in conformity with the IUPAC-IUBMB standards described in  Nucleic Acids Res.  13:3021-3030 (1985) and in the  Biochemical J.  219 (No. 2):345-373 (1984) which are herein incorporated by reference. The symbols and format used for nucleotide and amino acid sequence data comply with the rules set forth in 37 C.F.R. §1.822.  
     DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS  
     [0035] The disclosure of each reference set forth herein is incorporated herein by reference in its entirety.  
     [0036] In the context of this disclosure, a number of terms shall be utilized. The terms “polynucleotide”, “polynucleotide sequence”, “nucleic acid sequence”, and “nucleic acid fragment”/“isolated nucleic acid fragment” are used interchangeably herein. These terms encompass nucleotide sequences and the like. A polynucleotide may be a polymer of RNA or DNA that is single- or double-stranded, that optionally contains synthetic, non-natural or altered nucleotide bases. A polynucleotide in the form of a polymer of DNA may be comprised of one or more segments of cDNA, genomic DNA, synthetic DNA, or mixtures thereof. An isolated polynucleotide of the present invention may include at least 30 contiguous nucleotides, preferably at least 40 contiguous nucleotides, most preferably at least 60 contiguous nucleotides derived from SEQ ID NOs:1, 3, 5, 7, 9, 11 or 16, or the complement of such sequences.  
     [0037] The term “isolated” refers to materials, such as nucleic acid molecules and/or proteins, which are substantially free or otherwise removed from components that normally accompany or interact with the materials in a naturally occurring environment. Isolated polynucleotides may be purified from a host cell in which they naturally occur. Conventional nucleic acid purification methods known to skilled artisans may be used to obtain isolated polynucleotides. The term also embraces recombinant polynucleotides and chemically synthesized polynucleotides.  
     [0038] The term “recombinant” means, for example, that a nucleic acid sequence is made by an artificial combination of two otherwise separated segments of sequence, e.g., by chemical synthesis or by the manipulation of isolated nucleic acids by genetic engineering techniques. A “recombinant DNA construct” comprises any of the isolated polynucleotides of the present invention operably linked to at least one regulatory sequence.  
     [0039] As used herein, “contig” refers to a nucleotide sequence that is assembled from two or more constituent nucleotide sequences that share common or overlapping regions of sequence homology. For example, the nucleotide sequences of two or more nucleic acid fragments can be compared and aligned in order to identify common or overlapping sequences. Where common or overlapping sequences exist between two or more nucleic acid fragments, the sequences (and thus their corresponding nucleic acid fragments) can be assembled into a single contiguous nucleotide sequence.  
     [0040] As used herein, “substantially similar” refers to nucleic acid fragments wherein changes in one or more nucleotide bases results in substitution of one or more amino acids, but do not affect the functional properties of the polypeptide encoded by the nucleotide sequence. “Substantially similar” also refers to nucleic acid fragments wherein changes in one or more nucleotide bases does not affect the ability of the nucleic acid fragment to mediate alteration of gene expression by gene silencing through for example antisense or co-suppression technology. “Substantially similar” also refers to modifications of the nucleic acid fragments of the instant invention such as deletion or insertion of one or more nucleotides that do not substantially affect the functional properties of the resulting transcript vis-à-vis the ability to mediate gene silencing or alteration of the functional properties of the resulting protein molecule. It is therefore understood that the invention encompasses more than the specific exemplary nucleotide or amino acid sequences and includes functional equivalents thereof. The terms “substantially similar” and “corresponding substantially” are used interchangeably herein.  
     [0041] The “Clustal V method of alignment” corresponds to the alignment method labeled Clustal V (described by Higgins and Sharp (1989)  CABIOS.  5:151-153) and found in the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). The “default parameters” are the parameters pre-set by the manufacturer of the program and for multiple alignments they correspond to GAP PENALTY=10 and GAP LENGTH PENALTY=10, while for pairwise alignments they are KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. After alignment of the sequences, using the Clustal V program, it is possible to obtain a “percent identity” by viewing the “sequence distances” table on the same program.  
     [0042] Substantially similar nucleic acid fragments may be selected by screening nucleic acid fragments representing subfragments or modifications of the nucleic acid fragments of the instant invention, wherein one or more nucleotides are substituted, deleted and/or inserted, for their ability to affect the level of the polypeptide encoded by the unmodified nucleic acid fragment in a plant or plant cell. For example, a substantially similar nucleic acid fragment representing at least 30 contiguous nucleotides derived from the instant nucleic acid fragment can be constructed and introduced into a plant or plant cell. The level of the polypeptide encoded by the unmodified nucleic acid fragment present in a plant or plant cell exposed to the substantially similar nucleic fragment can then be compared to the level of the polypeptide in a plant or plant cell that is not exposed to the substantially similar nucleic acid fragment.  
     [0043] For example, it is well known in the art that antisense suppression and co-suppression of gene expression may be accomplished using nucleic acid fragments representing less than the entire coding region of a gene, and by using nucleic acid fragments that do not share 100% sequence identity with the gene to be suppressed. Moreover, alterations in a nucleic acid fragment which result in the production of a chemically equivalent amino acid at a given site, but do not effect the functional properties of the encoded polypeptide, are well known in the art. Thus, a codon for the amino acid alanine, a hydrophobic amino acid, may be substituted by a codon encoding another less hydrophobic residue, such as glycine, or a more hydrophobic residue, such as valine, leucine, or isoleucine. Similarly, changes which result in substitution of one negatively charged residue for another, such as aspartic acid for glutamic acid, or one positively charged residue for another, such as lysine for arginine, can also be expected to produce a functionally equivalent product. Nucleotide changes which result in alteration of the N-terminal and C-terminal portions of the polypeptide molecule would also not be expected to alter the activity of the polypeptide. Each of the proposed modifications is well within the routine skill in the art, as is determination of retention of biological activity, of the encoded products. Consequently, an isolated polynucleotide comprising a nucleotide sequence of at least 30 (preferably at least 40, most preferably at least 60) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11 or 16, and the complement of such nucleotide sequences may be used in methods of selecting an isolated polynucleotide that affects the expression of a phosphoribosylaminoimidazole carboxylase polypeptide in a host cell. A method of selecting an isolated polynucleotide that affects the level of expression of a polypeptide in a virus or in a host cell (eukaryotic, such as plant or yeast, prokaryotic such as bacterial) may comprise the steps of: constructing an isolated polynucleotide of the present invention or an isolated recombinant DNA construct or chimeric gene of the present invention; introducing the isolated polynucleotide or the isolated recombinant DNA construct or chimeric gene into a host cell; measuring the level of a polypeptide or enzyme activity in the host cell containing the isolated polynucleotide; and comparing the level of a polypeptide or enzyme activity in the host cell containing the isolated polynucleotide with the level of a polypeptide or enzyme activity in a host cell that does not contain the isolated polynucleotide.  
     [0044] Moreover, substantially similar nucleic acid fragments may also be characterized by their ability to hybridize. Estimates of such homology are provided by either DNA-DNA or DNA-RNA hybridization under conditions of stringency as is well understood by those skilled in the art (Hames and Higgins, Eds. (1985) Nucleic Acid Hybridisation, IRL Press, Oxford, U.K.). Stringency conditions can be adjusted to screen for moderately similar fragments, such as homologous sequences from distantly related organisms, to highly similar fragments, such as genes that duplicate functional enzymes from closely related organisms. Post-hybridization washes determine stringency conditions. One set of preferred conditions uses a series of washes starting with 6×SSC, 0.5% SDS at room temperature for 15 min, then repeated with 2×SSC, 0.5% SDS at 45° C. for 30 min, and then repeated twice with 0.2×SSC, 0.5% SDS at 50° C. for 30 min. A more preferred set of stringent conditions uses higher temperatures in which the washes are identical to those above except for the temperature of the final two 30 min washes in 0.2×SSC, 0.5% SDS was increased to 60° C. Another preferred set of highly stringent conditions uses two final washes in 0.1×SSC, 0.1% SDS at 65° C.  
     [0045] Substantially similar nucleic acid fragments of the instant invention may also be characterized by the percent identity of the amino acid sequences that they encode to the amino acid sequences disclosed herein, as determined by algorithms commonly employed by those skilled in this art. Suitable nucleic acid fragments (isolated polynucleotides of the present invention) encode polypeptides that are at least about 70% identical, preferably at least about 80% identical to the amino acid sequences reported herein. Preferred nucleic acid fragments encode amino acid sequences that are at least about 85% identical to the amino acid sequences reported herein. More preferred nucleic acid fragments encode amino acid sequences that are at least about 90% identical to the amino acid sequences reported herein. Most preferred are nucleic acid fragments that encode amino acid sequences that are at least about 95% identical to the amino acid sequences reported herein. Suitable nucleic acid fragments not only have the above identities but typically encode a polypeptide having at least 50 amino acids, preferably at least 100 amino acids, more preferably at least 150 amino acids, still more preferably at least 200 amino acids, and most preferably at least 250 amino acids.  
     [0046] It is well understood by one skilled in the art that many levels of sequence identity are useful in identifying related polypeptide sequences. Useful examples of percent identities are 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95%, or any integer percentage from 55% to 100%. Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal V method of alignment (Higgins and Sharp (1989)  CABIOS.  5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5.  
     [0047] A “substantial portion” of an amino acid or nucleotide sequence comprises an amino acid or a nucleotide sequence that is sufficient to afford putative identification of the protein or gene that the amino acid or nucleotide sequence comprises. Amino acid and nucleotide sequences can be evaluated either manually by one skilled in the art, or by using computer-based sequence comparison and identification tools that employ algorithms such as BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993)  J. Mol. Biol.  215:403-410). In general, a sequence of ten or more contiguous amino acids or thirty or more contiguous nucleotides is necessary in order to putatively identify a polypeptide or nucleic acid sequence as homologous to a known protein or gene. Moreover, with respect to nucleotide sequences, gene-specific oligonucleotide probes comprising 30 or more contiguous nucleotides may be used in sequence-dependent methods of gene identification (e.g., Southern hybridization) and isolation (e.g., in situ hybridization of bacterial colonies or bacteriophage plaques). In addition, short oligonucleotides of 12 or more nucleotides may be used as amplification primers in PCR in order to obtain a particular nucleic acid fragment comprising the primers. Accordingly, a “substantial portion” of a nucleotide sequence comprises a nucleotide sequence that will afford specific identification and/or isolation of a nucleic acid fragment comprising the sequence. The instant specification teaches amino acid and nucleotide sequences encoding polypeptides that comprise one or more particular plant proteins. The skilled artisan, having the benefit of the sequences as reported herein, may now use all or a substantial portion of the disclosed sequences for purposes known to those skilled in this art. Accordingly, the instant invention comprises the complete sequences as reported in the accompanying Sequence Listing, as well as substantial portions of those sequences as defined above.  
     [0048] “Codon degeneracy” refers to divergence in the genetic code permitting variation of the nucleotide sequence without effecting the amino acid sequence of an encoded polypeptide. Accordingly, the instant invention relates to any nucleic acid fragment comprising a nucleotide sequence that encodes all or a substantial portion of the amino acid sequences set forth herein. The skilled artisan is well aware of the “codon-bias” exhibited by a specific host cell in usage of nucleotide codons to specify a given amino acid. Therefore, when synthesizing a nucleic acid fragment for improved expression in a host cell, it is desirable to design the nucleic acid fragment such that its frequency of codon usage approaches the frequency of preferred codon usage of the host cell.  
     [0049] “Synthetic nucleic acid fragments” can be assembled from oligonucleotide building blocks that are chemically synthesized using procedures known to those skilled in the art. These building blocks are ligated and annealed to form larger nucleic acid fragments which may then be enzymatically assembled to construct the entire desired nucleic acid fragment. “Chemically synthesized”, as related to a nucleic acid fragment, means that the component nucleotides were assembled in vitro. Manual chemical synthesis of nucleic acid fragments may be accomplished using well established procedures, or automated chemical synthesis can be performed using one of a number of commercially available machines. Accordingly, the nucleic acid fragments can be tailored for optimal gene expression based on optimization of the nucleotide sequence to reflect the codon bias of the host cell. The skilled artisan appreciates the likelihood of successful gene expression if codon usage is biased towards those codons favored by the host. Determination of preferred codons can be based on a survey of genes derived from the host cell where sequence information is available.  
     [0050] “Gene” refers to a nucleic acid fragment that expresses a specific protein, including regulatory sequences preceding (5′ non-coding sequences) and following (3′ non-coding sequences) the coding sequence. “Native gene” refers to a gene as found in nature with its own regulatory sequences. “Chimeric gene” refers to any gene that is not a native gene, comprising regulatory and coding sequences that are not found together in nature. Accordingly, a chimeric gene may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source, but arranged in a manner different than that found in nature. “Endogenous gene” refers to a native gene in its natural location in the genome of an organism. A “foreign-gene” refers to a gene not normally found in the host organism, but that is introduced into the host organism by gene transfer. Foreign genes can comprise native genes inserted into a non-native organism, or chimeric genes. A “transgene” is a gene that has been introduced into the genome by a transformation procedure.  
     [0051] “Coding sequence” refers to a nucleotide sequence that codes for a specific amino acid sequence. “Regulatory sequences” refer to nucleotide sequences located upstream (5′ non-coding sequences), within, or downstream (3′ non-coding sequences) of a coding sequence, and which influence the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences may include promoters, translation leader sequences, introns, and polyadenylation recognition sequences.  
     [0052] “Promoter” refers to a nucleotide sequence capable of controlling the expression of a coding sequence or functional RNA. In general, a coding sequence is located 3′ to a promoter sequence. The promoter sequence consists of proximal and more distal upstream elements, the latter elements often referred to as enhancers. Accordingly, an “enhancer” is a nucleotide sequence which can stimulate promoter activity and may be an innate element of the promoter or a heterologous element inserted to enhance the level or tissue-specificity of a promoter. Promoters may be derived in their entirety from a native gene, or may be composed of different elements derived from different promoters found in nature, or may even comprise synthetic nucleotide segments. It is understood by those skilled in the art that different promoters may direct the expression of a gene in different tissues or cell types, or at different stages of development, or in response to different environmental conditions. Promoters which cause a nucleic acid fragment to be expressed in most cell types at most times are commonly referred to as “constitutive promoters”. New promoters of various types useful in plant cells are constantly being discovered; numerous examples may be found in the compilation by Okamuro and Goldberg (1989)  Biochemistry of Plants  15:1-82. It is further recognized that since in most cases the exact boundaries of regulatory sequences have not been completely defined, nucleic acid fragments of different lengths may have identical promoter activity.  
     [0053] “Translation leader sequence” refers to a nucleotide sequence located between the promoter sequence of a gene and the coding sequence. The translation leader sequence is present in the fully processed mRNA upstream of the translation start sequence. The translation leader sequence may affect processing of the primary transcript to mRNA, mRNA stability or translation efficiency. Examples of translation leader sequences have been described (Turner and Foster (1995)  Mol. Biotechnol.  3:225-236).  
     [0054] “3′ non-coding sequences” refer to nucleotide sequences located downstream of a coding sequence and include polyadenylation recognition sequences and other sequences encoding regulatory signals capable of affecting mRNA processing or gene expression. The polyadenylation signal is usually characterized by affecting the addition of polyadenylic acid tracts to the 3′ end of the mRNA precursor. The use of different 3′ non-coding sequences is exemplified by Ingelbrecht et al. (1989)  Plant Cell  1:671-680.  
     [0055] “RNA transcript” refers to the product resulting from RNA polymerase-catalyzed transcription of a DNA sequence. When the RNA transcript is a perfect complementary copy of the DNA sequence, it is referred to as the primary transcript or it may be a RNA sequence derived from posttranscriptional processing of the primary transcript and is referred to as the mature RNA. “Messenger RNA (mRNA)” refers to the RNA that is without introns and that can be translated into polypeptides by the cell. “cDNA” refers to DNA that is complementary to and derived from an mRNA template. The cDNA can be single-stranded or converted to double stranded form using, for example, the Klenow fragment of DNA polymerase I. “Sense-RNA” refers to an RNA transcript that includes the mRNA and so can be translated into a polypeptide by the cell. “Antisense RNA” refers to an RNA transcript that is complementary to all or part of a target primary transcript or mRNA and that blocks the expression of a target gene (see U.S. Pat. No. 5,107,065, incorporated herein by reference). The complementarity of an antisense RNA may be with any part of the specific nucleotide sequence, i.e., at the 5′ non-coding sequence, 3′ non-coding sequence, introns, or the coding sequence. “Functional RNA” 0  refers to sense RNA, antisense RNA, ribozyme RNA, or other RNA that may not be translated but yet has an effect on cellular processes.  
     [0056] The term “operably linked” refers to the association of two or more nucleic acid fragments on a single polynucleotide so that the function of one is affected by the other. For example, a promoter is operably linked with a coding sequence when it is capable of affecting the expression of that coding sequence (i.e., that the coding sequence is under the transcriptional control of the promoter). Coding sequences can be operably linked to regulatory sequences in sense or antisense orientation.  
     [0057] The term “expression”, as used herein, refers to the transcription and stable accumulation of sense (mRNA) or antisense RNA derived from the nucleic acid fragment of the invention. Expression may also refer to translation of mRNA into a polypeptide. “Antisense inhibition” refers to the production of antisense RNA transcripts capable of suppressing the expression of the target protein. “Overexpression” refers to the production of a gene product in transgenic organisms that exceeds levels of production in normal or non-transformed organisms. “Co-suppression” refers to the production of sense RNA transcripts capable of suppressing the expression of identical or substantially similar foreign or endogenous genes (U.S. Pat. No. 5,231,020, incorporated herein by reference).  
     [0058] A “protein” or “polypeptide” is a chain of amino acids arranged in a specific order determined by the coding sequence in a polynucleotide encoding the polypeptide. Each protein or polypeptide has a unique function.  
     [0059] “Altered levels” or “altered expression” refers to the production of gene product(s) in transgenic organisms in amounts or proportions that differ from that of normal or non-transformed organisms.  
     [0060] “Null mutant” refers here to a host cell which either lacks the expression of a certain polypeptide or expresses a polypeptide which is inactive or does not have any detectable expected enzymatic function.  
     [0061] “Mature protein” or the term “mature” when used in describing a protein refers to a post-translationally processed polypeptide; i.e., one from which any pre- or propeptides present in the primary translation product have been removed. “Precursor protein” or the term “precursor” when used in describing a protein refers to the primary product of translation of mRNA; i.e., with pre- and propeptides still present. Pre- and propeptides may be but are not limited to intracellular localization signals.  
     [0062] A “chloroplast transit peptide” is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the chloroplast or other plastid types present in the cell in which the protein is made. “Chloroplast transit sequence” refers to a nucleotide sequence that encodes a chloroplast transit peptide. A “signal peptide” is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the secretory system (Chrispeels (1991)  Ann. Rev. Plant Phys. Plant Mol. Biol.  42:21-53). If the protein is to be directed to a vacuole, a vacuolar targeting signal (supra) can further be added, or if to the endoplasmic reticulum, an endoplasmic reticulum retention signal (supra) may be added. If the protein is to be directed to the nucleus, any signal peptide present should be removed and instead a nuclear localization signal included (Raikhel (1992)  Plant Phys.  100:1627-1632).  
     [0063] “Transformation” refers to the transfer of a nucleic acid fragment into the genome of a host organism, resulting in genetically stable inheritance. Host organisms containing the transformed nucleic acid fragments are referred to as “transgenic” organisms. Examples of methods of plant transformation include Agrobacterium-mediated transformation (De Blaere et al. (1987)  Meth. Enzymol.  143:277) and particle-accelerated or “gene gun” transformation technology (Klein et al. (1987)  Nature  ( London ) 327:70-73; U.S. Pat. No. 4,945,050, incorporated herein by reference). Thus, isolated polynucleotides of the present invention can be incorporated into recombinant constructs, typically DNA constructs, capable of introduction into and replication in a host cell. Such a construct can be a vector that includes a replication system and sequences that are capable of transcription and translation of a polypeptide-encoding sequence in a given host cell. A number of vectors suitable for stable transfection of plant cells or for the establishment of transgenic plants have been described in, e.g., Pouwels et al., Cloning Vectors: A Laboratory Manual, 1985, supp. 1987; Weissbach and Weissbach, Methods for Plant Molecular Biology, Academic Press, 1989; and Flevin et al., Plant Molecular Biology Manual, Kluwer Academic Publishers, 1990. Typically, plant expression vectors include, for example, one or more cloned plant genes under the transcriptional control of 5′ and 3′ regulatory sequences and a dominant selectable marker. Such plant expression vectors also can contain a promoter regulatory region (e.g., a regulatory region controlling inducible or constitutive, environmentally- or developmentally-regulated, or cell- or tissue-specific expression), a transcription initiation start site, a ribosome binding site, an RNA processing signal, a transcription termination site, and/or a polyadenylation signal.  
     [0064] “Stable transformation” refers to the transfer of a nucleic acid fragment into a genome of a host organism, including both nuclear and organellar genomes, resulting in genetically stable inheritance. In contrast, “transient transformation” refers to the transfer of a nucleic acid fragment into the nucleus, or DNA-containing organelle, of a host organism resulting in gene expression without integration or stable inheritance. The term “transformation” as used herein refers to both stable transformation and transient transformation.  
     [0065] Standard recombinant DNA and molecular cloning techniques used herein are well known in the art and are described more fully in Sambrook et al.  Molecular Cloning: A Laboratory Manual ; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, 1989 (hereinafter “Maniatis”).  
     [0066] “Motifs” or “subsequences” refer to short regions of conserved sequences of nucleic acids or amino acids that comprise part of a longer sequence. For example, it is expected that such conserved subsequences would be important for function, and could be used to identify new homologues in plants. It is expected that some or all of the elements may be found in a homologue. Also, it is expected that one or two of the conserved amino acids in any given motif may differ in a true homologue.  
     [0067] “PCR” or “polymerase chain reaction” is well known by those skilled in the art as a technique used for the amplification of specific DNA segments (U.S. Pat. Nos. 4,683,195 and 4,800,159).  
     [0068] The present invention includes an isolated polynucleotide comprising: (a) a nucleotide sequence encoding a polypeptide comprising at least 100 amino acids, wherein the amino acid sequence of the polypeptide and the amino acid sequence of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17 have at least 70%, 75%, 80%, 85%, 90% or 95% identity based on the Clustal V method of alignment, or (b) the complement of the nucleotide sequence, wherein the complement and the nucleotide sequence contain the same number of nucleotides and are 100% complementary. The polypeptide preferably comprises the amino acid sequence of SEQ ID NO:2, 4, 6, 8, 10, 12 or 17. The nucleotide sequence preferably comprises the nucleotide sequence of SEQ ID NO:1, 3, 5, 7, 9, 11 or 16. The polypeptide preferably is a phosphoribosylaminoimidazole carboxylase.  
     [0069] This invention also includes the isolated complement of such polynucleotides, wherein the complement and the polynucleotide consist of the same number of nucleotides, and the nucleotide sequences of the complement and the polynucleotide have 100% complementarity.  
     [0070] Nucleic acid fragments encoding at least a portion of several phosphoribosylaminoimidazole carboxylases have been isolated and identified by comparison of random plant cDNA sequences to public databases containing nucleotide and protein sequences using the BLAST algorithms well known to those skilled in the art. The nucleic acid fragments of the instant invention may be used to isolate cDNAs and genes encoding homologous proteins from the same or other plant species. Isolation of homologous genes using sequence-dependent protocols is well known in the art. Examples of sequence-dependent protocols include, but are not limited to, methods of nucleic acid hybridization, and methods of DNA and RNA amplification as exemplified by various uses of nucleic acid amplification technologies (e.g., polymerase chain reaction, ligase chain reaction).  
     [0071] For example, genes encoding other phosphoribosylaminoimidazole carboxylases, either as cDNAs or genomic DNAs, could be isolated directly by using all or a portion of the instant nucleic acid fragments as DNA hybridization probes to screen libraries from any desired plant employing methodology well known to those skilled in the art. Specific oligonucleotide probes based upon the instant nucleic acid sequences can be designed and synthesized by methods known in the art (Maniatis). Moreover, an entire sequence can be used directly to synthesize DNA probes by methods known to the skilled artisan such as random primer DNA labeling, nick translation, end-labeling techniques, or RNA probes using available in vitro transcription systems. In addition, specific primers can be designed and used to amplify a part or all of the instant sequences. The resulting amplification products can be labeled directly during amplification reactions or labeled after amplification reactions, and used as probes to isolate full length cDNA or genomic fragments under conditions of appropriate stringency.  
     [0072] In addition, two short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols to amplify longer nucleic acid fragments encoding homologous genes from DNA or RNA. The polymerase chain reaction may also be performed on a library of cloned nucleic acid fragments wherein the sequence of one primer is derived from the instant nucleic acid fragments, and the sequence of the other primer takes advantage of the presence of the polyadenylic acid tracts to the 3′ end of the mRNA precursor encoding plant genes. Alternatively, the second primer sequence may be based upon sequences derived from the cloning vector. For example, the skilled artisan can follow the RACE protocol (Frohman et al. (1988)  Proc. Natl. Acad. Sci. USA  85:8998-9002) to generate cDNAs by using PCR to amplify copies of the region between a single point in the transcript and the 3′ or 5′ end. Primers oriented in the 3′ and 5′ directions can be designed from the instant sequences. Using commercially available 3′ RACE or 5′ RACE systems (BRL), specific 3′ or 5′ cDNA fragments can be isolated (Ohara et al. (1989)  Proc. Natl. Acad. Sci. USA  86:5673-5677; Loh et al. (1989)  Science  243:217-220). Products generated by the 3′ and 5′ RACE procedures can be combined to generate full-length cDNAs (Frohman and Martin (1989)  Techniques  1:165). Consequently, a polynucleotide comprising a nucleotide sequence of at least 30 (preferably at least 40, most preferably at least 60) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11 and 16, and the complement of such nucleotide sequences may be used in such methods to obtain a nucleic acid fragment encoding a substantial portion of an amino acid sequence of a polypeptide.  
     [0073] Availability of the instant nucleotide and deduced amino acid sequences facilitates immunological screening of cDNA expression libraries. Synthetic peptides representing portions of the instant amino acid sequences may be synthesized. These peptides can be used to immunize animals to produce polyclonal or monoclonal antibodies with specificity for peptides or proteins comprising the amino acid sequences. These antibodies can be then be used to screen cDNA expression libraries to isolate full-length cDNA clones of interest (Lerner (1984)  Adv. Immunol.  36:1-34; Maniatis).  
     [0074] In another embodiment, this invention includes viruses and host cells comprising either the recombinant DNA constructs or chimeric genes of the invention as described herein or an isolated polynucleotide of the invention as described herein. Examples of host cells which can be used to practice the invention include, but are not limited to, yeast, bacteria, and plants.  
     [0075] The nucleic acid fragments of the instant invention may be used to create transgenic plants in which the disclosed polypeptides are present at higher or lower levels than normal or in cell types or developmental stages in which they are not normally found. This would have the effect of altering the level of 5-amino-1-ribosyl-4-imidazolecarboxylic acid 5′-phosphate (carboxy-AIR) in those cells. Therefore, these genes may be used in plant cells to alter the activity of de novo nucleic acid biosynthetic pathways and salvage pathways which may alter efficient growth and development of plant cells. More specifically, the genes of the instant invention may useful as a herbicide target to inhibit the formation of carboxy-AIR and other compounds further along in the pathway that may be essential for growth and development.  
     [0076] Overexpression of the proteins of the instant invention may be accomplished by first constructing a recombinant DNA construct or chimeric gene in which the coding region is operably linked to a promoter capable of directing expression of a gene in the desired tissues at the desired stage of development. The recombinant DNA construct or chimeric gene may comprise promoter sequences and translation leader sequences derived from the same genes. 3′ Non-coding sequences encoding transcription termination signals may also be provided. The instant recombinant DNA construct or chimeric gene may also comprise one or more introns in order to facilitate gene expression.  
     [0077] Plasmid vectors comprising the instant isolated polynucleotide (or recombinant DNA construct or chimeric gene) may be constructed. The choice of plasmid vector is dependent upon the method that will be used to transform host plants. The skilled artisan is well aware of the genetic elements that must be present on the plasmid vector in order to successfully transform, select and propagate host cells containing the recombinant DNA construct or chimeric gene. The skilled artisan will also recognize that different independent transformation events will result in different levels and patterns of expression (Jones et al. (1985)  EMBO J.  4:2411-2418; De Almeida et al. (1989)  Mol. Gen. Genetics  218:78-86), and thus that multiple events must be screened in order to obtain lines displaying the desired expression level and pattern. Such screening may be accomplished by Southern analysis of DNA, Northern analysis of mRNA expression, Western analysis of protein expression, or phenotypic analysis.  
     [0078] For some applications it may be useful to direct the instant polypeptides to different cellular compartments, or to facilitate its secretion from the cell. It is thus envisioned that the recombinant DNA construct or chimeric gene described above may be further supplemented by directing the coding sequence to encode the instant polypeptides with appropriate intracellular targeting sequences such as transit sequences (Keegstra (1989)  Cell  56:247-253), signal sequences or sequences encoding endoplasmic reticulum localization (Chrispeels (1991)  Ann. Rev. Plant Phys. Plant Mol. Biol.  42:21-53), or nuclear localization signals (Raikhel (1992)  Plant Phys. 100:1627-1632) with or without removing targeting sequences that are already present. While the references cited give examples of each of these, the list is not exhaustive and more targeting signals of use may be discovered in the future.  
     [0079] It may also be desirable to reduce or eliminate expression of genes encoding the instant polypeptides in plants for some applications. In order to accomplish this, a recombinant DNA construct or chimeric gene designed for co-suppression of the instant polypeptide can be constructed by linking a gene or gene fragment encoding that polypeptide to plant promoter sequences. Alternatively, a recombinant DNA construct or chimeric gene designed to express antisense RNA for all or part of the instant nucleic acid fragment can be constructed by linking the gene or gene fragment in reverse orientation to plant promoter sequences. Either the co-suppression or antisense recombinant DNA constructs or chimeric genes could be introduced into plants via transformation wherein expression of the corresponding endogenous genes are reduced or eliminated.  
     [0080] The polypeptides of the instant invention were shown to be constitutively expressed using microbead arrays (data not shown; Brenner et al.,  Nat. Biotechnol.  18: 630-634 (2000); Brenner et al.,  Proc. Natl. Acad. Sci. USA  97(4):1665-1670 (2000)).  
     [0081] Molecular genetic solutions to the generation of plants with altered gene expression have a decided advantage over more traditional plant breeding approaches. Changes in plant phenotypes can be produced by specifically inhibiting expression of one or more genes by antisense inhibition or cosuppression (U.S. Pat. Nos. 5,190,931, 5,107,065 and 5,283,323). An antisense or cosuppression construct would act as a dominant negative regulator of gene activity. While conventional mutations can yield negative regulation of gene activity these effects are most likely recessive. The dominant negative regulation available with a transgenic approach may be advantageous from a breeding perspective. In addition, the ability to restrict the expression of a specific phenotype to the reproductive tissues of the plant by the use of tissue specific promoters may confer agronomic advantages relative to conventional mutations which may have an effect in all tissues in which a mutant gene is ordinarily expressed.  
     [0082] The person skilled in the art will know that special considerations are associated with the use of antisense or cosuppression technologies in order to reduce expression of particular genes. For example, the proper level of expression of sense or antisense genes may require the use of different recombinant DNA constructs or chimeric genes utilizing different regulatory elements known to the skilled artisan. Once transgenic plants are obtained by one of the methods described above, it will be necessary to screen individual transgenics for those that most effectively display the desired phenotype. Accordingly, the skilled artisan will develop methods for screening large numbers of transformants. The nature of these screens will generally be chosen on practical grounds. For example, one can screen by looking for changes in gene expression by using antibodies specific for the protein encoded by the gene being suppressed, or one could establish assays that specifically measure enzyme activity. A preferred method will be one which allows large numbers of samples to be processed rapidly, since it will be expected that a large number of transformants will be negative for the desired phenotype.  
     [0083] In another embodiment, the present invention includes a phosphoribosylaminoimidazole carboxylase polypeptide having an amino acid sequence that is at least 70% identical, based on the Clustal V method of alignment, to a polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 8, 10, 12 and 17.  
     [0084] The instant polypeptides (or portions thereof) may be produced in heterologous host cells, particularly in the cells of microbial hosts, and can be used to prepare antibodies to these proteins by methods well known to those skilled in the art. The antibodies are useful for detecting the polypeptides of the instant invention in situ in cells or in vitro in cell extracts. Preferred heterologous host cells for production of the instant polypeptides are microbial hosts. Microbial expression systems and expression vectors containing regulatory sequences that direct high level expression of foreign proteins are well known to those skilled in the art. Any of these could be used to construct a recombinant DNA construct or chimeric gene for production of the instant polypeptides. This recombinant DNA construct or chimeric gene could then be introduced into appropriate microorganisms via transformation to provide high level expression of the encoded phosphoribosylaminoimidazole carboxylase. An example of a vector for high level expression of the instant polypeptides in a bacterial host is provided (Example 6).  
     [0085] Additionally, the instant polypeptides can be used as a target to facilitate design and/or identification of inhibitors of those enzymes that may be useful as herbicides/pesticides. This is desirable because the polypeptides described herein catalyze a step in purine biosynthesis, more specifically the synthesis of 5-amino-1-ribosyl-4-imidazolecarboxylic acid 5′-phosphate (carboxy-AIR). Accordingly, inhibition of the activity of one or more of the enzymes described herein could lead to inhibition of plant growth. Thus, the instant polypeptides could be appropriate for new herbicide/pesticide discovery and design.  
     [0086] All or a substantial portion of the polynucleotides of the instant invention may also be used as probes for genetically and physically mapping the genes that they are a part of, and used as markers for traits linked to those genes. Such information may be useful in plant breeding in order to develop lines with desired phenotypes. For example, the instant nucleic acid fragments may be used as restriction fragment length polymorphism (RFLP) markers. Southern blots (Maniatis) of restriction-digested plant genomic DNA may be probed with the nucleic acid fragments of the instant invention. The resulting banding patterns may then be subjected to genetic analyses using computer programs such as MapMaker (Lander et al. (1987)  Genomics  1:174-181) in order to construct a genetic map. In addition, the nucleic acid fragments of the instant invention may be used to probe Southern blots containing restriction endonuclease-treated genomic DNAs of a set of individuals representing parent and progeny of a defined genetic cross. Segregation of the DNA polymorphisms is noted and used to calculate the position of the instant nucleic acid sequence in the genetic map previously obtained using this population (Botstein et al. (1980)  Am. J. Hum. Genet.  32:314-331).  
     [0087] The production and use of plant gene-derived probes for use in genetic mapping is described in Bernatzky and Tanksley (1986)  Plant Mol. Biol. Reporter  4:37-41. Numerous publications describe genetic mapping of specific cDNA clones using the methodology outlined above or variations thereof. For example, F2 intercross populations, backcross populations, randomly mated populations, near isogenic lines, and other sets of individuals may be used for mapping. Such methodologies are well known to those skilled in the art.  
     [0088] Nucleic acid probes derived from the instant nucleic acid sequences may also be used for physical mapping (i.e., placement of sequences on physical maps; see Hoheisel et al. In:  Nonmammalian Genomic Analysis: A Practical Guide , Academic press 1996, pp. 319-346, and references cited therein).  
     [0089] In another embodiment, nucleic acid probes derived from the instant nucleic acid sequences may be used in direct fluorescence in situ hybridization (FISH) mapping (Trask (1991)  Trends Genet.  7:149-154). Although current methods of FISH mapping favor use of large clones (several to several hundred KB; see Laan et al. (1995)  Genome Res.  5:13-20), improvements in sensitivity may allow performance of FISH mapping using shorter probes.  
     [0090] A variety of nucleic acid amplification-based methods of genetic and physical mapping may be carried out using the instant nucleic acid sequences. Examples include allele-specific amplification (Kazazian (1989)  J. Lab. Clin. Med.  11:95-96), polymorphism of PCR-amplified fragments (CAPS; Sheffield et al. (1993)  Genomics  16:325-332), allele-specific ligation (Landegren et al. (1988)  Science  241:1077-1080), nucleotide extension reactions (Sokolov (1990)  Nucleic Acid Res.  18:3671), Radiation Hybrid Mapping (Walter et al. (1997)  Nat. Genet.  7:22-28) and Happy Mapping (Dear and Cook (1989)  Nucleic Acid Res.  17:6795-6807). For these methods, the sequence of a nucleic acid fragment is used to design and produce primer pairs for use in the amplification reaction or in primer extension reactions. The design of such primers is well known to those skilled in the art. In methods employing PCR-based genetic mapping, it may be necessary to identify DNA sequence differences between the parents of the mapping cross in the region corresponding to the instant nucleic acid sequence. This, however, is generally not necessary for mapping methods.  
     [0091] Loss of function mutant phenotypes may be identified for the instant cDNA clones either by targeted gene disruption protocols or by identifying specific mutants for these genes contained in a maize population carrying mutations in all possible genes (Ballinger and Benzer (1989)  Proc. Natl. Acad. Sci USA  86:9402-9406; Koes et al. (1995)  Proc. Natl. Acad. Sci USA  92:8149-8153; Bensen et al. (1995)  Plant Cell  7:75-84). The latter approach may be accomplished in two ways. First, short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols in conjunction with a mutation tag sequence primer on DNAs prepared from a population of plants in which Mutator transposons or some other mutation-causing DNA element has been introduced (see Bensen, supra). The amplification of a specific DNA fragment with these primers indicates the insertion of the mutation tag element in or near the plant gene encoding the instant polypeptide. Alternatively, the instant nucleic acid fragment may be used as a hybridization probe against PCR amplification products generated from the mutation population using the mutation tag sequence primer in conjunction with an arbitrary genomic site primer, such as that for a restriction enzyme site-anchored synthetic adaptor. With either method, a plant containing a mutation in the endogenous gene encoding the instant polypeptide can be identified and obtained. This mutant plant can then be used to determine or confirm the natural function of the instant polypeptides disclosed herein.  
     EXAMPLES  
     [0092] The present invention is further illustrated in the following Examples, in which parts and percentages are by weight and degrees are Celsius, unless otherwise stated. It should be understood that these Examples, while indicating preferred embodiments of the invention, are given by way of illustration only. From the above discussion and these Examples, one skilled in the art can ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions. Thus, various modifications of the invention in addition to those shown and described herein will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims.  
     Example 1  
     Composition of cDNA Libraries; Isolation and Sequencing of cDNA Clones  
     [0093] cDNA libraries representing mRNAs from various brassica (Brassica), corn ( Zea mays ), rice ( Oiyza sativa ), soybean ( Glycine max ) and wheat ( Triticum aestivum ) tissues were prepared. The characteristics of the libraries are described below.  
               TABLE 2                          cDNA Libraries from Brassica, Corn, Rice, Soybean and Wheat                         Library   Tissue   Clone               ebp1f   Brassica ( Brassica ) (OGU+, Cyclone cultivar   ebp1f.pk002.g18:fis           containing Ogura restorer) 1-2 mm immature whole           bud       cpd1c   Corn ( Zea mays L. ) pooled BMS treated with   cpd1c.pk002.123:fis           chemicals related to protein kinases       cpi1c   Corn ( Zea mays L. ) pooled BMS treated with   cpi1c.pk011.g12:fis           chemicals related to biochemical compound           synthesis       cho1c   Corn ( Zea mays L. , Alexho Synthetic High Oil)   cho1c.pk003.f1:fis           embryo 20 DAP       rdi2c   Rice ( Otyza sativa , Nipponbare) developing   rdi2c.pk010.p22:fis           inflorescence at rachis branch-floral organ           primordia formation       sfl1   Soybean Immature Flower   sfl1.pk0118.d10:fis       wre1n   Wheat Root From 7 Day Old Etiolated Seedling*   wre1n.pk0004.g6:fis                          
 
     [0094] cDNA libraries may be prepared by any one of many methods available. For example, the cDNAs may be introduced into plasmid vectors by first preparing the cDNA libraries in Uni-ZAP™ XR vectors according to the manufacturer&#39;s protocol (Stratagene Cloning Systems, La Jolla, Calif.). The Uni-ZAP™ XR libraries are converted into plasmid libraries according to the protocol provided by Stratagene. Upon conversion, cDNA inserts will be contained in the plasmid vector pBluescript. In addition, the cDNAs may be introduced directly into precut Bluescript II SK(+) vectors (Stratagene) using T4 DNA ligase (New England Biolabs), followed by transfection into DH10B cells according to the manufacturer&#39;s protocol (GIBCO BRL Products). Once the cDNA inserts are in plasmid vectors, plasmid DNAs are prepared from randomly picked bacterial colonies containing recombinant pBluescript plasmids, or the insert cDNA sequences are amplified via polymerase chain reaction using primers specific for vector sequences flanking the inserted cDNA sequences. Amplified insert DNAs or plasmid DNAs are sequenced in dye-primer sequencing reactions to generate partial cDNA sequences (expressed sequence tags or “ESTs”; see Adams et al., (1991)  Science  252:1651-1656). The resulting ESTs are analyzed using a Perkin Elmer Model 377 fluorescent sequencer.  
     [0095] Full-insert sequence (FIS) data is generated utilizing a modified transposition protocol. Clones identified for FIS are recovered from archived glycerol stocks as single colonies, and plasmid DNAs are isolated via alkaline lysis. Isolated DNA templates are reacted with vector primed M13 forward and reverse oligonucleotides in a PCR-based sequencing reaction and loaded onto automated sequencers. Confirmation of clone identification is performed by sequence alignment to the original EST sequence from which the FIS request is made.  
     [0096] Confirmed templates are transposed via the Primer Island transposition kit (PE Applied Biosystems, Foster City, Calif.) which is based upon the  Saccharomyces cerevisiae  Ty1 transposable element (Devine and Boeke (1994)  Nucleic Acids Res.  22:3765-3772). The in vitro transposition system places unique binding sites randomly throughout a population of large DNA molecules. The transposed DNA is then used to transform DH10B electro-competent cells (Gibco BRL/Life Technologies, Rockville, Md.) via electroporation. The transposable element contains an additional selectable marker (named DHFR; Fling and Richards (1983)  Nucleic Acids Res.  11:5147-5158), allowing for dual selection on agar plates of only those subclones containing the integrated transposon. Multiple subclones are randomly selected from each transposition reaction, plasmid DNAs are prepared via alkaline lysis, and templates are sequenced (ABI Prism dye-terminator ReadyReaction mix) outward from the transposition event site, utilizing unique primers specific to the binding sites within the transposon.  
     [0097] Sequence data is collected (ABI Prism Collections) and assembled using Phred/Phrap (P. Green, University of Washington, Seattle). Phrep/Phrap is a public domain software program which re-reads the ABI sequence data, re-calls the bases, assigns quality values, and writes the base calls and quality values into editable output files. The Phrap sequence assembly program uses these quality values to increase the accuracy of the assembled sequence contigs. Assemblies are viewed by the Consed sequence editor (D. Gordon, University of Washington, Seattle).  
     [0098] In some of the clones the cDNA fragment corresponds to a portion of the 3′-terminus of the gene and does not cover the entire open reading frame. In order to obtain the upstream information one of two different protocols are used. The first of these methods results in the production of a fragment of DNA containing a portion of the desired gene sequence while the second method results in the production of a fragment containing the entire open reading frame. Both of these methods use two rounds of PCR amplification to obtain fragments from one or more libraries. The libraries some times are chosen based on previous knowledge that the specific gene should be found in a certain tissue and some times are randomly-chosen. Reactions to obtain the same gene may be performed on several libraries in parallel or on a pool of libraries. Library pools are normally prepared using from 3 to 5 different libraries and normalized to a uniform dilution. . In the first round of amplification both methods use a vector-specific (forward) primer corresponding to a portion of the vector located at the 5′-terminus of the clone coupled with a gene-specific (reverse) primer. The first method uses a sequence that is complementary to a portion of the already known gene sequence while the second method uses a gene-specific primer complementary to a portion of the 3′-untranslated region (also referred to as UTR). In the second round of amplification a nested set of primers is used for both methods. The resulting DNA fragment is ligated into a pBluescript vector using a commercial kit and following the manufacturer&#39;s protocol. This kit is selected from many available from several vendors including Invitrogen (Carlsbad, Calif.), Promega Biotech (Madison, Wis.), and Gibco-BRL (Gaithersburg, Md.). The plasmid DNA is isolated by alkaline lysis method and submitted for sequencing and assembly using Phred/Phrap, as above.  
     Example 2  
     Identification of cDNA Clones  
     [0099] cDNA clones encoding phosphoribosylaminoimidazole carboxylases were identified by conducting BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993)  J. Mol. Biol.  215:403-410) searches for similarity to sequences contained in the BLAST “nr” database (comprising all non-redundant GenBank CDS translations, sequences derived from the 3-dimensional structure Brookhaven Protein Data Bank, the last major release of the SWISS-PROT protein sequence database, EMBL, and DDBJ databases). The cDNA sequences obtained in Example 1 were analyzed for similarity to all publicly available DNA sequences contained in the “nr” database using the BLASTN algorithm provided by the National Center for Biotechnology Information (NCBI). The DNA sequences were translated in all reading frames and compared for similarity to all publicly available protein sequences contained in the “nr” database using the BLASTX algorithm (Gish and States (1993)  Nat. Genet.  3:266-272) provided by the NCBI. For convenience, the P-value (probability) of observing a match of a cDNA sequence to a sequence contained in the searched databases merely by chance as calculated by BLAST are reported herein as “plog” values, which represent the negative of the logarithm of the reported P-value. Accordingly, the greater the pLog value, the greater the likelihood that the cDNA sequence and the BLAST “hit” represent homologous proteins.  
     [0100] ESTs submitted for analysis are compared to the GenBank database as described above. ESTs that contain sequences more 5- or 3-prime can be found by using the BLASTn algorithm (Altschul et al (1997)  Nucleic Acids Res.  25:3389-3402.) against the Du Pont proprietary database comparing nucleotide sequences that share common or overlapping regions of sequence homology. Where common or overlapping sequences exist between two or more nucleic acid fragments, the sequences can be assembled into a single contiguous nucleotide sequence, thus extending the original fragment in either the 5 or 3 prime direction. Once the most 5-prime EST is identified, its complete sequence can be determined by Full Insert Sequencing as described in Example 1. Homologous genes belonging to different species can be found by comparing the amino acid sequence of a known gene (from either a proprietary source or a public database) against an EST database using the tBLASTn algorithm. The tBLASTn algorithm searches an amino acid query against a nucleotide database that is translated in all 6 reading frames. This search allows for differences in nucleotide codon usage between different species, and for codon degeneracy.  
     Example 3  
     Characterization of cDNA Clones Encoding Phosphoribosylaminoimidazole Carboxylase  
     [0101] The BLASTX search using the EST sequences from clones listed in Table 3 revealed similarity of the polypeptides encoded by the cDNAs to phosphoribosylaminoimidazole carboxylase from either  Vigna aconitifolia  (NCBI General Identification (GI) No. 1709930; SEQ ID NO:15),  Arabidopsis thaliana  (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) or  Nicotiana tabacum  (NCBI General Identification (GI) No.13173434; SEQ ID NO:14).  
     [0102] Shown in Table 3 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clones (“FIS”), the sequences of contigs assembled from two or more ESTs (“Contig”), sequences of contigs assembled from an FIS and one or more ESTs (“Contig*”), or sequences encoding an entire protein derived from an FIS, a contig, or an FIS and PCR (“CGS”):  
               TABLE 3                          BLAST Results for Sequences Encoding Polypeptides Homologous       to Phosphoribosylaminoimidazole Carboxylase                             Clone   Status   NCBI GI No.   BLAST pLog Score                                     ebp1f.pk002.g18:fis   CGS   7436526   180.00       cpd1c.pk002.I23:fis   CGS   7436526   180.00       cho1c.pk003.f1:fis   CGS   13173434   180.00       cpi1c.pk011.g12:fis   FIS   13173434   180.00       rdi2c.pk010.p22:fis   CGS   13173434   180.00       sfl1.pk0118.d10:fis   FIS   1709930   51.00       Contig of   FIS   7436526   31.22       wre1n.pk0004.g6:fis                  
 
     [0103] The nucleotide sequence of the entire cDNA insert in clone ebp1f.pk002.g18 is shown in SEQ ID NO:16. The amino acid sequence deduced from nucleotides 112 through 2025 of SEQ ID NO:16 is shown in SEQ ID NO:17 (stop codon encoded by nt 2026-2028). The nucleotide sequence of the entire cDNA insert in clone cpd1c.pk002.l23 is shown in SEQ ID NO:1. The amino acid sequence deduced from nucleotides 103 through 2004 of SEQ ID NO:1 is shown in SEQ ID NO:2 (stop codon encoded by nt 2005-2007). The nucleotide sequence of the entire cDNA insert in clone chol c.pk003.f1 is shown in SEQ ID NO:3. The amino acid sequence deduced from nucleotides 231 through 2138 of SEQ ID NO:3 is shown in SEQ ID NO:4 (stop codon encoded by nt 2139-2141). The nucleotide sequence of the entire cDNA insert in clone cpi1c.pk011.g12 is shown in SEQ ID NO:5. The amino acid sequence deduced from nucleotides 3 through 1571 of SEQ ID NO:5 is shown in SEQ ID NO:6 (stop codon encoded by nt 1572-1574). The nucleotide sequence of the entire cDNA insert in clone rdi2c.pk010.p22 is shown in SEQ ID NO:7. The amino acid sequence deduced from nucleotides 147 through 2039 of SEQ ID NO:7 is shown in SEQ ID NO:8 (stop codon encoded by nt 2040-2042). The nucleotide sequence of the entire cDNA insert in clone sfl1.pk0118.d10 is shown in SEQ ID NO:9. The amino acid sequence deduced from nucleotides 1 through 333 of SEQ ID NO:9 is shown in SEQ ID NO:10 (stop codon encoded by nt 334-336). The nucleotide sequence of the contig of clone wre1n.pk0004.g6 is shown in SEQ ID NO:11. The amino acid sequence deduced from nucleotides 1 through 348 of SEQ ID NO:11 is shown in SEQ ID NO:12.  
     [0104]FIGS. 2A, 2B,  2 C and  2 D present an alignment of the amino acid sequences set forth in SEQ ID Nos:2, 4, 6, 8, and 17, and the  Vigna aconitifolia  sequence (NCBI General Identification (GI) No.1709930; SEQ ID NO:15),  Arabidopsis thaliana  sequence (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) and  Nicotiana tabacum  sequence (NCBI General Identification (GI) No.13173434; SEQ ID NO:14). The data in Table 4 represents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID Nos:2, 4, 6, 8 and 17, and the  Vigna aconitifolia  sequence (NCBI General Identification (GI) No.1709930; SEQ ID NO:15),  Arabidopsis thaliana  sequence (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) or  Nicotiana tabacum  sequence (NCBI General Identification (GI) No. 13173434; SEQ ID NO:14).  
               TABLE 4                          Percent Identity of Amino Acid Sequences Deduced From the Nucleotide       Sequences of cDNA Clones Encoding Polypeptides Homologous to       Phosphoribosylaminoimidazole Carboxylase                                     Percent Identity to           SEQ ID   NOBI GI No. 1709930   NOBI GI No. 7436526   NOBI GI No. 13173434       NO.   (SEQ ID NO: 15)   (SEQ ID NO: 13)   (SEQ ID NQ: 14)                                     17   71.8   89.4   79.4       2   60.8   64.7   69.4       4   60.2   64.1   68.8       6   59.6   63.5   68.2       8   59.6   64.1   66.5                    
     [0105]FIG. 3 depicts the amino acid alignment of the catalytic region of  Vigna aconitifolia  (NCBI General Identification (GI) No. 1709930; SEQ ID NO:15) (nucleotides 387 to 557) with the amino acid sequences encoded by the following:  
     [0106] (a) nucleotide sequence derived from corn clone cpd1c.pk002.l23 (SEQ ID NO:2),  
     [0107] (b) nucleotide sequence derived from corn clone cho1c.pk003.f1 (SEQ ID NO:4),  
     [0108] (c) nucleotide sequence derived from corn clone cpi1c.pk011.g12 (SEQ ID NO:6),  
     [0109] (d) nucleotide sequence derived from rice clone rdi2c.pk010.p22 (SEQ ID NO:8),  
     [0110] (e) nucleotide sequence derived from brassica clone ebp1f.pk002.g18 (SEQ ID NO:17), (f) nucleotide sequence from  Arabidopsis thaliana  (NCBI General Identification (GI) No. 7436526; SEQ ID NO:13) and (g) nucleotide sequence from  Nicotiana tabacum  (NCBI General Identification (GI) No. 13173434; SEQ ID NO: 14).  
     [0111] Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal V method of alignment (Higgins and Sharp (1989)  CABIOS.  5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of a phosphoribosylaminoimidazole carboxylase. These sequences represent the first monocot species sequences encoding phosphoribosylaminoimidazole carboxylase known to Applicant.  
     Example 4  
     Expression of Recombinant DNA Constructs or Chimeric Genes in Monocot Cells  
     [0112] A recombinant DNA construct or chimeric gene comprising a cDNA encoding the instant polypeptide in sense orientation with respect to the maize 27 kD zein promoter that is located 5′ to the cDNA fragment, and the 10 kD zein 3′ end that is located 3′ to the cDNA fragment, can be constructed. The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites (NcoI or SmaI) can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the digested vector pML103 as described below. Amplification is then performed in a standard PCR. The amplified DNA is then digested with restriction enzymes NcoI and SmaI and fractionated on an agarose gel. The appropriate band can be isolated from the gel and combined with a 4.9 kb NcoI-SmaI fragment of the plasmid pML103. Plasmid pML103 has been deposited under the terms of the Budapest Treaty at ATCC (American Type Culture Collection, 10801 University Blvd., Manassas, Va. 20110-2209), and bears accession number ATCC 97366. The DNA segment from pML103 contains a 1.05 kb SaII-NcoI promoter fragment of the maize 27 kD zein gene and a 0.96 kb SmaI-SaII fragment from the 3′ end of the maize 10 kD zein gene in the vector pGem9Zf(+) (Promega). Vector and insert DNA can be ligated at 15° C. overnight, essentially as described (Maniatis). The ligated DNA may then be used to transform  E. coli  XL1-Blue ( Epicurian Coli  XL-1 Blue™; Stratagene). Bacterial transformants can be screened by restriction enzyme digestion of plasmid DNA and limited nucleotide sequence analysis using the dideoxy chain termination method (Sequenase™ DNA Sequencing Kit; U.S. Biochemical). The resulting plasmid construct would comprise a recombinant DNA construct or chimeric gene encoding, in the 5′ to 3′ direction, the maize 27 kD zein promoter, a cDNA fragment encoding the instant polypeptide, and the 10 kD zein 3′ region.  
     [0113] The recombinant DNA construct or chimeric gene described above can then be introduced into corn cells by the following procedure. Immature corn embryos can be dissected from developing caryopses derived from crosses of the inbred corn lines H99 and LH132. The embryos are isolated 10 to 11 days after pollination when they are 1.0 to 1.5 mm long. The embryos are then placed with the axis-side facing down and in contact with agarose-solidified N6 medium (Chu et al. (1975)  Sci. Sin. Peking  18:659-668). The embryos are kept in the dark at 27° C. Friable embryogenic callus consisting of undifferentiated masses of cells with somatic proembryoids and embryoids borne on suspensor structures proliferates from the scutellum of these immature embryos. The embryogenic callus isolated from the primary explant can be cultured on N6 medium and sub-cultured on this medium every 2 to 3 weeks.  
     [0114] The plasmid, p35S/Ac (obtained from Dr. Peter Eckes, Hoechst Ag, Frankfurt, Germany) may be used in transformation experiments in order to provide for a selectable marker. This plasmid contains the Pat gene (see European Patent Publication 0 242 236) which encodes phosphinothricin acetyl transferase (PAT). The enzyme PAT confers resistance to herbicidal glutamine synthetase inhibitors such as phosphinothricin. The pat gene in p35S/Ac is under the control of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985)  Nature  313:810-812) and the 3′ region of the nopaline synthase gene from the T-DNA of the Ti plasmid of  Agrobacterium tumefaciens.    
     [0115] The particle bombardment method (Klein et al. (1987)  Nature  327:70-73) may be used to transfer genes to the callus culture cells. According to this method, gold particles (1 μm in diameter) are coated with DNA using the following technique. Ten μg of plasmid DNAs are added to 50 μL of a suspension of gold particles (60 mg per mL). Calcium chloride (50 μL of a 2.5 M solution) and spermidine free base (20 μL of a 1.0 M solution) are added to the particles. The suspension is vortexed during the addition of these solutions. After 10 minutes, the tubes are briefly centrifuged (5 sec at 15,000 rpm) and the supernatant removed. The particles are resuspended in 200 μL of absolute ethanol, centrifuged again and the supernatant removed. The ethanol rinse is performed again and the particles resuspended in a final volume of 30 μL of ethanol. An aliquot (5 μL) of the DNA-coated gold particles can be placed in the center of a Kapton™ flying disc (Bio-Rad Labs). The particles are then accelerated into the corn tissue with a Biolistic™ PDS-1000/He (Bio-Rad Instruments, Hercules Calif.), using a helium pressure of 1000 psi, a gap distance of 0.5 cm and a flying distance of 1.0 cm.  
     [0116] For bombardment, the embryogenic tissue is placed on filter paper over agarose-solidified N6 medium. The tissue is arranged as a thin lawn and covered a circular area of about 5 cm in diameter. The petri dish containing the tissue can be placed in the chamber of the PDS-1000/He approximately 8 cm from the stopping screen. The air in the chamber is then evacuated to a vacuum of 28 inches of Hg. The macrocarrier is accelerated with a helium shock wave using a rupture membrane that bursts when the He pressure in the shock tube reaches 1000 psi.  
     [0117] Seven days after bombardment the tissue can be transferred to N6 medium that contains bialophos (5 mg per liter) and lacks casein or proline. The tissue continues to grow slowly on this medium. After an additional 2 weeks the tissue can be transferred to fresh N6 medium containing bialophos. After 6 weeks, areas of about 1 cm in diameter of actively growing callus can be identified on some of the plates containing the bialophos-supplemented medium. These calli may continue to grow when sub-cultured on the selective medium.  
     [0118] Plants can be regenerated from the transgenic callus by first transferring clusters of tissue to N6 medium supplemented with 0.2 mg per liter of 2,4-D. After two weeks the tissue can be transferred to regeneration medium (Fromm et al. (1990)  Bio/Technology  8:833-839).  
     Example 5  
     Expression of Recombinant DNA Constructs or Chimeric Genes in Dicot Cells  
     [0119] A seed-specific expression cassette composed of the promoter and transcription terminator from the gene encoding the β subunit of the seed storage protein phaseolin from the bean  Phaseolus vulgaris  (Doyle et al. (1986)  J. Biol. Chem.  261:9228-9238) can be used for expression of the instant polypeptides in transformed soybean. The phaseolin cassette includes about 500 nucleotides upstream (5′) from the translation initiation codon and about 1650 nucleotides downstream (3′) from the translation stop codon of phaseolin. Between the 5′ and 3′ regions are the unique restriction endonuclease sites Nco I (which includes the ATG translation initiation codon), Sma I, Kpn I and Xba I. The entire cassette is flanked by Hind III sites.  
     [0120] The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the expression vector. Amplification is then performed as described above, and the isolated fragment is inserted into a pUC18 vector carrying the seed expression cassette.  
     [0121] Soybean embryos may then be transformed with the expression vector comprising sequences encoding the instant polypeptides. To induce somatic embryos, cotyledons, 3-5 mm in length dissected from surface sterilized, immature seeds of the soybean cultivar A2872, can be cultured in the light or dark at 26° C. on an appropriate agar medium for 6-10 weeks. Somatic embryos which produce secondary embryos are then excised and placed into a suitable liquid medium. After repeated selection for clusters of somatic embryos which multiplied as early, globular staged embryos, the suspensions are maintained as described below.  
     [0122] Soybean embryogenic suspension cultures can be maintained in 35 mL liquid media on a rotary shaker, 150 rpm, at 26° C. with florescent lights on a 16:8 hour day/night schedule. Cultures are subcultured every two weeks by inoculating approximately 35 mg of tissue into 35 mL of liquid medium.  
     [0123] Soybean embryogenic suspension cultures may then be transformed by the method of particle gun bombardment (Klein et al. (1987)  Nature  (London) 327:70-73, U.S. Pat. No. 4,945,050). A DuPont Biolistic™ PDS1000/HE instrument (helium retrofit) can be used for these transformations.  
     [0124] A selectable marker gene which can be used to facilitate soybean transformation is a chimeric gene composed of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985)  Nature  313:810-812), the hygromycin phosphotransferase gene from plasmid pJR225 (from  E. coli ; Gritz et al.(1983)  Gene  25:179-188) and the 3′ region of the nopaline synthase gene from the T-DNA of the Ti plasmid of  Agrobacterium tumefaciens . The seed expression cassette comprising the phaseolin 5′ region, the fragment encoding the instant polypeptide and the phaseolin 3′ region can be isolated as a restriction fragment. This fragment can then be inserted into a unique restriction site of the vector carrying the marker gene.  
     [0125] To 50 μL of a 60 mg/mL 1 μm gold particle suspension is added (in order): 5 μL DNA (1 μg/μL), 20 μL spermidine (0.1 M), and 50 μL CaCl 2  (2.5 M). The particle preparation is then agitated for three minutes, spun in a microfuge for 10 seconds and the supernatant removed. The DNA-coated particles are then washed once in 400 μL 70% ethanol and resuspended in 40 μL of anhydrous ethanol. The DNA/particle suspension can be sonicated three times for one second each. Five μL of the DNA-coated gold particles are then loaded on each macro carrier disk.  
     [0126] Approximately 300-400 mg of a two-week-old suspension culture is placed in an empty 60×15 mm petri dish and the residual liquid removed from the tissue with a pipette. For each transformation experiment, approximately 5-10 plates of tissue are normally bombarded. Membrane rupture pressure is set at 1100 psi and the chamber is evacuated to a vacuum of 28 inches mercury. The tissue is placed approximately 3.5 inches away from the retaining screen and bombarded three times. Following bombardment, the tissue can be divided in half and placed back into liquid and cultured as described above.  
     [0127] Five to seven days post bombardment, the liquid media may be exchanged with fresh media, and eleven to twelve days post bombardment with fresh media containing 50 mg/mL hygromycin. This selective media can be refreshed weekly. Seven to eight weeks post bombardment, green, transformed tissue may be observed growing from untransformed, necrotic embryogenic clusters. Isolated green tissue is removed and inoculated into individual flasks to generate new, clonally propagated, transformed embryogenic suspension cultures. Each new line may be treated as an independent transformation event. These suspensions can then be subcultured and maintained as clusters of immature embryos or regenerated into whole plants by maturation and germination of individual somatic embryos.  
     Example 6  
     Expression of Recombinant DNA Constructs or Chimeric Genes in Microbial Cells  
     [0128] The cDNAs encoding the instant polypeptides can be inserted into the T7  E. coli  expression vector pBT430. This vector is a derivative of pET-3a (Rosenberg et al. (1987)  Gene  56:125-135) which employs the bacteriophage T7 RNA polymerase/T7 promoter system. Plasmid pBT430 was constructed by first destroying the EcoR I and Hind III sites in pET-3a at their original positions. An oligonucleotide adaptor containing EcoR I and Hind III sites was inserted at the BamH I site of pET-3a. This created pET-3aM with additional unique cloning sites for insertion of genes into the expression vector. Then, the Nde I site at the position of translation initiation was converted to an Nco I site using oligonucleotide-directed mutagenesis. The DNA sequence of pET-3aM in this region, 5′-CATATGG, was converted to 5′-CCCATGG in pBT430.  
     [0129] Plasmid DNA containing a cDNA may be appropriately digested to release a nucleic acid fragment encoding the protein. This fragment may then be purified on a 1% low melting agarose gel. Buffer and agarose contain 10 μg/ml ethidium bromide for visualization of the DNA fragment. The fragment can then be purified from the agarose gel by digestion with GELase™ (Epicentre Technologies, Madison, Wis.) according to the manufacturer&#39;s instructions, ethanol precipitated, dried and resuspended in 20 μL of water. Appropriate oligonucleotide adapters may be ligated to the fragment using T4 DNA ligase (New England Biolabs (NEB), Beverly, Mass.). The fragment containing the ligated adapters can be purified from the excess adapters using low melting agarose as described above. The vector pBT430 is digested, dephosphorylated with alkaline phosphatase (NEB) and deproteinized with phenol/chloroform as described above. The prepared vector pBT430 and fragment can then be ligated at 16° C. for 15 hours followed by transformation into DH5 electrocompetent cells (GIBCO BRL). Transformants can be selected on agar plates containing LB media and 100 μg/mL ampicillin. Transformants containing the gene encoding the instant polypeptide are then screened for the correct orientation with respect to the T7 promoter by restriction enzyme analysis.  
     [0130] For high level expression, a plasmid clone with the cDNA insert in the correct orientation relative to the T7 promoter can be transformed into  E. coli  strain BL21(DE3) (Studier et al. (1986)  J. Mol. Biol.  189:113-130). Cultures are grown in LB medium containing ampicillin (100 mg/L) at 25° C. At an optical density at 600 nm of approximately 1, IPTG (isopropylthio-β-galactoside, the inducer) can be added to a final concentration of 0.4 mM and incubation can be continued for 3 h at 25° C. Cells are then harvested by centrifugation and re-suspended in 50 μL of 50 mM Tris-HCl at pH 8.0 containing 0.1 mM DTT and 0.2 mM phenyl methylsulfonyl fluoride. A small amount of 1 mm glass beads can be added and the mixture sonicated 3 times for about 5 seconds each time with a microprobe sonicator. The mixture is centrifuged and the protein concentration of the supernatant determined. One μg of protein from the soluble fraction of the culture can be separated by SDS-polyacrylamide gel electrophoresis. Gels can be observed for protein bands migrating at the expected molecular weight.  
     Example 7  
     Evaluating Compounds for Their Ability to Inhibit the Activity of Phosphoribosylaminoimidazole Carboxylase  
     [0131] The polypeptides described herein may be produced using any number of methods known to those skilled in the art. Such methods include, but are not limited to, expression in bacteria as described in Example 6, or expression in eukaryotic cell culture, in planta, and using viral expression systems in suitably infected organisms or cell lines. The instant polypeptides may be expressed either as mature forms of the proteins as observed in vivo or as fusion proteins by covalent attachment to a variety of enzymes, proteins or affinity tags. Common fusion protein partners include glutathione S-transferase (“GST”), thioredoxin (“Trx”), maltose binding protein, and C- and/or N-terminal hexahistidine polypeptide (“(His) 6 ”). The fusion proteins may be engineered with a protease recognition site at the fusion point so that fusion partners can be separated by protease digestion to yield intact mature enzyme. Examples of such proteases include thrombin, enterokinase and factor Xa. However, any protease can be used which specifically cleaves the peptide connecting the fusion protein and the enzyme.  
     [0132] Purification of the instant polypeptides, if desired, may utilize any number of separation technologies familiar to those skilled in the art of protein purification. Examples of such methods include, but are not limited to, homogenization, filtration, centrifugation, heat denaturation, ammonium sulfate precipitation, desalting, pH precipitation, ion exchange chromatography, hydrophobic interaction chromatography and affinity chromatography, wherein the affinity ligand represents a substrate, substrate analog or inhibitor. When the instant polypeptides are expressed as fusion proteins, the purification protocol may include the use of an affinity resin which is specific for the fusion protein tag attached to the expressed enzyme or an affinity resin containing ligands which are specific for the enzyme. For example, the instant polypeptides may be expressed as a fusion protein coupled to the C-terminus of thioredoxin. In addition, a (His) 6  peptide may be engineered into the N-terminus of the fused thioredoxin moiety to afford additional opportunities for affinity purification. Other suitable affinity resins could be synthesized by linking the appropriate ligands to any suitable resin such as Sepharose-4B. In an alternate embodiment, a thioredoxin fusion protein may be eluted using dithiothreitol; however, elution may be accomplished using other reagents which interact to displace the thioredoxin from the resin. These reagents include β-mercaptoethanol or other reduced thiol. The eluted fusion protein may be subjected to further purification by traditional means as stated above, if desired. Proteolytic cleavage of the thioredoxin fusion protein and the enzyme may be accomplished after the fusion protein is purified or while the protein is still bound to the ThioBond™ affinity resin or other resin.  
     [0133] Crude, partially purified or purified enzyme, either alone or as a fusion protein, may be utilized in assays for the evaluation of compounds for their ability to inhibit enzymatic activation of the instant polypeptides disclosed herein. Assays may be conducted under well known experimental conditions which permit optimal enzymatic activity. For example, assays for phosphoribosylaminoimidazole carboxylase are presented by Lukens, L. N. and Buchanan, J. M.,  J. Biol. Chem.  234(7):1799-1805 (1959)).  
    
     
       
         1 
         
           
             17  
           
           
             1  
             2199  
             DNA  
             Zea mays  
           
            1 

gcacgaggca cgagcggcac gagctagaca cccatcgtct cccgttcccg gctaaaccct     60 

gcttgagccc tagacaccgc ccgcctccgg gaggtagaga gcatgcacgc caggttcctc    120 

agcgcgccgt cccctgcctc cgccgccccc tccccgtacg ttcgcttggc cttcacgggc    180 

gcccgcccgc gccgcgcgtg ttggaagccg cgaggccccg cgtccgcgtc cgcgccgccg    240 

cctcggcctc tccactcgct gtgcgcccgg gcctccatgc agcccgcctc tcccgcgcac    300 

gacgggcatg gtggtccgcc ggtgcacggc gtctccaaca ccgtcatcgg ggtcctgggg    360 

ggcggtcagc tcgggaagat gctctgccat gcagcgagtc aaatggggat tagaattgtc    420 

atcctcgacc ctcttcccgg ctgccccgcg agctcggttt gcgatgagca cgtaatcggg    480 

agcttcaacg atagggacac ggtccgggag ttcgccaaga ggtgtggggt cctaacagtg    540 

gagattgagc atgttgatgc cgccacactg gagaagctgg aaaaacaggg cgttgactgc    600 

gagcctaaag cctccacaat cacgattatt caggacaagt acaggcagaa aaagcatttc    660 

tcaagatgcg agatcccatt gcctgacttt atggaagtag atactttacg cagtatagag    720 

gaggctgggg aaaagtttgg ctatccccta atggtcaaaa gcaagagatt agcatatgat    780 

ggtcgaggaa atgctgtagc caagaacaga gaggagctac cttctgttgt tgcttcactg    840 

ggtgggtttg agcggggctt gtatgttgag agatggactc ctttcgtaaa ggagctttct    900 

gtaattgtgg caaggagcag agacaactct actgtctgct atcctgttgt ggaaacagtt    960 

cacaaggaaa atatatgcca tgttgttgaa gctcctgctg atttatctaa caaaataaag   1020 

aagttagcta ctagcgtggc tgaaaaagct atcaaatcat tagaaggagc tggtgtcttt   1080 

gctgtagagt tgtttttaac agaagatgat cagattttat tgaatgaggt agcccctagg   1140 

cctcacaata gtgggcatca cacaatagag tcatgctaca cctcacaata tgagcagcat   1200 

atacgcgcta ttcttggcct tcctcttggt gatccctcaa tgaaagcacc tgcagcaata   1260 

atgtacaaca tcctgggcga ggatgagggt gaagcagggt tctttctggc tcatcagctt   1320 

atcagtaggg cactaaccat tccaggcaca tcggtccatt ggtacgcaaa gccagagatg   1380 

cggaagcaaa ggaagatggg tcatattaca attgtggggc cttctaagat aagtgtaaaa   1440 

tcacgcttgg acaacttgct gcaaagaaac tcgtctgatc ccaaggaagt tagccctcgt   1500 

gttgctatta taatggggtc ccaatctgat cttcctgtga tgaaagatgc tgagagggtt   1560 

ttgaaagagt tcgacatacc ttgtgaggta actattgttt ctgcacatcg tacaccagag   1620 

cggatgtatg attatgcgaa gtctgctaaa gacaggggtt tcgaggtcat aattgcaggt   1680 

gcaggcggag cagctcattt accagggatg gtggcttcat tgactcctct tcctgtaatc   1740 

ggagttccca ttaagacttc aacactatca ggatttgatt ccctcctatc tattgtgcaa   1800 

atgccaaaag gtattcctgt tgcgactgtt gctatcggga atgcagaaaa tgcaggtttg   1860 

ctggcagcta ggattctggc tgcaagagat cctgagctcc aggacagggt aactaagtac   1920 

caggatgatc tgagggacat ggttttggag acggcagaaa ggctggagga ccaaggcccg   1980 

gaggaatttc tgaagggaat ggattgaccc cttttgaagc cctggctctg gggcctggga   2040 

gaggaatacg agtgtatggg tcggcgataa ctcactgtcc actcgattat ggtttaagat   2100 

ggggagtcat caagaagata ctttaatagg ctcggctgca ttcgattttc cttacaattc   2160 

ttttttgata aaaaaaaaaa aaaaaaaaaa aaaaaaaaa                          2199 

 
           
             2  
             634  
             PRT  
             Zea mays  
           
            2 

Met His Ala Arg Phe Leu Ser Ala Pro Ser Pro Ala Ser Ala Ala Pro 
  1               5                  10                  15 

Ser Pro Tyr Val Arg Leu Ala Phe Thr Gly Ala Arg Pro Arg Arg Ala 
             20                  25                  30 

Cys Trp Lys Pro Arg Gly Pro Ala Ser Ala Ser Ala Pro Pro Pro Arg 
         35                  40                  45 

Pro Leu His Ser Leu Cys Ala Arg Ala Ser Met Gln Pro Ala Ser Pro 
     50                  55                  60 

Ala His Asp Gly His Gly Gly Pro Pro Val His Gly Val Ser Asn Thr 
 65                  70                  75                  80 

Val Ile Gly Val Leu Gly Gly Gly Gln Leu Gly Lys Met Leu Cys His 
                 85                  90                  95 

Ala Ala Ser Gln Met Gly Ile Arg Ile Val Ile Leu Asp Pro Leu Pro 
            100                 105                 110 

Gly Cys Pro Ala Ser Ser Val Cys Asp Glu His Val Ile Gly Ser Phe 
        115                 120                 125 

Asn Asp Arg Asp Thr Val Arg Glu Phe Ala Lys Arg Cys Gly Val Leu 
    130                 135                 140 

Thr Val Glu Ile Glu His Val Asp Ala Ala Thr Leu Glu Lys Leu Glu 
145                 150                 155                 160 

Lys Gln Gly Val Asp Cys Glu Pro Lys Ala Ser Thr Ile Thr Ile Ile 
                165                 170                 175 

Gln Asp Lys Tyr Arg Gln Lys Lys His Phe Ser Arg Cys Glu Ile Pro 
            180                 185                 190 

Leu Pro Asp Phe Met Glu Val Asp Thr Leu Arg Ser Ile Glu Glu Ala 
        195                 200                 205 

Gly Glu Lys Phe Gly Tyr Pro Leu Met Val Lys Ser Lys Arg Leu Ala 
    210                 215                 220 

Tyr Asp Gly Arg Gly Asn Ala Val Ala Lys Asn Arg Glu Glu Leu Pro 
225                 230                 235                 240 

Ser Val Val Ala Ser Leu Gly Gly Phe Glu Arg Gly Leu Tyr Val Glu 
                245                 250                 255 

Arg Trp Thr Pro Phe Val Lys Glu Leu Ser Val Ile Val Ala Arg Ser 
            260                 265                 270 

Arg Asp Asn Ser Thr Val Cys Tyr Pro Val Val Glu Thr Val His Lys 
        275                 280                 285 

Glu Asn Ile Cys His Val Val Glu Ala Pro Ala Asp Leu Ser Asn Lys 
    290                 295                 300 

Ile Lys Lys Leu Ala Thr Ser Val Ala Glu Lys Ala Ile Lys Ser Leu 
305                 310                 315                 320 

Glu Gly Ala Gly Val Phe Ala Val Glu Leu Phe Leu Thr Glu Asp Asp 
                325                 330                 335 

Gln Ile Leu Leu Asn Glu Val Ala Pro Arg Pro His Asn Ser Gly His 
            340                 345                 350 

His Thr Ile Glu Ser Cys Tyr Thr Ser Gln Tyr Glu Gln His Ile Arg 
        355                 360                 365 

Ala Ile Leu Gly Leu Pro Leu Gly Asp Pro Ser Met Lys Ala Pro Ala 
    370                 375                 380 

Ala Ile Met Tyr Asn Ile Leu Gly Glu Asp Glu Gly Glu Ala Gly Phe 
385                 390                 395                 400 

Phe Leu Ala His Gln Leu Ile Ser Arg Ala Leu Thr Ile Pro Gly Thr 
                405                 410                 415 

Ser Val His Trp Tyr Ala Lys Pro Glu Met Arg Lys Gln Arg Lys Met 
            420                 425                 430 

Gly His Ile Thr Ile Val Gly Pro Ser Lys Ile Ser Val Lys Ser Arg 
        435                 440                 445 

Leu Asp Asn Leu Leu Gln Arg Asn Ser Ser Asp Pro Lys Glu Val Ser 
    450                 455                 460 

Pro Arg Val Ala Ile Ile Met Gly Ser Gln Ser Asp Leu Pro Val Met 
465                 470                 475                 480 

Lys Asp Ala Glu Arg Val Leu Lys Glu Phe Asp Ile Pro Cys Glu Val 
                485                 490                 495 

Thr Ile Val Ser Ala His Arg Thr Pro Glu Arg Met Tyr Asp Tyr Ala 
            500                 505                 510 

Lys Ser Ala Lys Asp Arg Gly Phe Glu Val Ile Ile Ala Gly Ala Gly 
        515                 520                 525 

Gly Ala Ala His Leu Pro Gly Met Val Ala Ser Leu Thr Pro Leu Pro 
    530                 535                 540 

Val Ile Gly Val Pro Ile Lys Thr Ser Thr Leu Ser Gly Phe Asp Ser 
545                 550                 555                 560 

Leu Leu Ser Ile Val Gln Met Pro Lys Gly Ile Pro Val Ala Thr Val 
                565                 570                 575 

Ala Ile Gly Asn Ala Glu Asn Ala Gly Leu Leu Ala Ala Arg Ile Leu 
            580                 585                 590 

Ala Ala Arg Asp Pro Glu Leu Gln Asp Arg Val Thr Lys Tyr Gln Asp 
        595                 600                 605 

Asp Leu Arg Asp Met Val Leu Glu Thr Ala Glu Arg Leu Glu Asp Gln 
    610                 615                 620 

Gly Pro Glu Glu Phe Leu Lys Gly Met Asp 
625                 630 

 
           
             3  
             1838  
             DNA  
             Zea mays  
           
            3 

gcacgagctg ccccgcgagc tcggtttgcg atgagcacgt cattgggagc ttcaatgatg     60 

aggacacggt ccgggagttc gccaagaggt gtggggtcct aacagtggag attgagcatg    120 

ttgatgtcac cacactggag aaactggaaa aacagggcgt tgactgcgag cctaacgcct    180 

ccacaatcat gattattcag gacaagtaca ggcagaaaaa gcatttctca agatgcgaga    240 

tcccattgcc tgacttcatg gaagtagata ctttacgcag tatagaggag gctggagaaa    300 

agtttggcta tcccctaatg gtcaaaagca agagattagc atatgatggt cgaggaaatg    360 

ctgtagccaa gaacaaagag gagctacctt ctgttgttgc ttcactaggt gggtttgagc    420 

ggggcttgta tgttgagaga tggactcctt tcgtaaagga gctttctgta atcgtggcaa    480 

ggagcagaga caactctact gtctgctatc ctgttgtgga aacagttcac aaggaaaata    540 

tatgccatgt tgttgaagct cctgctgatg tatctaacaa aataaagaag ttagctacta    600 

gcgtggctga aaaagctatt aaatcattag aaggagctgg tgtctttgct gtagagttgt    660 

ttttaacaga agatgatcag gttttattga atgaggtagc ccctaggcct cacaatagtg    720 

gacatcacac aatagagtca tgctacacct cacaatatga gcagcatata cgcgctattc    780 

ttggccttcc tcttggtgat ccctcaatga aagcacctgc agcaataatg tacaacatcc    840 

tgggcgagga tgagggtgaa gcagggttct ttctggctca tcagcttatc agtagggcac    900 

taaccattcc aggcacatcg gtccattggt acgcaaagcc agagatgcgg aagcaaagga    960 

agatgggtca tattacaatt gtggggcctt ctaagataag tgtaaaatca cgcttggaca   1020 

acttgctgca aagaaactcg tctgatccca aggaagttag ccctcgtgtt gctattataa   1080 

tggggtccca atctgatctt cctgtgatga aagatgctga gagggttttg aaagagttcg   1140 

acataccttg tgagctaact attgtttctg cacatcgtac accagagcgg atgtatgatt   1200 

atgcgaagtc tgctaaagac aggggtttcg aggtcataat tgcaggtgca ggcggagcag   1260 

ctcatttacc agggatggtg gcttcattga ctcctcttcc tgtaatcgga gttcccatta   1320 

agacttcaac actatcagga tttgattccc tcctatctat tgtgcaaatg ccaaaaggta   1380 

ttcctgttgc gactgttgct atcgggattg cagaaaatgc aggtttgctg gcagctagga   1440 

ttctggctgc aagagatcct gagctccagg acagggtaac taagtaccag gatgatctga   1500 

gggacatggt tttggagaca gcagaaaggc tggaggacca aggcccggag gaatttctga   1560 

agggaatgga ttgacccctt ttgaagccct ggctctgggg cctgggagga atacgaatat   1620 

atgggtcggc gataactcag tccactcgat tatggtttaa gatcgggagt catcaaaaag   1680 

atactttaag gactagttcc ggagctacaa aacctgaggg gattgggggc tagaatctcc   1740 

gtgactctta gagctgtttt gatgaccaag aatcacgaag ggatccatag gcttggctgc   1800 

cttattcaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaa                           1838 

 
           
             4  
             523  
             PRT  
             Zea mays  
           
            4 

Thr Ser Cys Pro Ala Ser Ser Val Cys Asp Glu His Val Ile Gly Ser 
  1               5                  10                  15 

Phe Asn Asp Glu Asp Thr Val Arg Glu Phe Ala Lys Arg Cys Gly Val 
             20                  25                  30 

Leu Thr Val Glu Ile Glu His Val Asp Val Thr Thr Leu Glu Lys Leu 
         35                  40                  45 

Glu Lys Gln Gly Val Asp Cys Glu Pro Asn Ala Ser Thr Ile Met Ile 
     50                  55                  60 

Ile Gln Asp Lys Tyr Arg Gln Lys Lys His Phe Ser Arg Cys Glu Ile 
 65                  70                  75                  80 

Pro Leu Pro Asp Phe Met Glu Val Asp Thr Leu Arg Ser Ile Glu Glu 
                 85                  90                  95 

Ala Gly Glu Lys Phe Gly Tyr Pro Leu Met Val Lys Ser Lys Arg Leu 
            100                 105                 110 

Ala Tyr Asp Gly Arg Gly Asn Ala Val Ala Lys Asn Lys Glu Glu Leu 
        115                 120                 125 

Pro Ser Val Val Ala Ser Leu Gly Gly Phe Glu Arg Gly Leu Tyr Val 
    130                 135                 140 

Glu Arg Trp Thr Pro Phe Val Lys Glu Leu Ser Val Ile Val Ala Arg 
145                 150                 155                 160 

Ser Arg Asp Asn Ser Thr Val Cys Tyr Pro Val Val Glu Thr Val His 
                165                 170                 175 

Lys Glu Asn Ile Cys His Val Val Glu Ala Pro Ala Asp Val Ser Asn 
            180                 185                 190 

Lys Ile Lys Lys Leu Ala Thr Ser Val Ala Glu Lys Ala Ile Lys Ser 
        195                 200                 205 

Leu Glu Gly Ala Gly Val Phe Ala Val Glu Leu Phe Leu Thr Glu Asp 
    210                 215                 220 

Asp Gln Val Leu Leu Asn Glu Val Ala Pro Arg Pro His Asn Ser Gly 
225                 230                 235                 240 

His His Thr Ile Glu Ser Cys Tyr Thr Ser Gln Tyr Glu Gln His Ile 
                245                 250                 255 

Arg Ala Ile Leu Gly Leu Pro Leu Gly Asp Pro Ser Met Lys Ala Pro 
            260                 265                 270 

Ala Ala Ile Met Tyr Asn Ile Leu Gly Glu Asp Glu Gly Glu Ala Gly 
        275                 280                 285 

Phe Phe Leu Ala His Gln Leu Ile Ser Arg Ala Leu Thr Ile Pro Gly 
    290                 295                 300 

Thr Ser Val His Trp Tyr Ala Lys Pro Glu Met Arg Lys Gln Arg Lys 
305                 310                 315                 320 

Met Gly His Ile Thr Ile Val Gly Pro Ser Lys Ile Ser Val Lys Ser 
                325                 330                 335 

Arg Leu Asp Asn Leu Leu Gln Arg Asn Ser Ser Asp Pro Lys Glu Val 
            340                 345                 350 

Ser Pro Arg Val Ala Ile Ile Met Gly Ser Gln Ser Asp Leu Pro Val 
        355                 360                 365 

Met Lys Asp Ala Glu Arg Val Leu Lys Glu Phe Asp Ile Pro Cys Glu 
    370                 375                 380 

Leu Thr Ile Val Ser Ala His Arg Thr Pro Glu Arg Met Tyr Asp Tyr 
385                 390                 395                 400 

Ala Lys Ser Ala Lys Asp Arg Gly Phe Glu Val Ile Ile Ala Gly Ala 
                405                 410                 415 

Gly Gly Ala Ala His Leu Pro Gly Met Val Ala Ser Leu Thr Pro Leu 
            420                 425                 430 

Pro Val Ile Gly Val Pro Ile Lys Thr Ser Thr Leu Ser Gly Phe Asp 
        435                 440                 445 

Ser Leu Leu Ser Ile Val Gln Met Pro Lys Gly Ile Pro Val Ala Thr 
    450                 455                 460 

Val Ala Ile Gly Ile Ala Glu Asn Ala Gly Leu Leu Ala Ala Arg Ile 
465                 470                 475                 480 

Leu Ala Ala Arg Asp Pro Glu Leu Gln Asp Arg Val Thr Lys Tyr Gln 
                485                 490                 495 

Asp Asp Leu Arg Asp Met Val Leu Glu Thr Ala Glu Arg Leu Glu Asp 
            500                 505                 510 

Gln Gly Pro Glu Glu Phe Leu Lys Gly Met Asp 
        515                 520 

 
           
             5  
             2434  
             DNA  
             Zea mays  
           
            5 

gcaccagaca aatcgctttc cgcgccgccg ccgcgccgcc gacctagaca cccatcgtct     60 

cccgttcccg gctaaaccct gcttgagccc tagacaccgc ccgcctccgg gaggtagaga    120 

gcatgcacgc caggttcctc agcgcgccgt cccctgcctc ccggaggctg atgagccggc    180 

taaaccctgc ttgagcccta gacaccgccc gcctccggga ggtagagagc atgcacgcca    240 

ggttcctcag cgcgccgtcc cctgcctccg ccgccccctc cccgtacgtt cgcttggcct    300 

tcacgggcgc ccgcccgcgc cgcgcgtgtt ggaagccggg gccgcgaggc cccgcgtccg    360 

cgtccgcgcc gccgcctcgg cctctccact cgctgtgcgc ccgggcctcc atgcaggccg    420 

cctctcccgc gcacgacggg catggtggtc cgccggtgca cggcgtctcc aacaccgtca    480 

tcggggtcct ggggggcggt cagctcggga agatgctctg ccatgcagcg agtcagatgg    540 

ggattagaat tgtcatcctc gaccctcttc ccggatgccc cgcgagctcg gtttgcgatg    600 

agcacgtcat cgggagcttc aacgatgggg acacggtccg ggagttcgcc aagaggtgtg    660 

gggtcctaac agtggaaatt gagcatgttg atgccgccac actggagaag ctggaaaaac    720 

agggcgttga ctgcgagcct aaagcctcca caatcacgat tattcaggac aagtacaggc    780 

agaaaaagca tttctcaaga tgcgagatcc cattgcctga cttcatggaa gtagatactt    840 

tacgcattat agaggaggct ggagaaaagt ttggctatcc cctaatggtc aaaagcaaga    900 

gattagcata tgatggtcga ggaaatgctg tagccaagaa caaagaggag ctaccttctg    960 

ttgttgcttc actgggtggg tttgagcggg gcttgtatgt tgagagatgg actcctttcg   1020 

taaaggagct ttctgtaatt gtggcaagga gcagagacaa ctctactgtc tgctatcctg   1080 

ttgtggaaac agttcacaag gaaaatatat gccatgttgt tgaagctcca gctgatgtat   1140 

ctaacaaaat aaataagtta gctactagcg tggctgaaaa agctatcaaa tcattagaag   1200 

gagctggtgt ctttgctgta gagttgtttt taacagaaga tgatcagatt ttattgaatg   1260 

aggtagcccc taggcctcac aatagtgggc atcacacaat agagtcatgc tacacctcac   1320 

aatatgagca gcatatacgc gctattcttg gccttcctct tggtgatccc tcaatgaaag   1380 

cacctgcagc aataatgtac aacatcctgg gcgaggatga gggtgaagca gggttctttc   1440 

tggctcatca gcttatcagt agggcactaa ccattccagg cacatcggtc cattggtacg   1500 

caaagccaga gatgcggaag caaaggaaga tgggtcatat tacaattgtg gggccttcta   1560 

agataagtgt aaaatcacgc ttggacaact tgctgcaaag aaactcgtct gatcccaagg   1620 

aagttagccc tcgtgttgct attataatgg ggtcccaatc tgatcttcct gtgatgaaag   1680 

atgctgagag ggttttgaaa gagttcgaca taccttgtga gttaactatt gtttctgcac   1740 

atcgtacacc agagcggatg tatgattatg cgaagtctgc taaagacagg ggtttcgagg   1800 

tcataattgc aggtgcaggc ggagcagctc atttaccagg gatggtggct tcattgactc   1860 

ctcttcctgt aatcggagtt cccattaaga cttcaacact atcaggattt gattccctcc   1920 

tatctattgt gcaaatgcca aaaggtattc ctgttgcgac tgttgctatc gggaatgcag   1980 

aaaatgcagg tttgctggca gctaggattc tggctgcaag agatcctgag ctccaggaca   2040 

gggtaactaa gtaccaggat gatctgaggg acatggtttt ggagacggca gaaaggctgg   2100 

aggaccaagg cccggaggaa tttctgaagg gaatggattg accccttttg aagccctggc   2160 

tctggggcct gggaggaata cgaatgtatg ggtcggcgat aactcactgt ccactcgatt   2220 

atggtttaag atcgggagtc atcaagaaga tactttaata ggctcggctg cattcgattt   2280 

tccttacaat tcttttttga taattgtgct atcttaaccc atctcaacat gccctttttc   2340 

ccttctaata tagggtatga attgggttta gagggggaaa aaagaatggg agtaaacgtt   2400 

gtacgtacga aaaataaaac ggttcgagta acgt                               2434 

 
           
             6  
             636  
             PRT  
             Zea mays  
           
            6 

Met His Ala Arg Phe Leu Ser Ala Pro Ser Pro Ala Ser Ala Ala Pro 
  1               5                  10                  15 

Ser Pro Tyr Val Arg Leu Ala Phe Thr Gly Ala Arg Pro Arg Arg Ala 
             20                  25                  30 

Cys Trp Lys Pro Gly Pro Arg Gly Pro Ala Ser Ala Ser Ala Pro Pro 
         35                  40                  45 

Pro Arg Pro Leu His Ser Leu Cys Ala Arg Ala Ser Met Gln Ala Ala 
     50                  55                  60 

Ser Pro Ala His Asp Gly His Gly Gly Pro Pro Val His Gly Val Ser 
 65                  70                  75                  80 

Asn Thr Val Ile Gly Val Leu Gly Gly Gly Gln Leu Gly Lys Met Leu 
                 85                  90                  95 

Cys His Ala Ala Ser Gln Met Gly Ile Arg Ile Val Ile Leu Asp Pro 
            100                 105                 110 

Leu Pro Gly Cys Pro Ala Ser Ser Val Cys Asp Glu His Val Ile Gly 
        115                 120                 125 

Ser Phe Asn Asp Gly Asp Thr Val Arg Glu Phe Ala Lys Arg Cys Gly 
    130                 135                 140 

Val Leu Thr Val Glu Ile Glu His Val Asp Ala Ala Thr Leu Glu Lys 
145                 150                 155                 160 

Leu Glu Lys Gln Gly Val Asp Cys Glu Pro Lys Ala Ser Thr Ile Thr 
                165                 170                 175 

Ile Ile Gln Asp Lys Tyr Arg Gln Lys Lys His Phe Ser Arg Cys Glu 
            180                 185                 190 

Ile Pro Leu Pro Asp Phe Met Glu Val Asp Thr Leu Arg Ile Ile Glu 
        195                 200                 205 

Glu Ala Gly Glu Lys Phe Gly Tyr Pro Leu Met Val Lys Ser Lys Arg 
    210                 215                 220 

Leu Ala Tyr Asp Gly Arg Gly Asn Ala Val Ala Lys Asn Lys Glu Glu 
225                 230                 235                 240 

Leu Pro Ser Val Val Ala Ser Leu Gly Gly Phe Glu Arg Gly Leu Tyr 
                245                 250                 255 

Val Glu Arg Trp Thr Pro Phe Val Lys Glu Leu Ser Val Ile Val Ala 
            260                 265                 270 

Arg Ser Arg Asp Asn Ser Thr Val Cys Tyr Pro Val Val Glu Thr Val 
        275                 280                 285 

His Lys Glu Asn Ile Cys His Val Val Glu Ala Pro Ala Asp Val Ser 
    290                 295                 300 

Asn Lys Ile Asn Lys Leu Ala Thr Ser Val Ala Glu Lys Ala Ile Lys 
305                 310                 315                 320 

Ser Leu Glu Gly Ala Gly Val Phe Ala Val Glu Leu Phe Leu Thr Glu 
                325                 330                 335 

Asp Asp Gln Ile Leu Leu Asn Glu Val Ala Pro Arg Pro His Asn Ser 
            340                 345                 350 

Gly His His Thr Ile Glu Ser Cys Tyr Thr Ser Gln Tyr Glu Gln His 
        355                 360                 365 

Ile Arg Ala Ile Leu Gly Leu Pro Leu Gly Asp Pro Ser Met Lys Ala 
    370                 375                 380 

Pro Ala Ala Ile Met Tyr Asn Ile Leu Gly Glu Asp Glu Gly Glu Ala 
385                 390                 395                 400 

Gly Phe Phe Leu Ala His Gln Leu Ile Ser Arg Ala Leu Thr Ile Pro 
                405                 410                 415 

Gly Thr Ser Val His Trp Tyr Ala Lys Pro Glu Met Arg Lys Gln Arg 
            420                 425                 430 

Lys Met Gly His Ile Thr Ile Val Gly Pro Ser Lys Ile Ser Val Lys 
        435                 440                 445 

Ser Arg Leu Asp Asn Leu Leu Gln Arg Asn Ser Ser Asp Pro Lys Glu 
    450                 455                 460 

Val Ser Pro Arg Val Ala Ile Ile Met Gly Ser Gln Ser Asp Leu Pro 
465                 470                 475                 480 

Val Met Lys Asp Ala Glu Arg Val Leu Lys Glu Phe Asp Ile Pro Cys 
                485                 490                 495 

Glu Leu Thr Ile Val Ser Ala His Arg Thr Pro Glu Arg Met Tyr Asp 
            500                 505                 510 

Tyr Ala Lys Ser Ala Lys Asp Arg Gly Phe Glu Val Ile Ile Ala Gly 
        515                 520                 525 

Ala Gly Gly Ala Ala His Leu Pro Gly Met Val Ala Ser Leu Thr Pro 
    530                 535                 540 

Leu Pro Val Ile Gly Val Pro Ile Lys Thr Ser Thr Leu Ser Gly Phe 
545                 550                 555                 560 

Asp Ser Leu Leu Ser Ile Val Gln Met Pro Lys Gly Ile Pro Val Ala 
                565                 570                 575 

Thr Val Ala Ile Gly Asn Ala Glu Asn Ala Gly Leu Leu Ala Ala Arg 
            580                 585                 590 

Ile Leu Ala Ala Arg Asp Pro Glu Leu Gln Asp Arg Val Thr Lys Tyr 
        595                 600                 605 

Gln Asp Asp Leu Arg Asp Met Val Leu Glu Thr Ala Glu Arg Leu Glu 
    610                 615                 620 

Asp Gln Gly Pro Glu Glu Phe Leu Lys Gly Met Asp 
625                 630                 635 

 
           
             7  
             2163  
             DNA  
             Oryza sativa  
           
            7 

gcacgaggga cgcgtcgctt tcctccgccg ccgccgcctc ctcctccgct ccgcgcggcc     60 

gtctcccacc tctcgcgcct cacctcctcc tcctcctaaa ccctcgctaa accctagcct    120 

cccacccccc caccccaccg gggagcatgc actccaggct cctcagcgcc ccctcccacg    180 

cctccgccgc ctcctcctcc cccctcccct tcgccgccgc ccacccttgc cgcgcctcct    240 

ggacgccgcg gcccacctcc ccgctgccgc ctctatcgct gctgcgcgcc accgcctcca    300 

tgcatccttc tcctcctccc gaggggcata gcgaccagcc tgtgcatggc gtcaccaaca    360 

cggtcgtcgg cgtgctgggg ggaggccagc tggggaagat gctgtgccag gcggccagcc    420 

agatgggggt caggatggcc atacttgatc ccctcgagga ctgcccggcg agctcggttt    480 

gccacgagca tgtcgtcggg agcttcaatg atggcgccac ggttagcgag ttcgcaaaga    540 

ggtgcggggt tttgacggtg gaaattgagc atgtcgacgc tgtaacactc gagaagcttg    600 

agaaacaggg catcgattgt gagcccaaag cctccaccat catgattatt caagacaagt    660 

acaggcagaa gactcatttc tcaaaatttg gaattccgtt acctgacttt gtggaagtag    720 

atactttaag tagcatagag aaagctgggg aaatgtttgg ttatcctcta atggtcaaaa    780 

gcaagagatt agcatatgat ggccgtggaa atgctgttgc tcacgacaaa aaagagctat    840 

cttctgttgt tgcttctctt ggtgggtttg agcatggctt gtatgttgag aggtggacat    900 

cttttgtaaa ggagctttct gtcattgtgg caaggagcag agacggttct acggtgtgct    960 

atcctgttgt tgaaaccatc cacaaggata acatctgcca tgttgttgag gctcctgccg   1020 

aggtgcctga taaaataaag aagttggcta ctaatgtagc tgaaaaggct atcaaatcat   1080 

tggaaggtgc tggtgttttc gctgtagaat tatttttaac acaagataat caggttttat   1140 

tgaatgaagt agctccaagg cctcacaaca gtgggcatca cacaattgag tcatgttata   1200 

cctcgcaata tgagcaacat ttacgtgcta ttcttggcct tcctcttggt gatccttcaa   1260 

tgaaagcgcc tgcatcaata atgtacaaca tcctgggtga ggatgagggt gaggcaggat   1320 

ttactcaagc tcatcagttg attgagagag ctttggacat ttcaggtgca tctgtccatt   1380 

ggtatgcaaa accagaaata cggaagcaga gaaagatggg ccatattaca attgtggggc   1440 

cttcaaagta cagtgtaaaa gcacgcttag ataagttgct gcaaagagac gcatatgacc   1500 

ccaagaaagt tgcagttaaa cctcgtgctg caataataat gggttctgat tctgatcttc   1560 

ctgtcatgaa agatgctgca gtagtattga agaaattcaa catacctttt gagcttacaa   1620 

ttgtttcggc tcatcgtaca ccagagagga tgtaccatta tgcattatct actaaagaaa   1680 

gaggcttaga ggtcataatt gcaggtgcag gtggagcggc tcacttacca gggatggtgg   1740 

cttcattgac ttctgtccca gtaataggag tacccatcat gacttcatct ttacatggaa   1800 

ctgattccct cctatctatt gtccagatgc cgaaaggtat tcctgttgct actgttgcaa   1860 

ttggaaatgc ggaaaatgca ggtttattgg cagttaggat gctggcctca agagatcctg   1920 

agttggggga caaggcaact gaataccagc atgatctgag ggatatggtg ttggagaaag   1980 

caaaaaggct cgaggaacta ggttgggagg aatataccga gctatacttg aagaagcatt   2040 

gatctctgct gcccgtttaa ttgacattct ttttgatcca gtgttgacat aagcagatat   2100 

ggtccatggt tgtggcaaat tcttaaaatc cttttcattc ttttcaaaaa aaaaaaaaaa   2160 

aaa                                                                 2163 

 
           
             8  
             631  
             PRT  
             Oryza sativa  
           
            8 

Met His Ser Arg Leu Leu Ser Ala Pro Ser His Ala Ser Ala Ala Ser 
  1               5                  10                  15 

Ser Ser Pro Leu Pro Phe Ala Ala Ala His Pro Cys Arg Ala Ser Trp 
             20                  25                  30 

Thr Pro Arg Pro Thr Ser Pro Leu Pro Pro Leu Ser Leu Leu Arg Ala 
         35                  40                  45 

Thr Ala Ser Met His Pro Ser Pro Pro Pro Glu Gly His Ser Asp Gln 
     50                  55                  60 

Pro Val His Gly Val Thr Asn Thr Val Val Gly Val Leu Gly Gly Gly 
 65                  70                  75                  80 

Gln Leu Gly Lys Met Leu Cys Gln Ala Ala Ser Gln Met Gly Val Arg 
                 85                  90                  95 

Met Ala Ile Leu Asp Pro Leu Glu Asp Cys Pro Ala Ser Ser Val Cys 
            100                 105                 110 

His Glu His Val Val Gly Ser Phe Asn Asp Gly Ala Thr Val Ser Glu 
        115                 120                 125 

Phe Ala Lys Arg Cys Gly Val Leu Thr Val Glu Ile Glu His Val Asp 
    130                 135                 140 

Ala Val Thr Leu Glu Lys Leu Glu Lys Gln Gly Ile Asp Cys Glu Pro 
145                 150                 155                 160 

Lys Ala Ser Thr Ile Met Ile Ile Gln Asp Lys Tyr Arg Gln Lys Thr 
                165                 170                 175 

His Phe Ser Lys Phe Gly Ile Pro Leu Pro Asp Phe Val Glu Val Asp 
            180                 185                 190 

Thr Leu Ser Ser Ile Glu Lys Ala Gly Glu Met Phe Gly Tyr Pro Leu 
        195                 200                 205 

Met Val Lys Ser Lys Arg Leu Ala Tyr Asp Gly Arg Gly Asn Ala Val 
    210                 215                 220 

Ala His Asp Lys Lys Glu Leu Ser Ser Val Val Ala Ser Leu Gly Gly 
225                 230                 235                 240 

Phe Glu His Gly Leu Tyr Val Glu Arg Trp Thr Ser Phe Val Lys Glu 
                245                 250                 255 

Leu Ser Val Ile Val Ala Arg Ser Arg Asp Gly Ser Thr Val Cys Tyr 
            260                 265                 270 

Pro Val Val Glu Thr Ile His Lys Asp Asn Ile Cys His Val Val Glu 
        275                 280                 285 

Ala Pro Ala Glu Val Pro Asp Lys Ile Lys Lys Leu Ala Thr Asn Val 
    290                 295                 300 

Ala Glu Lys Ala Ile Lys Ser Leu Glu Gly Ala Gly Val Phe Ala Val 
305                 310                 315                 320 

Glu Leu Phe Leu Thr Gln Asp Asn Gln Val Leu Leu Asn Glu Val Ala 
                325                 330                 335 

Pro Arg Pro His Asn Ser Gly His His Thr Ile Glu Ser Cys Tyr Thr 
            340                 345                 350 

Ser Gln Tyr Glu Gln His Leu Arg Ala Ile Leu Gly Leu Pro Leu Gly 
        355                 360                 365 

Asp Pro Ser Met Lys Ala Pro Ala Ser Ile Met Tyr Asn Ile Leu Gly 
    370                 375                 380 

Glu Asp Glu Gly Glu Ala Gly Phe Thr Gln Ala His Gln Leu Ile Glu 
385                 390                 395                 400 

Arg Ala Leu Asp Ile Ser Gly Ala Ser Val His Trp Tyr Ala Lys Pro 
                405                 410                 415 

Glu Ile Arg Lys Gln Arg Lys Met Gly His Ile Thr Ile Val Gly Pro 
            420                 425                 430 

Ser Lys Tyr Ser Val Lys Ala Arg Leu Asp Lys Leu Leu Gln Arg Asp 
        435                 440                 445 

Ala Tyr Asp Pro Lys Lys Val Ala Val Lys Pro Arg Ala Ala Ile Ile 
    450                 455                 460 

Met Gly Ser Asp Ser Asp Leu Pro Val Met Lys Asp Ala Ala Val Val 
465                 470                 475                 480 

Leu Lys Lys Phe Asn Ile Pro Phe Glu Leu Thr Ile Val Ser Ala His 
                485                 490                 495 

Arg Thr Pro Glu Arg Met Tyr His Tyr Ala Leu Ser Thr Lys Glu Arg 
            500                 505                 510 

Gly Leu Glu Val Ile Ile Ala Gly Ala Gly Gly Ala Ala His Leu Pro 
        515                 520                 525 

Gly Met Val Ala Ser Leu Thr Ser Val Pro Val Ile Gly Val Pro Ile 
    530                 535                 540 

Met Thr Ser Ser Leu His Gly Thr Asp Ser Leu Leu Ser Ile Val Gln 
545                 550                 555                 560 

Met Pro Lys Gly Ile Pro Val Ala Thr Val Ala Ile Gly Asn Ala Glu 
                565                 570                 575 

Asn Ala Gly Leu Leu Ala Val Arg Met Leu Ala Ser Arg Asp Pro Glu 
            580                 585                 590 

Leu Gly Asp Lys Ala Thr Glu Tyr Gln His Asp Leu Arg Asp Met Val 
        595                 600                 605 

Leu Glu Lys Ala Lys Arg Leu Glu Glu Leu Gly Trp Glu Glu Tyr Thr 
    610                 615                 620 

Glu Leu Tyr Leu Lys Lys His 
625                 630 

 
           
             9  
             576  
             DNA  
             Glycine max  
           
            9 

gcacgagctg gtgctggtgg tgcagctcac ttgcctggta tggttgctgc ccttactccc     60 

ttgcctgtta ttggcgttcc tgtgcgtgct tctaccttgg atgggattga ttcactcttg    120 

tcaattgtcc agatgccgag aggtgtccct gttgccactg ttgcagttaa taatgcaact    180 

aatgctggat tgctggcagt gaggatgttg ggtgttgcca atgataatct tctgtcaagg    240 

atgagtcaat atcaagaggc ccaaaaggaa agcgtattgg gcaaaggaga taagttagaa    300 

aaacatggct ggaaatccta cttaaacaat agttaattat ccccattaat ttggtgatta    360 

ttttttaggc tcatttctca tttttggtca actacaaaat tttgatagaa aagatttcac    420 

atggagggtg tatagaccca tggtatcaat aagtaggtta agaataattc gagctatatg    480 

ttttgttacc taaatcaaca ggttttctat tttctattgc taacatatag caacagatga    540 

aaattgtgta atctggcaaa aaaaaaaaaa aaaaaa                              576 

 
           
             10  
             111  
             PRT  
             Glycine max  
           
            10 

Ala Arg Ala Gly Ala Gly Gly Ala Ala His Leu Pro Gly Met Val Ala 
  1               5                  10                  15 

Ala Leu Thr Pro Leu Pro Val Ile Gly Val Pro Val Arg Ala Ser Thr 
             20                  25                  30 

Leu Asp Gly Ile Asp Ser Leu Leu Ser Ile Val Gln Met Pro Arg Gly 
         35                  40                  45 

Val Pro Val Ala Thr Val Ala Val Asn Asn Ala Thr Asn Ala Gly Leu 
     50                  55                  60 

Leu Ala Val Arg Met Leu Gly Val Ala Asn Asp Asn Leu Leu Ser Arg 
 65                  70                  75                  80 

Met Ser Gln Tyr Gln Glu Ala Gln Lys Glu Ser Val Leu Gly Lys Gly 
                 85                  90                  95 

Asp Lys Leu Glu Lys His Gly Trp Lys Ser Tyr Leu Asn Asn Ser 
            100                 105                 110 

 
           
             11  
             348  
             DNA  
             Triticum aestivum  
           
            11 

ctttctgtaa ttgtggcaag gtgcagagat ggttgtacag tcggttatcc tgtcattgaa     60 

accattcata aggataacat ctgtcatgtc gttgaagctc ctgctgagat acctgagaaa    120 

atgaagaagt tagctaccaa tgtagctgaa aaaactatca aatcattaga aggtttatcg    180 

aacgaagtag cccctagggc tcacattagt ggacaccata taatccagac atggcactct    240 

tcacaatacg agcaacatct acgtgctatt cttggacttc ctgttggtaa ccctgtaatt    300 

aaagcacccg caaccataat gaacaaaatc ctgggctatg atgaggtg                 348 

 
           
             12  
             116  
             PRT  
             Triticum aestivum  
           
            12 

Leu Ser Val Ile Val Ala Arg Cys Arg Asp Gly Cys Thr Val Gly Tyr 
  1               5                  10                  15 

Pro Val Ile Glu Thr Ile His Lys Asp Asn Ile Cys His Val Val Glu 
             20                  25                  30 

Ala Pro Ala Glu Ile Pro Glu Lys Met Lys Lys Leu Ala Thr Asn Val 
         35                  40                  45 

Ala Glu Lys Thr Ile Lys Ser Leu Glu Gly Leu Ser Asn Glu Val Ala 
     50                  55                  60 

Pro Arg Ala His Ile Ser Gly His His Ile Ile Gln Thr Trp His Ser 
 65                  70                  75                  80 

Ser Gln Tyr Glu Gln His Leu Arg Ala Ile Leu Gly Leu Pro Val Gly 
                 85                  90                  95 

Asn Pro Val Ile Lys Ala Pro Ala Thr Ile Met Asn Lys Ile Leu Gly 
            100                 105                 110 

Tyr Asp Glu Val 
        115 

 
           
             13  
             645  
             PRT  
             Arabidopsis thaliana  
           
            13 

Met Leu Leu Leu Lys Gln Ser Ser Ala Ala Val Leu Val Val Gly Asn 
  1               5                  10                  15 

Thr Thr Pro Val Leu His Thr Ser Arg Ser Thr Tyr Arg Val Gly Pro 
             20                  25                  30 

Phe Pro Val Thr Arg Thr Gln Ser Phe Gln Ser Leu Thr Met Ala Asn 
         35                  40                  45 

Leu Gln Lys Leu Pro Thr Ser Ser Ser Gly Lys Leu Asn Thr Ala Ser 
     50                  55                  60 

Ala Val Pro Cys Ser Ser His Asp Ala Ser Pro Ile Ser Glu Asn Arg 
 65                  70                  75                  80 

Glu Asn Lys His Val His Gly Val Ser Glu Lys Ile Val Gly Val Leu 
                 85                  90                  95 

Gly Gly Gly Gln Leu Gly Arg Met Leu Cys Gln Ala Ala Ser Gln Leu 
            100                 105                 110 

Ala Ile Lys Val Met Ile Leu Asp Pro Ser Lys Asn Cys Ser Ala Ser 
        115                 120                 125 

Ala Leu Ser Tyr Gly His Met Val Asp Ser Phe Asp Asp Ser Ala Thr 
    130                 135                 140 

Val Glu Glu Phe Ala Lys Arg Cys Gly Val Leu Thr Val Glu Ile Glu 
145                 150                 155                 160 

His Val Asp Val Asp Thr Leu Glu Lys Leu Glu Lys Gln Gly Val Asp 
                165                 170                 175 

Cys Gln Pro Lys Ala Ser Thr Ile Arg Ile Ile Gln Asp Lys Tyr Met 
            180                 185                 190 

Gln Lys Val His Phe Ser Gln His Gly Ile Pro Leu Pro Glu Phe Met 
        195                 200                 205 

Glu Ile Ser Asp Ile Glu Gly Ala Arg Lys Ala Gly Glu Leu Phe Gly 
    210                 215                 220 

Tyr Pro Leu Met Ile Lys Ser Lys Arg Leu Ala Tyr Asp Gly Arg Gly 
225                 230                 235                 240 

Asn Ala Val Ala Asn Asn Gln Asp Glu Leu Ser Ser Ala Val Thr Ala 
                245                 250                 255 

Leu Gly Gly Phe Ser Arg Gly Leu Tyr Ile Glu Lys Trp Ala Pro Phe 
            260                 265                 270 

Val Lys Glu Leu Ala Val Ile Val Ala Arg Gly Arg Asp Gly Ser Met 
        275                 280                 285 

Val Cys Tyr Pro Val Val Glu Thr Ile His Arg Asp Asn Ile Cys His 
    290                 295                 300 

Ile Val Lys Ala Pro Ala Asp Val Pro Trp Lys Ile Asn Lys Leu Ala 
305                 310                 315                 320 

Thr Asp Val Ala Gln Lys Ala Val Gly Ser Leu Glu Gly Ala Gly Val 
                325                 330                 335 

Phe Ala Val Glu Leu Phe Leu Thr Glu Asp Ser Gln Ile Leu Leu Asn 
            340                 345                 350 

Glu Val Ala Pro Arg Pro His Asn Ser Gly His Gln Thr Ile Glu Cys 
        355                 360                 365 

Cys Tyr Thr Ser Gln Phe Glu Gln His Leu Arg Ala Val Val Gly Leu 
    370                 375                 380 

Pro Leu Gly Asp Pro Ser Met Arg Thr Pro Ala Ser Ile Met Tyr Asn 
385                 390                 395                 400 

Ile Leu Gly Glu Asp Asp Val Ile Asp Gly Glu Ala Gly Phe Lys Leu 
                405                 410                 415 

Ala His Arg Leu Ile Ala Arg Ala Leu Cys Ile Pro Gly Ala Ser Val 
            420                 425                 430 

His Trp Tyr Asp Lys Pro Glu Met Arg Lys Gln Arg Lys Met Gly His 
        435                 440                 445 

Ile Thr Leu Val Gly Gln Ser Met Gly Ile Leu Glu Gln Arg Leu Gln 
    450                 455                 460 

Cys Ile Leu Ser Glu Gln Ser His Gln Val His Glu Thr Pro Arg Val 
465                 470                 475                 480 

Ala Ile Ile Met Gly Ser Asp Thr Asp Leu Pro Val Met Lys Asp Ala 
                485                 490                 495 

Ala Lys Ile Leu Asp Leu Phe Gly Val Thr His Glu Val Lys Ile Val 
            500                 505                 510 

Ser Ala His Arg Thr Pro Glu Met Met Tyr Thr Tyr Ala Thr Ser Ala 
        515                 520                 525 

His Ser Arg Gly Val Gln Val Ile Ile Ala Gly Ala Gly Gly Ala Ala 
    530                 535                 540 

His Leu Pro Gly Met Val Ala Ser Leu Thr Pro Leu Pro Val Ile Gly 
545                 550                 555                 560 

Val Pro Val Arg Ala Thr Arg Leu Asp Gly Val Asp Ser Leu Leu Ser 
                565                 570                 575 

Ile Val Gln Met Pro Arg Gly Val Pro Val Ala Thr Val Ala Ile Asn 
            580                 585                 590 

Asn Ala Thr Asn Ala Ala Leu Leu Ala Val Arg Met Leu Gly Ile Ser 
        595                 600                 605 

Asp Thr Asp Leu Val Ser Arg Met Arg Gln Tyr Gln Glu Asp Met Arg 
    610                 615                 620 

Asp Glu Asn Leu Asn Lys Gly Glu Lys Leu Glu Thr Glu Gly Trp Glu 
625                 630                 635                 640 

Ser Tyr Leu Asn Gln 
                645 

 
           
             14  
             623  
             PRT  
             Nicotiana tabacum  
           
            14 

Gly Pro Gln Arg Ser Ser Phe Ala Ser Pro Ile Leu Ala Val Asn Pro 
  1               5                  10                  15 

Gln Lys Ser Ile Ser Phe Leu Lys Asn His Ser Phe Val Phe Ser Ser 
             20                  25                  30 

Ser Leu Met Arg Gln Gln Ser Glu His Thr Pro Thr Met Leu Ser Cys 
         35                  40                  45 

Lys Ala Ser Leu Glu Val Val Thr Asp Ser Pro Gly Gly Leu Glu Val 
     50                  55                  60 

His Gly Ile Ser Glu Met Val Val Gly Val Leu Gly Gly Gly Gln Leu 
 65                  70                  75                  80 

Gly Arg Met Leu Cys Glu Ala Ala Ser Gln Met Ala Ile Lys Val Ile 
                 85                  90                  95 

Val Leu Asp Pro Met Asn Asn Cys Pro Ala Ser Ala Leu Ala His Gln 
            100                 105                 110 

His Val Val Gly Ser Tyr Asp Asp Ser Ala Thr Val Glu Glu Phe Gly 
        115                 120                 125 

Lys Arg Cys Gly Val Leu Thr Val Glu Ile Glu His Val Asp Val Val 
    130                 135                 140 

Thr Leu Glu Lys Leu Glu Gln Gln Gly Val Asp Cys Gln Pro Lys Ala 
145                 150                 155                 160 

Ser Thr Ile Arg Ile Ile Gln Asp Lys Tyr Leu Gln Lys Val His Phe 
                165                 170                 175 

Ser Arg His Ala Ile Pro Leu Pro Lys Phe Met Gln Ile Asp Asp Leu 
            180                 185                 190 

Glu Ser Ala Arg Arg Ala Gly Asp Leu Phe Gly Tyr Pro Leu Met Ile 
        195                 200                 205 

Lys Ser Arg Arg Leu Ala Tyr Asp Gly Arg Gly Asn Ala Val Ala Lys 
    210                 215                 220 

Ser Glu Glu Glu Leu Ser Ser Ala Val Asn Ala Leu Gly Gly Tyr Gly 
225                 230                 235                 240 

Arg Gly Leu Tyr Val Glu Lys Trp Ala Pro Phe Val Lys Glu Leu Ser 
                245                 250                 255 

Val Ile Val Pro Arg Gly Arg Asp Gly Ser Ile Ala Cys Tyr Pro Ala 
            260                 265                 270 

Val Glu Thr Ile His Arg Asp Asn Ile Cys His Ile Val Lys Ser Pro 
        275                 280                 285 

Ala Asn Val Ser Trp Lys Ser Met Lys Leu Ala Thr Asp Val Ala His 
    290                 295                 300 

Arg Ala Val Ser Ser Leu Glu Gly Ala Gly Val Phe Ala Val Glu Leu 
305                 310                 315                 320 

Phe Leu Thr Glu Asp Gly Gln Ile Leu Leu Asn Glu Val Ala Pro Arg 
                325                 330                 335 

Pro His Asn Ser Gly His His Thr Ile Glu Ala Cys Phe Thr Ser Gln 
            340                 345                 350 

Phe Glu Gln His Leu Arg Ala Val Val Gly Leu Pro Leu Gly Asp Pro 
        355                 360                 365 

Ser Met Lys Thr Pro Ala Ala Val Met Tyr Asn Ile Leu Gly Glu Asp 
    370                 375                 380 

Asp Gly Glu Pro Gly Phe Leu Leu Ala Asn Gln Leu Ile Glu Lys Ala 
385                 390                 395                 400 

Leu Gly Ile Pro Gly Val Ser Val His Trp Tyr Asp Lys Pro Glu Met 
                405                 410                 415 

Arg Arg Gln Arg Lys Met Gly His Ile Thr Ile Val Gly Pro Ser Met 
            420                 425                 430 

Gly Ile Val Glu Ala Gln Leu Arg Val Ile Leu Asn Glu Glu Ser Val 
        435                 440                 445 

Asn Gly His Pro Ala Val Ala Pro Arg Val Gly Ile Ile Met Gly Ser 
    450                 455                 460 

Asp Ser Asp Leu Pro Val Met Lys Asp Ala Ala Lys Ile Leu Asn Glu 
465                 470                 475                 480 

Phe Asp Val Pro Ala Glu Val Lys Ile Val Ser Ala His Arg Thr Pro 
                485                 490                 495 

Glu Met Met Phe Ser Tyr Ala Leu Ser Ala Arg Glu Arg Gly Ile Gln 
            500                 505                 510 

Val Ile Ile Ala Gly Ala Gly Gly Ala Ala His Leu Pro Gly Met Val 
        515                 520                 525 

Ala Ala Phe Thr Pro Leu Pro Val Ile Gly Val Pro Val Arg Ala Ser 
    530                 535                 540 

Thr Leu Asp Gly Leu Asp Ser Leu Leu Ser Ile Val Gln Met Pro Arg 
545                 550                 555                 560 

Gly Val Pro Val Ala Thr Val Ala Ile Asn Asn Ala Thr Asn Ala Gly 
                565                 570                 575 

Leu Leu Ala Val Arg Leu Leu Gly Ile Ser Asp Ile Lys Leu Gln Ala 
            580                 585                 590 

Arg Met Ala Gln Tyr Gln Glu Asp Arg Arg Asp Glu Val Leu Val Lys 
        595                 600                 605 

Gly Glu Arg Leu Glu Lys Ile Gly Phe Glu Glu Tyr Leu Asn Ser 
    610                 615                 620 

 
           
             15  
             557  
             PRT  
             Vigna aconitifolia  
           
            15 

Gly Leu Tyr Glu Val Val Val Gly Val Leu Gly Gly Gly Gln Leu Gly 
  1               5                  10                  15 

Arg Met Met Cys Gln Ala Ala Ser Gln Met Ala Ile Lys Val Met Val 
             20                  25                  30 

Leu Asp Pro Gln Glu Asn Cys Pro Ala Ser Ser Leu Ser Tyr His His 
         35                  40                  45 

Met Val Gly Ser Phe Asp Glu Ser Thr Lys Val Glu Glu Phe Ala Lys 
     50                  55                  60 

Arg Cys Gly Val Leu Thr Val Glu Ile Glu His Val Asp Val Asp Thr 
 65                  70                  75                  80 

Leu Glu Lys Leu Glu Lys Gln Gly Val Asp Cys Gln Pro Lys Ala Ser 
                 85                  90                  95 

Thr Val Arg Ile Ile Gln Asp Lys Tyr Gln Gln Lys Val Ala Leu Leu 
            100                 105                 110 

Pro Ala Trp Ile Pro Leu Pro Glu Phe Met Lys Ile Asp Asp Leu Lys 
        115                 120                 125 

Ala Lys Lys Trp Asp Ser Leu Asp Ile His Phe Met Ile Lys Ser Arg 
    130                 135                 140 

Arg Leu Ala Tyr Asp Gly Arg Gly Asn Phe Val Ala Lys Ser Glu Glu 
145                 150                 155                 160 

Glu Leu Ser Ser Ala Val Asp Ala Leu Gly Gly Phe Asp Arg Gly Leu 
                165                 170                 175 

Tyr Ala Glu Lys Trp Ala Pro Phe Val Lys Glu Leu Ala Val Ile Val 
            180                 185                 190 

Ala Arg Gly Arg Asp Asn Ser Ile Ser Cys Tyr Pro Val Val Glu Leu 
        195                 200                 205 

Phe Thr Gly His Ile Cys His Ile Val Lys Ser Pro Ala Asn Val Asn 
    210                 215                 220 

Trp Lys Thr Arg Glu Leu Ala Ile Glu Val Ala Phe Asn Ala Val Lys 
225                 230                 235                 240 

Ser Leu Glu Val Pro Gly Val Phe Ala Val Glu Leu Phe Leu Thr Lys 
                245                 250                 255 

Glu Gly Glu Ile Leu Leu Asn Glu Val Ala Pro Arg Pro His Asn Ser 
            260                 265                 270 

Gly His His Thr Ile Glu Ser Cys His Thr Ser Gln Phe Glu Gln His 
        275                 280                 285 

Leu Pro Ala Val Val Gly Leu Pro Leu Gly Asp Pro Ser Met Lys Thr 
    290                 295                 300 

Pro Ala Ala Ile Met Tyr Asn Ile Leu Gly Glu Glu Glu Gly Glu His 
305                 310                 315                 320 

Gly Phe Gln Leu Ala His Gln Leu Met Lys Arg Ala Met Thr Ile Pro 
                325                 330                 335 

Gly Ala Ser Val His Trp Tyr Asp Lys Pro Glu Met Arg Lys Gln Arg 
            340                 345                 350 

Lys Met Cys His Ile Thr Ile Val Gly Ser Ser Leu Ser Ser Ile Glu 
        355                 360                 365 

Ser Asn Leu Ala Ile Leu Leu Glu Gly Lys Gly Leu His Asp Lys Thr 
    370                 375                 380 

Ala Val Cys Ser Thr Leu Leu Gly Phe Ile Met Gly Ser Asp Ser Asp 
385                 390                 395                 400 

Leu Pro Val Met Lys Ser Ala Ala Glu Met Met Glu Met Phe Gly Val 
                405                 410                 415 

Pro His Glu Val Arg Ile Val Ser Ala His Arg Thr Pro Glu Leu Met 
            420                 425                 430 

Phe Cys Tyr Ala Ser Ser Ala His Glu Arg Gly Tyr Gln Val Ile Ile 
        435                 440                 445 

Ala Gly Ala Gly Gly Ala Ala His Leu Pro Gly Met Val Ala Ser Leu 
    450                 455                 460 

Thr Pro Leu Pro Val Val Gly Val Pro Val Arg Ala Ser Thr Leu Asp 
465                 470                 475                 480 

Gly Leu Asp Ser Leu Leu Ser Ile Val Gln Met Pro Arg Gly Val Pro 
                485                 490                 495 

Val Ala Thr Val Ala Val Asn Asn Ala Thr Asn Ala Gly Leu Leu Ala 
            500                 505                 510 

Val Arg Met Leu Gly Val Ala Asn Asp Asn Leu Leu Ser Arg Met Ser 
        515                 520                 525 

Gln Tyr Gln Glu Asp Gln Lys Glu Ala Val Leu Arg Glu Gly Asp Lys 
    530                 535                 540 

Leu Glu Lys His Gly Trp Glu Ser Tyr Leu Lys Asn Ser 
545                 550                 555 

 
           
             16  
             2329  
             DNA  
             Brassica  
           
            16 

gtgagttaat gggttttgtt tccctcaaat aaaaatcgaa ccttcgaagc ttaagttctc     60 

gcatcctctt tctccttgtc aagccacagc tcgaaaccag ctatgttcta gatgttgctt    120 

ctgaaacaga gctcagctgc tgttcttgtc tctgggaatc caagtcctgt cctttacact    180 

cctcgcttca cttccagagt tggatctctt ccagttagca aaacgaactc cttcaccatg    240 

gcgaatcttc agaagggtct tacttattct tcttctgaga aattcaaccc ggtgttagcg    300 

tgtagctctc acgaggcttc tcctatcagc gaggatacac atatcaaggg agtctctgag    360 

atcattgtgg gagtgttggg aggtggacag ttaggtcgca tgctttgcca agctgcttct    420 

caaatggcca tcaaggttat gattctagat ccttcaaaga actgttcagc aagctcatta    480 

gcttatggcc acatggttga tagctttgac gacagtgcta cagttgaaga gtttgcaaaa    540 

agatgtggag tcttgacagt agaaattgaa catgttgacg ttgaaacact agagaagctt    600 

gagaaacaag gagtagatgt ccaaccaaaa gcctctacta tcaggataat acaggataaa    660 

tacatacaaa aagttcattt ctctcggcat ggcatcccac ttccagagtt tatggagata    720 

agcgatattg aaggagctga aagagcaggt gaactttttg gttaccctct tatgatcaag    780 

agcaagagat tagcttatga tggacgagga aatgcagttg ctaatagcca agacgcgctt    840 

acttctgctg taactgctct tggaggtttc agtcgtggtt tgtacgttga gaaatgggca    900 

ccctttgtaa aggagttggc tgttattgtg gctaggggaa gagatggttc catggtttgt    960 

tatccagttg ttgaaactgt tcacagggat aacatatgcc atatagttaa agcaccagca   1020 

gatgtgcctt ggaagattaa caaacttgcc actgatgttg ctcaaaaggc tgttggttct   1080 

ttagaaggcg ctggtgtttt cgctgttgag ctgttcttga cagaggatgg tcagatcctg   1140 

ctgaatgaag ttgcacctag accacacaac agtggacatc agacgatcga gtcatgttac   1200 

acttcacagt ttgagcaaca cttgcgagct gtggttggtc ttccactcgg tgatccgtct   1260 

atgagaactc ctgcctccat tatgtacaat attctgggcg aagatgatgg agaagctggt   1320 

ttcagattgg cacatcggct cattgcaaga gctctgagtg ttccaggtgc atctgtgcat   1380 

tggtatgaca agccagaaat gagaaagcag cggaagatgg ggcacatcac tcttgttgga   1440 

aagtctattg gtgttttgga acaaaggttg cattgtatat taagtgaaca aagccatcaa   1500 

ctacatgaca ttgcagagat acctcgtgtt ggtatcatca tgggttcaga ctctgatctt   1560 

cctgttatga aagatgctgc aaaaattctt gacacgttta atgtaacata tgaggtgaag   1620 

atagtttcag cacatcggac accagagatg atgttttctt atgcaacatc agctcatagt   1680 

agaggaatcc aagtgataat tgcaggtgct ggtggtgctg ctcacttacc aggtatggtt   1740 

gcttcactca ctcccttacc tgtgattggt gtccctgtac gtgctacccg tttggatgga   1800 

gttgattcac ttctctccat tgttcagatg cctagaggtg ttcctgtagc cacagttgct   1860 

ataaacaact ccaccaacgc agccttgctt gctatcagga tgctggggat ctctgatact   1920 

gatctcgtct caaggataag tcagtaccag gaagacatga gagaagagaa catggttaaa   1980 

ggtgagaaac ttgagcgtca aggttgggaa tcatacttga accagtgaat attacttgtc   2040 

agaactcctc attggttacc gggaactcca tagtgatcga ttttgctgcc agtaagatct   2100 

tgttgagagt tagtttcgga tcttgagttg ttgtcacggc taattccggt ttagctttgt   2160 

caagttaagc cacaattgtt tgagactcaa gctaagtcct ggtgagggat ctgatagtta   2220 

gcttcagttc ttgagaaact attggatgag ttatccacac gatgttttat tttataatat   2280 

tatgtaactc ccatagagac atgggatata acatgttctt acaaaaaaa               2329 

 
           
             17  
             638  
             PRT  
             Brassica  
           
            17 

Met Leu Leu Leu Lys Gln Ser Ser Ala Ala Val Leu Val Ser Gly Asn 
  1               5                  10                  15 

Pro Ser Pro Val Leu Tyr Thr Pro Arg Phe Thr Ser Arg Val Gly Ser 
             20                  25                  30 

Leu Pro Val Ser Lys Thr Asn Ser Phe Thr Met Ala Asn Leu Gln Lys 
         35                  40                  45 

Gly Leu Thr Tyr Ser Ser Ser Glu Lys Phe Asn Pro Val Leu Ala Cys 
     50                  55                  60 

Ser Ser His Glu Ala Ser Pro Ile Ser Glu Asp Thr His Ile Lys Gly 
 65                  70                  75                  80 

Val Ser Glu Ile Ile Val Gly Val Leu Gly Gly Gly Gln Leu Gly Arg 
                 85                  90                  95 

Met Leu Cys Gln Ala Ala Ser Gln Met Ala Ile Lys Val Met Ile Leu 
            100                 105                 110 

Asp Pro Ser Lys Asn Cys Ser Ala Ser Ser Leu Ala Tyr Gly His Met 
        115                 120                 125 

Val Asp Ser Phe Asp Asp Ser Ala Thr Val Glu Glu Phe Ala Lys Arg 
    130                 135                 140 

Cys Gly Val Leu Thr Val Glu Ile Glu His Val Asp Val Glu Thr Leu 
145                 150                 155                 160 

Glu Lys Leu Glu Lys Gln Gly Val Asp Val Gln Pro Lys Ala Ser Thr 
                165                 170                 175 

Ile Arg Ile Ile Gln Asp Lys Tyr Ile Gln Lys Val His Phe Ser Arg 
            180                 185                 190 

His Gly Ile Pro Leu Pro Glu Phe Met Glu Ile Ser Asp Ile Glu Gly 
        195                 200                 205 

Ala Glu Arg Ala Gly Glu Leu Phe Gly Tyr Pro Leu Met Ile Lys Ser 
    210                 215                 220 

Lys Arg Leu Ala Tyr Asp Gly Arg Gly Asn Ala Val Ala Asn Ser Gln 
225                 230                 235                 240 

Asp Ala Leu Thr Ser Ala Val Thr Ala Leu Gly Gly Phe Ser Arg Gly 
                245                 250                 255 

Leu Tyr Val Glu Lys Trp Ala Pro Phe Val Lys Glu Leu Ala Val Ile 
            260                 265                 270 

Val Ala Arg Gly Arg Asp Gly Ser Met Val Cys Tyr Pro Val Val Glu 
        275                 280                 285 

Thr Val His Arg Asp Asn Ile Cys His Ile Val Lys Ala Pro Ala Asp 
    290                 295                 300 

Val Pro Trp Lys Ile Asn Lys Leu Ala Thr Asp Val Ala Gln Lys Ala 
305                 310                 315                 320 

Val Gly Ser Leu Glu Gly Ala Gly Val Phe Ala Val Glu Leu Phe Leu 
                325                 330                 335 

Thr Glu Asp Gly Gln Ile Leu Leu Asn Glu Val Ala Pro Arg Pro His 
            340                 345                 350 

Asn Ser Gly His Gln Thr Ile Glu Ser Cys Tyr Thr Ser Gln Phe Glu 
        355                 360                 365 

Gln His Leu Arg Ala Val Val Gly Leu Pro Leu Gly Asp Pro Ser Met 
    370                 375                 380 

Arg Thr Pro Ala Ser Ile Met Tyr Asn Ile Leu Gly Glu Asp Asp Gly 
385                 390                 395                 400 

Glu Ala Gly Phe Arg Leu Ala His Arg Leu Ile Ala Arg Ala Leu Ser 
                405                 410                 415 

Val Pro Gly Ala Ser Val His Trp Tyr Asp Lys Pro Glu Met Arg Lys 
            420                 425                 430 

Gln Arg Lys Met Gly His Ile Thr Leu Val Gly Lys Ser Ile Gly Val 
        435                 440                 445 

Leu Glu Gln Arg Leu His Cys Ile Leu Ser Glu Gln Ser His Gln Leu 
    450                 455                 460 

His Asp Ile Ala Glu Ile Pro Arg Val Gly Ile Ile Met Gly Ser Asp 
465                 470                 475                 480 

Ser Asp Leu Pro Val Met Lys Asp Ala Ala Lys Ile Leu Asp Thr Phe 
                485                 490                 495 

Asn Val Thr Tyr Glu Val Lys Ile Val Ser Ala His Arg Thr Pro Glu 
            500                 505                 510 

Met Met Phe Ser Tyr Ala Thr Ser Ala His Ser Arg Gly Ile Gln Val 
        515                 520                 525 

Ile Ile Ala Gly Ala Gly Gly Ala Ala His Leu Pro Gly Met Val Ala 
    530                 535                 540 

Ser Leu Thr Pro Leu Pro Val Ile Gly Val Pro Val Arg Ala Thr Arg 
545                 550                 555                 560 

Leu Asp Gly Val Asp Ser Leu Leu Ser Ile Val Gln Met Pro Arg Gly 
                565                 570                 575 

Val Pro Val Ala Thr Val Ala Ile Asn Asn Ser Thr Asn Ala Ala Leu 
            580                 585                 590 

Leu Ala Ile Arg Met Leu Gly Ile Ser Asp Thr Asp Leu Val Ser Arg 
        595                 600                 605 

Ile Ser Gln Tyr Gln Glu Asp Met Arg Glu Glu Asn Met Val Lys Gly 
    610                 615                 620 

Glu Lys Leu Glu Arg Gln Gly Trp Glu Ser Tyr Leu Asn Gln 
625                 630                 635