Cloning, sequencing and expression of a Pasteurella haemolytica A1 gene encoding a PurK-like protein.
A membrane protein antigen of Pasteurella haemolytica A1 encoded on the recombinant plasmid pYFC13 is isolated and characterized. Nucleotide sequence analysis of the insert DNA in pYFC13 identified the gene mpa1, which codes a protein of approximately 45 kDa without signal sequence. The deduced amino acids from the DNA sequence are homologous to Bacillus subtilis PurK by 29.4%; to Schizosaccharomyces pombe Pur6 by 29.34%, to Saccharomyces cerevisiae Pur6 by 25.867%; and to E. coli PurK by 25.223% identity, respectively. The purK and pur6 from these organisms are responsible for the activity of 5'-phosphoribosyl- 5-amino-4-imidazole carboxylase which is involved in de novo purine biosynthesis. The protein was over-expressed in E. coli by its own promoter. The antigen we designated as Mpa1, could be localized to the cytoplasmic membrane of both P. haemolytica A1 and E. coli TB1 harbored pYFC13. The Mpa1 was antigenic in rabbit and in cattle since both animals produced antibody against this protein.
Chang
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YF
Diagnostic Laboratory, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853.
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Young
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Struck
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D002262:Carboxy-Lyases; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D016978:Mannheimia haemolytica / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D014166:Transferases
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D014166:Transferases; C084340:Mpa1 protein, Pasteurella haemolytica; D002262:Carboxy-Lyases; C075015:purK protein, E coli