Peptide Deformylase in Staphylococcus aureus : Resistance to Inhibition Is Mediated by Mutations in the Formyltransferase Gene (original) (raw)

2000, Antimicrobial Agents and Chemotherapy

Abstract

Peptide deformylase, a bacterial enzyme, represents a novel target for antibiotic discovery. Two deformylase homologs, defA and defB , were identified in Staphylococcus aureus . The defA homolog, located upstream of the transformylase gene, was identified by genomic analysis and was cloned from chromosomal DNA by PCR. A distinct homolog, defB , was cloned from an S. aureus genomic library by complementation of the arabinose-dependent phenotype of a P BAD -def Escherichia coli strain grown under arabinose-limiting conditions. Overexpression in E. coli of defB , but not defA , correlated to increased deformylase activity and decreased susceptibility to actinonin, a deformylase-specific inhibitor. The defB gene could not be disrupted in wild-type S. aureus , suggesting that this gene, which encodes a functional deformylase, is essential. In contrast, the defA gene could be inactivated; the function of this gene is unknown. Actinonin-resistant mutants grew slowly in vitro and did not sh...

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References (33)

  1. Adams, J. M. 1968. On the release of the formyl group from nascent protein. J. Mol. Biol. 33:571-589.
  2. Altschul, S. F., T. L. Madden, A. A. Scha ¨ffer, J. Zhang, Z. Zhang, W. Miller, and D. J. Lipman. 1997. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25:3389-3402.
  3. Becker, A., I. Schlichting, W. Kabsch, D. Groche, S. Schultz, and A. F. Wagner. 1998. Iron center, substrate recognition and mechanism of peptide deformylase. Nat. Struct. Biol. 5:1053-1058.
  4. Becker, A., I. Schlichting, W. Kabsch, S. Schultz, and A. F. Wagner. 1998. Structure of peptide deformylase and identification of the substrate binding site. J. Biol. Chem. 273:11413-11416.
  5. Bianchetti, R., G. Lucchini, P. Crosti, and P. Tortora. 1977. Dependence of mitochondrial protein synthesis initiation on formylation of the initiator methionyl-tRNAf. J. Biol. Chem. 252:2519-2523.
  6. Breidt, F. J., and G. C. Stewart. 1987. Nucleotide and deduced amino acid sequences of the Staphylococcus aureus phospho-beta-galactosidase gene. Appl. Environ. Microbiol. 53:969-973.
  7. Bru ¨ckner, R. 1997. Gene replacement in Staphylococcus carnosus and Staph- ylococcus xylosus. FEMS Microbiol. Lett. 151:1-8.
  8. Chen, D. Z., P. Patel, C. J. Hackbarth, W. Wang, G. Dreyer, D. C. Young, P. S. Margolis, C. Wu, Z.-J. Ni, J. Trias, R. J. White, and Z. Yuan. 2000. Actinonin, a naturally occurring antibacterial agent, is a potent deformylase inhibitor. Biochemistry 39:1256-1262.
  9. Dardel, F., S. Ragusa, C. Lazennec, S. Blanquet, and T. Meinnel. 1998. Solution structure of nickel-peptide deformylase. J. Mol. Biol. 280:501-513.
  10. Ford, C. W., J. C. Hamel, D. Stapert, and R. J. Yancey. 1989. Establishment of an experimental model of a Staphylococcus aureus abscess in mice by use of dextran and gelatin microcarriers. J. Med. Microbiol. 28:259-266.
  11. Guillon, J. M., Y. Mechulam, J. M. Schmitter, S. Blanquet, and G. Fayat. 1992. Disruption of the gene for Met-tRNA(fMet) formyltransferase se- verely impairs growth of Escherichia coli. J. Bacteriol. 174:4294-4301.
  12. Kennedy, S., and H. F. Chambers. 1989. Daptomycin (LY146032) for pre- vention and treatment of experimental aortic valve endocarditis in rabbits. Antimicrob. Agents Chemother. 33:1522-1525.
  13. Kozak, M. 1999. Initiation of translation in prokaryotes and eukaryotes. Gene 234:187-208.
  14. Kreiswirth, B. N., S. Lo ¨fdahl, M. J. Betley, M. O'Reilly, P. M. Schlievert, M. S. Bergdoll, and R. P. Novick. 1983. The toxic shock syndrome exotoxin structural gene is not detectably transmitted by a prophage. Nature 305:709- 712.
  15. Lazennec, C., and T. Meinnel. 1997. Formate dehydrogenase-coupled spec- trophotometric assay of peptide deformylase. Anal. Biochem. 244:180-182.
  16. Link, A. J., D. Phillips, and G. M. Church. 1997. Methods for generating precise deletions and insertions in the genome of wild-type Escherichia coli: application to open reading frame characterization. J. Bacteriol. 179:6228- 6237.
  17. Mahler, H. R., K. Dawidowicz, and F. Feldman. 1972. Formate as a specific label for mitochondrial translational products. J. Biol. Chem. 247:7439-7442.
  18. Mazel, D., E. Cou ¨c, S. Blanchard, W. Saurin, and P. Marlie `re. 1997. A survey of polypeptide deformylase function throughout the eubacterial lin- eage. J. Mol. Biol. 266:939-949.
  19. Mazel, D., S. Pochet, and P. Marliere. 1994. Genetic characterization of polypeptide deformylase, a distinctive enzyme of eubacterial translation. EMBO J. 13:914-923.
  20. Meinnel, T., and S. Blanquet. 1994. Characterization of the Thermus ther- mophilus locus encoding peptide deformylase and methionyl-tRNA(fMet) formyltransferase. J. Bacteriol. 176:7387-7390.
  21. Meinnel, T., and S. Blanquet. 1993. Evidence that peptide deformylase and methionyl-tRNA (fMet) formyltransferase are encoded within the same operon in Escherichia coli. J. Bacteriol. 175:7737-7740.
  22. Meinnel, T., S. Blanquet, and F. Dardel. 1996. A new subclass of the zinc metalloproteases superfamily revealed by the solution structure of peptide deformylase. J. Mol. Biol. 262:375-386.
  23. Meinnel, T., C. Lazennec, and S. Blanquet. 1995. Mapping of the active site zinc ligands of peptide deformylase. J. Mol. Biol. 254:175-183.
  24. Meinnel, T., C. Lazennec, S. Villoing, and S. Blanquet. 1997. Structure- function relationships within the peptide deformylase family. Evidence for a conserved architecture of the active site involving three conserved motifs and a metal ion. J. Mol. Biol. 267:749-761.
  25. National Committee for Clinical Laboratory Standards. 1997. Methods for dilution antimicrobial tests for bacteria that grow aerobically. Approved standard M7-A4, 4th ed. National Committee for Clinical Laboratory Stan- dards, Wayne, Pa.
  26. Newton, D. T., C. Creuzenet, and D. Mangroo. 1999. Formylation is not essential for initiation of protein synthesis in all eubacteria. J. Biol. Chem. 274:22143-22146.
  27. Niemeyer, D. M., M. J. Pucci, J. A. Thanassi, V. K. Sharma, and G. L. Archer. 1996. Role of mecA transcriptional regulation in the phenotypic expression of methicillin resistance in Staphylococcus aureus. J. Bacteriol. 178:5464-5471.
  28. Novick, R. P. 1991. Genetic systems in staphylococci. Methods Enzymol. 204:587-636.
  29. Ochman, H., A. S. Gerber, and D. L. Hartl. 1988. Genetic applications of an inverse polymerase chain reaction. Genetics 120:621-623.
  30. Rajagopalan, P. T., A. Datta, and D. Pei. 1997. Purification, characterization, and inhibition of peptide deformylase from Escherichia coli. Biochemistry 36:13910-13918.
  31. Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, Plainview, N.Y.
  32. Thomas, W. D. J., and G. L. Archer. 1989. Identification and cloning of the conjugative transfer region of Staphylococcus aureus plasmid pGO1. J. Bac- teriol. 171:684-691.
  33. Wada, A., and H. Watanabe. 1998. Penicillin-binding protein 1 of Staphylo- coccus aureus is essential for growth. J. Bacteriol. 180:2759-2765.