Skip to main content
Log in

Rapid screening for putative exported proteins from staphylococcus aureus using alkaline phosphatase as a reporter molecule

  • Research
  • Published:
Molecular Biotechnology Aims and scope Submit manuscript

Abstract

Staphylococcus aureus causes a wide range of infections in humans, ranging from superficial skin infections to the more serious toxin-mediated diseases such as toxic shock syndrome. Owing to the increasing resistance of this bacterium to a wide range of antibiotics, the need to determine the virulence factors involved in infection is becoming more important as these molecules are potential therapeutic targets. In this study, we have screened for putative exported proteins from S. aureus on the basis that these proteins are likely to be the first point of contact between the bacterium and host during infection. We have constructed gene fusions between S. aureus DNA and a truncated version of the Escherichia coli phoA gene, and we report on the characterization of the recombinants exhibiting alkaline phosphatase activity. As well as known S. aureus proteins, we have identified a number of putative open reading frames that encode proteins similar to those from nonstaphylococcal species and also unique proteins that do not have any homologues on the current databases.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Archer, G.L. and Crossley, K.B. (1997) The Staphylococci in Human Disease, Churchill. Livingstone, NY.

    Google Scholar 

  2. Hiramatsu K., Hanaki, H., Ino, T., Yabuta, K., Oguri, T., and Tenover, F.C. (1997) Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J. Antimicrob. Chemother. 40, 135–136.

    Article  PubMed  CAS  Google Scholar 

  3. Bohach, G.A., Dinges, M.M., Mitchell, D.T., Ohlendorf, D.H., and Schlievert, P.M. (1997) Exotoxins, in The Staphylococci in Human Disease (Crossley, K.B. and Archer, G.L., eds.), Churchill Livingstone, NY, pp. 83–111.

    Google Scholar 

  4. Faith, J.M., Jonsson, H., Guss, B., Swilalski, L.M., Wiberg, K., Lindberg, M., and Hook, M. (1992) Molecular characterization and expression of a gene encoding a Staphylococcus aureus collagen adhesin. J. Biol. Chem. 267, 4766–4772.

    Google Scholar 

  5. McDevitt, D., Francois, P., Vaudaux, P., and Foster, T.J. (1994). Molecular characterization of the clumping factor (fibrinogen receptor) of Staphylococcus aureus. Mol. Microbiol. 11, 237–248.

    Article  PubMed  CAS  Google Scholar 

  6. Nucifora, G., Chu, L., Misra, T.K., and Silver, S. (1989) Cadmium resistance from Staphylococcus aureus plasmid pI258 cadA gene results from a cadmium-efflux ATPase. Proc. Natl. Acad. Sci. USA 86, 3544–3548.

    Article  PubMed  CAS  Google Scholar 

  7. Nair, S., Song, Y., Meghji, S., Reddi, K., Harris, M., Ross, A., Pool, S., Wilson, M., and Henderson, B. (1995) Surface-associated proteins from Staphylococcus aureus demonstrate potent bone resorbing activity. J. Bone and Miner. Res. 10, 726–734.

    Article  CAS  Google Scholar 

  8. Meghji, S., Crean, S.J., Hill, P.A., Sheikh, M., Nair, S.P., Heron, K., Henderson, B., Mawer, E.B., and Harris, M. (1998) Surface-associated protein from Staphylococcus aureus stimulates osteoclastogenesis: Possible role in S. aureus-induced bone pathology. Br. J. Rheumatol. 37, 1095–1101.

    Article  PubMed  CAS  Google Scholar 

  9. Arora, M., Shah, N., Meghji, S., Henderson, B., Harris, M., Nair, S., Wilson, M., Gray, C.M., Jones, S.J., and Boyde, A. (1998) Effect of Staphylococcus aureus extracellular proteinaceous fraction in an isolated osteoclastic resorption assay. J. Bone Miner. Metab. 16, 158–161.

    Article  CAS  Google Scholar 

  10. Manoil, C., Makalanos, J.J., and Beckwith, J. (1990) Alkaline phosphatase fusions: sensors of subcellular location. J. Bacteriol. 172, 515–518.

    PubMed  CAS  Google Scholar 

  11. Calamia, J. and Manoil, C. (1990) Lac permease of Escherichia coli: Topology and sequence elements promoting membrane insertion. Proc. Natl. Acad. Sci. USA 87, 4937–4941.

    Article  PubMed  CAS  Google Scholar 

  12. Pearce, B.J., Yin, Y.B., and Masure, H.R. (1993) Genetic identification of exported proteins in Streptococcus pneumoniae. Mol. Microbiol. 9, 1037–1050.

    Article  PubMed  CAS  Google Scholar 

  13. Bergdoll, M.S., Borja, C.R., Robbins, R., and Weiss, K.F. (1971) Identification of enterotoxin E. Infect. Immun. 4, 593–595.

    PubMed  CAS  Google Scholar 

  14. Manoil, C. and Beckwith, J. (1985) Tnpho A: A transposon probe for protein export signals. Proc. Natl. Acad. Sci. USA 82, 8129–8133.

    Article  PubMed  CAS  Google Scholar 

  15. Sanger, F., Nicklen, S., and Coulson, A.R. (1977) DNA sequencing with chain-terminating inhibitors. Proc. Natl. Acad. Sci. USA 74, 5463–5467.

    Article  PubMed  CAS  Google Scholar 

  16. Roe, B.A., Yudong Qian, Dorman, A., Najar, F.Z., Clifton, S., and Iandolo, J. Staphylococcus aureus genome sequencing project. www.genome.ou.edu/staph.htlm. Funded by NIH and Merck Research Foundation.

  17. Hofmann, K. and Stoffel, W. (1993) TMbase-A database of membrane spanning proteins segments. Biol. Chem. Hoppe-Seyler 347, 166.

    Google Scholar 

  18. Nielsen, H., Engelbrecht, J., Brunak, S., and von Heijne, G. (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng. 10, 1–6.

    Article  PubMed  CAS  Google Scholar 

  19. Gravet, A., Colin, D.A., Keller, D., Giradot, R., Monteil, H., and Prevost, G. (1998) Characterisation of a novel structural member, LukE-LukD, of the bicomponent staphylococcal leucotoxins family. FEBS Lett. 436, 202–208.

    Article  PubMed  CAS  Google Scholar 

  20. Farrell, A.M., Taylor, D., and Holland, K.T. (1995) Cloning, nucleotide sequence determination and expression of the Staphylococcus aureus hyaluronate lyase gene. FEMS Microbiol. Letts. 130, 81–85.

    CAS  Google Scholar 

  21. Wada, A. and Watanabe, H. (1998) Penicillin-binding protein 1 of Staphylococcus aureus is essential for growth. J. Bacteriol. 180, 2759–2765.

    PubMed  CAS  Google Scholar 

  22. Ramadurai, L. and Jayaswal, R.K. (1997) Molecular cloning, sequencing, and expression of lytM, a unique autolytic gene of Staphylococcus aureus. J. Bacteriol. 179, 3625–3631.

    PubMed  CAS  Google Scholar 

  23. Lowe, A.M., Beattie, D.T. and Deresiewicz, R.L. (1998) Identification of novel staphylococcal virulence genes by in vivo expression technology. Mol. Microbiol. 27, 967–976.

    Article  PubMed  CAS  Google Scholar 

  24. Ni Eidhin, D., Perkins, S., Francois, P., Vaudaux, P., Hook, M. and Foster, T.J. (1998) Clumping factor B (ClfB), a new surface-located fibrinogenbinding adhesin of Staphylococcus aureus. Mol. Microbiol. 30, 245–57

    Article  PubMed  CAS  Google Scholar 

  25. Patti, J.M., Allen, B.L., McGavin, M.J., and Hook, M. (1994) MSCRAMM-mediated adherence of microorganisms to host tissues. Annu. Rev. Microbiol. 48, 585–617.

    Article  PubMed  CAS  Google Scholar 

  26. Schwan, W.R., Coulter, S.N., Ng, E.Y.W., Langhorne, M.H., Ritchie, H.D., Brody, L.L., Westbrock-Wadman, S., Bayer, A.S., Folger, K.R., and Stover, C.K. (1998) Identification and characterization of the PutP proline permease that contributes to in vivo survival of Staphylococcus aureus in animal models. Infect. Immun. 66, 567–572.

    PubMed  CAS  Google Scholar 

  27. Yousif, Y., Schiltz, E., Okada, K., Batsford, S., and Vogt, A. (1994) Staphylococcal neutral phosphatase. A highly cationic molecule with binding properties for immunoglobulin. APMIS 102, 891–900.

    Article  PubMed  CAS  Google Scholar 

  28. Roberson, J.R., Fox, L.K., Hancock, D.D., and Besser, T.E. (1992) Evaluation of methods for differentiation of coagulase-positive staphylococci. J. Clin. Micro. 30, 3217–3219.

    CAS  Google Scholar 

  29. Jackson, M.P. and Iandolo, J.J. (1986) Cloning and expression of the exfoliative toxin B gene from Staphylococcus aureus. J. Bacteriol. 166, 574–580.

    PubMed  CAS  Google Scholar 

  30. Ranelli, D.M., Jones, C.L., Johns, M.B., Mussey, G.J., and Khan, S.A. (1985) Molecular cloning of staphylococcal enterotoxin B gene in Escherichia coli and Staphylococcus aureus. Proc. Natl. Acad. Sci. USA 82, 5850–5854.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to S. P. Nair.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Williams, R.J., Ward, J.M., Henderson, B. et al. Rapid screening for putative exported proteins from staphylococcus aureus using alkaline phosphatase as a reporter molecule. Mol Biotechnol 15, 11–20 (2000). https://doi.org/10.1385/MB:15:1:11

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1385/MB:15:1:11

Index Entries

Navigation