Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Structural and biochemical characterization of the type III secretion chaperones CesT and SigE

Nature Structural Biology volume 8, pages 1031–1036 (2001)Cite this article

Abstract

Several Gram-negative bacterial pathogens have evolved a type III secretion system to deliver virulence effector proteins directly into eukaryotic cells, a process essential for disease. This specialized secretion process requires customized chaperones specific for particular effector proteins. The crystal structures of the enterohemorrhagic Escherichia coli O157:H7 Tir-specific chaperone CesT and the Salmonella enterica SigD-specific chaperone SigE reveal a common overall fold and formation of homodimers. Site-directed mutagenesis suggests that variable, delocalized hydrophobic surfaces observed on the chaperone homodimers are responsible for specific binding to a particular effector protein. Isothermal titration calorimetry studies of Tir–CesT and enzymatic activity profiles of SigD–SigE indicate that the effector proteins are not globally unfolded in the presence of their cognate chaperones.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Electron density with MAD phases improved by solvent modification.
Figure 2: Monomeric structures
Figure 3: Variable homodimers and hydrophobic effector binding surfaces.
Figure 4: Functional analysis of effector–chaperone complexes.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Lee, C.A. Trends Microbiol. 5, 148–156 (1997).

    Article  CAS  PubMed  Google Scholar 

  2. Hueck, C.J. Microbiol. Mol. Biol. Rev. 62, 379–433 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Kubori, T. et al. Science 280, 602–605 (1998).

    Article  CAS  PubMed  Google Scholar 

  4. Wattiau, P., Woestyn, S. & Cornelis, G.R. Mol. Microbiol. 20, 255–262 (1996).

    Article  CAS  PubMed  Google Scholar 

  5. Bennett, J.C. & Hughes, C. Trends Microbiol. 8, 202–204 (2000).

    Article  CAS  PubMed  Google Scholar 

  6. Woestyn, S., Sory, M.P., Boland, A., Lequenne, O. & Cornelis, G.R. Mol. Microbiol. 20, 1261–1271 (1996).

    Article  CAS  PubMed  Google Scholar 

  7. Abe, A. et al. Mol. Microbiol. 33, 1162–1175 (1999).

    Article  CAS  PubMed  Google Scholar 

  8. Elliott, S.J. et al. Mol. Microbiol. 33, 1176–1189 (1999).

    Article  CAS  PubMed  Google Scholar 

  9. Darwin, K.H., Robinson, L.S. & Miller, V.L. J. Bacteriol. 183, 1452–1454 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Hendrickson, W.A. Science 254, 51–58 (1991).

    Article  CAS  PubMed  Google Scholar 

  11. Xu, Z., Knafels, J.D. & Yoshino, K. Nature Struct. Biol. 7, 1172–1177 (2000).

    Article  CAS  PubMed  Google Scholar 

  12. Janin, J., Miller, S. & Chothia, C. J. Mol. Biol. 204, 155–164 (1988).

    Article  CAS  PubMed  Google Scholar 

  13. Vijayalakshmi, J., Mukhergee, M.K., Graumann, J., Jakob, U. & Saper, M.A. Structure (Camb.) 9, 367–375 (2001).

    Article  CAS  Google Scholar 

  14. Bennett, M.J., Schlunegger, M.P. & Eisenberg, D. Protein Sci. 4, 2455–2468 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Vogel, H.J. Biochem. Cell Biol. 72, 357–376.

  16. Zhu, X. et al. Science 272, 1606–1614 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Braig, K. et al. Nature 371, 578–586 (1994).

    Article  CAS  PubMed  Google Scholar 

  18. Abe, A. & Nagano, H. Microbiol. Immunol. 44, 857–861 (2000).

    Article  CAS  PubMed  Google Scholar 

  19. Gauthier, A., de Grado, M. & Finlay, B.B. Infect. Immun. 68, 4344–4348 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Hartland, E.L. et al. Mol. Microbiol. 32, 151–158 (1999).

    Article  CAS  PubMed  Google Scholar 

  21. de Grado, M. et al. Cell Microbiol. 1, 7–17 (1999).

    Article  CAS  PubMed  Google Scholar 

  22. Goosney, D.L. et al. Curr. Biol. 10, 735–738 (2000).

    Article  CAS  PubMed  Google Scholar 

  23. Goosney, D.L., DeVinney, R. & Finlay, B.B. Infect. Immun. 69, 3315–3322 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Marcus, S.L., Wenk, M.R., Steele-Mortimer, O. & Finlay, B.B. FEBS Lett. 494, 201–207 (2001).

    Article  CAS  PubMed  Google Scholar 

  25. Doublie, S. Methods Enzymol. 276, 523–530 (1997).

    Article  CAS  PubMed  Google Scholar 

  26. Otwinowski, Z. & Minor, W. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  PubMed  Google Scholar 

  27. Terwilliger, T.C. & Berendzen, J. Acta Crystallogr. D 55, 849–861 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Abrahams, J.P. & Leslie, A.G. Acta Crystallogr. D 52, 30–42 (1996).

    Article  CAS  PubMed  Google Scholar 

  29. Terwilliger, T.C. Acta Crystallogr. D 55, 1863–1871 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. McRee, D.E. J. Struct. Biol. 125, 156–165 (1999).

    Article  CAS  PubMed  Google Scholar 

  31. Brunger, A.T. et al. Acta Crystallogr. D 54, 905–921 (1998).

    Article  CAS  PubMed  Google Scholar 

  32. Laskowski, R.A., MacArthur, M.W. & Thornton, J.M. Curr. Opin. Struct. Biol. 8, 631–639 (1998).

    Article  CAS  PubMed  Google Scholar 

  33. Johnson, M.L., Correia, J.J., Yphantis, D.A. & Halvorson, H.R. Biophys. J. 36, 575–588 (1981).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Kraulis, P.J. J. Appl. Crystallogr. 24, 946–950 (1991).

    Article  Google Scholar 

  35. Bacon, D.J. & Anderson, W.F. J. Mol. Graph. 6, 219–220 (1988).

    Article  Google Scholar 

  36. Honig, B. & Nicholls, A. Science 268, 1144–1149 (1995).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank L. Knodler for critical reading of this manuscript and L. Hicks for excellent technical assistance in the analytical ultracentrifugation experiments. Work in our laboratories was supported by the Burroughs Wellcome Foundation, the Howard Hughes Medical Institute (HHMI), Canadian Institute of Health Research (CIHR), the Canadian Bacterial Disease Network Center of Excellence and the Toronto Hospital for Sick Children. N.C.J.S. is a CIHR Scholar, a BWF New Investigator and a HHMI International Scholar. B.B.F. is a CIHR Distinguished Investigator and a HHMI International Scholar. We are thankful to excellent technical support at beamline 9-2 at SSRL and beamline X-12C at NSLS.

Author information

Author notes

  1. Yu Luo, Michela G. Bertero and Cyril Kay: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, V6T 1Z3, Canada

    Yu Luo, Michela G. Bertero, Elizabeth A. Frey, Richard A. Pfuetzner, Daniel Lim, B. Brett Finlay & Natalie C.J. Strynadka

  2. Biotechnology Laboratory, University of British Columbia, Vancouver, V6T 1Z3, Canada

    Elizabeth A. Frey, Louise Creagh, Sandra L. Marcus, Charles Haynes & B. Brett Finlay

  3. Department of Cell Biology, Yale University School of Medicine, New Haven, 06510, Connecticut, USA

    Markus R. Wenk

  4. Department of Molecular and Medical Genetics, University of Toronto, Toronto, M5G 1X5, Canada

    Frank Sicheri

  5. Department of Biochemistry, Protein Engineering Network of Centres of Excellence, University of Alberta, Edmonton, T6G 2H7, Alberta, Canada

    Cyril Kay

Authors
  1. Yu Luo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Michela G. Bertero
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Elizabeth A. Frey
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Richard A. Pfuetzner
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Markus R. Wenk
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Louise Creagh
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sandra L. Marcus
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Daniel Lim
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Frank Sicheri
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Cyril Kay
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Charles Haynes
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. B. Brett Finlay
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Natalie C.J. Strynadka
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Natalie C.J. Strynadka.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Luo, Y., Bertero, M., Frey, E. et al. Structural and biochemical characterization of the type III secretion chaperones CesT and SigE. Nat Struct Mol Biol 8, 1031–1036 (2001). https://doi.org/10.1038/nsb717

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsb717

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing