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:

Structure of α-lytic protease complexed with its pro region

Nature Structural Biology volume 5, pages 945–950 (1998)Cite this article

Abstract

While the majority of proteins fold rapidly and spontaneously to their native states, the extracellular bacterial protease α-lytic protease (αLP) has a t 1/2 for folding of ~2,000 years, corresponding to a folding barrier of 30 kcal mol –1 . αLP is synthesized as a pro-enzyme where its pro region (Pro) acts as a foldase to stabilize the transition state for the folding reaction. Pro also functions as a potent folding catalyst when supplied as a separate polypeptide chain, accelerating the rate of αLP folding by a factor of 3 × 10 9 . In the absence of Pro, αLP folds only partially to a stable molten globule-like intermediate state. Addition of Pro to this intermediate leads to rapid formation of native αLP. Here we report the crystal structures of Pro and of the non-covalent inhibitory complex between Pro and native αLP. The C-shaped Pro surrounds the C-terminal ß-barrel domain of the folded protease, forming a large complementary interface. Regions of extensive hydration in the interface explain how Pro binds tightly to the native state, yet even more tightly to the folding transition state. Based on structural and functional data we propose that a specific structural element in αLP is largely responsible for the folding barrier and suggest how Pro can overcome this barrier.

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: Free energy diagram for the in vitro folding pathway of αLP, adapted from refs 9,10.
Figure 2: a, Ribbon diagram of unbound Pro. Disordered residues are indicated by dots.
Figure 3: a, Detailed view of the C-tail of wild-type Pro (green) inserted into the active site of αLP (blue).
Figure 4: Proposed mechanism for Pro-assisted folding of αLP.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Zhu, X., Ohta, Y., Jordan, F. & Inouye, M. Nature 339, 483–484 (1989).

    Article  CAS  Google Scholar 

  2. Baker, D., Silen, J.L. & Agard, D.A. Proteins Struct. Funct. Genet. 12, 339–344 (1992).

    Article  CAS  Google Scholar 

  3. Strausberg, S., Alexander, P., Wang, L., Schwarz, F. & Bryan, P. Biochemistry 32, 8112– 8119 (1993).

    Article  CAS  Google Scholar 

  4. Winther, J.R. & Sorensen, P. Proc. Natl. Acad. Sci. USA 88, 9330–9334 ( 1991).

    Article  CAS  Google Scholar 

  5. Baker, D., Shiau, A.K. & Agard, D.A. Curr. Opin. Cell Biol. 5, 966 –970 (1993).

    Article  CAS  Google Scholar 

  6. Sohl, J.L. & Agard, D.A., In Intramolecular Chaperones and Protein Folding (eds Shinde, U. & Inouye, M.) 61– 83 (R.G. Landes Co., Austin, Texas; 1995).

    Google Scholar 

  7. Silen, J.L. & Agard, D.A. Nature 341, 462–464 (1989).

    Article  CAS  Google Scholar 

  8. Baker, D., Sohl, J.L. & Agard, D.A. Nature 356, 263– 265 (1992).

    Article  CAS  Google Scholar 

  9. Peters, R.J. et al. Biochemistry, 37,12058– 12067 (1998).

    Article  CAS  Google Scholar 

  10. Sohl, J.L., Jaswal, S.S. & Agard, D.A. Nature, in the press ( 1998).

  11. Sohl, J.L., Shiau, A.K., Rader, S.D., Wilk, B.J. & Agard, D.A. Biochemistry 36, 3894– 3902 (1997).

    Article  CAS  Google Scholar 

  12. Leahy, D.J., Hendrickson, W.A., Aukhil, I. & Erickson, H.P. Science 258, 987–991 ( 1992).

    Article  CAS  Google Scholar 

  13. Bone, R., Shenvi, A.B., Kettner, C.A. & Agard, D.A. Biochemistry 26, 7609–7614 (1987).

    Article  CAS  Google Scholar 

  14. Fersht, A. Enzyme structure and mechanism (W.H. Freeman, New York; 1985).

  15. Gewirth, D.T. & Sigler, P.B. Nature Struct. Biol. 2, 386–394 (1995).

    Article  CAS  Google Scholar 

  16. Schirmer, T. & Evans, P.R. Nature 343, 140–145 (1990).

    Article  CAS  Google Scholar 

  17. Royer, W.E. Jr., Pardanani, A., Gibson, Q.H., Peterson, E.S. & Friedman, J.M. Proc. Natl. Acad. Sci. USA 93, 14526–14531 (1996).

    Article  CAS  Google Scholar 

  18. Allaire, M., Chernaia, M.M., Malcolm, B.A. & James, M.N.G. Nature 369, 72–76 ( 1994).

    Article  Google Scholar 

  19. Gallagher, T., Gilliland, G., Wang, L. & Bryan, P. Structure 3, 907–914 (1995).

    Article  CAS  Google Scholar 

  20. Silen, J.L., McGrath, C.N., Smith, K.R. & Agard, D.A. Gene 69, 237–244 ( 1988).

    Article  CAS  Google Scholar 

  21. LeMaster, D.M. & Richards, F.M. Biochemistry 24, 7263–7268 ( 1985).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  23. Hendrickson, W.A., Smith, J.L., Phizackerly, R.P. & Merritt, E.A. Proteins Struct. Funct. Genet. 4, 77– 88 (1988).

    Article  CAS  Google Scholar 

  24. French, S. & Wilson, K. Acta Crystallogr. A 34 , 517–525 (1978).

    Article  Google Scholar 

  25. de La Fortelle, E. & Bricogne, G. Meth. Enz. 276, 472–494 (1997).

    Article  CAS  Google Scholar 

  26. Collaborative Computational Project, Number 4 Acta Crystallogr. D 50, 760–776 (1994).

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

    Article  CAS  Google Scholar 

  28. Kleywegt, G.J. & Jones, T.A. Acta Crystallogr. D 53, 179–185 ( 1997).

    Article  CAS  Google Scholar 

  29. Pannu, N.S. & Read, R.J. Acta Crystallogr. A 52 , 659–668 (1996).

    Article  Google Scholar 

  30. Rice, L.M. & Brünger, A.T. Proteins Struct. Funct. Genet. 19, 277–290 ( 1994).

    Article  CAS  Google Scholar 

  31. Brünger, A.T., Kuriyan, J. & Karplus, M. Science 235, 458– 460 (1987).

    Article  Google Scholar 

  32. Brünger, A.T., Krukowski, A. & Erickson, J. Acta Crystallogr. A 46, 585– 593 (1990).

    Article  Google Scholar 

  33. Brünger, A.T. X-PLOR version 3.1: a system for X-ray crystallography and NMR (Yale Univ. Press, New Haven, Connecticut; 1992).

    Google Scholar 

  34. Bone, R., Silen, J.L. & Agard, D.A. Nature 339, 191– 195 (1989).

    Article  CAS  Google Scholar 

  35. Mau, I.-F.T. Ph. D. Thesis, University of California, San Francisco (1998).

  36. Rader, S.D. Ph.D. Thesis, University of California, San Francisco (1996).

  37. Read, R.J. Acta Crystallogr. A 42, 140–149 (1986).

    Article  Google Scholar 

Download references

Acknowledgements

This work was supported by the Howard Hughes Medical Institute (HHMI). N.K.S. was supported in part by a Damon Runyon-Walter Winchell postdoctoral fellowship, T.M. by an HHMI predoctoral fellowship, and S.D.R. by an NIH training grant. We thank D. King for mass spectroscopy analysis; C. Ogata (NSLS), M. Soltis and H. Bellamy (SSRL) for beamline support; P. David for logistical help; C. Wilson for the occasional use of his Raxis II; A. Shiau and A. Derman for assistance with data collection; and A. Derman, S. Gillmor, A. Shiau and J. Sohl for critical comments on the manuscript. Some data were collected at SSRL, which is operated by the Department of Energy, Office of Basic Energy Sciences. The SSRL Biotechnology Program is supported by the National Institutes of Health, National Center for Research Resources, Biomedical Technology Program, and by the Department of Energy, Office of Biological and Environmental Research.

Author information

Author notes

  1. Nicholas K. Sauter, Ted Mau, Stephen D. Rader and David A. Agard: These authors contributed equally to this work. Howard Hughes Medical Institute, San Francisco, San Francisco , California 94143-0448, USA.

Authors and Affiliations

  1. the Graduate Group in Biophysics and the Department of Biochemistry and Biophysics, University of California, San Francisco , San Francisco, 94143-0448, California, USA

    Ted Mau, Stephen D. Rader & David A. Agard

Authors
  1. Nicholas K. Sauter
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ted Mau
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Stephen D. Rader
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. David A. Agard
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to David A. Agard.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Sauter, N., Mau, T., Rader, S. et al. Structure of α-lytic protease complexed with its pro region. Nat Struct Mol Biol 5, 945–950 (1998). https://doi.org/10.1038/2919

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/2919

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