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Crystal structure of the class D β-lactamase OXA-10

Nature Structural Biology volume 7, pages 918–925 (2000)Cite this article

Abstract

We report the crystal structure of a class D β-lactamase, the broad spectrum enzyme OXA-10 from Pseudomonas aeruginosa at 2.0 Å resolution. There are significant differences between the overall fold observed in this structure and those of the evolutionarily related class A and class C β-lactamases. Furthermore, the structure suggests the unique, cation mediated formation of a homodimer. Kinetic and hydrodynamic data shows that the dimer is a relevant species in solution and is the more active form of the enzyme. Comparison of the molecular details of the active sites of the class A and class C enzymes with the OXA-10 structure reveals that there is no counterpart in OXA-10 to the residues proposed to act as general bases in either of these enzymes (Glu 166 and Tyr 150, respectively). Our structures of the native and chloride inhibited forms of OXA-10 suggest that the class D enzymes have evolved a distinct catalytic mechanism for β-lactam hydrolysis. Clinical variants of OXA-10 are also discussed in light of the structure.

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Figure 1: The protein fold of the class D β-lactamase OXA-10.
Figure 2: A sequence alignment of the class D (group 2d33) β-lactamases and related enzymes.
Figure 3: Superpositions of the overall fold of class D OXA-10 with related enzymes.
Figure 4: The OXA-10 homodimer.
Figure 5: The active site of OXA-10.

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References

  1. Frère, J.M. Beta-lactamases and bacterial resistance to antibiotics. Mol. Microbiol. 16, 385–395 (1995).

    Article  Google Scholar 

  2. Joris, B., et al. Comparison of the sequences of class A beta-lactamases and of the secondary structure elements of penicillin-recognizing proteins. Antimicrob. Agents Chemother. 35, 2294–2301 (1991).

    Article  CAS  Google Scholar 

  3. Frère, J.M., Dubus, A., Galleni, M., Matagne, A. & Amicosante, G. Mechanistic diversity of beta-lactamases. Biochem. Soc. Trans. 27, 58–63 (1999).

    Article  Google Scholar 

  4. Bush, K. Metallo-beta-lactamases: a class apart. Clin. Infect. Dis. 27 (Suppl 1), S48–53 (1998).

    Article  CAS  Google Scholar 

  5. Knox, J.R., Moews, P.C. & Frère, J.M. Molecular evolution of bacterial beta-lactam resistance. Chem. Biol. 3, 937–947 (1996).

    Article  CAS  Google Scholar 

  6. Gordon, E., Mouz, N., Duée, E. & Dideberg, O. The crystal structure of the penicillin-binding protein 2x from Streptoccus pneumoniae and its acyl-enzyme form: implication in drug resistance. J. Mol. Biol. 299, 477–485 (2000).

    Article  CAS  Google Scholar 

  7. Nass, T. & Nordmann, P. OXA-type beta-lactamases. Curr. Pharm. Design 5, 865–879 (1999).

    Google Scholar 

  8. Mugnier, P., Casin, I., Bouthers, A.T. & Collatz, E. Novel OXA-10-derived extended-spectrum beta-lactamases selected in vivo or in vitro. Antimicrob. Agents Chemother. 42, 3113–3116 (1998).

    Article  CAS  Google Scholar 

  9. Bush, K. The evolution of beta-lactamases. Ciba Found. Symp. 207, 152–163 (1997).

    CAS  PubMed  Google Scholar 

  10. Strynadka, N.C.J., et al. Molecular structure of the acyl-enzyme intermediate in beta-lactam hydrolysis at 1.7 Å resolution. Nature 359, 700–705 (1992).

    Article  CAS  Google Scholar 

  11. Jelsch, C., Mourey, L., Masson, J-M. & Samama, J-P. Crystal structure of Escherichia coli TEM1 beta-lactamase at 1.8 Å resolution. Proteins 16, 364–383 (1993).

    Article  CAS  Google Scholar 

  12. Oefner, C., et al. Refined crystal structure of beta-lactamase from Citrobacter freundii indicates a mechanism for beta-lactam hydrolysis. Nature 343, 284–289 (1990).

    Article  CAS  Google Scholar 

  13. Monaghan, C., Holland, S. & Dale, J.W. The interaction of anthraquinone dyes with the plasmid-mediated OXA-2 beta-lactamase. Biochem. J. 205, 413–417 (1982).

    Article  CAS  Google Scholar 

  14. Sanschagrin, F., Couture, F. & Levesque, R.C. Primary structure of OXA-3 and phylogeny of oxacillin-hydrolyzing class D beta-lactamases. Antimicrob. Agents Chemother. 39, 887–893 (1995).

    Article  CAS  Google Scholar 

  15. Glusker, J.P. Structural aspects of metal liganding to functional groups in proteins. Adv. Protein Chem. 42, 1–76 (1991).

    Article  CAS  Google Scholar 

  16. Dale J.W. & Smith, J.T. The dimeric nature of an R-factor mediated beta-lactamase. Biochem. Biophys. Res. Commun. 68, 1000–1005 (1976).

    Article  CAS  Google Scholar 

  17. Danel, F., Hall, L.M., Gur, D. & Livermore, D.M. OXA-16, a further extended-spectrum variant of OXA-10 beta-lactamase, from two Pseudomonas aeruginosa isolates. Antimicrob. Agents Chemother. 42, 3117–3122 (1998).

    Article  CAS  Google Scholar 

  18. Lobkovsky, E., et al. Crystallographic structure of a phosphonate derivative of the Enterobacter cloacae P99 cephalosporinase: mechanistic interpretation of a beta-lactamase transition-state analog. Biochemistry 33, 6762–6772 (1994).

    Article  CAS  Google Scholar 

  19. Maveyraud, L., Pratt, R.F. & Samama, J.P. Crystal structure of an acylation transition-state analog of the TEM-1 beta-lactamase. Mechanistic implications for class A beta-lactamases. Biochemistry 37, 2622–2628 (1998).

    Article  CAS  Google Scholar 

  20. Zhu, Y., et al. Structure, function, and fate of the BlaR signal transducer involved in induction of beta-lactamase in Bacillus licheniformis. J. Bacteriol. 174, 6171–6178 (1992).

    Article  CAS  Google Scholar 

  21. Jacob-Dubuisson, F., Lamotte-Brasseur J., Dideberg, O., Joris, B. & Frere, J.M. Arginine 220 is a critical residue for the catalytic mechanism of the Streptomyces albus G beta-lactamase. Protein Eng. 4, 811–819 (1991).

    Article  CAS  Google Scholar 

  22. Dubus, A., Normark, S., Kania, K. & Page, M.G.P. The role of tyrosine 150 in catalysis of β-lactam hydrolysis by AmpC β-lactamase from Escherichia coli investigated by site-directed mutagenesis. Biochemistry 33, 8577–8586 (1994).

    Article  CAS  Google Scholar 

  23. Ledent, P. Raquet, X., Joris, B., Van Beeumen, J. & Frere J.M. A comparative study of class-D beta-lactamases. Biochem. J. 292, 555–562 (1993).

    Article  CAS  Google Scholar 

  24. Little, J.W., et al. Cleavage of LexA repressor. Methods Enzymol. 244, 266–284 (1994).

    Article  CAS  Google Scholar 

  25. Paetzel, M., et al. Use of site-directed chemical modification to study an essential lysine in Escherichia coli leader peptidase. J. Biol. Chem. 272, 9994–10003 (1997).

    Article  CAS  Google Scholar 

  26. Patricelli, M.P. & Cravatt, B.F. Fatty acid amide hydrolase competitively degrades bioactive amides and esters through a nonconventional catalytic mechanism. Biochemistry 38, 14125–14130 (1999).

    Article  CAS  Google Scholar 

  27. Birghan, C., Mundt, E. & Gorbalenya, A.E. A non-canonical lon proteinase lacking the ATPase domain employs the Ser-Lys catalytic dyad to exercise broad control over the life cycle of a double-stranded RNA virus. EMBO J. 19, 114–123 (2000).

    Article  CAS  Google Scholar 

  28. Keiler, K.C. & Sauer, R.T. Identification of active site residues of the Tsp protease. J. Biol. Chem. 270, 28864–28868 (1995).

    Article  CAS  Google Scholar 

  29. Haase, J. & Lanka, E. A specific protease encoded by the conjugative DNA transfer systems of IncP and Ti plasmids is essential for pilus synthesis. J. Bacteriol. 179, 5728–5735 (1997).

    Article  CAS  Google Scholar 

  30. Lietz, E.J., Truher, H., Kahn, D., Hokenson, M.J. & Fink, A.L. Lysine-73 is involved in the acylation and deacylation of beta-lactamase. Biochemistry 39, 4971–4981 (2000).

    Article  CAS  Google Scholar 

  31. Damblon, C., et al. The catalytic mechanism of beta-lactamases: NMR titration of an active-site lysine residue of the TEM-1 enzyme. Proc. Natl. Acad. Sci. USA 93, 1747–1752 (1996).

    Article  CAS  Google Scholar 

  32. Philippon, A.M., Paul G. & Jacoby, G.A. Properties of PSE-2 beta-lactamase and genetic basis for its production in Pseudomonas aeruginosa. Antimicrob. Agents Chemother. 24, 362–369 (1983).

    Article  CAS  Google Scholar 

  33. Bush, K., Jacoby G.A., Medeiros, A.A. A functional classification scheme for beta-lactamases and its correlation with molecular structure. Antimicrob. Agents Chemother. 39, 1211–1233 (1995).

    Article  CAS  Google Scholar 

  34. Bilwes, A., Rees, B., Moras, D., Menez, R. & Menez, A. X-ray structure at 1.55 Å of toxin gamma, a cardiotoxin from Naja nigricollis venom. Crystal packing reveals a model for insertion into membranes. J. Mol. Biol. 239,122–136 (1994).

    Article  CAS  Google Scholar 

  35. Otwinowski, Z. In Denzo (eds, Sawyer, L., Isaacs, N. & Baily, S.) 56–62 (SERC Daresbury Laboratory, Warrington, UK; 1993).

    Google Scholar 

  36. Terwilliger, T.C. & Berendzen, J. Automated structure solution for MIR and MAD. Acta Crystallogr. D 55, 849–861 (1999).

    Article  CAS  Google Scholar 

  37. La Fortelle, E. & Bricogne, G. Maximum-likelihood heavy-atom parameter refinement in the MIR and MAD methods Methods Enzymol. 276, 472–494 (1997).

    Article  Google Scholar 

  38. Jones, T.A., Zou, J.-Y., Cowan, S.W. & Kieldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  Google Scholar 

  39. Brünger, A.T. et al. Crystallography & NMR system: a new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998).

    Article  Google Scholar 

  40. Jones, S. & Thornton, J.M. Protein-protein interactions: a review of protein dimer structure. Prog. Biophys. Mol. Biol. 63, 31–165 (1995).

    Article  CAS  Google Scholar 

  41. Cohn, E.J. & Edsall, J.T. Density and apparent specific volume of proteins. In Proteins, amino acids and peptides as ions and dipolar ions (ed. E. J. Cohn), 370–381 (Reinhold, New York; 1943).

    Google Scholar 

  42. Schuck, P. Simultaneous radial and wavelength analysis with the Optima XL-A analytical ultracentrifuge. Prog. Colloid Polymer Sci. 94, 1–13 (1994).

    Article  CAS  Google Scholar 

  43. Kraulis, P. G. Molscript: a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallog. 24, 946–950 (1991).

    Article  Google Scholar 

  44. Nicholls, A., Sharp, K.A. & Honig, B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 11, 281–296 (1991).

    Article  CAS  Google Scholar 

  45. Couture, F., Lachapelle, J. & Levesque, R.C. Phylogeny of LCR-1 and OXA-5 with class A and class D beta-lactamases. Mol. Microbiol. 6, 1693–1705 (1992).

    Article  CAS  Google Scholar 

  46. Thompson, J.D., Higgins, D.G. & Gibson, T.J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22, 4673–4680 (1994).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank the Medical Research Council of Canada (M.P. is a MRC fellow, N.C.J.S. is a MRC scholar), the Burroughs Wellcome Foundation (New Investigator Award to N.C.J.S.) and Hoffman-La Roche Pharmaceuticals (to F.D. and M.G.P.P.) for support. We thank H. Bellamy of beamline 1-5 at the SSRL for data collection access and R. Sweet for access to beamline X12C at the NSLS (Brookhaven National Laboratory).

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of British Columbia, 2146 Health Sciences Mall, Vancouver, V6T 1Z3, British Columbia, Canada

    Mark Paetzel, Liza de Castro, Steven C. Mosimann & Natalie C.J. Strynadka

  2. Pharma Division, F. Hoffman-La Roche Ltd., Preclinical Research, Basel, CH-4070, Switzerland

    Franck Danel & Malcolm G.P. Page

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Correspondence to Natalie C.J. Strynadka.

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Paetzel, M., Danel, F., de Castro, L. et al. Crystal structure of the class D β-lactamase OXA-10. Nat Struct Mol Biol 7, 918–925 (2000). https://doi.org/10.1038/79688

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