We thank the staff of beamline BM16 at the European Synchrotron Radiation Facility for their support, C. Fernández-Cabrera for excellent technical assistance, G. Rivas for ultracentrifugation studies, and P. Grooves and M. Sefton for critical reading of the manuscript.
Combating bacterial resistance to β-lactams, the most widely used antibiotics, is an emergent and clinically important challenge. OXA-24 is a class D β-lactamase isolated from a multiresistant epidemic clinical strain of Acinetobacter baumannii. We have investigated how OXA-24 specifically hydrolyzes the last resort carbapenem antibiotic, and we have determined the crystal structure of OXA-24 at a resolution of 2.5 Å. The structure shows that the carbapenem's substrate specificity is determined by a hydrophobic barrier that is established through the specific arrangement of the Tyr-112 and Met-223 side chains, which define a tunnel-like entrance to the active site. The importance of these residues was further confirmed by mutagenesis studies. Biochemical and microbiological analyses of specific point mutants selected on the basis of structural criteria significantly reduced the catalytic efficiency (kcat/Km) against carbapenems, whereas the specificity for oxacillin was noticeably increased. This is the previously unrecognized crystal structure that has been obtained for a class D carbapenemase enzyme. Accordingly, this information may help to improve the development of effective new drugs to combat β-lactam resistance. More specifically, it may help to overcome carbapenem resistance in A. baumannii, probably one of the most worrying infectious threats in hospitals worldwide.
This work was supported by Grant BFU2005-05055 from the Spanish Ministry of Science; Grant PGIDI4BTF916028PR from Dirección Xeral I+D, Xunta de Galicia; and Grants PI040514, PI061368, and RD06/0008 from the Fondo de Investigaciones Sanitarias. E.S. holds a fellowship from the Spanish Ministry of Science. A.B. is a recipient of a scholarship from the Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica.
Peer reviewed