Structural basis of activity against aztreonam and extended spectrum cephalosporins for two carbapenem-hydrolyzing class D β-lactamases from Acinetobacter baumannii.
Joshua M Mitchell, Jozlyn R Clasman, Cynthia M June, Kip-Chumba J Kaitany, James R LaFleur, Magdalena A Taracila, Neil V Klinger, Robert A Bonomo, Troy Wymore, Agnieszka Szarecka, Rachel A Powers, David A Leonard
Author Information
Magdalena A Taracila: ∥Departments of Medicine, Pharmacology, Biochemistry, and Molecular Biology and Microbiology, Case Western Reserve University and Research Service, and Louis Stokes Cleveland Department of Veterans Affairs Medical Center, Cleveland, Ohio 44106, United States.
Robert A Bonomo: ∥Departments of Medicine, Pharmacology, Biochemistry, and Molecular Biology and Microbiology, Case Western Reserve University and Research Service, and Louis Stokes Cleveland Department of Veterans Affairs Medical Center, Cleveland, Ohio 44106, United States.
Troy Wymore: ⊥UT/ORNL Center for Molecular Biophysics, Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831, United States.
The carbapenem-hydrolyzing class D β-lactamases OXA-23 and OXA-24/40 have emerged worldwide as causative agents for β-lactam antibiotic resistance in Acinetobacter species. Many variants of these enzymes have appeared clinically, including OXA-160 and OXA-225, which both contain a P → S substitution at homologous positions in the OXA-24/40 and OXA-23 backgrounds, respectively. We purified OXA-160 and OXA-225 and used steady-state kinetic analysis to compare the substrate profiles of these variants to their parental enzymes, OXA-24/40 and OXA-23. OXA-160 and OXA-225 possess greatly enhanced hydrolytic activities against aztreonam, ceftazidime, cefotaxime, and ceftriaxone when compared to OXA-24/40 and OXA-23. These enhanced activities are the result of much lower Km values, suggesting that the P → S substitution enhances the binding affinity of these drugs. We have determined the structures of the acylated forms of OXA-160 (with ceftazidime and aztreonam) and OXA-225 (ceftazidime). These structures show that the R1 oxyimino side-chain of these drugs occupies a space near the β5-β6 loop and the omega loop of the enzymes. The P → S substitution found in OXA-160 and OXA-225 results in a deviation of the β5-β6 loop, relieving the steric clash with the R1 side-chain carboxypropyl group of aztreonam and ceftazidime. These results reveal worrying trends in the enhancement of substrate spectrum of class D β-lactamases but may also provide a map for β-lactam improvement.
