Insights into class D beta-lactamases are revealed by the crystal structure of the OXA10 enzyme from Pseudomonas aeruginosa

Background: beta -lactam antibiotic therapies are commonly challenged by th e hydrolytic activities of beta -lactamases in bacteria. These enzymes have been grouped into four classes: A, B, C, and D. Class B beta -lactamases a re zinc dependent, and enzymes of classes A, C, and D are transiently acyla ted on a serine residue in the course of the turnover chemistry. While clas s A and C beta -lactamases have been extensively characterized by biochemic al and structural methods, class D enzymes remain the least studied despite their increasing importance in the clinic. Results: The crystal structure of the OXA10 class D beta -lactamase has bee n solved to 1.66 A resolution from a gold derivative and MAD phasing. This structure reveals that beta -lactamases from classes D and A, despite very poor sequence similarity, share a similar overall fold. An additional beta strand in OXA10 mediates the association into dimers characterized by analy tical ultracentrifugation. Major differences are found when comparing the m olecular details of the active site of this class D enzyme to the correspon ding regions in class A and C B-lactamases. In the native structure of the OXA10 enzyme solved to 1.8 Angstrom, Lys-70 is carbamylated. Conclusions: Several features were revealed by this study: the dimeric stru cture of the OXA10 beta -lactamase, an extension of the substrate binding s ite which suggests that class D enzymes may bind other substrates beside be ta -lactams, and carbamylation of the active site Lys-70 residue. The CO2-d ependent activity of the OXA10 enzyme and the kinetic properties of the nat ural OXA17 mutant protein suggest possible relationships between carbamylat ion, inhibition of the enzyme by anions, and biphasic behavior of the enzym e.