Crystal structure of a D-aminopeptidase from Ochrobactrum anthropi, a new member of the 'penicillin-recognizing enzyme' family

Background: beta-Lactam compounds are the most widely used antibiotics. The y inactivate bacterial DD-transpeptidases, also called penicillin-binding p roteins (PBPs), involved in cell-wall biosynthesis. The most common bacteri al resistance mechanism against beta-lactam compounds is the synthesis of b eta-lactamases that hydrolyse beta-lactam rings. These enzymes are believed to have evolved from cell-wall DD-peptidases. Understanding the biochemica l and mechanistic features of the beta-lactam targets is crucial because of the increasing number of resistant bacteria. DAP is a D-aminopeptidase pro duced by Ochrobactrum anthropi. It is inhibited by various beta-lactam comp ounds and shares similar to 25% sequence identity with the R61 DD-carboxype ptidase and the class C beta-lactamases. Results: The crystal structure of DAP has been determined to 1.9 Angstrom r esolution using the multiple isomorphous replacement (MIR) method. The enzy me folds into three domains, A, B and C. Domain A, which contains conserved catalytic residues, has the classical fold of serine beta-lactamases, wher eas domains B and C are both antiparallel eight-stranded beta barrels. A lo op of domain C protrudes into the substrate-binding site of the enzyme. Conclusions: Comparison of the biochemical properties and the structure of DAP with PBPs and serine beta-lactamases shows that although the catalytic site of the enzyme is very similar to that of beta-lactamases, its substrat e and inhibitor specificity rests on residues of domain C, DAP is a new mem ber of the family of penicillin-recognizing proteins (PRPs) and, at the pre sent time, its enzymatic specificity is clearly unique.