DdlN from Vancomycin-Producing <i>Amycolatopsis orientalis</i> C329.2 Is a VanA Homologue with <scp>d</scp> -Alanyl- <scp>d</scp> -Lactate Ligase Activity

ABSTRACT Vancomycin-resistant enterococci acquire high-level resistance to glycopeptide antibiotics through the synthesis of peptidoglycan terminating in d -alanyl- d -lactate. A key enzyme in this process is a d -alanyl- d -alanine ligase homologue, VanA or VanB, which preferentially catalyzes the synthesis of the depsipeptide d -alanyl- d -lactate. We report the overexpression, purification, and enzymatic characterization of DdlN, a VanA and VanB homologue encoded by a gene of the vancomycin-producing organism Amycolatopsis orientalis C329.2. Evaluation of kinetic parameters for the synthesis of peptides and depsipeptides revealed a close relationship between VanA and DdlN in that depsipeptide formation was kinetically preferred at physiologic pH; however, the DdlN enzyme demonstrated a narrower substrate specificity and commensurately increased affinity for d -lactate in the C-terminal position over VanA. The results of these functional experiments also reinforce the results of previous studies that demonstrated that glycopeptide resistance enzymes from glycopeptide-producing bacteria are potential sources of resistance enzymes in clinically relevant bacteria.

DdlN from Vancomycin-Producing Amycolatopsis orientalis C329.2 Is a VanA Homologue with d -Alanyl- d -Lactate Ligase Activity Journal Articles