Active-site mutants of class B beta-lactamases: substrate binding and mechanistic study

Increased resistance to β-lactam antibiotics is mainly due to β-lactamases. X-ray structures of zinc β-lactamases unraveled the coordination of the metal ions, but their mode of action remains unclear. Recently, enzymes in which one of the zinc ligands was mutated have been characterized and their catalytic activity against several β-lactam antibiotics measured. A molecular modeling study of these enzymes was performed here to explain the catalytic activity of the mutants. Coordination around the zinc ions influences the way the tetrahedral intermediate is bound; any modification influences the first recognition of the substrate by the enzyme. For all the studied mutants, at least one of the interactions fails, inducing a loss of catalytic efficiency compared to the wild type. The present studies show that the enzyme cavity is a structure of high plasticity both structurally and mechanistically and that local modifications may propagate its effects far from the mutated amino acid. Received 28 August 2002; received after revision 22 October 2002; accepted 24 October 2002 RID="*" ID="*"Corresponding author.