Mechanism of Ampicillin Hydrolysis by New Delhi Metallo-β-Lactamase 1: Insight From QM/MM MP2 Calculation.

The New Delhi metallo-β-lactamase 1 (NDM-1) can hydrolyze nearly all clinically important β-lactam antibiotics, narrowing the options for effective treatment of bacterial infections. QM/MM MP2 calculations are performed to reveal the mechanism of ampicillin hydrolysis catalyzed by NDM-1. It is found that the rate-determining step is the dissociation of hydrolyzed ampicillin from the NDM-1 active site, which requires a proton transfer from the bridging neutral water molecule to the newly formed carboxylate group. The precedent reaction steps, including the hydroxide nucleophilic addition, CN bond cleavage, and the protonation of the negative lactam N atom by a solvent water molecule, all require insignificant activation free energies. The calculated activation free energy for this rate-determining proton transfer step is 16.0 kcal/mol, in good agreement with experimental values of 13.7 ~ 14.7 kcal/mol. This proton transfer step exhibits a solvent hydrogen-deuterium kinetic isotope effect of 3.4, consistent with several experimental kinetic results.