Multiscale Simulations Identify Origins of Differential Carbapenem Hydrolysis by the OXA-48 β-Lactamase

[Image: see text] OXA-48 β-lactamases are frequently encountered in bacterial infections caused by carbapenem-resistant Gram-negative bacteria. Due to the importance of carbapenems in the treatment of healthcare-associated infections and the increasingly wide dissemination of OXA-48-like enzymes on plasmids, these β-lactamases are of high clinical significance. Notably, OXA-48 hydrolyzes imipenem more efficiently than other commonly used carbapenems, such as meropenem. Here, we use extensive multiscale simulations of imipenem and meropenem hydrolysis by OXA-48 to dissect the dynamics and to explore differences in the reactivity of the possible conformational substates of the respective acylenzymes. Quantum mechanics/molecular mechanics (QM/MM) simulations of the deacylation reaction for both substrates demonstrate that deacylation is favored when the 6α-hydroxyethyl group is able to hydrogen bond to the water molecule responsible for deacylation but disfavored by the increasing hydration of either oxygen of the carboxylated Lys73 general base. Differences in free energy barriers calculated from the QM/MM simulations correlate well with the experimentally observed differences in hydrolytic efficiency between meropenem and imipenem. We conclude that the impaired breakdown of meropenem, compared to imipenem, which arises from a subtle change in the hydrogen bonding pattern between the deacylating water molecule and the antibiotic, is most likely induced by the meropenem 1β-methyl group. In addition to increased insights into carbapenem breakdown by OXA β-lactamases, which may aid in future efforts to design antibiotics or inhibitors, our approach exemplifies the combined use of atomistic simulations in determining the possible different enzyme–substrate substates and their influence on enzyme reaction kinetics.