Acylation of class a beta-lactamases by penicillins: A theoretical examination of the role of serine 130 and the beta-lactam carboxylate group

Herein we present results of a computational study on the benzylpenicillin acylation of the class A TEM1 beta -lactamase via hydroxyl-only and hydroxy l and carboxylate assisted processes. These mechanisms correspond to a one- step Ser130-assisted process and a second route in which the beta -lactam c arboxylate and the Ser130 hydroxyl group help the proton transfer from the hydroxyl group of Ser70 to the beta -lactam leaving N atom. The internal ge ometry of the reactive part of the TEM1-benzylpenicillin system is taken fr om a B3LYP/6-31 +G* computational study on the methanol-assisted methanolys is reaction of a penicillin model compound (3 alpha -carboxypenam). The 6-a cylamino side chain and the 2-methyl groups of benzylpenicillin, together w ith the closer residues around the essential Ser70, are relaxed by carrying out geometry optimizations with a hybrid QM/MM method. The corresponding r elative energies in the protein combine the B3LYP/6-31+G* electronic energi es of the reactive subsystem with semiempirical PM3 energies of the TEM1-be nzylpenicillin systems both in vacuo and in solution. The PM3 calculations on the TEM1-benzylpenicillin systems are performed with a Divide and Conque r linear-scaling method. The hydroxyl and carboxylate assisted pathway, whi ch is the most favored one, is in agreement with the experimentally observe d kinetic isotope effects and is also compatible with the effects of mutage nesis experiments on the Ser130 residue. These results suggest that a simil ar mechanism for the formation of acylenzyme intermediates could be relevan t to other active-site serine penicillin-recognizing enzymes.