Catalytic Mechanism of Class A β-Lactamase: Role of Lysine 73 and C3-Carboxyl Group of the Substrate Pen G in the Deacylation Step

Abstract

The deacylation mechanism in hydrolysis of β-lactam antibiotics by class A β-lactamase was investigated by the density functional theory. The model compound was composed of a substrate penicillin G, four catalytic residues(Ser70, Lys73, Ser130, and Glu166), and a water molecule. We have found that the deacylation reaction proceeds via four elementary reactions. First, Lys73 is deprotonated by a concerted double proton transfer from Lys73Nζ to Ser130Oγ and from Ser130Oγ to C3-carboxylate in the substrate. Second, the acyl-enzyme tetrahedral intermediate (TI) is formed by an assistance of Glu166 which acts as a general base catalyst. Third, Lys73 is protonated by concerted double proton transfer from C3-carboxylic acid to Ser130Oγ and from Ser130Oγ to Lys73Nζ. Finally, the degraded substrate is detached from the enzyme in concert with a single proton transfer from Lys73Nζ to Ser70Oγ. It is remarkably noted that the deacylation is not proceeded only by Glu166, but Lys73 also participates in the reaction. Mor...