Crystal Structure of 6α-(Hydroxymethyl)penicillanate Complexed to the TEM-1 β-Lactamase from Escherichia coli :  Evidence on the Mechanism of Action of a Novel Inhibitor Designed by a Computer-Aided Process

The crystal structure of the complex of the TEM-1 β-lactamase from Escherichia coli inhibited by 6α-(hydroxymethyl)penicillanic acid ( 1 ) is reported herein. This is the first structure for an acyl-enzyme intermediate with a substrate reported for a native class A β-lactamase. This compound was designed and synthesized as a molecule that would acylate the active site of the enzyme, but would resist deacylation by virtue of the fact that its C 6 α hydroxymethyl moiety was expected to occupy the space near the hydrolytic water molecule ( J. Am. Chem. Soc. 1995, 117, 11055). The crystal structure of the acyl-enzyme species is closely similar to one of the two energy-minimized acyl-enzyme models generated in the course of the design aspect of the work. The crystal structure provides evidence for a number of mechanistic features for the inhibition process and the ultimate recovery of the activity. Our results reported herein are consistent with the side-chain carboxylate of Glu-166 being the active-site basic function that activates the hydrolytic water for the deacylation step in the course of catalysis by class A β-lactamases. The design principles applied for compound 1 hold the promise of general utility for development of novel inhibitors for other hydrolytic enzymes.