Engineering the synthetic potential of β-lactam synthetase and the importance of catalytic loop dynamics

Abstract

The 2-azetidinone ring of the Class A and D β-lactamase inhibitor clavulanic acid (1) is synthesized by the ATP-utilizing enzyme β-lactam synthetase (βLS). A hydroxyethyl group attached to C-6 of 1 in the (S) configuration markedly enhances the efficacy of this compound against Class C β-lactamases. Guided by a series of X-ray structures of βLS, we have engineered this enzyme to act upon a methylated substrate analogue to give selectively the (3S)-methyl β-lactam core, which, upon closure of the second ring of the bicyclic system of 1, would lead to the (6S)-methylated clavulanic acid derivative.

Biocatalytic amide bond formation

“…90 β-LS has been engineered using rational design to accept methyl and ethyl substituted substrates yielding clavulanic derivatives. 92 Carbanepam synthase (CarA) also utilizes an ATP-dependent mechanism to produce a lactam ring but is besides four-membered rings also able to form 5- and 6-membered lactam rings. 93 This enzyme has subsequently been used in a two-enzyme cascade to prepare a range of bicyclic β-lactams in excellent diastereomeric ratios.…”