Tebipenem, a New Carbapenem Antibiotic, Is a Slow Substrate That Inhibits the β-Lactamase from Mycobacterium tuberculosis

Abstract

The genome of Mycobacterium tuberculosis contains a gene, blaC, which encodes a highly active β-lactamase (BlaC). We have previously shown that BlaC has an extremely broad spectrum of activity against penicillins and cephalosporins but weak activity against newer carbapenems. We have shown that carbapenems such as meropenem, doripenem, and ertapenem react with the enzyme to form enzyme–drug covalent complexes that are hydrolyzed extremely slowly. In the current study, we have determined apparent Km and kcat values of 0.8 μM and 0.03 min–1, respectively, for tebipenem, a novel carbapenem whose prodrug form, the pivalyl ester, is orally available. Tebipenem exhibits slow tight-binding inhibition at low micromolar concentrations versus the chromogenic substrate nitrocefin. FT-ICR mass spectrometry demonstrated that the tebipenem acyl–enzyme complex remains stable for greater than 90 min and exists as mixture of the covalently bound drug and the bound retro-aldol cleavage product. We have also determined the high-resolution crystal structures of the BlaC–tebipenem covalent acylated adduct (1.9 Å) with wild-type BlaC and the BlaC–tebipenem Michaelis–Menten complex (1.75 Å) with the K73A BlaC variant. These structures are compared to each other and to other carbapenem–BlaC structures.