In vitro cross-linking of peptidoglycan by Mycobacterium tuberculosis L,D-transpeptidases and inactivation of these enzymes by carbapenems.

In vitro cross-linking of peptidoglycan by Mycobacterium tuberculosis L,D-transpeptidases and inactivation of these enzymes by carbapenems.

Antimicrob Agents Chemother. 2013 Sep 16;

Authors: Cordillot M, Dubée V, Triboulet S, Dubost L, Marie A, Hugonnet JE, Arthur M, Mainardi JL

Abstract
The Mycobacterium tuberculosis peptidoglycan is mainly cross-linked by l,d-transpeptidases (LDTs), which are efficiently inactivated by a single β-lactam class, the carbapenems. Development of carbapenems for tuberculosis treatment has recently raised considerable interest since these drugs, in association with the β-lactamase inhibitor clavulanic acid, are uniformly active against extensively drug-resistant M. tuberculosis and kill both exponentially growing and dormant forms of the bacilli. We have purified the five l,d-transpeptidase paralogues of M. tuberculosis (Mt1 to 5) and compared their activities with peptidoglycan fragments and carbapenems. The five LDTs were functional in vitro since they were active in the peptidoglycan cross-linking (Mt5), β-lactam acylation (Mt3), or both (Mt1, Mt2 and Mt4) assays. Mt3 was the only LDT that was inactive in the cross-linking assay suggesting that this enzyme might be involved in other cellular functions such as the anchoring of proteins to peptidoglycan as shown in Escherichia coli. Inactivation of LDTs by carbapenems is a two-step reaction comprising reversible formation of a tetrahedral intermediate, the oxyanion, followed by irreversible rupture of the β-lactam ring that leads to formation of a stable acylenzyme. Determination of the rate constants for these two steps revealed important differences (up to 460 fold) between carbapenems, which affected the velocity of oxyanion and acylenzyme formation. Imipenem inactivated LDTs more rapidly than ertapenem and both drugs were more efficient than meropenem and doripenem indicating that modification of the carbapenem side chain could be used to optimize their anti-mycobacterial activity.

PMID: 24041897 [PubMed - as supplied by publisher]