Clostridioides difficile canonical L,D-transpeptidases catalyze a novel type of peptidoglycan cross-links and are not required for beta-lactam resistance

作者：Nicola F. Galley, Darren Greetham, Marcel G. Alamán-Zárate, Michael P. Williamson, Caroline A. Evans, William Spittal, Jessica E. Buddle, Jane Freeman, Georgina Davis, Mark J. Dickman, Mark H. Wilcox, Andrew L. Lovering, Robert P. Fagan, Stéphane Mesnage · 发表于：Journal of Biological Chemistry · 年份：2023 · DOI：10.1016/j.jbc.2023.105529 · 被引用次数：13 · 研究领域：Clostridium difficile and Clostridium perfringens research、Antimicrobial Resistance in Staphylococcus、Antibiotic Resistance in Bacteria

Clostridioides difficile is the leading cause of antibiotic-associated diarrhoea worldwide with significant morbidity and mortality. This organism is naturally resistant to several beta-lactam antibiotics that inhibit the polymerisation of peptidoglycan, an essential component of the bacteria cell envelope. Previous work has revealed that C. difficile peptidoglycan has an unusual composition. It mostly contains 3-3 cross-links, catalysed by enzymes called L,D-transpeptidases (Ldts) that are poorly inhibited by beta-lactams. It was therefore hypothesized that peptidoglycan polymerization by these enzymes could underpin antibiotic resistance. Here, we investigated the catalytic activity of the three canonical Ldts encoded by C. difficile (Ldt Cd1 , Ldt Cd2 and Ldt Cd3 ) in vitro and explored their contribution to growth and antibiotic resistance. We show that two of these enzymes catalyse the formation of novel types of peptidoglycan cross-links using meso-diaminopimelic (DAP) acid both as a donor and an acceptor, also observed in peptidoglycan sacculi. We demonstrate that the simultaneous deletion of these three genes only has a minor impact on both peptidoglycan structure and resistance to beta-lactams. This unexpected result therefore implies that the formation of 3-3 peptidoglycan cross-links in C. difficile is catalysed by as yet unidentified non-canonical Ldt enzymes.