By the year 2050, it is projected that deaths attributed to antimicrobial resistant infections will surpass that of cancer as a leading cause of death. Mycobacterial species, including <i>Mycobacterium tuberculosis</i>, cause deadly infections and require lengthy, costly treatments. Mycobacterial species are highly resistant to commonly prescribed antibiotics in part because of their unique cell wall containing a differentially cross-linked peptidoglycan (PG) layer compared to other bacterial species. The L,D transpeptidase (Ldt) enzyme class is associated with cross-linking and remodeling the PG layer. Six monophyletic clades of Ldts have been identified, with each of these classes predicted to possess a different role in PG biosynthesis. However, the specific reactions catalyzed by these enzymes are largely unknown. By better understanding each class’ role and the reactions catalyzed by paralogous Ldts, we can design antibiotics that therapeutically target mycobacterial infections more effectively and in doing so we can better treat deadly infections like tuberculosis. <br/><br/>Accessing the putative Ldt PG substrates is synthetically challenging, but in order to understand the reactions catalyzed by evolutionarily distinct Ldts, we need to perform substrate activity relationship studies. To this end, we are enzymatically generating Ldt substrates via PG biosynthetic enzymes. Here, we present the cloning and purification of these biosynthetic enzymes and preliminary characterization of putative Ldt substrates. We anticipate the data from these studies will provide insight into the physiological roles of these enzymes, thereby informing future drug strategies toward combating mycobacterial infections.

Enzymatic synthesis of mycobacterial L,D-transpeptidase substrates

ACS Spring 2022

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