In most well-studied rod-shaped bacteria, peptidoglycan is primarily crosslinked by penicillin binding proteins (PBPs). However, in mycobacteria L,D-transpeptidase (LDT)-mediated crosslinks are highly abundant. To elucidate the role of these unusual crosslinks, we characterized mycobacterial cells lacking all LDTs. We find that LDT-mediated crosslinks are required for rod shape maintenance specifically at sites of aging cell wall, a byproduct of polar elongation. Asymmetric polar growth leads to a non-uniform distribution of these two types of crosslinks in a single cell. Consequently, in the absence of LDT-mediated crosslinks, PBP-catalyzed crosslinks become more important. Because of this, Mycobacterium tuberculosis is more rapidly killed using a combination of drugs capable of PBP and LDT inhibition. Thus, knowledge about the single-cell distribution of drug targets can be exploited to more effectively treat this pathogen.

Maturing Mycobacterial Peptidoglycan Requires Non-canonical Crosslinks to Maintain Shape