Targeting Mycobacterial Transpeptidases: Evaluating the Roles of Ldt and PBP Inhibition in Suppressing Mycobacterium smegmatis

Abstract

{beta}-Lactams demonstrate promising in vitro activity against Mycobacterium species and are being explored for tuberculosis treatment; however, evidence of their in vivo efficacy versus Mycobacterium tuberculosis remains limited. To achieve broad clinically relevant potency, optimisation of the classical {beta}-lactam scaffolds or development of new or non-{beta}-lactam inhibitors for mycobacterial transpeptidases is likely required. In mycobacteria, potential targets of {beta}-lactams include L,D-transpeptidases (Ldts) and penicillin-binding proteins (PBPs). Reports suggest that dual inhibition of Ldts and PBPs may be necessary to achieve effective anti-mycobacterial activity, yet the specific contributions of Ldt and PBP inhibition to the {beta}-lactam antibacterial mechanisms are not understood. We used fluorogenic substrate mimics to investigate the effects of {beta}-lactams and reported LdtMt2 inhibitors on Mycobacterium smegmatis (Msm), assessing their impacts on the transpeptidase activities of Ldts and PBPs in living cells. The results reveal a statistically significant correlation between both Ldt and PBP inhibition and Msm growth suppression; though under the tested conditions a stronger correlation between Ldt inhibition and Msm growth suppression was observed. Notably, inhibition of both PBPs and Ldts was observed in all active inhibitors, though {beta}-lactams manifest increased potency of PBP inhibition. The combination of the {beta}-lactams meropenem and faropenem with selected LdtMt2 inhibitors manifested an additive inhibitory effect against Msm. Our results highlight the importance of further optimising PBPs and Ldt transpeptidase inhibition, particularly of Ldts, to improve {beta}-lactam efficacy versus mycobacteria.