pH-dependent direct sulfhydrylation pathway is required for the pathogenesis of <i>Mycobacterium tuberculosis</i>

Abstract Methionine is essential for the survival of Mycobacterium tuberculosis ( M. tuberculosis ) inside the host. However, the transsulfuration pathway, a major contributor of methionine, is dispensable for the growth of M. tuberculosis suggesting redundancy in the methionine biosynthesis pathway. Orthologues of MetZ TB in other bacterial species are known to operate a redundant single-step methionine biosynthesis pathway called direct sulfhydrylation. In this study, we demonstrate that genetic disruption of the metZ -mediated direct sulfhydrylation pathway in M. tuberculosis hinders growth at low pH, an effect mitigated by methionine supplementation. Computational analyses, including in-silico molecular docking and molecular dynamics (MD) simulations, reveal enhanced binding of the MetZ substrate, O-succinyl homoserine (OSH), to the active site of MetZ at acidic pH. Intriguingly, despite increased intracellular ATP levels, a relative decrease in the frequency of Bedaquiline (BDQ)-induced persisters is observed in metZ -deficient strain, suggesting a role of direct sulfhydrylation pathway in modulating BDQ sensitivity. Finally, we demonstrated that the absence of metZ impedes the ability of M. tuberculosis to grow inside the host.