Catalytic Mechanism of Mycobacterium tuberculosis Methionine Sulfoxide Reductase A

Abstract

The oxidation of Met to methionine sulfoxide(MetSO) by oxidants such as hydrogen peroxide, hypochlorite, orperoxynitrite has profound effects on protein function. Thismodification can be reversed by methionine sulfoxide reductases(msr). In the context of pathogen infection, the reduction ofoxidized proteins gains significance due to microbial oxidativedamage generated by the immune system. For example,Mycobacterium tuberculosis (Mt) utilizes msrs (MtmsrA andMtmsrB) as part of the repair response to the host-inducedoxidative stress. The absence of these enzymes makes Mycobacteriaprone to increased susceptibility to cell death, pointing them out aspotential therapeutic targets. This study provides a detailedcharacterization of the catalytic mechanism of MtmsrA using acomprehensive approach, including experimental techniques andtheoretical methodologies. Confirming a ping-pong type enzymatic mechanism, we elucidate the catalytic parameters for sulfoxideand thioredoxin substrates (k cat /K M = 2656 ± 525 M −1 s −1 and 1.7 ± 0.8 × 10 6 M −1 s −1 , respectively). Notably, the entropic natureof the activation process thermodynamics, representing ∼85% of the activation free energy at room temperature, is underscored.Furthermore, the current study questions the plausibility of a sulfurane intermediate, which may be a transition-state-like structure,suggesting the involvement of a conserved histidine residue as an acid−base catalyst in the MetSO reduction mechanism. Thismechanistic insight not only advances our understanding of Mt antioxidant enzymes but also holds implications for future drugdiscovery and biotechnological applications.