The oxidation of Met to methionine sulfoxide (MetSO) by oxidants such as hydrogen peroxide, hypochlorite, or peroxynitrite has profound effects on protein function. This modification can be reversed by methionine sulfoxide reductases (msr). In the context of pathogen infection, the reduction of oxidized proteins gains significance due to microbial oxidative damage generated by the immune system. For example, Mycobacterium tuberculosis (Mt) utilizes msrs (MtmsrA and MtmsrB) as part of the repair response to the host-induced oxidative stress. The absence of these enzymes makes Mycobacteria prone to increased susceptibility to cell death, pointing them out as potential therapeutic targets. This study provides a detailed characterization of the catalytic mechanism of MtmsrA using a comprehensive approach, including experimental techniques and theoretical methodologies. Confirming a ping-pong type enzymatic mechanism, we elucidate the catalytic parameters for sulfoxide and thioredoxin substrates (k_cat/K_M = 2656 ± 525 M^–1 s^–1 and 1.7 ± 0.8 × 10⁶ M^–1 s^–1, respectively). Notably, the entropic nature of 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. This mechanistic insight not only advances our understanding of Mt antioxidant enzymes but also holds implications for future drug discovery and biotechnological applications.

中文翻译：

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结核分枝杆菌甲硫氨酸亚硫酸还原酶A的催化机制

过氧化氢、次氯酸盐或过氧亚硝酸盐等氧化剂将 Met 氧化为甲硫氨酸亚砜 (MetSO)，这对蛋白质功能具有深远的影响。这种修饰可以通过甲硫氨酸亚砜还原酶 (msr) 逆转。在病原体感染的情况下，由于免疫系统产生的微生物氧化损伤，氧化蛋白质的减少变得重要。例如，结核分枝杆菌( Mt ) 利用 msrs（Mt msrA 和Mt msrB）作为对宿主诱导的氧化应激的修复反应的一部分。这些酶的缺乏使得分枝杆菌更容易发生细胞死亡，从而将它们视为潜在的治疗靶点。本研究采用综合方法（包括实验技术和理论方法）详细描述了Mt msrA 的催化机制。为了确认乒乓型酶促机制，我们阐明了亚砜和硫氧还蛋白底物的催化参数（分别为k _cat / K _M = 2656 ± 525 M ^–1 s ^–1和 1.7 ± 0.8 × 10 ⁶ M ^–1 s ^–1） ）。值得注意的是，强调了活化过程热力学的熵性质，代表室温下活化自由能的~85%。此外，目前的研究对硫烷中间体的合理性提出质疑，该中间体可能是一种过渡态结构，表明保守的组氨酸残基作为酸碱催化剂参与 MetSO 还原机制。这种机制的见解不仅增进了我们对Mt抗氧化酶的理解，而且对未来的药物发现和生物技术应用具有重要意义。