Porathoor Sini, Choudhury Asha Rani, Chakrabarti Rajarshi, Anand Ruchi
Department of Chemistry, Indian Institute of Technology Bombay, Mumbai, Maharashtra 400076, India.
Nucleic Acids Res. 2025 Mar 20;53(6). doi: 10.1093/nar/gkaf171.
Nucleobase deaminases are important players in maintaining a stringent nucleobase pool and enhancing genetic diversity via judicious base editing. Here, we delineate the mechanism of Mycobacterium smegmatis deaminase, Msd, found predominantly in Mycobacterium species, that selectively catalyzes the deamination of mutagenic bases. Molecular dynamic studies reveal the dynamic nature of unique structural insertions that cycle between 'closed' and 'open' states, enabling zinc-assisted deamination by occluding the solvent. Corroborating X-ray crystallographic and biochemical studies assert that the appropriate length of the two gating loops and proper positioning of the di-proline motif they harbor are paramount to effective closure. Analysis reveals that although both natural base deaminases, guanine and cytosine deaminase, operate via a similar gating mechanism to Msd, they employ topologically differentially placed structural elements to achieve the 'closed' form. The comparison shows that Mycobacterium deaminases lack the dual-proton shuttle, which renders them ineffective for the deamination of natural bases but allows them to selectively target mutagenic s-triazine scaffolds, thereby imparting innate immunity against these drugs. The study highlights how topologically unique insertions in the cytidine deaminase fold play a critical role in harnessing evolutionary versatility, responsible in imparting fidelity for a nucleobase deamination reaction.
核碱基脱氨酶在维持严格的核碱基库以及通过明智的碱基编辑增强遗传多样性方面发挥着重要作用。在此，我们阐述了耻垢分枝杆菌脱氨酶（Msd）的作用机制，该酶主要存在于分枝杆菌属物种中，可选择性地催化诱变碱基的脱氨反应。分子动力学研究揭示了独特结构插入片段的动态特性，这些片段在“闭合”和“开放”状态之间循环，通过封闭溶剂实现锌辅助脱氨。X射线晶体学和生化研究证实，两个门控环的适当长度以及它们所包含的双脯氨酸基序的正确定位对于有效闭合至关重要。分析表明，尽管天然碱基脱氨酶，即鸟嘌呤脱氨酶和胞嘧啶脱氨酶，通过与Msd类似的门控机制发挥作用，但它们采用拓扑位置不同的结构元件来实现“闭合”形式。比较结果显示，分枝杆菌脱氨酶缺乏双质子穿梭机制，这使得它们对天然碱基的脱氨反应无效，但却能选择性地靶向诱变的三嗪支架，从而赋予对这些药物的天然免疫性。该研究突出了胞苷脱氨酶折叠中拓扑独特的插入片段如何在利用进化多样性方面发挥关键作用，这种多样性负责赋予核碱基脱氨反应的保真度。
