Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560 012, India.
Acta Crystallogr D Struct Biol. 2017 Apr 1;73(Pt 4):349-364. doi: 10.1107/S2059798317002534. Epub 2017 Mar 31.
Mycobacterium smegmatis MutT1, which is made up of a Nudix domain (domain 1) and a histidine phosphatase domain (domain 2), efficiently hydrolyses 8-oxo-GTP and 8-oxo-dGTP to the corresponding nucleoside diphosphates and phosphate in the presence of magnesium ions. Domain 1 alone hydrolyses nucleoside triphosphates less efficiently. Under high concentrations and over long periods, the full-length enzyme as well as domain 1 catalyses the hydrolysis of the nucleoside triphosphates to the respective nucleoside monophosphates and pyrophosphate. The role of domain 2 appears to be limited to speeding up the reaction. Crystal structures of the apoenzyme and those of ligand-bound enzyme prepared in the presence of 8-oxo-GTP or 8-oxo-dGTP and different concentrations of magnesium were determined. In all of the structures except one, the molecules arrange themselves in a head-to-tail fashion in which domain 1 is brought into contact with domain 2 (trans domain 2) of a neighbouring molecule. The binding site for NTP (site A) is almost exclusively made up of residues from domain 1, while those for NDP (site B) and NMP (site C) are at the interface between domain 1 and trans domain 2 in an unusual instance of intermolecular interactions leading to binding sites. Protein-ligand interactions at site A lead to a proposal for the mechanism of hydrolysis of NTP to NDP and phosphate. A small modification in site A in the crystal which does not exhibit the head-to-tail arrangement appears to facilitate the production of NMP and pyrophosphate from NTP. The two arrangements could be in dynamic equilibrium in the cellular milieu.
分枝杆菌 MutT1 由 Nudix 结构域（结构域 1）和组氨酸磷酸酶结构域（结构域 2）组成，在镁离子存在的情况下，能有效地将 8-氧代-GTP 和 8-氧代-dGTP 水解为相应的核苷二磷酸和磷酸。单独的结构域 1 对核苷三磷酸的水解效率较低。在高浓度和长时间下，全长酶以及结构域 1 催化核苷三磷酸水解为相应的核苷一磷酸和焦磷酸。结构域 2 的作用似乎仅限于加快反应速度。测定了apo 酶和配体结合酶的晶体结构，这些酶是在 8-氧代-GTP 或 8-氧代-dGTP 和不同浓度的镁存在下制备的。除了一个结构外，所有结构中的分子都以头对尾的方式排列，其中结构域 1 与相邻分子的结构域 2（跨结构域 2）接触。NTP 的结合位点（位点 A）几乎完全由结构域 1 的残基组成，而 NDP（位点 B）和 NMP（位点 C）的结合位点位于结构域 1 和跨结构域 2 之间的界面上，这种界面上的相互作用形成了不寻常的结合位点。位于位点 A 的蛋白-配体相互作用导致了 NTP 水解为 NDP 和磷酸的机制的提出。在晶体中，位点 A 发生了一个小的修饰，这种修饰不表现出头对尾的排列，似乎促进了 NTP 生成 NMP 和焦磷酸。这两种排列可能在细胞环境中处于动态平衡状态。
