Plasticity, ligand conformation and enzyme action of Mycobacterium smegmatis MutT1

Mycobacterium smegmatis MutT1 (MsMutT1) is a sanitation enzyme made up of an N-terminal Nudix hydrolase domain and a C-terminal domain resembling a histidine phosphatase. It has been established that the action of MutT1 on 8-oxo-dGTP, 8-oxo-GTP and diadenosine polyphosphates is modulated by intermolecular interactions. In order to further explore this and to elucidate the structural basis of its differential action on 8-oxo-NTPs and unsubstituted NTPs, the crystal structures of complexes of MsMutT1 with 8-oxo-dGTP, GMPPNP and GMPPCP have been determined. Replacement soaking was used in order to ensure that the complexes were isomorphous to one another. Analysis of the structural data led to the elucidation of a relationship between the arrangements of molecules observed in the crystals, molecular plasticity and the action of the enzyme on nucleotides. The dominant mode of arrangement involving a head-to-tail sequence predominantly leads to the generation of NDPs. The other mode of packing arrangement appears to preferentially generate NMPs. This work also provides interesting insights into the dependence of enzyme action on the conformation of the ligand. The possibility of modulating the enzyme action through differences in intermolecular interactions and ligand conformations makes MsMutT1 a versatile enzyme.

耻垢分枝杆菌MutT1（Mycobacterium smegmatis MutT1, MsMutT1）是一种核苷酸清洁酶，由N端Nudix水解酶结构域（N-terminal Nudix hydrolase domain）与类似组氨酸磷酸酶（histidine phosphatase）的C端结构域构成。已有研究证实，该酶对8-氧代-2'-脱氧鸟苷三磷酸（8-oxo-dGTP）、8-氧代鸟苷三磷酸（8-oxo-GTP）以及多磷酸腺苷二酯（diadenosine polyphosphates）的催化活性受分子间相互作用调控。为进一步探究该调控机制，并阐明其对8-氧代核苷三磷酸（8-oxo-NTPs）与未取代核苷三磷酸（unsubstituted NTPs）产生差异化催化作用的结构基础，本研究解析了MsMutT1分别与8-氧代-2'-脱氧鸟苷三磷酸（8-oxo-dGTP）、5'-[α,β-亚氨基]三磷酸鸟苷（GMPPNP）以及5'-[β,γ-亚甲基]三磷酸鸟苷（GMPPCP）形成复合物的晶体结构。为确保各复合物彼此间为同晶型结构，实验采用了置换浸泡法（Replacement soaking）。对结构数据的分析揭示了晶体中分子排布方式、分子可塑性（molecular plasticity）与该酶对核苷酸的催化作用之间的内在关联。其中以头尾相接序列为主导的主要排布模式主要催化生成核苷二磷酸（NDPs）；另一种堆积排布模式则优先催化生成核苷一磷酸（NMPs）。本研究还为酶催化活性依赖于配体构象（ligand conformation）这一观点提供了新颖的见解。通过分子间相互作用与配体构象差异即可调控该酶的催化活性，这一特性使MsMutT1成为一种多功能酶。