Unraveling the stereochemical and dynamic aspects of the catalytic site of bacterial peptidyl-tRNA hydrolase.
Unraveling the stereochemical and dynamic aspects of the catalytic site of bacterial peptidyl-tRNA hydrolase.
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DOI：
10.1261/rna.057620.116
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发表时间：
2017-02
期刊：
影响因子：
--
通讯作者：
Arora A
中科院分区：
文献类型：
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作者：
Kabra A;Shahid S;Pal RK;Yadav R;Pulavarti SV;Jain A;Tripathi S;Arora A
细菌肽基-tRNA 水解酶（Pth；EC 3.1.1.29）水解细胞质中积累的肽基-tRNA，从而通过减轻 tRNA 饥饿来防止细胞死亡。对霍乱弧菌 Pth (VcPth) 及其参与催化的关键残基突变体的 X 射线和 NMR 研究表明，该蛋白质的活性和选择性取决于残基 H24、D97、N118 和 N14 的立体化学和动力学。 D97-H24 相互作用对于活性至关重要，因为它增加了 H24 的亲核性。 N118 和 N14 与 H24 具有正交竞争相互作用，两者都会降低 H24 的亲核性，并且可能通过肽基-tRNA 底物的定位来抵消。靠近 H24 的区域和盖子区域表现出缓慢的运动，这可以帮助容纳基板。螺旋α3表现出缓慢的摆动，其N-帽残基N118的中间时间尺度运动，其可以作为捕蝇纸来定位底物的可断裂酯键。总体而言，N14、H24、D97 和 N118 侧链之间相互作用的动力学控制该酶对底物的催化。
Bacterial peptidyl-tRNA hydrolase (Pth; EC 3.1.1.29) hydrolyzes the peptidyl-tRNAs accumulated in the cytoplasm and thereby prevents cell death by alleviating tRNA starvation. X-ray and NMR studies of Vibrio cholerae Pth (VcPth) and mutants of its key residues involved in catalysis show that the activity and selectivity of the protein depends on the stereochemistry and dynamics of residues H24, D97, N118, and N14. D97-H24 interaction is critical for activity because it increases the nucleophilicity of H24. The N118 and N14 have orthogonally competing interactions with H24, both of which reduce the nucleophilicity of H24 and are likely to be offset by positioning of a peptidyl-tRNA substrate. The region proximal to H24 and the lid region exhibit slow motions that may assist in accommodating the substrate. Helix α3 exhibits a slow wobble with intermediate time scale motions of its N-cap residue N118, which may work as a flypaper to position the scissile ester bond of the substrate. Overall, the dynamics of interactions between the side chains of N14, H24, D97, and N118, control the catalysis of substrate by this enzyme.