Exploring the Catalytic Mechanism of Human Glutamine Synthetase by Computer Simulations.
Exploring the Catalytic Mechanism of Human Glutamine Synthetase by Computer Simulations.
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通过计算机模拟探索人谷氨酰胺合成酶的催化机制
DOI：
10.1021/acs.biochem.6b00822
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发表时间：
2016
期刊：
影响因子：
2.9
通讯作者：
Bartesaghi S
中科院分区：
文献类型：
--
作者：
Issoglio FM;Campolo N;Zeida A;Grune T;Radi R;Estrin DA;Bartesaghi S
谷氨酰胺合成酶是一种重要的酶，可催化谷氨酸和氨依赖 ATP 形成谷氨酰胺。在哺乳动物中，它在预防大脑兴奋性毒性和肝脏解氨毒方面发挥着关键作用。在植物和细菌中，它是氮代谢的基础，对于生物体的生存至关重要。在这项工作中，我们展示了如何使用经典分子动力学模拟和多尺度量子力学/分子力学模拟来检查人类谷氨酰胺合成酶（HsGS）的结构特性和动力学，以及原子级细节催化过程中涉及的反应机制。我们的结果表明，谷氨酰胺的形成通过两步机制进行，其中包括第一步，其中γ-谷氨酰磷酸中间体形成，具有5 kcal/mol自由能垒和-8 kcal/mol反应自由能，然后是涉及氨亲核攻击的第二个限速步骤，具有19 kcal/mol自由能垒和几乎为零的反应自由能。对每个步骤中结构特征的详细分析揭示了反应热力学和动力学中与蛋白质残基和底物相关的酸碱平衡的相关性。这些结果提供了对 HsGS 动力学的全面研究，并为进一步分析自然变异和翻译后修饰中发生的 HsGS 活性变化奠定了基础。
Glutamine synthetase is an important enzyme that catalyzes the ATP-dependent formation of glutamine from glutamate and ammonia. In mammals, it plays a key role in preventing excitotoxicity in the brain and detoxifying ammonia in the liver. In plants and bacteria, it is fundamental for nitrogen metabolism, being critical for the survival of the organism. In this work, we show how the use of classical molecular dynamics simulations and multiscale quantum mechanics/molecular mechanics simulations allowed us to examine the structural properties and dynamics of human glutamine synthetase (HsGS), as well as the reaction mechanisms involved in the catalytic process with atomic level detail. Our results suggest that glutamine formation proceeds through a two-step mechanism that includes a first step in which the γ-glutamyl phosphate intermediate forms, with a 5 kcal/mol free energy barrier and a −8 kcal/mol reaction free energy, and then a second rate-limiting step involving the ammonia nucleophilic attack, with a free energy barrier of 19 kcal/mol and a reaction free energy of almost zero. A detailed analysis of structural features within each step exposed the relevance of the acid–base equilibrium related to protein residues and substrates in the thermodynamics and kinetics of the reactions. These results provide a comprehensive study ofHsGS dynamics and establish the groundwork for further analysis regarding changes inHsGS activity, as occur in natural variants and post-translational modifications.
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影响因子：
7.4
作者：
Melo, Paula M.;Silva, Liliana S.;Carvalho, Helena G.
通讯作者：
Carvalho, Helena G.
影响因子：
5.7
作者：
van Rooyen, Jason M.;Abratt, Valerie R.;Sewell, Trevor
通讯作者：
Sewell, Trevor
影响因子：
15
作者：
Crespo, A;Martí, MA;Roitberg, AE
通讯作者：
Roitberg, AE
谷氨酰胺合成酶的催化机制。
影响因子：
4.8
作者：
F. Wedler;B. Horn
通讯作者：
B. Horn
DOI：
10.1042/bj3280159
发表时间：
1997-11
期刊：
The Biochemical journal
影响因子：
--
作者：
Chad D. Listrom;H. Morizono;Basavapatna S. Rajagopal;Mark T. McCANN;Mendel Tuchman;N. Allewell
通讯作者：
Chad D. Listrom;H. Morizono;Basavapatna S. Rajagopal;Mark T. McCANN;Mendel Tuchman;N. Allewell