烟酰胺酶催化水解脱氨反应的QM/MM分子动力学模拟

doi:10.7503/cjcu20150365

摘要/Abstract

摘要：

采用QM(PM3)/MM分子动力学(MD)方法模拟了烟酰胺酶催化烟酰胺水解脱氨形成烟碱酸的反应过程. 计算结果表明, 硫的亲核进攻是整个反应的速率控制步骤. 当改变Ala155所在Loop区的位置, 在亲核进攻时, 底物能够自由旋转, 可以加速亲核进攻过程并降低整个催化反应的能垒. 讨论了氨分子的离去过程和质子传递过程的相关细节. 为烟酰胺酶的定点突变以及脱氨酶的设计提供了有益的参考.

关键词: 烟酰胺酶, 脱氨, 反应机理, QM/MM方法, 分子动力学模拟

Abstract:

The deamination process of nicotinamide to nicotinic acid by nicotinamidase simulated with combined QM(PM3)/MM molecular dynamics(MD) method. Our calculated results show that the whole mechanism includes four steps, transition of H_S, nucleophilic attack by sulfur group, nucleophilic attack by hydroxyl group and proton transfer from Asp16 to Cys159. The rate-determined step is the nucleophilic attack by sulfur group. Through conformational analyses, we are able to indicate that high energy consumption mainly because the nucleophilic attack needs to break the hydrogen bond between substrate and Ala155. We suggest that mutation of the loop, which make Ala155 far from nicotinamide, could weaken the hydrogen bond and increase the reactivity of nicotinamidase. Based on this ground, release of ammonia and proton transfer processes are discussed in details. The results could give some useful suggestions to enzyme design and site-directed mutagenesis studies.

Key words: Nicotinamidase, Deamination, Mechanism, Quantum mechanics/molecular mechanics(QM/MM) method, Molecular dynamics simulation(Ed.: Y,Z)

中图分类号:

O641

TrendMD:

段新丽, 张欣, 雷鸣. 烟酰胺酶催化水解脱氨反应的QM/MM分子动力学模拟. 高等学校化学学报, 2015, 36(12): 2491.

DUAN Xinli, ZHANG Xin, LEI Ming. QM/MM Molecular Dynamics Simulation on the Mechanisms for the Hydrolytic Deamination of Nicotinamidase^†. Chem. J. Chinese Universities, 2015, 36(12): 2491.

图/表 8

Scheme 1 Catalytic reaction of nicotinamidase

Fig.1 Crystal structure of nicotinamidase(A)(PDB entry: 2WTA) and details in QM part(B) (A) The active site with nicotinamide is highlighted in yellow; (B) QM/MM boundary is shown by the red break lines.

Fig.2 Reaction pathways for the deamination process in QM/MM MD simulations

Fig.3 Free energy profile from the binding state B to the intermediate state(I1)(A) and fluctuations of the distances of the breaking and forming bonds in HS transfer process(B) RC1 refers to the distance difference between the HS—S@Cys159 and HS—OD3@Asp16.

Fig.4 Free energy profile from the intermediate state(I1) to the intermediate state(I2) via a transition state(TS2)(A) and fluctuations of the distances of the breaking and forming bonds in nucleophilic attack of sulfur(B) RC2 refers to the distance difference between the C7—S@Cys159 and C7—N8.

Fig.5 Snapshots of the micro-environments for QM part at the I1(A), TS2(B) and I2 states(C) Hydrogen bond lengths(nm) in step 2 are shown by the dash lines. The values in parentheses are standard deviations.

Fig.6 Free energy profile from the intermediate state(I2') to the intermediate state(I3) via a transition state(TS3)(A) and fluctuations of the distances of the breaking and forming bonds in nucleophilic attack of hydroxyl group(B) RC3 refers to the minus distance between the OW3—C7 and HW6—OD3.

Fig.7 Free energy profile from the intermediate state(I3) to the product state(P) via a transition state(TS4)(A) and fluctuations of the distances of the breaking and forming bonds in nicotinic acid formation process(B) RC4 refers to the minus distance between the O9—HW6 and HW5—S.

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