The 伪-Amino Group of the Threonine Substrate as The General Base During tRNA Aminoacylation: A New Version of Substrate-Assisted Catalysis Predicted by Hybrid DFT

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• 作者：WenJuan Huang ; Eric A. C. Bushnell ; Christopher S. Francklyn ; James W. Gauld
• 刊名：Journal of Physical Chemistry A
• 出版年：2011
• 出版时间：November 17, 2011
• 年：2011
• 卷：115
• 期：45
• 页码：13050-13060
• 全文大小：1330K
• 年卷期：v.115,no.45(November 17, 2011)
• ISSN：1520-5215

文摘

Density functional theory-based methods in combination with large chemical models have been used to investigate the mechanism of the second half-reaction catalyzed by Thr-tRNA synthetase: aminoacyl transfer from Thr-AMP onto the _A763鈥睴H of the cognate tRNA. In particular, we have examined pathways in which an active site His309 residue is either protonated or neutral (i.e., potentially able to act as a base). In the protonated His309-assisted mechanism, the rate-limiting step is formation of the tetrahedral intermediate. The barrier for this step is 155.0 kJ mol^鈥?, and thus, such a pathway is concluded to not be enzymatically feasible. For the neutral His309-assisted mechanism, two models were used with the difference being whether Lys465 was included. For either model, the barrier of the rate-limiting step is below the upper thermodynamic enzymatic limit of 125 kJ mol^鈥?. Specifically, without Lys465, the rate-limiting barrier is 122.1 kJ mol^鈥? and corresponds to a rotation about the tetrahedral intermediate C_carb鈥揙H bond. For the model with Lys465, the rate-limiting barrier is slightly lower and corresponds to the formation of the tetrahedral intermediate. Importantly, for both 鈥渘eutral His309鈥?models, the neutral amino group of the threonyl substrate directly acts as the proton acceptor; in the formation of the tetrahedral intermediate, the _A763鈥睴H proton is directly transferred onto the Thr-NH₂. Therefore, the overall mechanism follows a general substrate-assisted catalytic mechanism.