Mechanistic Insights into the Rate-Limiting Step in Purine-Specific Nucleoside Hydrolase
Mechanistic Insights into the Rate-Limiting Step in Purine-Specific Nucleoside Hydrolase
复制标题
嘌呤特异性核苷水解酶限速步骤的机制见解
DOI：
10.1021/acs.jctc.5b00045
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发表时间：
2015-07-01
影响因子：
5.5
通讯作者：
Cao, Zexing
中科院分区：
文献类型：
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作者：
Chen, Nanhao;Zhao, Yuan;Cao, Zexing
肌苷腺苷鸟苷特异性核苷水解酶（IA（-NH））的一个完整的酶催化循环被认为包括四个步骤：底物结合、化学反应、碱基释放和核糖释放。然而，整个酶促反应的限速步骤的机制细节仍然是未知的，即使核糖释放可能是最困难的阶段。基于最新的量子力学和分子力学（QM/MM）分子动力学（MD）模拟，核糖释放过程可以分为两个步骤：“核糖解离”和“核糖释放”。“核糖解离”包括“切割”和“交换”阶段，其中亚稳的6倍中间体将恢复为Ca 2+的8倍配位壳，如在apo-IAG-NH中观察到的。广泛的随机加速分子动力学和MD模拟已被用来验证合理的释放通道，估计的势垒为整个反应的速率决定步骤是13.0千卡/摩尔，这是可比的实验值16.7千卡/摩尔。此外，已经发现由loop 1和loop 2以及活性口袋周围的关键残基产生的门控机制在操纵核糖释放中起重要作用。
A full enzymatic catalysis cycle in the inosine adenosine guanosine specific nucleoside hydrolase (IA(-NH) was assumed to be comprised of four steps: substrate binding, chemical reaction, base release, and ribose release. Nevertheless, the mechanistic details for the rate-limiting step of the entire enzymatic reaction are still unknown, even though the ribose release was likely to be the most difficult stage. Based on state-of-the-art quantum mechanics and molecular mechanics (QM/MM) molecular dynamics (MD) simulations, the ribose release process can be divided into two steps: "ribose dissociation" and "ribose release". The "ribose dissociation" includes "cleavage" and "exchange" stages, in which a metastable 6-fold intermediate will recover to an 8-fold coordination shell of Ca2+ as observed in apo- IAG-NH. Extensive random acceleration molecular dynamics and MD simulations have been employed to verify plausible release channels, and the estimated barrier for the rate-determining step of the entire reaction is 13.0 kcal/mol, which is comparable to the experimental value of 16.7 kcal/mol. Moreover, the gating mechanism arising from loop1 and loop2, as well as key residues around the active pocket, has been found to play an important role in manipulating the ribose release.