Computational study of the phosphoryl transfer catalyzed by a cyclin-dependent kinase
Computational study of the phosphoryl transfer catalyzed by a cyclin-dependent kinase
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DOI：
10.1002/chem.200700044
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发表时间：
2007-01-01
影响因子：
4.3
通讯作者：
Recanatini, Maurizio
中科院分区：
文献类型：
--
作者：
De Vivo, Marco;Cavalli, Andrea;Recanatini, Maurizio
细胞周期蛋白依赖性激酶CDK2催化从三磷酸腺苷到其多肽底物的苏氨酸或丝氨酸残基的γ-磷酸转移。在这里，我们通过气相密度泛函计算、经典分子动力学和Car-Parrinello QM/MM模拟研究了CdK2的反应机理。我们重点研究了保守的Asp127的作用以及CDK2催化的磷酰化转移反应的本质。我们的发现表明，Asp127通过帮助底物丝氨酸或苏氨酸的近攻击取向的形成，以其去质子化的形式活跃。因此，在催化过程中，残基不起到一般碱的作用。磷酰化转移的机理是一个类S(N)2的协同步骤，呈现类磷烷的过渡态几何构型。尽管得到的反应机理与先前对相同催化反应机理的密度泛函研究是一致的(Cavalli等人，Chem.通讯。2003,1308-1309)，当进行QM/MM计算时，反应势垒相当低，如本研究中(约为42kcalol(-1)qm，而不是约24kcalol(-1)qm/MM)；这表明蛋白质环境和溶剂水在催化中发挥了重要作用。由于细胞周期蛋白依赖性蛋白激酶(CDK)家族的氨基酸序列高度保守，这些结果可能对CDK家族具有普遍意义。
A cyclin-dependent kinase, Cdk2, catalyzes the transfer of the gamma-phosphate from ATP to a threonine or serine residue of its polypeptide substrates. Here, we investigate aspects of the reaction mechanism of Cdk2 by gas-phase density functional calculations, classical molecular dynamics, and Car-Parrinello QM/MM simulations. We focus on the role of the conserved Asp127 and on the nature of the phosphoryl transfer reaction mechanism catalyzed by Cdk2. Our findings suggest that Asp127 is active in its deprotonated form by assisting the formation of the near-attack orientation of the substrate serine or threonine. Therefore, the residue does not act as a general base during the catalysis. The mechanism for the phosphoryl transfer is a single S(N)2-like concerted step, which shows a phosphorane-like transition state geometry. Although the resulting reaction mechanism is in agreement with a previous density functional study of the same catalytic reaction mechanism (Cavalli et al., Chem. Comm. 2003, 1308-1309), the reaction barrier is considerably lower when QM/MM calculations are performed, as in this study (approximate to 42 kcalmol(-1) QM vs approximate to 24 kcalmol(-1) QM/MM); this indicates that important roles for the catalysis are played by the protein environment and solvent waters. Because of the high amino acid sequence conservation among the whole family of cyclin-dependent kinases (CDKs), these results could be general for the CDK family.