Characterization of the active site of DNA polymerase β by molecular dynamics and quantum chemical calculation
Characterization of the active site of DNA polymerase β by molecular dynamics and quantum chemical calculation
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通过分子动力学和量子化学计算表征 DNA 聚合酶 β 的活性位点
DOI：
10.1002/prot.10451
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
T. Straatsma
中科院分区：
文献类型：
--
作者：
R. Rittenhouse;W. Apostoluk;John H. Miller;T. Straatsma
已经确定，DNA修复酶聚合酶β（pol β）的完全形成的聚合酶活性位点（包括两个结合的Mg 2+阳离子和三磷酸核苷（dNTP）底物）仅存在于化学核苷酸转移步骤之前的催化循环中的一个点。活性构象的结构一直是非常感兴趣的主题，因为它涉及到化学步骤的机制，也涉及到保真度保证的问题。尽管三元聚合物β-（引物-模板）DNA-dNTP复合物的晶体结构提供了活性位点的主要结构特征，但由于有意的改变（例如，从引物和底物中除去3′OH基团），以获得循环中期的结构。从最接近完全形成的活性位点的晶体结构[蛋白质数据库（PDB）代码：1bpy]开始，对溶剂化三元复合物进行了两次分子动力学（MD）模拟：一次通过建模将缺失的3′ OH恢复到引物和底物中，另一次没有恢复3′ OH。模拟结果以及对简化的活性中心模型的从头算优化表明，晶体结构中缺失的引物3′OH是催化阳离子配位层中的显著扰动的原因，并允许我们对晶体学观察到的活性中心结构提出几项修正和补充。此外，计算有助于解决有关配位体的质子化状态所提出的问题。蛋白质2003;53：000-000.© 2003 Wiley利斯公司
It is well established that the fully formed polymerase active site of the DNA repair enzyme, polymerase β (pol β), including two bound Mg2+ cations and the nucleoside triphosphate (dNTP) substrate, exists at only one point in the catalytic cycle just prior to the chemical nucleotidyl transfer step. The structure of the active conformation has been the subject of much interest as it relates to the mechanism of the chemical step and also to the question of fidelity assurance. Although crystal structures of ternary pol β–(primer‐template) DNA–dNTP complexes have provided the main structural features of the active site, they are necessarily incomplete due to intentional alterations (e.g., removal of the 3′OH groups from primer and substrate) needed to obtain a structure from midcycle. Working from the crystal structure closest to the fully formed active site [Protein Data Bank (PDB) code: 1bpy], two molecular dynamics (MD) simulations of the solvated ternary complex were performed: one with the missing 3′OHs restored, via modeling, to the primer and substrate, and the other without restoration of the 3′OHs. The results of the simulations, together with ab initio optimizations on simplified active‐site models, indicate that the missing primer 3′OH in the crystal structure is responsible for a significant perturbation in the coordination sphere of the catalytic cation and allow us to suggest several corrections and additions to the active‐site structure as observed by crystallography. In addition, the calculations help to resolve questions raised regarding the protonation states of coordinating ligands. Proteins 2003;53:000–000. © 2003 Wiley‐Liss, Inc.
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影响因子：
2.9
作者：
Larsen, TM;Wedekind, JE;Reed, GH
通讯作者：
Reed, GH
影响因子：
2.9
作者：
B. G. Werneburg;J. Ahn;X. Zhong;R. Hondal;V. Kraynov;M. Tsai
通讯作者：
B. G. Werneburg;J. Ahn;X. Zhong;R. Hondal;V. Kraynov;M. Tsai
DOI：
10.1016/0027-5107(95)00117-4
发表时间：
1996-02-19
影响因子：
2.3
作者：
Loeb, LA;Christians, FC
通讯作者：
Christians, FC
影响因子：
2.9
作者：
Pelletier, H;Sawaya, MR;Kraut, J
通讯作者：
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影响因子：
56.9
作者：
PELLETIER, H;SAWAYA, MR;KRAUT, J
通讯作者：
KRAUT, J