Magnesium-induced assembly of a complete DNA polymerase catalytic complex
Magnesium-induced assembly of a complete DNA polymerase catalytic complex
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DOI：
10.1016/j.str.2006.01.011
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发表时间：
2006-04-01
期刊：
影响因子：
5.7
通讯作者：
Wilson, SH
中科院分区：
文献类型：
--
作者：
Batra, VK;Beard, WA;Wilson, SH
核苷酸转移酶反应的分子细节仍然是推测性的，因为捕获用于结构测定的催化中间体的策略利用缺乏引物末端3 '-OH和催化Mg 2+的底物，导致不完整和扭曲的活性位点几何形状。由于这些基本原子的几何排列将影响化学，因此对保真度策略的结构洞察受到阻碍。在这里，我们提出了一个晶体结构的预催化复合物的DNA聚合酶与结合的基板，包括引物3 '-OH和催化Mg 2+。该催化中间体用可去甲羟基化的脱氧核苷酸类似物捕获。比较了两种新的DNA聚合酶β-缺少3 '-OH或催化Mg ~（2+）的结构。这些结构提供了直接的证据，即需要两个原子来实现L3'对进入的核苷酸的α P的在线亲核攻击所必需的适当几何形状。
The molecular details of the nucleotidyl transferase reaction have remained speculative, as strategies to trap catalytic intermediates for structure determination utilize substrates lacking the primer terminus 3'-OH and catalytic Mg2+, resulting in an incomplete and distorted active site geometry. Since the geometric arrangement of these essential atoms will impact chemistry, structural insight into fidelity strategies has been hampered. Here, we present a crystal structure of a precatalytic complex of a DNA polymerase with bound substrates that include the primer 3'-OH and catalytic Mg2+. This catalytic intermediate was trapped with a northydrolyzable deoxynucleotide analog. Comparison with two new structures of DNA polymerase beta-lacking the 3'-OH or catalytic Mg2+ is described. These structures provide direct evidence that both atoms are required to achieve a proper geometry necessary for an in-line nucleophilic attack of L3' on the alpha P of the incoming nucleotide.