Structures of KlenTaq DNA Polymerase Caught While Incorporating C5-Modified Pyrimidine and C7-Modified 7-Deazapurine Nucleoside Triphosphates
Structures of KlenTaq DNA Polymerase Caught While Incorporating C5-Modified Pyrimidine and C7-Modified 7-Deazapurine Nucleoside Triphosphates
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DOI：
10.1021/ja3017889
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发表时间：
2012-07-25
影响因子：
15
通讯作者：
Marx, Andreas
中科院分区：
文献类型：
--
作者：
Bergen, Konrad;Steck, Anna-Lena;Marx, Andreas
DNA聚合酶接受化学修饰核苷酸的能力对于许多生物技术应用是至关重要的。虽然这些类似物被广泛使用，但人们对非天然核苷酸替代物的结构基础的探索还很少。在本论文中，我们提出了6种修饰的2‘-脱氧核苷-5’-O-三磷酸盐(DNTPs)的晶体结构，这些dNTPs与DNA聚合酶和引物/模板络合物中的嘧啶的CS位或7-去氮嘌呤的C7位进行了修饰。修饰的dNTPs处于催化作用的位置，从而导致掺入。这些结构数据提供了对加入和接受修改后的dNTPs的机制的洞察。我们的结果为修饰核苷酸的合理设计打开了大门，这将为未来的应用提供巨大的机会。
The capability of DNA polymerases to accept chemically modified nucleotides is of paramount importance for many biotechnological applications. Although these analogues are widely used, the structural basis for the acceptance of the unnatural nucleotide surrogates has been only sparsely explored. Here we present in total six crystal structures of modified 2'-deoxynucleoside-5'-O-triphosphates (dNTPs) carrying modifications at the CS positions of pyrimidines or C7 positions of 7-deazapurines in complex with a DNA polymerase and a primer/template complex. The modified dNTPs are in positions poised for catalysis leading to incorporation. These structural data provide insight into the mechanism of incorporation and acceptance of modified dNTPs. Our results open the door for rational design of modified nucleotides, which should offer great opportunities for future applications.