Crystal structure of a thermostable type B DNA polymerase from Thermococcus gorgonarius
Crystal structure of a thermostable type B DNA polymerase from Thermococcus gorgonarius
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DOI：
10.1073/pnas.96.7.3600
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发表时间：
1999-03-30
影响因子：
11.1
通讯作者：
Angerer, B
中科院分区：
文献类型：
--
作者：
Hopfner, KP;Eichinger, A;Angerer, B
大多数已知的古生菌DNA聚合酶属于B型家族，也包括真核生物的DNA复制聚合酶，但在极端条件下保持高保真。我们在这里描述了古细菌DNA聚合酶的2.5埃分辨率晶体结构，并确定了折叠的结构特征和可能与其耐热功能有关的活性部位。与噬菌体RB69的中温B型DNA聚合酶gp43相比，突出了嗜热适应，包括在DNA-蛋白质界面存在两个二硫键和增强的静电互补。与gp43相反，核酸外切酶和拇指结构域中的几个环被更紧密地包装；这显然阻止了引物与核酸外切酶活性部位的结合。这种“封闭”构象的生理作用尚不清楚，但可能代表一种聚合酶模式，而不是开放的核酸外切酶位点的编辑模式。这种古老的13DNA聚合酶结构为基于结构的聚合酶或配体的设计提供了起点，这些聚合酶或配体在生物技术和抗病毒或抗癌药物的开发中应用。
Most known archaeal DNA polymerases belong to the type B family, which also includes the DNA replication polymerases of eukaryotes, but maintain high fidelity at extreme conditions. We describe here the 2.5 Angstrom resolution crystal structure of a DNA polymerase from the Archaea Thermococcus gorgonarius and identify structural features of the fold and the active site that are likely responsible for its thermostable function. Comparison with the mesophilic B type DNA polymerase gp43 of the bacteriophage RB69 highlights thermophilic adaptations, which include the presence of two disulfide bonds and an enhanced electrostatic complementarity at the DNA-protein interface. In contrast to gp43, several loops in the exonuclease and thumb domains are more closely packed; this apparently blocks primer binding to the exonuclease active site. A physiological role of this "closed" conformation is unknown but may represent a polymerase mode, in contrast to an editing mode with an open exonuclease site. This archaeal 13 DNA polymerase structure provides a starting point for structure-based design of polymerases or ligands with applications in biotechnology and the development of antiviral or anticancer agents.