Crystal structure of DNA polymerase from hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1
Crystal structure of DNA polymerase from hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1
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DOI：
10.1006/jmbi.2000.4403
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发表时间：
2001-02-23
影响因子：
5.6
通讯作者：
Kai, Y
中科院分区：
文献类型：
--
作者：
Hashimoto, H;Nishioka, M;Kai, Y
确定了来自超嗜热古菌 Kodakaraensis KOD1 的 B 族 DNA 聚合酶（KOD DNA 聚合酶）的晶体结构。 KOD DNA 聚合酶具有已知的最高延伸率、持续合成能力和保真度。我们对 KOD DNA 聚合酶进行了结构分析，以阐明这些酶特征的机制。来自超嗜热古菌的 DNA 聚合酶的结构比较突出了 Thumb 结构域的构象差异。 KOD DNA 聚合酶的 Thumb 结构域显示“开放”构象。手指子结构域在聚合酶活性位点的侧面具有许多碱性残基。这些残基被认为可以通过静电相互作用接触到传入的 dNTP。 β-发夹基序（残基 242-249）从核酸外切酶 (Exo) 结构域延伸，如噬菌体 RB69 的 RB69 DNA 聚合酶的编辑复合物所示。许多精氨酸残基位于KOD DNA聚合酶的分叉点（模板结合和编辑裂口的连接处），表明碱性环境适合引物和模板DNA双链体的分配以及在高温环境下稳定部分熔化的DNA结构。分叉点处熔融 DNA 结构的稳定可能与 KOD DNA 聚合酶的高 PCR 性能相关，这是由于低错误率、高延伸率和持续加工能力。 (C) 2001 年学术出版社。
The crystal structure of family B DNA polymerase from the hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1 (KOD DNA polymerase) was determined. KOD DNA polymerase exhibits the highest known extension rate, processivity and fidelity. We carried out the structural analysis of KOD DNA polymerase in order to clarify the mechanisms of those enzymatic features. Structural comparison of DNA polymerases from hyperthermophilic archaea highlighted the conformational difference in Thumb domains. The Thumb domain of KOD DNA polymerase shows an "opened" conformation. The fingers subdomain possessed many basic residues at the side of the polymerase active site. The residues are considered to be accessible to the incoming dNTP by electrostatic interaction. A beta -hairpin motif (residues 242-249) extends from the Exonuclease (Exo) domain as seen in the editing complex of the RB69 DNA polymerase from bacteriophage RB69. Many arginine residues are located at the forked-point (the junction of the template-binding and editing clefts) of KOD DNA polymerase, suggesting that the basic environment is suitable for partitioning of the primer and template DNA duplex and for stabilizing the partially melted DNA structure in the high-temperature environments. The stabilization of the melted DNA structure at the forked-point may be correlated with the high PCR performance of KOD DNA polymerase, which is due to low error rate, high elongation rate and processivity. (C) 2001 Academic Press.