Mutation of the little finger domain in human DNA polymerase η alters fidelity when copying undamaged DNA.
Mutation of the little finger domain in human DNA polymerase η alters fidelity when copying undamaged DNA.
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DOI：
10.1002/em.21807
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发表时间：
2013-10
影响因子：
2.8
通讯作者：
McCulloch, Scott D.
中科院分区：
文献类型：
--
作者：
Beardslee, Renee A.;Suarez, Samuel C.;Toffton, Shannon M.;McCulloch, Scott D.
DNA 聚合酶 η (pol η) 在 DNA 复制过程中合成过去的环丁烷嘧啶二聚体和可能的 7,8-二氢-8-氧代鸟嘌呤 (8-oxoG) 损伤。 pol η 的丢失与突变率的增加相关，这表明它在突变抑制中不可或缺的作用。最近有报道称，小指区域的 β 链 12（氨基酸 316-324）将模板链与酶的催化核心正确定位。作者假设，β-链 12 残基的修饰会破坏正确的酶-DNA 比对并改变 pol η 的活性和保真度。为了研究这一点，作者纯化了含有聚合酶催化核心的蛋白质，合并单个氨基酸变化以选择 β-链 12 残基，并评估每个 pol η 的 DNA 合成活性。确定复制含有 DNA 的顺式-顺式环丁烷胸腺嘧啶-胸腺嘧啶二聚体和 8-oxoG 损伤时的损伤旁路效率和复制保真度，并与野生型聚合酶的相应值进行比较。结果证实了 β 链在聚合酶功能中的重要性，并表明当复制未损坏的 DNA 时，保真度最常发生改变。此外，研究表明，远离活性位点的 DNA-蛋白质接触会显着影响合成的保真度。
DNA polymerase η (pol η) synthesizes past cyclobutane pyrimidine dimer and possibly 7,8-dihydro-8-oxoguanine (8-oxoG) lesions during DNA replication. Loss of pol η is associated with an increase in mutation rate, demonstrating its indispensable role in mutation suppression. It has been recently reported that β-strand 12 (amino acids 316–324) of the little finger region correctly positions the template strand with the catalytic core of the enzyme. The authors hypothesized that modification of β-strand 12 residues would disrupt correct enzyme–DNA alignment and alter pol η’s activity and fidelity. To investigate this, the authors purified proteins containing the catalytic core of the polymerase, incorporated single amino acid changes to select β-strand 12 residues, and evaluated DNA synthesis activity for each pol η. Lesion bypass efficiencies and replication fidelities when copying DNA-containing cis-syn cyclobutane thymine-thymine dimer and 8-oxoG lesions were determined and compared with the corresponding values for the wild-type polymerase. The results confirm the importance of the β-strand in polymerase function and show that fidelity is most often altered when undamaged DNA is copied. Additionally, it is shown that DNA–protein contacts distal to the active site can significantly affect the fidelity of synthesis.
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