Structural insights into the promutagenic bypass of the major cisplatin-induced DNA lesion.
Structural insights into the promutagenic bypass of the major cisplatin-induced DNA lesion.
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DOI：
10.1042/bcj20190906
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发表时间：
2020-03-13
期刊：
影响因子：
--
通讯作者：
Lee S
中科院分区：
文献类型：
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作者：
Ouzon-Shubeita H;Vilas CK;Lee S
顺铂-1，2-d（GpG）（Pt-GG）链内交联是顺铂产生的主要DNA损伤。顺铂已被证明主要诱导G至T突变，Pt-GG允许人DNA聚合酶β（polβ）显著错误掺入dATP。一致的是，在癌细胞中经常观察到的polβ过表达与顺铂耐药性和突变表型有关。然而，dATP与Pt-GG相反的错误掺入的结构基础是未知的。在这里，我们报告的第一个结构的DNA聚合酶不准确地绕过Pt-GG。在Mg ~（2+）或Mn ~（2+）存在下，我们解决了polβ在Pt-GG的5′-dG对面错误掺入dATP的两种结构。Mg 2+结合的结构表现出催化的次优构象，而Mn 2+结合的结构是在催化更有利的半封闭构象。在这两种结构中，dATP不与Pt-GG形成共面碱基配对。在polβ活性位点中，与Pt-GG相对的syn-dATP似乎通过蛋白质模板和pi堆积相互作用而稳定，这类似于polβ介导的与脱碱基位点相对的dATP掺入。总体而言，我们的结果表明，模板Pt-GG在polβ活性位点的行为像一个脱碱基位点，促进插入dATP在非指令的方式。
The cisplatin-1,2-d(GpG) (Pt-GG) intrastrand cross-link is the predominant DNA lesion generated by cisplatin. Cisplatin has been shown to predominantly induce G to T mutations and Pt-GG permits significant misincorporation of dATP by human DNA polymerase β (polβ). In agreement, polβ overexpression, which is frequently observed in cancer cells, is linked to cisplatin resistance and a mutator phenotype. However, the structural basis for the misincorporation of dATP opposite Pt-GG is unknown. Here, we report the first structures of a DNA polymerase inaccurately bypassing Pt-GG. We solved two structures of polβ misincorporating dATP opposite the 5′-dG of Pt-GG in the presence of Mg2+ or Mn2+. The Mg2+-bound structure exhibits a sub-optimal conformation for catalysis, while the Mn2+-bound structure is in a catalytically more favorable semi-closed conformation. In both structures, dATP does not form a coplanar base pairing with Pt-GG. In the polβ active site, the syn-dATP opposite Pt-GG appears to be stabilized by protein templating and pi stacking interactions, which resembles the polβ-mediated dATP incorporation opposite an abasic site. Overall, our results suggest that the templating Pt-GG in the polβ active site behaves like an abasic site, promoting the insertion of dATP in a non-instructional manner.