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首页> 外文学位 >Lesion bypass by human DNA polymerase iota: Molecular modeling and dynamics simulations of a Y-family polymerase.
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Lesion bypass by human DNA polymerase iota: Molecular modeling and dynamics simulations of a Y-family polymerase.

机译：人类DNA聚合酶iota的病变绕过：Y族聚合酶的分子建模和动力学模拟。

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摘要

Human DNA polymerase iota is a lesion bypass polymerase of the Y family, capable of incorporating nucleotides opposite a variety of lesions. With undamaged templating purines polymerase iota normally favors Hoogsteen base pairing with syn template. This unusual base pairing is well suited to insertion opposite guanine minor groove lesions, placing the lesion on the spacious major groove side of the polymerase. However, counterintuitively, certain major groove lesions are also bypassed by poli. We have used molecular modeling and dynamics simulations to investigate this phenomenon. Primer extension studies have shown that poli is capable of near-error-free nucleotide incorporation opposite the bulky major groove adduct N-(deoxyguanosin-8-yl)-2-acetyl-aminofluorene (dG-AAF). Our results suggest that normal Watson-Crick base pairing with template anti could be employed in poli for bypass of dG-AAF. In poli with Hoogsteen paired dG-AAF the bulky AAF moiety would reside on the cramped minor groove side of the template. The Hoogsteen-capable conformation distorts the active site, disrupting interactions necessary for incorporation of dC opposite the lesion. Watson-Crick pairing places the AAF rings on the spacious major groove side, similar to the position of minor groove adducts observed with Hoogsteen pairing. Watson-Crick paired structures show a well-ordered active site, with a near reaction-ready ternary complex. Futhermore, polymerase iota can incorporate nucleotides opposite a 10S-(+)-trans-anti -[BP]-N6-dA lesion (dA*); while mainly error-free, the identity of misincorporated bases is influenced by local sequence context. Our results suggest that hydrogen bonds between the benzo[ a]pyrenyl moiety and nearby bases encourage the templating base to maintain the anti glycosidic bond conformation in the binary complex in a 5'-CAGA*TT-3' sequence. This facilitates correct incorporation of dT via a Watson-Crick pair. In a 5'-TTTA*GA-3' sequence the lesion does not form these hydrogen bonds, permitting dA* to rotate around the glycosidic bond to syn and incorporate dT via a Hoogsteen pair. With syn dA* there is also an opportunity for increased misincorporation of dGTP. Together these results expand our understanding of polymerase iota's lesion bypass capabilities. Specifically, our results suggest that polymerase iota can bypass not only minor groove lesions utilizing Hoogsteen base pairing but also major groove lesions utilizing Watson-Crick base pairing.
机译：人DNA聚合酶iota是Y家族的一种病变旁路聚合酶，能够掺入与多种病变相对的核苷酸。使用未损坏的模板嘌呤时，聚合酶iota通常偏爱带有Syn模板的Hoogsteen碱基配对。这种不寻常的碱基配对非常适合插入鸟嘌呤小沟相对的病灶，将病灶置于聚合酶的宽大沟一侧。但是，与直觉相反，某些主要的凹槽病变也被poli绕过。我们已经使用分子建模和动力学模拟来研究这种现象。引物延伸研究表明，poli具有与大主沟加合物N-（脱氧鸟苷-8-基）-2-乙酰基氨基芴（dG-AAF）对立的几乎无错误的核苷酸掺入能力。我们的结果表明，正常的Watson-Crick碱基与模板抗的配对可用于poli，以绕过dG-AAF。与Hoogsteen配对的dG-AAF结合使用时，庞大的AAF部分将位于模板狭窄的小凹槽侧。具有Hoogsteen功能的构象扭曲了活性位点，破坏了在病变对面掺入dC所必需的相互作用。沃森-克里克（Watson-Crick）配对将AAF环放置在宽大的主凹槽一侧，类似于在Hoogsteen配对中观察到的小凹槽加合物的位置。沃森-克里克（Watson-Crick）的配对结构显示出一个有序的活性位点，具有接近反应的三元复合物。此外，聚合酶iota可以掺入与10S-（+）-反式-反-[-BP] -N6-dA病变（dA *）相对的核苷酸。尽管基本没有错误，但并入错误的碱基的身份受本地序列上下文影响。我们的结果表明，苯并[a]苯甲酰基部分与附近碱基之间的氢键鼓励模板化碱基在5'-CAGA * TT-3'序列中保持二元复合物中的抗糖苷键构象。这有助于通过Watson-Crick对正确地合并dT。在5'-TTTA * GA-3'序列中，病变不形成这些氢键，从而使dA *围绕糖苷键旋转以合成并通过Hoogsteen对掺入dT。使用syn dA *，还有机会增加dGTP的错误掺入。这些结果共同扩展了我们对聚合酶iota病变旁路功能的理解。具体而言，我们的结果表明，聚合酶Iota不仅可以利用Hoogsteen碱基配对绕过小沟纹病变，而且还可以利用Watson-Crick碱基配对绕过大沟纹病变。

著录项

  • 作者

    Donny-Clark, Kerry.;

  • 作者单位

    New York University.;

  • 授予单位 New York University.;
  • 学科 Biology Molecular.
  • 学位 Ph.D.
  • 年度 2009
  • 页码 329 p.
  • 总页数 329
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

  • 入库时间 2022-08-17 11:38:26

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