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首页> 外文学位 >Synthesis of fluorinated analogs of oxidative DNA lesions and their use to probe features of recognition and repair by base excision repair glycosylases.
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Synthesis of fluorinated analogs of oxidative DNA lesions and their use to probe features of recognition and repair by base excision repair glycosylases.

机译：氧化性DNA损伤的氟化类似物的合成及其在探测碱基切除修复糖基化酶识别和修复特征中的用途。

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摘要

Damage to DNA occurs through various sources, both exogenous and endogenous. Base lesions resulting from the oxidation of guanine include 8-oxo-7,8-dihydroguanine (OG), guanidinohydantoin (Gh), and spiroiminodihydantoin (Sp). These modified bases can form mismatches and lead to a G → C and G → T transversion mutations in DNA. Fortunately, these lesions are recognized and removed by DNA repair glycosylases found in all organisms. This work explores the features of recognition and repair of these oxidized guanine lesions by a diverse set of base excision glycosylases (MutY, Fpg, hOGG1, Nei, hNEIL1) using synthesized 2' fluorinated nucleosides such as 2'-fluoro-adenosine (FA), 2'-fluoro-8-oxo-guanosine (FOG), 2'-fluoro-guanidinohydan-toin (FGh), and 2'-fluoro-spiroiminodihydantoin (FSp).;FA and FOG nucleosides in both ribo and arabino sugar conformations have been synthesized and incorporated into oligonucleotide duplexes. Furthermore, oxidation of FOG within an oligonucleotide strand yielded FGh and FSp, both with the ribo and arabino sugar conformation. FA is not a substrate for the adenine glycosylase MutY. However, FA with the fluorine in arabino conformation binds to MutY about 4 fold tighter than the fluorine in the ribo conformation. Similar studies with FOG, FGh, and FSp analogues showed Fpg and its functional homologous human endonuclease VIII-like DNA glycosylase (hNEIL1) can recognize all three lesion containing oligos efficiently. FOG is not a substrate for Fpg, Nei and hNEIL1, and FGh and FSp have increased resistance to cleavage compared to Gh and Sp with all these glycosylases. The sugar conformation of F-nucleotide influences the extent of cleavage as well as binding affinity of the glycosyalse is an important factor to affect the recognition and repair of these lesions.;Duplex stability with unmodified and modified nucleosides have been evaluated by melting temperature measurements, and results revealed duplex destabilization caused by introducing these lesions may be a factor leading to improved recognition of these analogues. X-ray crystallography studies about Fpg and NEIL1 bound to DNA duplexes containing these analogues are ongoing.;In addition, transition state analogue 1-aza-deoxyribose (1N) was synthesized and the crystal structures of Bacillus stearothermophilus (Bs) MutY bound to DNA duplexes containing 1N were determined. These structures provide the first view of an intermediate step of the MutY reaction and necessitate modification of previously proposed models for the mechanism of MutY.
机译：对DNA的损害是通过外源和内源的多种来源发生的。鸟嘌呤氧化产生的基础病变包括8-oxo-7,8-dihydroguanine（OG），胍基乙内酰脲（Gh）和spiroiminodihydantoin（Sp）。这些修饰的碱基可形成错配并导致DNA中的G→C和G→T颠换突变。幸运的是，所有生物中都存在的DNA修复糖基化酶可以识别并清除这些病变。这项工作探索了使用合成的2'氟化核苷（例如2'-氟-腺苷（FA）），通过多种碱基切除糖基化酶（MutY，Fpg，hOGG1，Nei，hNEIL1）识别和修复这些氧化的鸟嘌呤损伤的特征。 ，2'-氟-8-氧代鸟苷（FOG），2'-氟-胍基乙内酰脲（FGh）和2'-氟-螺二胺基乙内酰脲（FSp）.;核糖和阿拉伯糖构象中的FA和FOG核苷已合成并掺入寡核苷酸双链体中。此外，寡核苷酸链内的FOG氧化产生FGh和FSp，具有核糖和阿拉伯糖构象。 FA不是腺嘌呤糖基化酶MutY的底物。然而，具有阿拉伯糖构象的氟的FA与MutY的结合比核糖构象的氟紧密约4倍。对FOG，FGh和FSp类似物的类似研究表明，Fpg及其功能同源的人类核酸内切酶VIII样DNA糖基化酶（hNEIL1）可以有效识别所有三个病变的寡核苷酸。 FOG不是Fpg，Nei和hNEIL1的底物，并且与所有这些糖基化酶的Gh和Sp相比，FGh和FSp对切割的抗性增强。 F-核苷酸的糖构象影响切割的程度以及糖基化酶的结合亲和力是影响这些损伤的识别和修复的重要因素。通过熔融温度测量评估了未修饰和修饰核苷的双链体稳定性，结果表明，由引入这些病变引起的双链不稳定可能是导致这些类似物识别能力提高的一个因素。有关与包含这些类似物的DNA双链体结合的Fpg和NEIL1的X射线晶体学研究正在进行中;此外，合成了过渡态类似物1-氮杂脱氧核糖（1N），并且嗜热脂肪芽孢杆菌（Bs）MutY的晶体结构与DNA结合确定含有1N的双链体。这些结构提供了MutY反应中间步骤的第一个视图，并且有必要修改先前提出的MutY机制的模型。

著录项

  • 作者

    Cao, Sheng.;

  • 作者单位

    The University of Utah.;

  • 授予单位 The University of Utah.;
  • 学科 Chemistry Biochemistry.;Chemistry Organic.
  • 学位 Ph.D.
  • 年度 2010
  • 页码 292 p.
  • 总页数 292
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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