双重DNA和DNA复制过程中的常见突变性氧化DNA损伤的结构动力学
Benjamin J Ryan1, Haozhe Yang2, Jan Henric T Bacurio3
1Department of Biochemistry and Molecular Biology, and Department of Cancer Biology, University of Kansas Medical Center, Kansas City, Kansas 66160, United States.
Journal of the American Chemical Society
|May 2, 2022
概括
甲基胺 (Fapy•dG) 的DNA损伤比8-oxodGuo更具有突变性. 它的分子动力学和与DNA聚合酶β的相互作用影响了突变性和聚合酶忠实性.
科学领域:
- 生物化学
- 分子生物学
- 结构生物学
背景情况:
- N6-(2-Deoxy-α,β-d-erythro-pentofuranosyl) -2,6-diamino-4-hydroxy-5-formamido pyrimidine (Fapy•dG) 是一个显著的基因组DNA损伤,特别是在无氧条件下.
- 与相关的7,8-二-8-oxo-2'-deoxyguanosine (8-oxodGuo) 相比,Fapy•dG在哺乳动物细胞中具有更高的突变性.
- Fapy•dG的易于表皮化对理解其化学行为和生物影响提出了挑战.
研究的目的:
- 在双重DNA中的Fapy•dG及其与DNA聚合酶β (Polβ) 的相互作用的结晶学特征.
- 阐明Fapy•dG异体动力学 (α和β) 在基配对和聚合酶活性中的作用.
- 确定Fapy•dG的突变性潜力及其加工中的Pol β的贡献.
主要方法:
- 用X射线结晶学测定双重DNA和Polβ中的Fapy•dG结构.
- 硫浸泡实验以捕获环开放的Fapy•dG.
- 在HEK 293T细胞 (野生类型和Pol β knockdown) 中进行突变性测试.
- 通过Polβ对抗Fapy•dG的核酸结合的动力学研究.
主要成果:
- 根据配对的基质 (细胞因子或腺因) 的不同,Fapy•dG采用不同的异构形式 (α和β).
- 结晶学数据显示,Fapy•dG的环开放和表皮化可以在水分子的帮助下发生在晶体状态下.
- 在Fapy•dG突变中,Pol β起着双重作用,促进G → T转换,同时抑制G → A转换.
- 动力分析显示dCMP与β- 异构体相比 (∼90倍) 的整合显著更快,影响了聚合酶的忠实性.
结论:
- 对于了解这种病变的核酸结合和DNA修复,Fapy•dG的异构动态至关重要.
- Pol β与Fapy•dG的相互作用影响其突变性结果,突出显示了该酶在病变处理中的作用.
- 这项研究提供了Fapy•dG独特的化学特性及其生物后果的结构和动态见解.
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