Wiederholt Carissa J, Delaney Michael O, Pope Mary Ann, David Sheila S, Greenberg Marc M
Department of Chemistry, Johns Hopkins University, 3400 North Charles Street, Baltimore, Maryland 21218, USA.
Biochemistry. 2003 Aug 19;42(32):9755-60. doi: 10.1021/bi034844h.
Fapy.dG is produced in DNA as a result of oxidative stress. Under some conditions Fapy.dG is formed in greater yields than 8-oxodG from a common chemical precursor. Recently, Fapy.dG and its C-nucleoside analogue were incorporated in chemically synthesized oligonucleotides at defined sites. Like 8-oxodG, Fapy.dG instructs DNA polymerase to misincorporate dA opposite it in vitro. The interactions of DNA containing Fapy.dG or the nonhydrolyzable analogue with Fpg and MutY are described. Fpg excises Fapy.dG (K(M) = 2.0 nM, k(cat) = 0.14 min(-1)) opposite dC approximately 17-fold more efficiently than when mispaired with dA, which is misinserted by DNA polymerase in vitro. Fpg also prefers to bind duplexes containing Fapy.dG.dC or beta-C-Fapy.dG.dC compared to those in which the lesion is opposite dA. MutY incises dA when it is opposite Fapy.dG and strongly binds duplexes containing the lesion or beta-C-Fapy.dG. Incision from Fapy.dG.dA is faster than from dG.dA mispairs but slower than from DNA containing 8-oxodG opposite dA. These data demonstrate that Fapy.dG closely resembles the interactions of 8-oxodG with two members of the GO repair pathway in vitro. The similar effects of Fapy.dG and 8-oxodG on DNA polymerase and repair enzymes in vitro raise the question as to whether Fapy.dG elicits similar effects in vivo.
由于氧化应激，Fapy.dG在DNA中产生。在某些条件下，Fapy.dG从常见化学前体形成的产量高于8-氧代脱氧鸟苷（8-oxodG）。最近，Fapy.dG及其C-核苷类似物被掺入化学合成的寡核苷酸中的特定位点。与8-oxodG一样，Fapy.dG在体外会指导DNA聚合酶在其对面错误掺入dA。本文描述了含有Fapy.dG或其不可水解类似物的DNA与Fpg和MutY的相互作用。Fpg切除与dC相对的Fapy.dG（K(M)=2.0 nM，k(cat)=0.14 min(-1)），其效率比与在体外被DNA聚合酶错误插入的dA错配时高约17倍。与损伤与dA相对的双链体相比，Fpg也更倾向于结合含有Fapy.dG.dC或β-C-Fapy.dG.dC的双链体。当dA与Fapy.dG相对时，MutY会切割dA，并强烈结合含有该损伤或β-C-Fapy.dG的双链体。从Fapy.dG.dA进行的切割比从dG.dA错配进行的切割更快，但比含有与dA相对的8-oxodG的DNA进行的切割更慢。这些数据表明，Fapy.dG在体外与GO修复途径的两个成员的相互作用与8-oxodG非常相似。Fapy.dG和8-oxodG在体外对DNA聚合酶和修复酶的类似作用提出了一个问题，即Fapy.dG在体内是否会引发类似的作用。
