Separation-of-Function Mutants Unravel the Dual-Reaction Mode of Human 8-Oxoguanine DNA Glycosylase
Separation-of-Function Mutants Unravel the Dual-Reaction Mode of Human 8-Oxoguanine DNA Glycosylase
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DOI：
10.1016/j.str.2010.09.023
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发表时间：
2011-01-12
期刊：
影响因子：
5.7
通讯作者：
Bjoras, Magnar
中科院分区：
文献类型：
--
作者：
Dalhus, Bjorn;Forsbring, Monika;Bjoras, Magnar
7,8-二氢-8-氧鸟嘌呤（8oxoG）是DNA中活性氧与鸟嘌呤反应时形成的主要诱变碱基。人类8oxoG DNA糖基化酶（hOgg1）识别并启动8oxoG的修复。hOgg1被认为是一种双功能的DNA糖基酶，它催化去除受损的碱基，然后切割中间碱基DNA的主干（AP裂解酶/6-消除酶）。当作用于含有8oxog的DNA时，hOgg1催化的这两个步骤被认为是耦合的，其中Lys249是一个关键残基。然而，一些证据表明，同时独立的n -糖基键的单功能水解是hOgg1在体内的相关反应模式。在这里，我们通过设计功能分离突变体提供了hOgg1单功能模式的生化和结构证据。Asp268被鉴定为催化残基，而Lys249在水解反应中对8oxoG的特异性识别和最终定位至关重要。
7,8-Dihydro-8-oxoguanine (8oxoG) is a major mutagenic base lesion formed when reactive oxygen species react with guanine in DNA. The human 8oxoG DNA glycosylase (hOgg1) recognizes and initiates repair of 8oxoG. hOgg1 is acknowledged as a bifunctional DNA glycosylase catalyzing removal of the damaged base followed by cleavage of the backbone of the intermediate abasic DNA (AP lyase/6-elimination). When acting on 8oxoG-containing DNA, these two steps in the hOgg1 catalysis are considered coupled, with Lys249 implicated as a key residue. However, several lines of evidence point to a concurrent and independent monofunctional hydrolysis of the N-glycosylic bond being the in vivo relevant reaction mode of hOgg1. Here, we present biochemical and structural evidence for the monofunctional mode of hOgg1 by design of separation-of-function mutants. Asp268 is identified as the catalytic residue, while Lys249 appears critical for the specific recognition and final alignment of 8oxoG during the hydrolysis reaction.