STRUCTURAL STUDIES OF DNA GLYCOSYLASES WITH DAMAGED DNA

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。DNA四个碱基的自发损伤是引起癌症的突变的主要原因。大多数这些遗传病变是通过一种被称为碱基切除DNA修复（BER）的途径纠正的。BER的关键成分是DNA糖基酶，这种酶扫描基因组寻找特定种类的碱基损伤，然后通过催化从DNA主干上切除受损碱基来启动修复过程。我们研究的长期目标是了解它们利用的确切反应途径。对DNA损伤识别和去除的全面、基本的理解代表了解决肿瘤发生之谜的一个主要方面。在本研究中，我们将重点关注DNA糖基化酶修复DNA中8-氧鸟嘌呤碱基的详细反应途径。关于DNA糖基酶（尤其是hOgg1）对底物识别和催化的结构生物学，文献中有大量的工作。在这里提出的工作中，我们试图从结构上表征hOgg1反应途径中的早期中间体，其中hOgg1对受损DNA进行采样。通过设计合适的突变体并使用共价捕获策略，我们能够在催化途径的不同状态下捕获酶。通过获得这些中间体的高分辨率结构，我们的目标是了解整个催化途径的不同组成部分。参考文献：Nat Struct Biol. 2003年3月；生物化学。2003年2月18日；42(6):1564-72。《自然》2000年2月24日；403 (6772): 859 - 66

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Spontaneous damage to the four bases of DNA is a major cause of the mutations that give rise to cancer. Most of these genetic lesions are corrected by a pathway known as base-excision DNA repair (BER). The key components of BER are DNA glycosylases, enzymes that scan the genome in search of particular kinds of base damage, then initiate the repair process by catalyzing excision of the damaged base from the DNA backbone. The long-term goals of our studies are to understand the precise reaction pathways that they utilize. A comprehensive, fundamental understanding of DNA damage recognition and removal represents the solution to a major aspect of the tumorigenesis puzzle. In the proposed studies, we will focus on the detailed reaction pathway of base-excision repair by the DNA glycosylase, human Ogg1 protein which repairs 8-oxoguanine bases in DNA. There is an ample body of work in the literature on the structural biology of substrate recognition and catalysis by DNA glycosylases, especially hOgg1. In the proposed work here, we are trying to structurally characterize early intermediates in the reaction pathway of hOgg1 where it is sampling damaged DNA. By designing suitable mutants and using a covalent trapping strategy, we are able to trap the enzyme in different states of the catalytic pathway. By obtaining high resolution structures of these intermediates, we aim to understand the different components of the entire catalytic pathway. References: Nat Struct Biol. 2003 Mar;10(3):204-11 Biochemistry. 2003 Feb 18;42(6):1564-72. Nature. 2000 Feb 24;403(6772):859-66