Elucidating the Chemistry and Targets of Cross-linking by Endogenous DNA Damage

阐明内源性 DNA 损伤交联的化学原理和靶点

• 批准号: 8532857
• 负责人: Sarah C. Shuck
• 金额: $ 3.86万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8532857
• 关键词: Amino Acids; Area; Cell physiology; Characteristics; Chemistry; Chronic; DNA; DNA Adduction; DNA Adducts; DNA Damage; DNA lesion; DNA-Binding Proteins; Detection; Development; Digestion; Erythro; Etiology; Exhibits; Exposure to; Genome Stability; Genomics; Goals; Inflammation; Investigation; Lead; Link; Lipid Peroxidation; Lysine; Maintenance; Malondialdehyde; Mammalian Cell; Metabolism; Methodology; Methods; Modeling; Modification; Mutagenesis; Nuclear Extract; Nuclear Proteins; Oligonucleotides; Oxygen; Phospholipids; Post-Translational Protein Processing; Production; Proteins; Proteomics; Reaction; Reactive Oxygen Species; Reagent; Research; Site; Source; Techniques; adduct; base; carcinogenesis; crosslink; environmental agent; insight; novel; oxidation

DESCRIPTION (provided by applicant): Reactive oxygen species (ROS) produced from cellular metabolism and exposure to environmental agents pose a significant threat to genomic stability. Direct DNA oxidation by ROS or modification by products of lipid peroxidation result in the formation of mutagenic DNA adducts that are implicated in carcinogenesis. A variety of DNA adducts are produced following DNA oxidation including 3-(2!-deoxy- "-D-erythro-pentofuranosyl)- pyrimido[1,2-a]purin-10(3H)-one (M1dG) and N6-oxopropenyl-dA (OPdA). M1dG and OPdA possess the unique characteristic of electrophilic reactivity, which enables them to react with a variety of nucleophiles. OPdA is the more reactive adduct and forms covalent cross-links with N#-acetyl lysine. This raises the possibility that OPdA forms cross-links with DNA-binding proteins. The chemistry of this DNA-protein cross-link formation is not well understood and presents an exciting and novel area for discovery. I propose to use EcoRI as a model DNA-binding protein to elucidate the chemistry of cross-link formation and to optimize the methods for detection and identification of adducted proteins. I will then identify the proteins cross-linked to OPdA adducts in nuclear extracts of mammalian cells. The formation of covalent cross-links between DNA damage and protein has not been extensively investigated despite the detrimental cellular effects these conjugates can induce. The studies outlined in this proposal will increase our understanding of how cross-links are formed as well as what proteins are able to form cross-links with OPdA. This investigation represents the first study to elucidate the interaction of OPdA with DNA-binding proteins.

描述（由申请人提供）：由细胞代谢产生的活性氧（ROS）和暴露于环境药物对基因组稳定性构成重大威胁。通过ROS进行直接DNA氧化或通过脂质过氧化的产物修饰导致与癌变有关的诱变DNA加合物的形成。 DNA氧化后产生多种DNA加合物，包括3-（2！-deoxy-“ -d-d-d-derthro-戊戊酰基）-Fyrimido [1,2-A] Purin-100（3H）-ON（M1DG） - 一个（M1DG）和N6-氧丙基苯基苯基 - DA（OPDA）。多种核粒子是反应性的，与n＃乙酰基赖氨酸的交叉链接增加了OPDA形成与DNA结合蛋白的交叉链接的可能性。交叉链接形成的化学和优化蛋白质的检测和鉴定，我将在哺乳动物细胞的核提取物中识别出与OPDA加合物的交叉链接。该提案中概述的研究将增加我们对如何形成交联的理解，以及哪些蛋白质能够与OPDA形成交联。这项研究是阐明OPDA与DNA结合蛋白相互作用的首次研究。