The role of cysteines in the response of 8-oxoguanine glycosylase (OGG1) to oxidative stress

半胱氨酸在 8-氧代鸟嘌呤糖基化酶 (OGG1) 氧化应激反应中的作用

• 批准号: 9980416
• 负责人: Khadijeh Alnajjar
• 金额: $ 9.88万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980416
• 关键词: 8-Oxoguanine DNA Glycosylase; 8-hydroxyguanosine; Affect; Aging; Arsenic; Autoimmune Diseases; Award; B-Cell Activation; BCL2 gene; Binding; Biochemical; Biochemistry; Biological; Biological Assay; Biological Markers; Cell Culture Techniques; Cell physiology; Cells; Chromatin; Comet Assay; Complement; Cysteine; DNA; DNA Binding; DNA Damage; DNA Repair; DNA Repair Gene; DNA glycosylase; DNA lesion; Data; Disease; Disease Outcome; Drug Design; Drug Targeting; Ecology; Educational workshop; Electrophoretic Mobility Shift Assay; Enhancers; Excision; Exposure to; Foundations; Frequencies; Future; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic Transcription; Goals; Grant; Guanine; Heavy Metals; Herbicides; Histones; Homeostasis; Hydrogen Peroxide; Immunofluorescence Immunologic; Immunoprecipitation; Individual; Inflammatory; Lead; Lesion; Ligands; Light; Lyase; Lysine; Malignant Neoplasms; Manuscripts; Mass Spectrum Analysis; Measures; Mediating; Mentors; Modification; Molecular; Molecular Conformation; Mutagenesis; Mutate; Mutation; NEIL3 gene; Neurodegenerative Disorders; Nuclear; Oxidation-Reduction; Oxidative Stress; Oxides; POLB gene; Paraquat; Pharmacotherapy; Phase; Post-Translational Protein Processing; Postdoctoral Fellow; Preparation; Process; Promoter Regions; Proteins; Proteomics; Reactive Oxygen Species; Research; Role; Running; Signal Transduction; Single Nucleotide Polymorphism; Site; Site-Directed Mutagenesis; Specificity; Structure; Substrate Specificity; Sulfhydryl Compounds; TNF gene; Technical Expertise; Testing; Therapeutic; Toxic Environmental Substances; Toxin; Training; Universities; Vascular Endothelial Growth Factor C; Vascular Endothelial Growth Factors; Work; Writing; XRCC1 gene; biochemical tools; biophysical tools; career development; chromatin remodeling; demethylation; design; histone demethylase; human disease; in vitro testing; in vivo; insight; mutant; oxidation; oxidative DNA damage; prevent; programs; promoter; protein protein interaction; recruit; repaired; response; stopped-flow fluorescence; student mentoring; symposium; therapy design; tool; transcription factor; transversion mutation

Project Summary/Abstract: Oxidative stress caused from exposure to environmental toxins results in DNA damage that is repaired by the DNA repair machinery. 8-oxoguanine (8-oxoG) is the most commonly formed DNA lesion, which is recognized and removed by 8-oxoguanine glycosylase (OGG1). The function of OGG1 is important to prevent propagation of DNA error and to mediate transcription of genes responsible for the response to oxidative stress. The inability of cells to perform these tasks can lead to autoimmune and neurodegenerative diseases, cancer, and aging. In addition to DNA oxidation, cysteines can also be targeted for oxidative modification under oxidative stress. OGG1 contains 8 cysteine residues which are potentially targeted for modification in the presence of environmental toxins. This proposal seeks to elucidate the role of cysteine in OGG1 in DNA repair and transcription in response to oxidative stress. Biochemical characterization of OGG1 cysteine mutants is proposed in specific aim 1 to look at the function of cysteine in DNA binding, enzymatic activity, conformational changes, and in protein-protein interactions. These studies will be initiated during the K99 mentored phase and then continued independently during the R00 phase. The second aim will focus on in vivo cell culture work to understand the role of cysteine in DNA repair and in gene regulation and also to identify OGG1 modifications resulting from environmental toxin treatment. The proposed work in this aim will be initiated during the mentored phase, and the technical expertise gained during the mentored phase will be employed during my independent work to accomplish the proposed goals. This work will clarify the role of OGG1 modification in disease initiation and progression and will help in predicting the biological effect of toxin exposure. This work will also provide a foundation for targeted drug design. The long-term goal for this award is to transition into an independent academic research program to explore the comprehensive molecular mechanism of the DNA damage response under oxidative stress and how modifications in the DNA repair machinery lead to disease. To achieve the goals of this award, I assembled a team of mentors with expertise in environmental science, proteomics, redox biochemistry, DNA glycosylases, DNA repair, and gene transcription. This team will also provide me with mentoring in career development for transitioning into independence and establishing and running a successful lab. Further training will be acquired from attending special topic workshops and courses on grant writing and career development offered both at and outside of Yale University. Additional opportunities to attend and present at conferences and to mentor students and postdocs along with continued preparation of manuscripts will be complementary for my long-term goal of establishing an independent research program. These tools will be essential to gain technical training and for career development to establish a successful research program to study the molecular mechanism of the DNA damage response under oxidative stress and its role in disease.

项目摘要/摘要：暴露于环境毒素引起的氧化应激导致DNA 由DNA修复机器修复的损伤。8-氧鸟嘌呤(8-oxoG)是最常见的形式 DNA损伤，由8-氧鸟嘌呤糖基酶(OGG1)识别和去除。OGG1的功能是 对于防止DNA错误的传播和调节负责基因转录非常重要 对氧化应激的反应。细胞无法执行这些任务可导致自身免疫和 神经退行性疾病、癌症和衰老。除了DNA氧化，半胱氨酸也可以成为靶点 用于氧化应激下的氧化修饰。OGG1含有8个半胱氨酸残基，它们可能是 在存在环境毒素的情况下进行目标修改。这项建议旨在阐明 半胱氨酸在氧化应激下DNA修复和转录中的作用。生化特性 OGG1半胱氨酸突变体是在特定目标1中提出的，以考察半胱氨酸在DNA结合中的功能， 酶活性、构象变化和蛋白质-蛋白质相互作用。这些研究将启动 在K99指导阶段，然后在R00阶段独立继续。第二个目标是 关注体内细胞培养工作，以了解半胱氨酸在DNA修复和基因调控中的作用 也是为了确定环境毒素处理导致的OGG1修饰。拟在这方面开展的工作 AIM将在指导阶段启动，并在指导阶段获得技术专业知识 将受雇于我的独立工作期间，以完成所提出的目标。这项工作将澄清这一角色 OGG1修饰在疾病发生和发展中的作用，并将有助于预测OGG1的生物学效应 暴露在毒素下。这项工作也将为靶向药物设计提供基础。这方面的长期目标是 该奖项是向独立的学术研究计划过渡，探索全面的分子 氧化应激下DNA损伤反应的机制及DNA修复中的修饰 机器会导致疾病。为了实现这个奖项的目标，我组建了一个具有以下专业知识的导师团队 环境科学、蛋白质组学、氧化还原生物化学、DNA糖基酶、DNA修复和基因 抄写。这个团队还将为我提供职业发展方面的指导，帮助我过渡到 独立自主，建立和运营一个成功的实验室。进一步的培训将通过参加 在境内外举办关于赠款撰写和职业发展的专题讲习班和课程 耶鲁大学。更多出席会议并在会议上发言的机会，以及指导学生和 博士后和手稿的持续准备将对我的长期目标起到补充作用 建立独立的研究计划。这些工具对于获得技术培训和 职业发展建立了一个成功的研究计划，研究DNA的分子机制 氧化应激下的损伤反应及其在疾病中的作用。

项目成果

The role of cysteines in the structure and function of OGG1.

• DOI: 10.1074/jbc.ra120.016126
• 发表时间: 2021-01
• 期刊: The Journal of biological chemistry
• 作者: Wang K;Maayah M;Sweasy JB;Alnajjar KS
• 通讯作者: Alnajjar KS