Biochemical Insight of Hyperphosphorylation of Replication Protein A

复制蛋白 A 过度磷酸化的生化见解

• 批准号: 8005163
• 负责人: Yue Zou
• 金额: $ 7.4万
• 依托单位: EAST TENNESSEE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-05 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8005163
• 关键词: A-Form DNA; Academic Research Enhancement Awards; Address; Affect; Affinity; Binding; Biochemical; Biochemical Pathway; Biological; Biomedical Research; Cancer Biology; Cell Cycle Progression; Cell physiology; Cells; Characteristics; Chemicals; DNA; DNA Binding Domain; DNA Damage; DNA Repair; DNA biosynthesis; DNA damage checkpoint; DNA lesion; Data; Defect; Disease; Environment; Environmental Hazards; Fluorescence Resonance Energy Transfer; Fluorescence Spectroscopy; Genetic; Genetic Recombination; Genomic Instability; Goals; Human; Laboratories; Lead; Malignant Neoplasms; Mediating; Metabolic Pathway; Metabolism; Methods; Molecular; Mutagenesis; Mutation; Nerve Degeneration; Phosphorylation; Play; Prevention; Process; Property; Proteins; Regulation; Replication Initiation; Research; Role; SS DNA BP; Series; Signal Transduction; Site; Site-Directed Mutagenesis; Source; Stress; Surface Plasmon Resonance; Testing; Thermodynamics; Time; Work; base; carcinogenesis; career; coping; experience; graduate student; insight; medical schools; repaired; replication factor A; response; ultraviolet irradiation

DESCRIPTION (provided by applicant): Summary The goal of this project is to elucidate the biochemical basis of the functions of human replication protein A (RPA) in DNA damage responses, and to better understand the damage-induced mutagenesis and carcinogenesis. DNA damage has been widely implicated in human cancer induction. Human RPA, the major single-stranded DNA binding protein, is essential for all DNA metabolic pathways such as DNA replication, DNA repair, recombination and DNA damage checkpoints. Given that RPA undergoes hyperphosphorylation in cells in response to DNA damage, it is hypothesized that the protein may play a modulatory role in cellular DNA damage responses mediated by the hyperphosphorylation at the N- terminus of its RPA32 subunit. How the hyperphosphorylation affect the cellular activities of RPA in terms of biochemical mechanism remains unknown. In this study, using an array of rigorous biochemical methods we will: (1) identify domain-domain interactions and structural characteristic of RPA upon DNA damage-induced hyperphosphorylation; (2) characterize the effects of hyperphosphorylation on RPA interaction with DNA intermediates of biological significance; and (3) determine binding affinity and thermodynamics of the hyperphosphorylation-induced domain-domain interaction of RPA. The proposed study is highly relevant to cancer biology as human defects in DNA damage responses lead to genome instability and thus a series of diseases with high potential to evolve cancer. This Academic Research Enhancement Award will allow undergraduate and graduate students to gain biomedical research experience in a medical school environment that will prepare them for careers in scientific research. Specifically, the proposed study will address an important question regarding the biochemical basis of RPA hyperphosphorylation in cellular DNA damage responses. This project is highly relevant to cancer biology as DNA damage is the major cause to human cancer.

本项目的目的是阐明人复制蛋白A（RPA）在DNA损伤反应中功能的生化基础，并更好地理解损伤诱导的突变和致癌作用。DNA损伤在人类癌症诱导中已被广泛涉及。人RPA是主要的单链DNA结合蛋白，在DNA复制、DNA修复、重组和DNA损伤检查点等所有DNA代谢途径中都必不可少。鉴于RPA在细胞中响应于DNA损伤而经历过度磷酸化，假设该蛋白质可能在由其RPA 32亚基的N-末端处的过度磷酸化介导的细胞DNA损伤应答中起调节作用。RPA的过度磷酸化是如何影响RPA的细胞活性的，其生化机制尚不清楚。在本研究中，我们将使用一系列严格的生物化学方法：（1）确定结构域之间的相互作用和结构特征的RPA在DNA损伤诱导的过度磷酸化;（2）表征的影响，RPA与DNA的生物学意义的中间体的相互作用;和（3）确定的结合亲和力和热力学的过度磷酸化诱导的RPA的结构域之间的相互作用。这项研究与癌症生物学高度相关，因为人类在DNA损伤反应中的缺陷会导致基因组不稳定，从而导致一系列具有高度发展癌症潜力的疾病。该学术研究增强奖将允许本科生和研究生在医学院环境中获得生物医学研究经验，为他们的科学研究生涯做好准备。具体来说，拟议的研究将解决一个重要的问题，RPA过度磷酸化的生化基础细胞DNA损伤反应。该项目与癌症生物学高度相关，因为DNA损伤是人类癌症的主要原因。