Ribonucleotide Reductase Regulation: Diferric Y* assembly/maintenance and Sml1

核糖核苷酸还原酶调节：Diferric Y* 组装/维护和 Sml1

• 批准号: 7802291
• 负责人: JOANNE STUBBE
• 金额: $ 40.07万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-02 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7802291
• 关键词: Affinity Chromatography; Amaze; Anabolism; Arabinose; Binding; Biochemical Genetics; C-terminal; Candidate Disease Gene; Cell Death; Cells; Chemicals; Chemistry; Complex; Crystallography; Cytochromes b5; DNA; DNA Repair; DNA biosynthesis; Drug usage; Electrons; Ensure; Escherichia coli; Escherichia coli Proteins; Ferredoxin; Fission Yeast; Flavodoxin; Free Radicals; Genetic; Goals; Growth; Hematologic Neoplasms; Homologous Gene; Housing; Human; In Vitro; Iron; Kinetics; Label; Lead; Link; Maintenance; Malignant Neoplasms; Modeling; Monitor; Mossbauer Spectroscopy; Mutagenesis; Nucleotides; Organism; Outcome; Pathway interactions; Phase II Clinical Trials; Phosphorylation; Phosphorylation Site; Phosphotransferases; Play; Predisposition; Process; Proteins; Reaction; Recycling; Reducing Agents; Regulation; Relative (related person); Ribonucleotide Reductase; Role; Saccharomyces cerevisiae; Site; Source; Substrate Specificity; Testing; Triapine; Viral; Yeasts; cofactor; crosslink; drug development; hydroxyurea; in vivo; inhibitor/antagonist; interest; mutant; new therapeutic target; nucleic acid metabolism; protein degradation; public health relevance; repaired; research study; tumor

DESCRIPTION (provided by applicant): Ribonucleotide reductases (RNRs) catalyze the conversion of nucleotides to deoxynucleotides in all organisms and provide the monomeric precursors required for DNA replication and DNA repair. The class I RNRs are composed of two subunits: the ?n subunit binds the four NDP substrates and the allosteric effectors (NTPs and dATP) that govern substrate specificity and turnover rate. The ?? subunit houses the essential diferric-tyrosyl radical (Y) cofactor required to initiate the chemically difficult reduction process on ?n. The active RNR complex is ?n?2. Regulation of RNRs is largely responsible for controlling the relative ratios and amounts of the dNTP pools, which is critical to the fidelity of DNA replication and repair. Loss of this control can lead to cell death, genetic instability, and in humans, a predisposition to cancer. RNR's central role in nucleic acid metabolism has made them the successful target in the treatment of a number of malignancies. Regulation of RNR activity occurs at multiple levels: transcriptionally, by control of the subcellular localization of ?n and ?2, by the binding of allosteric effectors (ATP, dNTPs to ?n), by control of protein degradation, by control of the concentration of the Y generated by the di-iron metallo-cofactor, and by small protein inhibitors. This proposal focuses on the regulation of the class I RNRs from E. coli and S. cerevisiae. Two regulatory mechanisms will be examined using an integration of biochemical and genetic approaches. The first and second specific aims are to elucidate the biosynthetic and maintenance (repair) pathways of the essential diferric-Y cofactor of ?2 in E. coli and S. cerevisiae. Experiments are presented to identify the assembly factors required for iron and reducing equivalent delivery. The Y of the cofactor is the target of hydroxyurea used in the treatment of hematologic malignancies and of triapine in phase II clinical trials. Thus understanding whether the clusters can be repaired can have dramatic outcomes clinically. The third specific aim in S. cerevisiae is to understand mechanism of the small proteins: Sml1 and the newly discovered Spd1, in RNR inhibition. This understanding could identify a new therapeutic target. The long-range goal is to understand quantitatively how all of the regulatory mechanisms are integrated to control cellular dNTPs pools under different growth conditions. PUBLIC HEALTH RELEVANCE: Ribonucleotide reductases catalyze the conversion of nucleotides to deoxynucleotides in all organisms. Their regulation is essential for controlling dNTP pools, critical for the fidelity of DNA replication and repair. Two regulatory mechanisms are examined in this proposal; understanding these mechanisms could lead to new therapeutic targets.

描述（由申请人提供）：核糖核苷酸还原酶（RNR）催化核苷酸向所有生物中的脱氧核苷酸的转化，并提供DNA复制和DNA修复所需的单体前体。 I类RNR由两个亚基组成：？N亚基结合了四个NDP底物和控制基板特异性和周转率的变构效应子（NTP和DATP）。这 ？？亚基包含启动化学困难还原过程所需的必需差异 - 自由基（y）辅助因子。活性RNR复合物是？n？2。 RNR的调节主要负责控制DNTP池的相对比率和量，这对于DNA复制和修复的保真度至关重要。失去这种控制会导致细胞死亡，遗传不稳定性和人类对癌症的倾向。 RNR在核酸代谢中的核心作用使它们成为治疗多种恶性肿瘤的成功靶标。 RNR活性的调节在多个级别上发生：转录，通过控制蛋白质降解，通过控制蛋白质降解，通过控制蛋白质降解的结合，通过控制蛋白质降解的结合，通过控制蛋白质降解的结合，通过控制di-riron Metallo-Metallo-Metallo-cof-cofactactor和By-by-bysyby protein protein nibnibitors nibitors n bysiration。该提议着重于大肠杆菌和酿酒酵母的I类RNR的调节。将使用生化和遗传方法的整合来检查两种调节机制。第一个和第二个特定的目的是阐明基本差异辅助因子的生物合成和维持（修复）途径是大肠杆菌和酿酒酵母中的。提出了实验，以确定铁和减少等效输送所需的组装因子。辅因子的Y是用于治疗血液系统恶性肿瘤和三丁丁氏菌的羟基脲的靶标。因此，了解是否可以修复簇可以在临床上产生巨大的结果。酿酒酵母中的第三个具体目的是了解小蛋白质的机制：SML1和新发现的SPD1，在RNR抑制中。这种理解可以确定一个新的治疗靶点。远程目标是定量地了解所有调节机制如何在不同的生长条件下整合以控制细胞DNTPS池。公共卫生相关性：核糖核苷酸还原酶催化核苷酸向所有生物体中脱氧核苷酸的转化。它们的调节对于控制DNTP池至关重要，这对于DNA复制和修复的保真度至关重要。该提案中检查了两种监管机制。了解这些机制可能导致新的治疗靶标。