Structural Studies of SUMO Protein Modification

SUMO蛋白修饰的结构研究

• 批准号: 7641113
• 负责人: CHRISTOPHER D. LIMA
• 金额: $ 35.82万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7641113
• 关键词: Address; Apoptosis; Binding; Biochemical; Biochemical Genetics; Biochemistry; Biological; C-terminal; Cell Cycle; Cell Cycle Progression; Cell Cycle Regulation; Cells; Chromosome Segregation; Complex; Cysteine; Cytokinesis; DNA Repair; Dissection; Ensure; Enzymes; Evolution; Funding; Genes; Genetic; Genetic Techniques; Genetic Transcription; Glycine; Health; Human; In Vitro; Kinetics; Ligase; Ligation; Lysine; Malignant Neoplasms; Mediating; Metabolism; Mission; Modification; Molecular; Molecular Weight; Natural regeneration; Nuclear; Pathway interactions; Peptide Hydrolases; Physiological; Play; Post-Translational Protein Processing; Process; Protein Family; Protein Isoforms; Protein Precursors; Proteins; Reagent; Recruitment Activity; Regulation; Research; Research Personnel; Research Proposals; Role; Saccharomyces cerevisiae; Site-Directed Mutagenesis; Small Ubiquitin-Related Modifier Proteins; Specificity; Stress; Structure; Surface; System; Transcriptional Regulation; Translating; Ubiquitin; Ubiquitination; United States National Institutes of Health; Yeasts; adduct; amino group; base; cofactor; in vivo; inhibitor/antagonist; insight; multicatalytic endopeptidase complex; nucleocytoplasmic transport; programs; protein degradation; reconstitution; repaired; response; thioester; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): Modification of cellular proteins by the Small Ubiquitin-like Modifier SUMO is essential for eukaryotic nuclear processes and cell cycle progression in yeast. Unlike ubiquitin conjugation, which can target proteins for degradation by the proteosome, SUMO modification targets proteins for changes in activity and localization. SUMO is translated as a precursor protein that is processed by proteases to generate a mature form which is then activated via C-terminal adenylation and transferred to an intramolecular cysteine by the SUMO activating enzyme E1 to form a thioester bond between SUMO and E1. The E1-SUMO thioester is then transferred to a cysteine residue within the E2 conjugating protein to form a thioester bond between E2 and SUMO. While the E2-SUMO complex is competent to transfer SUMO to lysine residues on the protein target, E3 ligases facilitate this process by binding substrates and E2-SUMO or by activating the E2-SUMO complex for conjugation. SUMO modification can be reversed by proteases that liberate both substrate and SUMO for additional rounds of conjugation. Established functions for SUMO are growing and now include regulating nucleocytoplasmic transport, nuclear body metabolism, transcription, cellular localization, chromosome segregation, DNA repair and replication, and regulation of cell cycle progression in so much as Saccharomyces cerevisiae genes for SUMO, a protease, E1, and E2 are all required for cell cycle progression through G2-M. Taken together, these observations suggest that SUMO conjugation plays a central role in regulatory processes involved in eukaryotic nuclear metabolism and cell cycle control, processes that are of direct relevance to human health, cancer, and the mission of the NIH. This proposal seeks to address the functional and structural significance for components of the SUMO conjugation apparatus. We will utilize structural, biochemical and genetic techniques to establish the basis for 1) SUMO isoform recognition by SUMO enzymes, 2) interactions between E1, SUMO, and E2 3) activities catalyzed by SUMO E3s and factors that recognize SUMO via distinct SUMO binding domains, 4) the kinetic and structural basis for RING-type E3 SUMO ligases. In addition to contributing to our understanding of SUMO conjugation, a thorough structure-function analysis of the SUMO pathway will yield insights into conserved mechanisms utilized in other ubiquitin and ubiquitin-like conjugation pathways.

描述(由申请人提供)：用小的泛素样修饰物SUMO修饰细胞蛋白质对酵母中真核细胞的核过程和细胞周期进程是必不可少的。与泛素结合不同的是，泛素结合可以针对蛋白质体降解的蛋白质，相扑修饰针对的是蛋白质的活性和定位的变化。SUMO被翻译为一种前体蛋白，被蛋白酶处理产生成熟的形式，然后通过C-末端的腺化作用被激活，并被相扑激活酶E1转移到分子内的半胱氨酸，在相扑和E1之间形成硫酯键。然后，将E1-SUMO硫酯转移到E2结合蛋白中的半胱氨酸残基上，在E2和SUMO之间形成硫酯键。虽然E2-SUMO复合体能够将相扑转移到蛋白质靶标上的赖氨酸残基，但E3连接酶通过结合底物和E2-SUMO或激活E2-相扑复合体进行结合来促进这一过程。相扑修饰可以被释放底物和相扑的蛋白酶逆转，以便进行额外的几轮接合。相扑的既定功能正在增长，现在包括调节核细胞质运输、核体新陈代谢、转录、细胞定位、染色体分离、DNA修复和复制以及细胞周期进展的调节，就像酿酒酵母的相扑、一种蛋白酶、E1和E2的基因都是细胞周期通过G2-M所必需的一样。综上所述，这些观察表明，相扑结合在涉及真核细胞核代谢和细胞周期控制的调节过程中发挥核心作用，这些过程与人类健康、癌症和NIH的使命直接相关。这项提案旨在解决相扑共轭装置部件的功能和结构意义。我们将利用结构、生化和遗传技术为1)相扑酶识别相扑异构体建立基础，2)相扑E3和相扑E3之间的相互作用，3)相扑E3和通过不同相扑结合区识别相扑的因子的活性，4)环型E3相扑连接酶的动力学和结构基础。除了有助于我们对相扑结合的理解外，对相扑途径的彻底结构-功能分析将有助于我们深入了解其他泛素和泛素样共轭途径中使用的保守机制。