Function and regulation of the proteasome

蛋白酶体的功能和调节

基本信息

批准号：
7558282
负责人：
George N. DeMartino
金额：
$ 32.19万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
1994
资助国家：
美国
起止时间：
1994-07-01 至 2013-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7558282
关键词：
26S proteasome ATP Hydrolysis ATP phosphohydrolase ATP-Dependent Proteases Accounting Achievement Address Affect Binding Binding Sites Biochemical Biochemistry Bioinformatics Carboxypeptidase Cell physiology Chemicals Collaborations Complex Consumption Data Disease Doctor of Philosophy Elements Ensure Escherichia coli Eukaryotic Cell Event Foundations Functional disorder Goals Hand Health Human Resources Hydrolysis In Vitro Individual Left Louisiana Mass Spectrum Analysis Mediating Modeling Modification Molecular Mutagenesis Mutation Peptide Hydrolases Peptides Play Polyubiquitin Preparation Principal Investigator Process Proteasome Binding Proteins Proteolysis Proteomics Publications Published Comment Recombinant Proteins Recombinants Regulation Relative (related person)Research Research Personnel Role Saint Jude Children&apos s Research Hospital Staging Structure Supervision System Testing Training Ubiquitin Ubiquitination Universities Variant base crosslink experience improved malignant muscle neoplasm monomer multicatalytic endopeptidase complex mutant neuropathology programs protein degradation protein function reconstitution research study response success wasting

项目摘要

DESCRIPTION (provided by applicant): The long term goal of this research is to comprehensively understand the mechanisms and regulation of intracellular protein degradation. Protein degradation regulates nearly every aspect of normal cellular function. Dysfunction of protein degradation underlies many diseases including cancer, muscle wasting, and neuropathologies. Most protein degradation in eukaryotic cells is catalyzed by the 26S proteasome, the ATP dependent protease of the ubiquitin system. The immediate goal of this project is to determine the mechanisms by which ATP binding and hydrolysis mediate proteasome function. Specific Aim 1 will characterize biochemical features of ATP binding and hydrolysis by the 26S proteasome and define the relative qualitative and quantitative contributions of six different AAA ATPase subunits of the 26S proteasome to these processes. These experiments will provide essential new information about these subunits and support subsequent mechanistic studies on the roles of ATP and the AAA subunits in proteasome function. Specific Aim 2 will determine molecular mechanisms by which ATP binding to AAA subunits mediates both the assembly of the 26S proteasome from protease (20S proteasome) and regulatory (PA700/19S) sub complexes, and the activation of protease activity upon assembly, using biochemically defined in vitro systems. These experiments will test the hypothesis that proteasome assembly and activation require separate ATP binding events and define roles of individual AAA subunits in these processes. These experiments will distinguish between alternative models in which the six AAA subunits have either distinct/dedicated roles or multiple/redundant roles. Specific Aim 3 will define the role of ATP hydrolysis in 26S proteasome catalyzed protein degradation. These experiments will compare relative rates of degradation of variants of a model substrate that differ systematically in structural stability and proteasome targeting elements, such as structurally defined polyubiquitin chains. By comparing the qualitative and quantitative requirements for ATP hydrolysis and proteolysis, these experiments will deconvolute the multiple roles of ATP consumption in sub processes of proteolysis such as substrate binding, unfolding, translocation, and deubiquitylation. Completion of these aims will provide detailed information about mechanisms of proteolysis by the 26S proteasome and elucidate the fundamental role of ATP in mediating proteasome function. The long term goal of this research is to comprehensively understand the mechanisms and regulation of intracellular protein degradation. Protein degradation regulates nearly every aspect of normal cellular function. Dysfunction of protein degradation underlies many diseases including cancer, muscle wasting, and neuropathologies. Most protein degradation in eukaryotic cells is catalyzed by the 26S proteasome, the ATP dependent protease of the ubiquitin system. The immediate goal of this project is to determine the mechanisms by which ATP binding and hydrolysis mediate proteasome function.

描述（由申请人提供）：这项研究的长期目标是全面了解细胞内蛋白质降解的机制和调节。蛋白质降解几乎调节正常细胞功能的各个方面。蛋白质降解的功能障碍是许多疾病，包括癌症，肌肉浪费和神经病理学。真核细胞中的大多数蛋白质降解是由26S蛋白酶体（泛素系统的ATP依赖性蛋白酶）催化的。该项目的直接目标是确定ATP结合和水解介导的质体功能的机制。具体目标1将表征26S蛋白酶体的ATP结合和水解的生化特征，并定义26S蛋白酶体对这些过程的六种不同AAA ATPase亚基的相对定性和定量贡献。这些实验将提供有关这些亚基的基本新信息，并支持有关ATP和AAA亚基在蛋白酶体功能中的作用的后续机械研究。具体的目标2将确定ATP与AAA亚基结合的分子机制，从蛋白酶（20S蛋白酶体）和调节型（PA700/19S）亚络合物中介导26S蛋白酶体的组装，以及使用生物学定义的VITY IN VITRO INTICRO SYSTICE的组装时蛋白酶的激活。这些实验将检验以下假设：蛋白酶体组装和激活需要单独的ATP结合事件，并在这些过程中定义单个AAA亚基的作用。这些实验将区分六个AAA亚基具有独特/专用角色或多个/冗余角色的替代模型。具体目标3将定义ATP水解在26S蛋白酶体催化蛋白降解中的作用。这些实验将比较模型底物变体降解的相对降解速率，该变体在结构稳定性和蛋白酶体靶向元件（例如结构定义的多泛素链）上有系统地差异。通过比较ATP水解和蛋白水解的定性和定量需求，这些实验将在蛋白水解的亚过程（例如底物结合，展开，易位，易位和去泛素化和去泛素化）中反应ATP消耗的多重作用。这些目标的完成将提供有关26S蛋白酶体蛋白水解机制的详细信息，并阐明ATP在介导蛋白酶体功能中的基本作用。这项研究的长期目标是全面了解细胞内蛋白质降解的机制和调节。蛋白质降解几乎调节正常细胞功能的各个方面。蛋白质降解的功能障碍是许多疾病，包括癌症，肌肉浪费和神经病理学。真核细胞中的大多数蛋白质降解是由26S蛋白酶体（泛素系统的ATP依赖性蛋白酶）催化的。该项目的直接目标是确定ATP结合和水解介导的质体功能的机制。