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Protein unfolding in a physiological system

蛋白质在生理系统中的展开

基本信息

批准号：
7619492
负责人：
ANDREAS MATOUSCHEK
金额：
$ 32.17万
依托单位：
NORTHWESTERN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-04-01 至 2011-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7619492
关键词：
Active Sites Adaptor Signaling Protein Address Affect Antigen Presentation Binding Biochemical Bioinformatics Biological Catalytic Domain Cell Nucleus Cell physiology Cells Cleaved cell Complex Cyclin B Cyclins Cytosol Discrimination Drosophila genus Enzymes Eukaryotic Cell Genetic Genetic Screening Goals Homologous Gene Human In Vitro Investigation Lead Length Malignant Neoplasms Methods Modification Molecular Molecular Conformation Nature Neurodegenerative Disorders Peptides Physiological Play Polyubiquitin Process Proteasome Binding Protein Databases Protein Fragment Proteins Proteolysis Regulation Research Role Signal Transduction Site Specificity Structure System Testing Ubiquitin Ubiquitination Work Xenopus base cell growth cell growth regulation computerized data processing fighting fly genetic regulatory protein genetic selection human disease in vivo insight multicatalytic endopeptidase complex novel particle protein aggregate protein complex protein degradation protein misfolding research study transcription factor yeast protein

项目摘要

DESCRIPTION (provided by applicant): The proteasome, located in the cytosol and nucleus of eukaryotic cells, plays a central role in cellular regulation by controlling the concentrations of hundreds of regulatory proteins, removing misfolded proteins, and producing some peptides displayed in antigen presentation. It is an approx. 2MDa cylindrical particle with its active sites of proteolysis buried deep inside the structure where they are accessible only through a narrow channel. Substrate proteins must be unfolded to fit the channel and to be degraded. The long-term goals of the research proposed here are to understand protein unfolding by the proteasome using a combination of modern biochemical, computational, genetic, and cell biological methods. These experiments will provide an in-depth understanding of proteasome function and may reveal unexpected regulatory mechanisms. The immediate goal is to understand how the unfolding step affects the specificity of the proteasome for its substrates and how it affects the nature of its degradation products. We will first define how degradation is initiated after substrate binding by the proteasome. Proteins are targeted to the proteasome by the covalent attachment of a ubiquitin tag but degradation initiates at a different site in the substrate. The initiation step contributes to the specificity of degradation and the sequence and spatial requirements for efficient initiation will be determined in vitro. The effect of these rules on the specificity of degradation of natural proteins will be tested in vivo. Second, we will investigate how the proteasome unfolds and degrades only part of a protein in a process called processing. The proteasome degrades most proteins completely to small peptides but a novel type of signal can lead to partial protein degradation. The processing signal and mechanism will be characterized and physiological examples of processing will be detected by a bioinformatics search and a genetic screen. This work is relevant to human diseases; many of the proteins degraded by the proteasome regulate cell growth and division. Since proteins must be unfolded before they are cleaved by the proteasome, understanding unfolding during degradation may open new avenues to fight cancer, for example by stabilizing specific proteins against unfolding and degradation. Some neurodegenerative diseases are associated with the accumulation of protein aggregates, which the proteasome fails to remove for unknown reasons. The investigation of partial protein degradation may lend insight into this question.

描述（由申请人提供）：蛋白酶体位于真核细胞的胞质溶胶和细胞核中，通过控制数百种调节蛋白的浓度、去除错误折叠的蛋白质以及产生在抗原呈递中展示的一些肽，在细胞调节中发挥核心作用。这是一个近似。2 MDa圆柱形颗粒，其蛋白水解活性位点深埋在结构内部，只能通过狭窄的通道进入。底物蛋白必须解折叠以适应通道并被降解。这里提出的研究的长期目标是使用现代生物化学，计算，遗传学和细胞生物学方法的组合来理解蛋白酶体的蛋白质展开。这些实验将提供对蛋白酶体功能的深入了解，并可能揭示意想不到的调控机制。当前的目标是了解解折叠步骤如何影响蛋白酶体对其底物的特异性以及它如何影响其降解产物的性质。我们将首先定义如何降解后，底物结合蛋白酶体启动。蛋白质通过泛素标签的共价连接靶向蛋白酶体，但降解起始于底物中的不同位点。起始步骤有助于降解的特异性，将在体外确定有效起始的序列和空间要求。将在体内测试这些规则对天然蛋白质降解特异性的影响。其次，我们将研究蛋白酶体如何在一个称为加工的过程中展开和降解蛋白质的一部分。蛋白酶体将大多数蛋白质完全降解为小肽，但一种新型信号可以导致部分蛋白质降解。处理信号和机制将被表征，并且处理的生理实例将通过生物信息学搜索和遗传筛选来检测。这项工作与人类疾病有关;许多被蛋白酶体降解的蛋白质调节细胞生长和分裂。由于蛋白质在被蛋白酶体切割之前必须解折叠，因此了解降解过程中的解折叠可能会为对抗癌症开辟新的途径，例如通过稳定特定蛋白质对抗解折叠和降解。一些神经退行性疾病与蛋白质聚集体的积累有关，蛋白酶体由于未知的原因未能去除蛋白质聚集体。对部分蛋白质降解的研究可能有助于深入了解这个问题。