STRUCTURAL CORRELATES OF PROTEASOME REGULATORS

蛋白酶体调节剂的结构相关性

• 批准号: 2910405
• 负责人: EDWARD Peter GOGOL
• 金额: $ 12.18万
• 依托单位: UNIVERSITY OF MISSOURI KANSAS CITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2002-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2910405
• 关键词: binding sites; cryoelectron microscopy; electron microscopy; enzyme activity; enzyme mechanism; enzyme substrate complex; image enhancement; protease inhibitor; proteasome; protein binding; protein structure; proteolysis; stoichiometry; ubiquitin

DESCRIPTION (Adapted from applicant's abstract): The controlled degradation of most cellular proteins is performed by large protein complexes known as proteasomes that are found in both the nucleus and cytoplasm of an cells. Proteasome activity is central to almost all cell processes, including aspects of protein synthesis and turnover, cell cycle control, metabolic response to environment and immune response, and is implicated in a number of disease states, including cancer and Alzheimer's disease. While the catalytic mechanism of the proteolytic sites has been clarified by recent structural work, there is only a preliminary upstanding of the factors involved in regulation and targeting of proteasome activity. This proposal seeks to determine the interactions of the proteasome with three of the best-characterized regulators of its activity. The first of these is the major activator, PA700, which is responsible for targeting the proteasome to ubiquitinated substrates. Hydrolysis of this class of substrates is believed to be the single most important cellular role of the proteasome. Using electron of negatively stained specimens, we have shown that PA700 binds to the 20S proteasome and converts in to another previously-isolated form (26), in an apparently cooperative manner. Moreover, another protein complex that increases the activation of proteasome by PA700, called modulator, promotes the formation of the PA700 caps without itself forming any stable association with the resulting 26S proteasome. They now seek to correlate the observed structural modification with proteolytic activity, and to better define the organization of the multiple subunits and activities of the PA700 proteasome regulator, by extending our electron microscopy and image analysis studies to include cryo-electron microscopy and three-dimensional reconstructions. A second major component of this proposal is the structural examination of an alternative activator of the proteasome. PA28, which is responsible for the role of the proteasome in antigen presentation. They will define its structural organization, and determine its interaction with PA700 in mixed complexes. The third component of this proposal is a parallel study of the binding site and structural correlates of a specific protein inhibitor of the proteasome, P131, and its effects on formation of activated proteasomes.

描述（改编自申请人的摘要）：受控退化 大多数细胞蛋白的大型蛋白质复合物（称为 在细胞的细胞核和细胞质中发现的蛋白酶体。 蛋白酶体活动对于几乎所有细胞过程都是至关重要的，包括 蛋白质合成和周转，细胞周期控制，代谢方面 对环境和免疫反应的反应，并与 疾病状态，包括癌症和阿尔茨海默氏病。 而 蛋白水解位点的催化机制已通过最近的 结构性工作，只有初步的因素 参与蛋白酶体活动的调节和靶向。 该建议旨在确定蛋白酶体的相互作用 具有其活动的三个最佳调节器。 第一个 其中是主要的激活剂PA700，负责定位 蛋白酶体至泛素化的底物。 这类水解 据认为底物是 蛋白酶体。 使用带负染色标本的电子，我们显示了 PA700与20S蛋白酶体结合并转换为另一个 以前分离的形式（26），显然是合作的。 此外，另一种增加激活的蛋白质复合物 PA700的蛋白酶体，称为调制器，促进了PA700的形成 上限没有自身与结果26s形成任何稳定的关联 蛋白酶体。 他们现在寻求将观察到的结构修饰相关联 具有蛋白水解活性，并更好地定义 PA700蛋白酶体调节剂的多个亚基和活动， 扩展我们的电子显微镜和图像分析研究以包括 冷冻电子显微镜和三维重建。 该提案的第二个主要组成部分是结构检查 蛋白酶体的替代激活剂。 PA28，负责 蛋白酶体在抗原表现中的作用。 他们将定义 它的结构组织，并确定其与PA700的互动 混合复合物。 该提案的第三个组成部分是对结合的平行研究 特定蛋白抑制剂的位点和结构相关 蛋白酶体P131及其对活化蛋白酶体形成的影响。