FUNCTION AND REGULATION OF THE PROTEASOME

描述：这项研究计划的长期目标是 全面了解生物化学机制和 细胞内蛋白质降解的生理调节。这 目标对于理解众多细胞具有广泛的意义 利用蛋白质降解作为一种机制的过程 调节，并用于减轻以组织萎缩为特征的疾病。 这个项目的总体目标是建立 PA28是蛋白酶体的一种蛋白激活蛋白。蛋白酶体是一种 所有泛素依赖所需的细胞内蛋白酶--以及一种未知的 真核细胞中泛素非依赖性蛋白质的降解量。 体外和体内的蛋白酶体功能都是由多个 调控蛋白，其相对的细胞作用不明确。PA28 是这些蛋白质中最鲜为人知的一种。细胞的作用 PA28将通过在哺乳动物细胞中过度表达该蛋白来定义。 这些研究将检验PA28提高总体死亡率的假设 细胞内蛋白质的降解。PA28的细胞功能将 将通过密切相关的生化实验进行进一步检验。这些 实验将确定PA28可逆磷酸化的作用 作为一种控制其活动的机制，通过检验以下假设 PA28的磷酸化抑制其蛋白酶体刺激活性。 将在#年研究PA28对模型蛋白质降解的影响 PA28-蛋白酶体体外降解蛋白质的动力学分析 单独和与其他蛋白酶体调节的复合体 蛋白质复合体。这些实验将检验这样的假设：PA28的S 细胞功能是促进两种大肽的降解 直接和由其他蛋白酶体的作用产生的大肽 复合体最后，PA28蛋白酶体复合体的细胞底物 将通过使用表达克隆的筛选策略进行鉴定。 这些高度集成的研究将定义PA28的细胞作用和 可能是它们的生化基础。因此，这个项目将极大地 增加对生物多样性的机制和调控的理解。 胞内蛋白质降解。

DESCRIPTION: The long-term objective of this research program is to provide a comprehensive understanding of the biochemical mechanisms and physiological regulation of intracellular protein degradation. This objective has broad significance for understanding numerous cellular processes that utilize protein degradation as a mechanism for their regulation, and for alleviating diseases characterize by tissue atrophy. The general goal of this project is to establish the physiological role of PA28, a protein activation of the proteasome. The proteasome is an intracellular protease required for all ubiquitin-dependent-and an unknown amount of ubiquitin-independent protein degradation in eukaryotic cells. Proteasome function both in vitro and in vivo is mediated by multiple regulatory proteins whose relative cellular roles are poorly defined. PA28 is one of the least understood of these proteins. The cellular role of PA28 will be defined by over-expressing the protein in mammalian cells. These studies will test the hypothesis that PA28 increases overall rates of intracellular protein degradation. The cellular function of PA28 will be examined further by closely related biochemical experiments. These experiments will establish the role of reversible phosphorylation of PA28 as a mechanism for control of its activity by testing the hypothesis that phosphorylation of the PA28 inhibits its proteasome-stimulatory activity. The effect of PA28 on the degradation of model proteins will be studied in vitro by analyzing the kinetics of protein degradation by PA28-proteasome complexes alone and in combination with other proteasome-regulatory protein complexes. These experiments will test the hypothesis that PA28's cellular function is to promote degradation both of large peptides directly and of large peptides generated by the action of other proteasome complexes Finally, cellular substrates for the PA28 proteasome complex will be identified with a screening strategy using expression cloning. These highly integrated studies will define cellular role of PA28 and the possible biochemical bases for them. Therefore, this project will greatly increase the understanding of the mechanisms and regulation of intracellular protein degradation.