ROLE OF HEAT SHOCK PROTEINS IN PROTEIN FOLDING

热休克蛋白在蛋白质折叠中的作用

• 批准号: 3304746
• 负责人: ANTHONY L FINK
• 金额: $ 21.13万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-01-01 至 1993-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3304746
• 关键词: SDS polyacrylamide gel electrophoresis; adenosine triphosphate; binding proteins; biophysics; catalyst; chemical binding; chemical kinetics; chemical models; chemical stability; circular dichroism; conformation; crosslink; crystallization; fluorescent dye /probe; gel filtration chromatography; immunoprecipitation; infrared spectrometry; monoclonal antibody; phosphorylation; protein denaturation; protein folding; protein purification; protein sequence; protein structure; proteolysis; stoichiometry; stress proteins

Mammalian (and other) cells respond to stress by increasing expression of a class of proteins known as heat shock proteins. Most of these proteins are produced constitutively under normal conditions and play essential roles in the life of the cell. Recent studies have shown that the mammalian hsp 70 family of heatshock proteins, and related polypeptide chain binding proteins and molecular chaperones (e.g. GroEL) from other sources, can bind to short peptides, and to nascent polypeptides in cells, and are involved in various aspects of protein assembly, translocation and folding. The relatively large fraction of cell protein, especially under conditions of stress, present as this type of protein, and the potential lethality of mutant forms, indicates their critical importance to cell function. Even though it is clear that proteins fold spontaneously in vitro, it is not clear that this is the case in vivo. In fact, a good case can be made for the need for some form of assistance in folding and assembly in the cell in order to minimize competing side reactions. The goal of the proposed research is to determine the molecular details of how hsp 70 and related polypeptide chain binding proteins facilitate the folding and assembly of substrate proteins. Biophysical characterization of the hsp 70 proteins will provide information on their structure and stability. A major part of the proposal concerns elucidation of the molecular details of the interaction of the hsp 70 with its substrate protein, and the role of ATP in this process. These studies will be carried out under in vitro conditions. The stoichiometry and binding constants for the interactions will be determined. A range of proteins from small monomeric to large oligomeric will be used as substrate proteins. The effect of hsp 70 on the kinetics of protein folding, and the affinity of hsp 70 for intermediates in folding will be determined. The structural and sequence specificity requirements for hsp 70 binding to proteins will be ascertained. From these investigations we expect to be able to provide a detailed molecular description of the function of this important class of cellular agent.

哺乳动物（和其他）细胞通过增加a的表达来应对压力 称为热休克蛋白的一类蛋白质。 这些蛋白质大部分是 在正常条件下组成型产生并发挥重要作用 细胞的生命。 最近的研究表明，哺乳动物 hsp 70 热休克蛋白家族和相关的多肽链结合 来自其他来源的蛋白质和分子伴侣（例如 GroEL）可以结合 短肽和细胞中的新生多肽，并且涉及 蛋白质组装、易位和折叠的各个方面。 这 相对较大部分的细胞蛋白，特别是在 压力，以这种类型的蛋白质存在，以及潜在的致命性 突变形式表明它们对细胞功能至关重要。 甚至 尽管很明显蛋白质在体外会自发折叠，但事实并非如此 显然体内也是如此。 事实上，可以提出一个很好的案例 在细胞中折叠和组装时需要某种形式的帮助 以尽量减少竞争性副反应。 拟议的目标 研究的目的是确定 hsp 70 及其相关分子的分子细节 多肽链结合蛋白促进折叠和组装 底物蛋白。 hsp 70 蛋白的生物物理表征 将提供有关其结构和稳定性的信息。 很大一部分是 该提案涉及阐明分子细节 hsp 70 与其底物蛋白的相互作用以及 ATP 的作用 在这个过程中。 这些研究将在体外进行 状况。 相互作用的化学计量和结合常数 将被确定。 一系列蛋白质，从小单体到大蛋白质 寡聚体将用作底物蛋白。 hsp 70 对 蛋白质折叠动力学以及 hsp 70 对中间体的亲和力 折叠时将被确定。 结构和序列特异性 将确定 hsp 70 与蛋白质结合的要求。 从 我们希望这些研究能够提供详细的分子信息 这类重要细胞因子的功能描述。