Functional specification of HSP70 by J-domain protein co-chaperones

J 结构域蛋白共伴侣对 HSP70 的功能规范

• 批准号: 504257241
• 负责人: Professor Dr. Bernd Bukau
• 金额: --
• 依托单位: Zentrum für Molekulare Biologie (ZMBH)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/504257241?language=en
• 关键词: Functional; specification; HSP70; domain; protein

Members of the heat shock protein 70 (HSP70) family are ATP-dependent molecular chaperones that act as central hub of cellular protein homeostasis. They ensure proteins attain and maintain their correct three-dimensional fold, become disassembled when aggregated, or cleared by cellular degradation pathways. ATP hydrolysis promotes substrate trapping by Hsp70s and therefore is the critical step that initiates the functional chaperone cycle. It is allosterically regulated both by substrate binding and the action of co-chaperones of the J-domain protein (JDP) family, also referred to as Hsp40. JDPs are an extended family of proteins (>40 members in human cells) that contain a conserved J-domain which promotes ATP hydrolysis in the HSP70 nucleotide binding domain via distal interactions with the flexible linker connecting this domain with the substrate binding domain. Class A and B JDP members share a common architecture with the N-terminal J-domain, a disordered GF rich linker, 2 C-terminal substrate binding domains (CTDs) and a dimerisation domain. The key differentiating feature is the composition of the GF linker, which in the case of class A JDPs is extended by a zinc finger domain. Despite their high structural similarities, class A and class B JDPs have been implicated in distinct cellular activities, with class A JDPS primarily interacting with monomeric misfolded proteins or small oligomeric species, while class B proteins of both type I (DNAJB1/4) and type II (DNAJB6/8) preferentially interact with oligomeric species and larger aggregates. Despite three decades of research into the HSP70 chaperone machinery, the roles and diversity within the JDP co-chaperone family remains largely unexplored on a mechanistic level. In recent years, JDPs have come to the foreground as potential therapeutic targets for the selective tuning of specific HSP70 activities without perturbing (other) essential housekeeping activities. We therefore aim to perform a comprehensive analysis of JDP-mediated recruitment of the human HSP70 chaperone to cellular substrates to answer the fundamental question: how do canonical class A and class B JDPs get HSP70 to act in so many different folding processes in the human cell? Our specific aims are: 1/ Describe biochemical properties that drive substrate specificity between JDP classes and member within these classes, 2/ Identify the trigger for activation and differences in regulatory switches within the class B JDP family, 3/ Characterise how different regulatory mechanisms determine the assembly of HSP70 complexes that perform different cellular activities and, 4/ Probe the interplay between JDP classes in tuning HSP70 function.Overall, the proposed project will generate complementary datasets that combined will build a comprehensive mechanistic understanding of JDP mediated stimulation of HSP70 chaperone activity.

热休克蛋白70（HSP 70）家族成员是ATP依赖的分子伴侣，作为细胞蛋白质稳态的中心枢纽。它们确保蛋白质获得并保持其正确的三维折叠，在聚集时分解，或通过细胞降解途径清除。ATP水解促进Hsp 70的底物捕获，因此是启动功能性伴侣循环的关键步骤。它通过底物结合和J结构域蛋白（JDP）家族（也称为Hsp 40）的共分子伴侣的作用进行变构调节。JDP是一个扩展的蛋白质家族（在人类细胞中>40个成员），其含有保守的J结构域，其通过与连接该结构域与底物结合结构域的柔性接头的远端相互作用促进HSP 70核苷酸结合结构域中的ATP水解。A类和B类JDP成员具有共同的结构，包括N-末端J-结构域、无序的富含GF的接头、2个C-末端底物结合结构域（CTD）和一个二聚化结构域。关键的区别特征是GF接头的组成，在A类JDP的情况下，GF接头通过锌指结构域延伸。尽管A类和B类JDP具有高度的结构相似性，但它们涉及不同的细胞活性，其中A类JDP主要与单体错误折叠蛋白或小的寡聚物种类相互作用，而I型（DNAJB 1/4）和II型（DNAJB 6/8）的B类蛋白优先与寡聚物种类和较大的聚集体相互作用。尽管对HSP 70分子伴侣机制的研究已经进行了三十年，但JDP共分子伴侣家族中的角色和多样性在机制水平上仍然很大程度上未被探索。近年来，JDP已经成为选择性调节特定HSP 70活性而不干扰（其他）基本管家活动的潜在治疗靶点。因此，我们的目标是进行一个全面的分析JDP介导的招聘的人HSP 70分子伴侣的细胞基板，以回答基本的问题：如何典型的A类和B类JDP让HSP 70在这么多不同的折叠过程中在人类细胞中发挥作用？我们的具体目标是：1/描述驱动JDP类别和这些类别内的成员之间的底物特异性的生物化学性质，2/识别激活的触发因素和B类JDP家族内调节开关的差异，3/表征不同的调节机制如何确定执行不同细胞活性的HSP 70复合物的组装，4/探索JDP类之间在调节HSP 70功能方面的相互作用。总体而言，所提出的项目将生成互补的数据集，这些数据集将建立对JDP介导的HSP 70伴侣活性刺激的全面机制理解。