Protein Folding in the Eukaryotic Cytosol

描述(申请人提供)：该研究计划的长期目标是了解新翻译的蛋白质如何在真核细胞中折叠。拟议的研究将集中在多肽合成过程中发生在核糖体上的折叠事件，并将检查分子伴侣在折叠过程中的作用。理解从头蛋白质折叠的概念框架源于我们在上一个资金周期中的工作，该工作表明一个名为CLIPS(链接到蛋白质合成的伴侣)的伴侣网络在物理上和功能上与翻译机制有关。我们的工作假设是，CLIPS伴侣的任务是引导新合成的多肽形成它们的折叠构象。伴侣介导的折叠途径似乎涉及不同类别的片段之间的合作，包括在折叠过程中起早期作用的伴侣，如新生的链相关复合体(NAC)、Hsp70蛋白和Gim/PreFoldin复合体，以及在折叠过程中似乎作用较晚的机械上不同的伴侣Trc/CCT和Hsp90。我们阐明伴侣如何调节新合成蛋白质折叠的一般策略依赖于体外和体内方法的紧密结合。我们提出的实验旨在获得对伴侣在从头折叠中的作用的功能、机制和结构的见解。

DESCRIPTION (provided by applicant): The long term goal of this Research Program is to understand how newly translated proteins fold in eukaryotic cells. The proposed research will focus on folding events as they occur at the ribosome during synthesis of a polypeptide and will examine the role of molecular chaperones in their folding process. The conceptual framework for understanding de novo protein folding originates from our work in the previous funding cycle, which showed that a network of chaperones named CLIPS (Chaperones Linked to Protein Synthesis) is physically and functionally linked to the translation machinery. Our working hypothesis is that the CLIPS chaperones are tasked with guiding newly synthesized polypeptides to their folded conformation. Chaperone-mediated folding pathways appear to involve the cooperation of different classes of CLIPS, including chaperones that act early in the folding process, such as the Nascent Chain Associated Complex (NAC), the Hsp70 proteins and the GIM/prefoldin complex, and the mechanistically distinct chaperones TRiC/CCT and Hsp90, which appear to act later in the folding process. Our general strategy to elucidate how chaperones mediate the folding of newly synthesized proteins relies on the close integration of in vitro and in vivo approaches. Our proposed experiments are aimed at obtaining functional, mechanistic and structural insights into the role of chaperones in de novo folding.