Folding and degradation of membrane proteins

摘要 （项目名称：膜蛋白的折叠和降解） 我的研究计划的目标是阐明化学和物理原理的基础， 膜蛋白的折叠和降解。细胞应该 保持生理上的最佳水平 功能性蛋白质。这是通过分子伴侣的平衡作用来实现的， 蛋白酶降解错误折叠和多余的蛋白质，以及应激反应信号 调节伴侣蛋白和蛋白酶水平的途径。这种蛋白质质量控制机器 使用多层机制来监测蛋白质组的折叠状态。一个人的命运， 给定的蛋白质，它是否会折叠或降解，很大程度上取决于内在的折叠特性 蛋白质。虽然大多数折叠和降解研究都集中在水溶性 虽然膜蛋白的内在折叠特性影响了膜蛋白的功能，但目前还不清楚膜蛋白的内在折叠特性如何影响膜蛋白的功能。 他们的退化。知识差距主要源于研究膜的固有困难 蛋白质在其天然脂质双层环境中的折叠以及对蛋白质折叠的理解不足。 折叠和降解的序列决定因素。我们使用一个组合模型， 膜整合ATP依赖蛋白酶FtsH。大肠杆菌作为模型降解机器， 和E.大肠杆菌作为模型底物，这两种都是广泛 在原核和真核细胞中保守。我们开发了一系列的方法来研究 膜蛋白折叠的双层的基础上新的空间捕获原理。我们也 开发了一个体外双层系统，以研究FtsH介导的膜蛋白降解。 利用这些方法上的创新，我对未来研究的愿景是学习可概括的 折叠，蛋白质相互作用和细胞膜蛋白质的质量控制的教训 膜。我们将深入研究三个尚未回答的问题：1）什么是详细的分子 FtsH介导的膜蛋白降解机制？2)脂双层是良溶剂吗 对于膜蛋白的变性状态或促进其非特异性的不良溶剂， 崩溃？3)来自嗜热生物的膜蛋白是如何实现其 热稳定性和活性？如果成功，这项研究的结果将推动我们的基本 了解膜蛋白降解的机制和能量学，确定新的 控制膜蛋白折叠和相互作用的脂质双层的物理性质， 并发现新的膜蛋白稳定基序，这将提供一个有用的设计， 工程原理

ABSTRACT (Project Title: Folding and Degradation of Membrane Proteins) The goal of my research program is to elucidate the chemical and physical principles underlying the folding and degradation of membrane proteins. Cells should maintain physiologically optimal levels of functional proteins. This is achieved by the balanced actions of molecular chaperones to facilitate folding, proteases to degrade misfolded and superfluous proteins, and stress-response signaling pathways to regulate levels of chaperones and proteases. This protein quality control machinery uses multiple layers of mechanisms to monitor the folding status of a proteome. Thus, the fate of a given protein, whether it will fold or be degraded, strongly depends on the intrinsic folding properties of the protein. While a majority of folding and degradation studies have focused on water-soluble proteins, it is not well understood how the intrinsic folding properties of membrane proteins affect their degradation. The knowledge gap mainly stems from inherent difficulties in studying membrane protein folding in their native lipid bilayer environment as well as an insufficient understanding of folding and sequence determinants for degradation. We use a combined model employing the membrane-integrated ATP-dependent protease FtsH of E. coli as a model degradation machine, and the intramembrane protease GlpG of E. coli as a model substrate, both of which are widely conserved in prokaryotic and eukaryotic cells. We developed an array of methods to study membrane protein folding in the bilayers based on the novel steric-trapping principle. We also developed an in vitro bilayer system to study FtsH-mediated degradation of a membrane protein. Using these methodological innovations, my vision for future research is to learn the generalizable lessons of folding, protein-protein interactions and quality control of membrane proteins in the cell membranes. We will delve into three unanswered problems: 1) What is the detailed molecular mechanisms of FtsH-mediated membrane protein degradation? 2) Is the lipid bilayer a good solvent for the denatured states of membrane protein or a poor solvent that promotes their nonspecific collapse? 3) How do membrane proteins from thermophilic organisms achieve their unusual thermostability and activity? If successful, the outcome of this study will advance our fundamental understanding of mechanisms and energetics of membrane protein degradation, identify new physical properties of the lipid bilayer that control folding and interactions of membrane proteins, and discover new stabilizing motifs for membrane proteins that will provide a useful design and engineering principle.