Molecular Basis of Tail-Anchored Membrane Protein Targeting

尾锚定膜蛋白靶向的分子基础

基本信息

批准号：
8830981
负责人：
Robert J Keenan
金额：
$ 38.7万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-04-05 至 2018-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8830981
关键词：
ATP phosphohydrolase Accounting Architecture Binding Biochemical Biochemical Genetics Biogenesis Biological Process Biophysical Process Biophysics C-terminal Catalysis Cell membrane Cell physiology Cells Chemicals Complex Crystallography Cytosol Data Defect Development Diabetes Mellitus Disease Endoplasmic Reticulum Environment Enzymes Eukaryotic Cell Event Fluorescence Spectroscopy Foundations Functional disorder Goals Grant Growth Health Heart Diseases Human Pathology In Vitro Integral Membrane Protein Lead Link Lipid Bilayers Malignant Neoplasms Maps Mediating Membrane Membrane Proteins Modeling Molecular Molecular Chaperones Monitor N-terminal Neurodegenerative Disorders Pathway interactions Play Positioning Attribute Process Property Protein translocation Proteins Quality Control Reaction Recombinants Research Resolution Role Site Staging Structure System Tail Transmembrane Domain Ursidae Family Work Yeasts base fight against genetic analysis high throughput screening human disease innovation insight interdisciplinary approach novel novel therapeutics protein complex receptor reconstitution spectroscopic survey stoichiometry tool trafficking

项目摘要

DESCRIPTION (provided by applicant): The goal of this research is to develop a detailed molecular understanding of a newly discovered post-translational targeting pathway that directs insertion of tail-anchored (TA) membrane proteins into the endoplasmic reticulum (ER) membrane. TA proteins, which account for nearly 5% of all eukaryotic membrane proteins, are anchored to the lipid bilayer by a single C-terminal transmembrane domain (TMD) and have a cytosolic-facing amino-terminal domain. These proteins are found in virtually all cell membranes where they mediate numerous essential cellular processes. Post-translational targeting and insertion of TA proteins is mediated by the newly discovered 'guided entry of tail-anchored protein' (GET) pathway. Over the past few years, a basic framework for the pathway has been defined. First, a 'pre-targeting' factor captures a newly synthesized TA protein in the cytosol, and transfers it to a soluble ATPase, called Get3. This targeting complex is directed to the ER membrane via an interaction with the Get1/2 receptor complex which is both necessary and sufficient to drive TA substrate release and insertion into the membrane. Our research goal is to define the biochemical and biophysical principles that underlie TA protein insertion. First, we will elucidate the molecular identity of the Get3-TA substrate targeting complex at defined steps along the targeting pathway (Aim 1). Second, we will determine whether TA substrates are integrated directly into the bilayer or if insertion requires a chaperoning role for the the Get1/2 receptor complex (Aim 2). Finally, we will define how the static and dynamic properties of the Get1/2 complex allow it to orchestrate the essential steps of TA substrate recruitment, release and insertion (Aim 3). These Aims will be accomplished using a powerful interdisciplinary approach that combines structure-function analyses with state-of- the-art spectroscopic studies. By defining common themes between known insertion pathways, this work will deepen our understanding of the fundamental cell biological and biophysical process of TMD insertion. By developing new tools and experimental strategies, this work promises to enable analysis of other complex membrane-associated processes. Finally, because defects in TA protein biogenesis are linked to much human pathology, these studies promise insight that may lead to new therapeutic strategies for use in the fight against human disease.

描述（由申请人提供）：本研究的目标是对新发现的翻译后靶向途径进行详细的分子理解，该途径指导尾部锚定（TA）膜蛋白插入内质网（ER）膜。 TA 蛋白占所有真核细胞膜蛋白的近 5%，通过单个 C 端跨膜结构域 (TMD) 锚定在脂质双层上，并具有面向胞质的氨基端结构域。这些蛋白质几乎存在于所有细胞膜中，它们介导许多重要的细胞过程。 TA 蛋白的翻译后靶向和插入是由新发现的“尾锚定蛋白引导进入”(GET) 途径介导的。在过去的几年里，该路径的基本框架已经确定。首先，“预靶向”因子捕获细胞质中新合成的 TA 蛋白，并将其转移至可溶性 ATP 酶（称为 Get3）。该靶向复合物通过与 Get1/2 受体复合物的相互作用定向至 ER 膜，这对于驱动 TA 底物释放和插入膜来说是必要且充分的。我们的研究目标是确定 TA 蛋白插入的生化和生物物理原理。首先，我们将在靶向途径的确定步骤中阐明 Get3-TA 底物靶向复合物的分子身份（目标 1）。其次，我们将确定 TA 底物是否直接整合到双层中，或者插入是否需要 Get1/2 的陪伴作用受体复合物（目标 2）。最后，我们将定义 Get1/2 复合物的静态和动态特性如何使其能够协调 TA 底物招募、释放和插入的基本步骤（目标 3）。这些目标将使用强大的跨学科方法来实现，该方法将结构功能分析与最先进的光谱研究相结合。通过定义已知插入途径之间的共同主题，这项工作将加深我们对 TMD 插入的基本细胞生物学和生物物理过程的理解。通过开发新的工具和实验策略，这项工作有望能够分析其他复杂的膜相关过程。最后，由于 TA 蛋白生物发生的缺陷与许多人类病理学有关，这些研究有望带来新的见解，可能会带来用于对抗人类疾病的新治疗策略。