Molecular Basis of Tail-Anchored Membrane Protein Targeting

尾锚定膜蛋白靶向的分子基础

• 批准号: 8245723
• 负责人: Robert J Keenan
• 金额: $ 43.38万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-05 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8245723
• 关键词: ADP-AlF4; ATP Hydrolysis; ATP phosphohydrolase; Active Sites; Binding; Binding Sites; Biochemical; Biochemical Genetics; Biogenesis; Biological; Cell Death; Cell physiology; Cells; Cellular biology; Complex; Data; Development; Endoplasmic Reticulum; Ensure; Family; Goals; Health; In Vitro; Integral Membrane Protein; Lead; Libraries; Malignant Neoplasms; Maps; Mediating; Membrane; Membrane Proteins; Molecular; Molecular Conformation; Names; Neurodegenerative Disorders; Nucleotides; Outer Mitochondrial Membrane; Pathology; Pathway interactions; Physiological; Play; Process; Property; Proteins; Reagent; Recombinants; Regulation; Resolution; Role; Signal Recognition Particle; Signal Transduction; Site; Staging; Structure; Substrate Interaction; Surface; Tail; Testing; Viral; Work; base; chronic liver disease; crosslink; directed evolution; genetic analysis; in vivo; insight; interdisciplinary approach; mutant; novel; novel therapeutics; programs; receptor; receptor binding; trafficking

DESCRIPTION (provided by applicant): The overall goal of this program is to develop a detailed molecular understanding of a novel post- translational targeting pathway that directs insertion of tail-anchored (TA) membrane proteins into the endoplasmic reticulum (ER) membrane. TA proteins constitute a large family of integral membrane proteins that play critical roles in virtually all aspects of cell biology, ranging from intracellular trafficking and viral replication to regulation of cell death. Despite the physiological importance of these proteins, the machinery and molecular mechanisms that mediate recognition, targeting, and insertion of TA proteins into the correct organellar membrane are poorly understood. The targeting and insertion of newly synthesized proteins into the ER membrane is an essential cellular process. For most membrane proteins, this is achieved co-translationally in a process mediated by the signal recognition particle. However, for the nearly 5% of all eukaryotic membrane proteins that are tail-anchored, targeting is achieved post-translationally. This process is mediated by a newly discovered and evolutionarily conserved ATPase, termed TRC40, which interacts with an ER-bound receptor to ensure efficient and accurate TA protein targeting. In an important breakthrough, we have recently determined high-resolution crystal structures of TRC40. Based upon this structural information, the following specific hypotheses will be tested. 1) Recognition of specific physiochemical properties of the targeting signal directs selective TA protein targeting by TRC40. 2) Conformational changes in TRC40 regulate its ATPase activity and TA substrate interactions. 3) The TRC receptor catalyzes these conformational changes through specific interactions with the TRC40-TA substrate complex. Using a powerful interdisciplinary approach, we will establish the fundamental biochemical and biophysical principles that underlie the process of TA protein targeting and insertion. The specific aims of this project are: 1. To determine the molecular basis of selective TA substrate recognition by TRC40 2. To determine how conformational changes in TRC40 regulate ATPase activity and TA substrate interactions 3. To establish how the TRC receptor coordinates targeting and release of TA proteins at the ER membrane PUBLIC HEALTH RELEVANCE: Tail-anchored (TA) membrane proteins play critical roles in virtually all aspects of cell biology. Given the role of TA proteins in pathologies ranging from cancer, neurodegenerative disease and chronic liver diseases (to name only a few), it is clear that a detailed mechanistic understanding of the targeting, membrane insertion and regulation of TA proteins is of fundamental cell biological and physiological significance. The studies described here will provide insight into the molecular details of TA membrane protein biogenesis, and may ultimately lead to the development of new therapeutic strategies that work by modulating the targeting of TA membrane proteins.

描述（由申请人提供）：本项目的总体目标是对指导尾锚定（TA）膜蛋白插入内质网（ER）膜的新型翻译后靶向途径进行详细的分子理解。TA蛋白构成了一个完整的膜蛋白的大家族，其在细胞生物学的几乎所有方面发挥关键作用，从细胞内运输和病毒复制到细胞死亡的调节。尽管这些蛋白质的生理重要性，机械和分子机制，介导的识别，靶向和插入TA蛋白到正确的细胞器膜知之甚少。 新合成的蛋白质靶向和插入ER膜是一个重要的细胞过程。对于大多数膜蛋白，这是在信号识别颗粒介导的过程中协同实现的。然而，对于所有尾部锚定的真核细胞膜蛋白中的近5%，靶向是在翻译后实现的。这个过程是由一个新发现的和进化上保守的ATP酶介导的，称为TRC 40，它与ER结合的受体相互作用，以确保有效和准确的TA蛋白靶向。在一个重要的突破中，我们最近确定了TRC 40的高分辨率晶体结构。 根据该结构信息，将检验以下特定假设。1)靶向信号的特定生理化学性质的识别指导TRC 40的选择性TA蛋白靶向。2)TRC 40的构象变化调节其ATP酶活性和TA底物相互作用。3)TRC受体通过与TRC 40-TA底物复合物的特异性相互作用催化这些构象变化。使用强大的跨学科的方法，我们将建立基本的生物化学和生物物理学原理的基础上的TA蛋白靶向和插入的过程。该项目的具体目标是：1。确定TRC 40选择性识别TA底物的分子基础。确定TRC 40的构象变化如何调节ATP酶活性和TA底物相互作用3.确定TRC受体如何协调TA蛋白在ER膜上的靶向和释放 公共卫生相关性：尾锚定（TA）膜蛋白在细胞生物学的几乎所有方面都起着关键作用。鉴于TA蛋白在癌症、神经退行性疾病和慢性肝病（仅举几例）等病理学中的作用，很明显，对TA蛋白的靶向、膜插入和调节的详细机制理解具有基本的细胞生物学和生理学意义。这里描述的研究将提供深入了解TA膜蛋白生物发生的分子细节，并可能最终导致新的治疗策略的发展，通过调节TA膜蛋白的靶向工作。