Structural Basis of Quantal Release

量子释放的结构基础

• 批准号: 9900872
• 负责人: Fei Li
• 金额: $ 12.12万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9900872
• 关键词: Address; Affinity; Allosteric Regulation; Award; Biological; Brain; Brain Diseases; Carrier Proteins; Cells; Chemicals; Communication; Communities; Complex; Coupling; Cryo-electron tomography; Cryoelectron Microscopy; Disease; Electron Microscopy; Electrophysiology (science); Endocytosis; Environment; Equilibrium; Event; Exhibits; Exocytosis; Foundations; Future; Glutamate Transporter; Glutamates; Goals; Institution; Ions; Knowledge; Learning; Lysosomes; Membrane Potentials; Membrane Proteins; Membrane Transport Proteins; Mentors; Methodology; Mitochondria; Molecular; Neurologic; Neurons; Neurosciences; Neurotransmitters; Organelles; Pathway interactions; Physiological; Physiology; Play; Process; Property; Proteins; Proton Pump; Proton-Translocating ATPases; Rattus; Receptor Activation; Regulation; Research; Research Personnel; Research Project Grants; Research Training; Resolution; Resources; Role; Scientist; Signal Transduction; Structure; Structure-Activity Relationship; Synapses; Synaptic Transmission; Synaptic Vesicles; Techniques; Technology; Testing; Therapeutic Intervention; Time; Training; Training Programs; Vesicle; Work; X-Ray Crystallography; Xenopus oocyte; brain health; career; driving force; insight; interest; mutant; neurotransmission; neurotransmitter transport; particle; protein complex; protein purification; protein structure function; quantum; receptor; skills; synaptic function; tool; training opportunity; uptake; vacuolar H+-ATPase

PROJECT SUMMARY/ABSTRACT Vesicular glutamate transporters (VGLUTs) package the major excitatory neurotransmitter glutamate into synaptic vesicles, thus play a critical role in quantal release and neurotransmission. Despite decades of research, the mechanism of glutamate transport by the VGLUTs and its regulation under different ionic and pH conditions during exocytosis and endocytosis remains poorly understood. The lack of molecular level understanding of the structure and function of VGLUTs severely hinders our ability to understand their role in normal brain function, as well as in many psychiatric and neurological conditions where malfunction of VGLUTs has been implicated. This proposal aims to bridge this gap by providing a molecular blue print of a mammalian VGLUT protein with atomic details. Specifically, the candidate Dr. Fei Li will determine high-resolution structures of the rat VGLUT1 (rVGLUT1) by cryo electronmicrosocpy (cryo-EM) (Aim 1, mentored) and characterize its function by electrophysiology (Aim 2, mentored). Dr. Fei Li will further determine the structure of the native synaptic vesicle by cryo electrontomography (cryo-ET) (Aim 3A, mentored) and to reveal the higher order organization and interactions of synaptic vesicle membrane proteins (Aim 3B, independent). The goal of this K99/R00 Pathway to Independence Award proposal is to enhance Dr. Li’s knowledge in neuroscience and to provide opportunity of additional training in several key techniques that are critical to launching her independent research career focusing on the structure and function of neuronal membrane proteins in a leading research institution. The proposed aims allow Dr. Li to explore new biological questions that she can continue to investigate in her independent career while providing opportunities for training at the same time. Dr. Li has a strong interest in membrane protein structure and function in the context of brain physiology, and a strong background in membrane protein and X-ray crystallography. To guide the proposed project and to enhance her career prospects, Dr. Li has selected three leading scientists in, respectively, membrane proteins (Dr. Robert Stroud), electron microscopy (Dr. Yifan Cheng), as well as electrophysiology and neuroscience (Dr. Robert Edwards) as her co-mentors. Training through this project will allow Dr. Li to build a highly versatile and integrative skill set extending her background in membrane proteins and X-ray crystallography to the state-of-the-art cryo-EM and cryo-ET technology. Skills in electrophysiology will provide her with the tools required for functional analysis. Together, the research project and training program proposed in this award will play a critical role in preparing Dr. Li for a successful independent research career.

项目摘要/摘要 囊泡谷氨酸转运蛋白（VGLUTS）包装主要兴奋神经递质谷氨酸 突触蔬菜，因此在量化和神经传递中起关键作用。尽管数十年 研究，vgluts谷氨酸转运的机制及其在不同离子和pH下的调节 胞吐作用和内吞作用期间的状况仍然很少了解。缺乏分子水平 了解Vgluts的结构和功能，严重阻碍了我们了解其在 正常的大脑功能以及在许多精神病和神经系统状况下，vgluts的故障 已暗示。该建议旨在通过提供哺乳动物的分子蓝色印刷品来弥合这一差距 带有原子细节的VGLUT蛋白。具体而言，候选人Fei Li博士将确定高分辨率 Cryo Electronicsicrosocpy（Cryo-EM）（AIM 1，指导）和 通过电生理学来表征其功能（AIM 2，指导）。 Fei Li博士将进一步确定 通过冷冻电显影（Cryo-ET）（AIM 3A，指导）和 揭示高阶组织和突触囊泡膜蛋白的相互作用（目标 3B，独立）。这项K99/R00独立奖提案的目标是增强Li博士 关于神经科学的知识，并为几种关键技术提供其他培训的机会 关注神经元的结构和功能的独立研究职业至关重要 领先的研究机构中的膜蛋白。拟议的目的使李博士能够探索新的生物学 她可以在独立职业中继续调查的问题，同时为 同时训练。 Li博士对膜蛋白结构和功能有浓厚的兴趣 脑生理学，以及膜蛋白和X射线晶体学方面的强烈背景。指导 提议的项目并为了提高她的职业前景，李博士选择了三位领先的科学家， 分别是膜蛋白（Robert Stroud博士），电子显微镜（Yifan Cheng博士）以及 电生理学和神经科学（Robert Edwards博士）作为她的联合主张。通过这个项目培训将 允许Li博士建立一个高度通用和集成的技能集，以扩展其在膜蛋白上的背景 以及最先进的冷冻EM和Cryo-ET技术的X射线晶体学。电生理学的技能 将为她提供功能分析所需的工具。一起研究和培训 该奖项中提出的计划将在为成功的独立的李博士准备李博士方面发挥关键作用 研究职业。