Biophysical Studies of Macromolecules and Molecular Assemblies

大分子和分子组装体的生物物理研究

• 批准号: 9069538
• 负责人: STEVEN G. BOXER
• 金额: $ 37.39万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9069538
• 关键词: Active Sites; Address; Applications Grants; Architecture; Area; Award; Biological; Biological Process; Biology; Biophysics; Biotechnology; Catalysis; Cell physiology; Development; Dissociation; Drug usage; Electrophysiology (science); Electrostatics; Enzymes; Event; Evolution; Exhibits; Funding Opportunities; Goals; Grant; Green Fluorescent Proteins; Image; Integral Membrane Protein; Interferometry; Lateral; Light; Lipids; Maps; Mass Spectrum Analysis; Measurement; Membrane; Membrane Fusion; Membrane Proteins; Methods; Motion; National Institute of General Medical Sciences; Optics; Process; Proteins; Protons; Reaction; Research; Research Personnel; Resolution; Spectrum Analysis; Time; Variant; Vesicle; Viral; Vision; Work; analytical method; biological systems; biophysical analysis; chromophore; design; electric field; imaging modality; macromolecule; membrane model; model development; molecular assembly/self assembly; novel; optogenetics; public health relevance

 DESCRIPTION (provided by applicant): This MIRA grant proposal briefly summarizes the activities currently supported by two NIGMS grants and my vision for the evolution of this research over the next five years. The common theme that unifies this research is a unique combination of the development and application of new physical methods that can be broadly applied to the quantitative analysis of biological systems. The long-term goals of GM27738 (Electrostatics and Dynamics in Proteins) are to develop spectroscopic methods for probing electric fields in proteins and to apply these methods to obtain quantitative information on fields and their effects on function at the active sites of several enzymes. We have led the development of vibrational Stark spectroscopy as a general approach to map these fields. Recent and continuing work emphasizes the connection between electrostatics and catalysis where we can, for the first time, estimate the electrostatic contribution to the catalytic proficiecy of enzymes. We also study dynamics in green fluorescent protein (GFP), which we began to study reasoning that the native chromophore could be used to probe solvation dynamics (time dependent electric fields) in proteins. Instead we discovered that GFP exhibits excited state proton transfer. Recent work emphasizes novel applications of "split" GFP where light-driven association and dissociation reactions have been discovered. We seek to understand the basic mechanism(s) of these processes and optimize them for optogenetic and imaging applications. The long-term goals of GM069630 (Membrane Fusion and Dynamics Using Supported Bilayers) are to develop methods to probe the organization and dynamic reorganization of lipids and proteins in biological membranes and to apply these methods to problems of broad biological importance. Our lab has pioneered the development of model membrane architectures, along with imaging and analytical methods, that probe fundamental aspects of membrane organization and dynamics. We use these approaches to address open questions in three areas of current biological significance. (i) The mechanism of vesicle and enveloped viral membrane fusion is studied using model membrane architectures we developed as targets to precisely probe the steps of fusion at the single event level. (ii) The organization of lipids and membrane proteins are characterized with unprecedented lateral resolution using imaging mass spectrometry, emphasizing the correlated motion of lipid and protein components believed to be associated in rafts. (iii) A novel "membrane interferometer" has been designed with the ultimate goal of correlated measurements of integral membrane protein conformational changes by optical interferometry and electrical activity by electrophysiological methods. Each of these areas represents a major current challenge in membrane biophysics and biology.

 描述（由申请人提供）：本MIRA赠款提案简要总结了目前由两个NIGMS赠款支持的活动，以及我对未来五年这项研究发展的愿景。统一这项研究的共同主题是新物理方法的开发和应用的独特组合，可广泛应用于生物系统的定量分析。 GM27738（蛋白质中的静电学和动力学）的长期目标是开发用于探测蛋白质中电场的光谱方法，并应用这些方法获得场的定量信息 以及它们对几种酶活性位点功能的影响。我们领导了振动斯塔克光谱的发展，作为一种通用的方法来映射这些领域。最近和持续的工作强调了静电和催化之间的联系，我们可以第一次估计静电对酶催化效率的贡献。我们还研究了绿色荧光蛋白（GFP）中的动力学，我们开始研究天然发色团可用于探测蛋白质中的溶剂化动力学（时间依赖性电场）的推理。相反，我们发现GFP表现出激发态质子转移。最近的工作强调了新的应用“分裂”GFP光驱动的协会和解离反应已被发现。我们试图了解这些过程的基本机制，并优化它们用于光遗传学和成像应用。 GM069630的长期目标（使用支撑双层的膜融合和动力学）是开发方法来探测生物膜中脂质和蛋白质的组织和动态重组，并将这些方法应用于具有广泛生物重要性的问题。我们的实验室率先开发了模型膜结构，沿着成像和分析方法，探索膜组织和动力学的基本方面。我们使用这些方法来解决目前生物学意义的三个领域的开放问题。(i)囊泡和包膜病毒膜融合的机制进行了研究，使用模型膜结构，我们开发的目标，以精确探测融合的步骤，在单事件水平。(ii)脂质和膜蛋白的组织的特点是前所未有的横向分辨率使用成像质谱，强调相关的运动的脂质和蛋白质成分被认为是相关的筏。(iii)设计了一种新型的“膜干涉仪”，其最终目标是通过光学干涉术和电生理学方法对膜蛋白的构象变化和电活性进行相关测量。这些领域中的每一个都代表了膜生物物理学和生物学中当前的主要挑战。