A Helical Crystallization Pipeline for Membrane Protein Structure by TEM

通过 TEM 分析膜蛋白结构的螺旋结晶流程

• 批准号: 8517141
• 负责人: BRIDGET Olivia CARRAGHER
• 金额: $ 36.44万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-30 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8517141
• 关键词: Accounting; Affinity; Area; Binding; Binding Proteins; Biological Factors; Caliber; Characteristics; Classification; Complement; Complex; Crystallization; Data; Databases; Detergents; Development; Dialysis procedure; Electrons; Ensure; Evaluation; Filler; Goals; Growth; Head; Image; Integral Membrane Protein; Licensing; Ligands; Lipids; Maps; Membrane Proteins; Methods; Modeling; Molecular Machines; Nickel; Online Systems; Organism; Outcome Assessment; Pharmaceutical Preparations; Play; Procedures; Process; Production; Property; Protein Binding; Proteins; Research; Resolution; Scoring Method; Solutions; Structure; Surface; Therapeutic; Tube; Tubular formation; Vesicle; X-Ray Crystallography; abstracting; electron crystallography; graphical user interface; insight; interest; light microscopy; novel; novel strategies; open source; particle; protein complex; protein structure; reconstitution; reconstruction; screening; three dimensional structure; tool; transmission process

Carragher, Bridget Abstract Electron crystallography has a key role to play in understanding integral membrane proteins (IMPs) as it provides one of the most direct means of providing insight into the functional state of these molecular machines in their lipid-associated forms, and also has the potential to facilitate examination of physiologically relevant transitional states and complexes. Helical or tubular crystals, which are the natural product of proteins crystallizing on the surface of a vesicle, offer some unique advantages over other crystalline forms. The goal of this proposal is to develop an integrated pipeline to enable structure determination by transmission electron microcopy of IMPs in the form of tubular crystals. Given the very high number of IMPs that need to be solved, helical crystallization should be available as a standard method in the armamentarium of approaches for determining the structure of IMPs. However, its use is currently limited by a lack of standardized approaches and the enormous effort and specialized expertise required that form a barrier to entry. New or streamlined approaches are needed in three areas: crystal growth, screening, and analysis. An integrated approach is essential to ensure that progress in any one area is not throttled by bottlenecks at significant other steps in the process. Our goal is to establish TEM of tubular crystals as a standard approach to complement X-ray crystallography and NMR for characterizing IMPs.

卡拉格，布里奇特 摘要 电子晶体学在理解膜整合蛋白（IMP）方面发挥着关键作用，因为它 提供了一种最直接的手段，提供洞察这些分子的功能状态， 机器在其脂质相关的形式，也有可能促进检查生理 相关的过渡状态和复合体。螺旋状或管状晶体，是蛋白质的天然产物 在囊泡的表面上结晶，提供了一些优于其他结晶形式的独特优势。目标 这一建议的一个重要方面是开发一个集成的管道，使结构确定的透射电子 管状晶体形式的IMP显微镜。鉴于需要解决的IMP数量非常多， 螺旋结晶应该是一个标准的方法，在医疗设备的方法， 确定IMP的结构。然而，由于缺乏标准化的方法，其使用目前受到限制 以及所需的巨大努力和专业知识构成了进入的障碍。新的或精简的 需要在三个方面采取措施：晶体生长、筛选和分析。采取综合办法 必须确保任何一个领域的进展不因其他重大步骤的瓶颈而受到阻碍， 过程我们的目标是建立管状晶体的TEM作为标准方法，以补充X射线 用于表征IMP的晶体学和NMR。