Midwest Consortium for High Resolution Cryoelectron Microscopy

中西部高分辨率冷冻电子显微镜联盟

• 批准号: 9257031
• 负责人: ESTHER BULLITT
• 金额: $ 134.88万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9257031
• 关键词: 3-Dimensional; Adhesions; Alabama; Antiviral Agents; Back; Bacterial Proteins; Biological Process; Boston; Cell physiology; Chemoreceptors; Collaborations; Collection; Colorado; Complex; Cryoelectron Microscopy; Data; Data Collection; Detection; Development; Disease; Early Diagnosis; Electron Beam; Electron Microscope; Electrons; Engineering; Ensure; Equipment; Faculty; Fostering; Freezing; Funding; Home environment; Human; Huntington Disease; Image; Institution; Interruption; Kansas; Laboratories; Light; Macromolecular Complexes; Manuals; Medicine; Methods; Michigan; Microscope; Midwestern United States; Missouri; Motion; Movement; Performance; Phase; Productivity; Proteins; Recovery; Research; Research Personnel; Resolution; Resources; Sampling; Schedule; Services; Shipping; Signal Transduction; Site; Structure; Synaptic Vesicles; Techniques; Titan; Training; United States National Institutes of Health; Universities; Vermont; Virginia; Virus; Virus Diseases; Visit; college; contrast imaging; cost; detector; experience; faculty support; flexibility; high resolution imaging; image processing; improved; insight; instrument; inter-institutional; medical schools; method development; movie; operation; particle; pathogen; portability; protein complex; public health relevance; quantum; reconstruction; repaired; supercomputer; symposium; three dimensional structure; tomography; tool; transmission process

PROJECT SUMMARY Recently, single particle cryo-electron microscopy (cryo-EM) combined with 3-D reconstruction has emerged as a revolutionary tool for solving high-resolution 3-D structures of viruses and macromolecular complexes. The rapidly increasing number of near-atomic resolution (3-4Ǻ) structures solved using cryo-EM has allowed unprecedented atomic level understanding of fundamental cellular processes and viral infections. To obtain near-atomic resolution cryo-EM structures, requires the collection of high-resolution image data using state-of- the-art imaging resources, including both a high-end transmission electron microscope and a direct electron detector. The direct electron detectors not only improve image resolution and contrast, but also record movies for subsequent computational correction of electron beam-induced, sample movements during exposure. The increased image contrast and resolution are essential for solving near-atomic resolution structures of small protein complexes. However, the high cost to purchase and maintain a state-of-the-art cryo-EM resource, with both a high-end electron microscope and a direct electron detector, precludes many cryo-EM investigators from having access to these new techniques. Here, the creation of a Midwest Consortium for High- Resolution Cryo-electron Microscopy is proposed to provide access to such high-resolution data collection capability for cryo-EM laboratories without access to such resources. This consortium will consist of 4 investigators from the host institution (Purdue Univ.) which will maintain the high-resolution data collection resource (Titan Krios 300kV FEG microscope with phase plate, energy filter, and direct electron detector) and will provide services to 11 investigators from 10 partnering institutions (Boston Univ. School of Medicine, Michigan State Univ., Penn State College of Medicine, Rutgers Univ., Univ. of Alabama at Birmingham, Univ. of Colorado Boulder, Univ. of Kansas, Univ. of Missouri, Univ. of Vermont, and Virginia Commonwealth Univ.). The collective experience of the Purdue facility staff, faculty and onsite service engineer, in high-resolution cryo-EM imaging, will ensure that the facility operates at peak performance with minimal service interruptions. The high-resolution data collection capabilities established at Purdue and accessible to the partnering labs, will allow these investigators, who are all, except for the 3 new investigators, NIH-funded, to overcome the resolution barrier and facilitate discovery within their own cryo-EM projects on a range of structures, such as, bacterial pathogen adhesion proteins, human viruses, Huntington's Disease proteins, synaptic vesicle proteins, chemoreceptor signaling complex, etc. The Consortium will provide comprehensive support to the cryo-EM projects, including shipping samples, preparing sample grids, collecting high-resolution single particle and tomography images, processing raw movies, and transferring data back to the partners' labs. The data collection services will range from full data collection where partners simply mail in the samples and then receive the image data, to hands-on operations of the cryo-EM by partners trained by the host facility's staff.

项目概要 最近，单粒子冷冻电子显微镜 (cryo-EM) 与 3-D 重建相结合已经出现 作为解决病毒和大分子复合物高分辨率 3D 结构的革命性工具。 使用冷冻电镜解决的近原子分辨率 (3-4Ǻ) 结构数量迅速增加，使得 对基本细胞过程和病毒感染的前所未有的原子水平理解。获得 近原子分辨率冷冻电镜结构，需要使用状态收集高分辨率图像数据 最先进的成像资源，包括高端透射电子显微镜和直接电子显微镜 探测器。直接电子探测器不仅可以提高图像分辨率和对比度，还可以录制电影 用于随后对曝光期间电子束引起的样品运动进行计算校正。这 增加图像对比度和分辨率对于解决小尺寸的近原子分辨率结构至关重要 蛋白质复合物。然而，购买和维护最先进的冷冻电镜资源的成本很高， 高端电子显微镜和直接电子探测器让许多冷冻电镜研究人员望而却步 从获得这些新技术。在这里，创建了中西部高中联盟 分辨率冷冻电子显微镜被提议提供对这种高分辨率数据收集的访问 无法获得此类资源的冷冻电镜实验室的能力。该财团将由 4 家组成 来自主办机构（普渡大学）的研究人员将维护高分辨率数据收集 资源（Titan Krios 300kV FEG 显微镜，带相位板、能量滤波器和直接电子探测器）和 将为来自 10 个合作机构（波士顿大学医学院、 密歇根州立大学、宾夕法尼亚州立大学医学院、罗格斯大学、大学。阿拉巴马州伯明翰大学。 科罗拉多博尔德大学堪萨斯大学密苏里大学佛蒙特州和弗吉尼亚联邦大学）。 普渡大学设施工作人员、教师和现场服务工程师的集体经验，高分辨率 冷冻电镜成像将确保该设施以最佳性能运行，并将服务中断降至最低。 普渡大学建立的高分辨率数据收集能力可供合作实验室使用，将 允许这些研究人员（除了 NIH 资助的 3 名新研究人员之外）克服 解决分辨率障碍并促进他们自己的冷冻电镜项目中对一系列结构的发现，例如， 细菌病原体粘附蛋白、人类病毒、亨廷顿病蛋白、突触小泡蛋白、 化学感受器信号复合体等，联盟将为冷冻电镜提供全面支持 项目，包括运输样品、准备样品网格、收集高分辨率单粒子和 断层扫描图像、处理原始电影并将数据传输回合作伙伴的实验室。数据 收集服务的范围包括完整的数据收集，合作伙伴只需邮寄样本，然后 接收图像数据，由受过主机设施工作人员培训的合作伙伴亲自操作冷冻电镜。