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.

项目摘要 近年来，结合三维重建的单粒子冷冻电子显微镜（cryo-EM）出现了 作为解决病毒和大分子复合物的高分辨率3D结构的革命性工具。 近原子分辨率（3- 4 nm）结构的快速增长，使用冷冻EM解决了允许 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 近原子分辨率的冷冻EM结构，需要收集高分辨率的图像数据， 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。 检测器直接电子探测器不仅可以提高图像的分辨率和对比度，而且还可以记录电影 用于曝光期间电子束引起的样品移动的后续计算校正。的 增加的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是必要的。 蛋白质复合物然而，购买和维护最先进的冷冻EM资源的高成本， 高端电子显微镜和直接电子探测器，排除了许多冷冻EM研究人员 使用这些新技术。在这里，建立一个中西部高- 分辨率冷冻电子显微镜被提议提供对这样的高分辨率数据收集的访问 在无法获得此类资源的情况下，低温EM实验室的能力。该联盟将由4名 来自主办机构（普渡大学）的研究者这将保持高分辨率的数据收集 资源（Titan Krios 300 kV FEG显微镜，具有相位板、能量过滤器和直接电子检测器）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，罗格斯大学宾夕法尼亚州立医学院，亚拉巴马大学伯明翰分校 （科罗拉多博尔德大学、堪萨斯大学、密苏里州大学、佛蒙特大学和弗吉尼亚联邦大学）。 普渡设施的工作人员，教师和现场服务工程师的集体经验，在高分辨率 cryo-EM成像将确保该设施以最低限度的服务中断以最高性能运行。 在普渡大学建立的高分辨率数据收集能力和合作实验室的访问能力，将 允许这些研究者，除了3名新的研究者，NIH资助的，克服 分辨率障碍，并促进在他们自己的cryo-EM项目中对一系列结构的发现，例如， 细菌病原体粘附蛋白，人病毒，亨廷顿病蛋白，突触囊泡蛋白， 化学感受器信号复合物等。该联盟将提供全面的支持，冷冻EM 项目，包括运送样品，准备样品网格，收集高分辨率的单颗粒， 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 收集服务的范围将从合作伙伴只需邮寄样品的完整数据收集， 接收图像数据，由主机机构工作人员培训的合作伙伴动手操作cryo-EM。