Acquisition of 200keV FEG Cryo-Electron Microscope

购置 200keV FEG 冷冻电子显微镜

• 批准号: 0002805
• 负责人: Edward Egelman
• 金额: $ 22.5万
• 依托单位: University of Virginia Main Campus
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2002-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0002805&HistoricalAwards=false
• 关键词: Acquisition; 200keV; FEG; Cryo; Electron

ABSTRACT0002805Edward H. EgelmanUniversity of VirginiaAcquisition of 200 KeV FEG Cryo-Electron Microscope Electron microscopy (EM) is an extremely powerful technique for studying the structure of macromolecular complexes, such as protein polymers, protein-DNA complexes and viruses. Significant advances in electron microscopy have been made in specimen preparation, instrumentation and image. These advances make it possible to use an electron microscopic to determine the three-dimensional structure of a protein. In addition, they also allow for lower-resolution studies that are complementary to those obtained by such techniques as x-ray crystallography and nuclear magnetic resonance. A 200 KeV Field Emission Gun cryo-electron microscope, manufactured by Philips/FEI, will significantly enhance the research programs of a group of six investigators at the University of Virginia Medical School. This microscope will have a highly coherent electron source that will allow for resolutions much higher than obtainable with conventional instruments. The microscope will be equipped for the imaging of frozen-hydrated specimens, maintained in the microscope at liquid nitrogen temperatures. It will also have image acquisition capabilities, using a CCD camera, that will allow for on-line image analysis. The projects that will take advantage of the increased resolution provided by this instrument are: 1) studies of F-actin, and complexes of F-actin with actin-binding proteins; 2) studies of protein-DNA complexes, particularly helical recombination filaments such as RecA/Rad51/UvsX, and ring helicases such as rho, DnaB and T7 gp4; 3) studies of the icosahedral Herpes Simplex virus capsid; 4) studies of Ca2+-dependent membrane binding proteins such as annexin, using two dimensional crystals formed on lipid monolayers; 5) studies of Protein Kinase C isozymes using two dimensional crystals formed on lipid monolayers; and 6) studies of ion channels, such as VacA hexamers, using cryo-EM of two-dimensional crystals to complement ongoing studies with Atomic Force Microscopy. Many of the projects have already produced results that are at the limit of reliable resolution obtainable by conventional TEM of negatively stained specimens. The new instrument will not only enhance these individual research programs, but will greatly strengthen the environment for structural biology at the University of Virginia and the training of graduate students and postdoctoral fellows.

摘要0002805爱德华H. Egelman美国埃格尔曼大学获得200 KeV FEG低温电子显微镜电子显微镜（EM）是研究大分子复合物（如蛋白质聚合物、蛋白质-DNA复合物和病毒）结构的一种极其强大的技术。 电子显微镜在样品制备、仪器和图像方面取得了重大进展。 这些进展使得使用电子显微镜来确定蛋白质的三维结构成为可能。 此外，它们还允许低分辨率的研究，这些研究与通过X射线晶体学和核磁共振等技术获得的研究互补。由飞利浦/FEI制造的200 KeV场发射枪低温电子显微镜将显着增强弗吉尼亚大学医学院六名研究人员的研究项目。 这种显微镜将有一个高度相干的电子源，将允许分辨率比传统仪器高得多。 显微镜将配备用于冷冻水合标本成像的设备，保存在液氮温度下的显微镜中。 它还将具有图像采集能力，使用CCD照相机，可进行在线图像分析。将利用该仪器提供的更高分辨率的项目有：1）F-肌动蛋白和F-肌动蛋白与肌动蛋白结合蛋白的复合物的研究; 2）蛋白质-DNA复合物的研究，特别是螺旋重组丝如RecA/Rad 51/UvsX和环解旋酶如rho、DnaB和T7 gp 4; 3）二十面体单纯疱疹病毒衣壳的研究; 4）使用在脂质单层上形成的二维晶体研究Ca 2+依赖性膜结合蛋白，例如膜联蛋白; 5）使用在脂质单层上形成的二维晶体研究蛋白激酶C同工酶;以及6）使用二维晶体的冷冻-EM研究离子通道，例如VacA六聚体，以补充使用原子力显微镜进行的研究。 许多项目已经产生的结果是在可靠的分辨率的限制，可通过传统的TEM负染色标本。 新仪器不仅将增强这些个人研究计划，而且将大大加强弗吉尼亚大学的结构生物学环境以及研究生和博士后研究员的培训。