Improved data collection for FEI TF20 and Philips CM12 electron microscopes

改进了 FEI TF20 和 Philips CM12 电子显微镜的数据收集

• 批准号: 7591369
• 负责人: ESTHER BULLITT
• 金额: $ 26.33万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7591369
• 关键词: Apoptosis; Area; Arts; Astigmatism; Bacteria; Bacterial Adhesion; Biogenesis; Cardiovascular Diseases; Cell Death; Cholesterol; Comparative Study; Complex; Data; Data Collection; Data Set; Dose; Electron Microscope; Film; Funding; Generations; Human; Human poliovirus; Image; Low-Density Lipoproteins; Mammals; Mediating; Membrane; Microscope; Minor; Mitochondria; Muscle; Muscle Contraction; Organ; Pathogenesis; Pathway interactions; Pilum; Polioviruses; Polymerase; Prevention; Productivity; Protein Biosynthesis; Proteins; Regulation; Request for Proposals; Research; Research Project Grants; Resolution; Ribosomes; Role; Sampling; Skeletal Muscle; Structure; System; Thin Filament; Time; Training; Viral; Work; base; charge coupled device camera; digital imaging; human disease; image reconstruction; improved; macromolecular assembly; nitrogen metabolism; public health relevance; receptor binding; respiratory; tool

DESCRIPTION (provided by applicant): This proposal requests funding for the purchase of a TVIPS 4K x 4K CCD camera for our FEI Tecnai F20 (TF20) electron microscope, with a 1K x 1K Fastscan CCD accessory. Research by the major users of the TF20, including six NIH-funded projects, provide cutting edge, state-of-the-art advances in the fields of bacterial pathogenesis, apoptosis, cardiovascular disease, viral replication, protein biogenesis, and muscle regulation. Minor users broaden our research further with four NIH-funded studies on the glycolytic pathway, nitrogen metabolism in bacteria, and the mitochondrial respiratory pathway. The purchase of a new CCD camera for the TF20 will allow us to collect data efficiently and accurately for the following major projects: 1) comparative studies on the specialization of pathogenic bacterial adhesion pili for colonization of specific organs in their human hosts, 2) studies on the apoptosome, a macromolecular assembly that is critical to the mitochondrial apoptosis pathway, 3) studies on the role of `bad cholesterol' in cardiovascular disease: the structure and function of LDL low density lipoproteins alone and in complex with its binding receptor, LDLR, 4) studies on the role of protein oligomerization for the replication of poliovirus RNA, including studies on the 3Dpol polymerase in isolation and tethered to membranes via the protein 3AB, 5) comparative studies of ribosome-Sec61 and ribosome-SecY complexes that mediate co-translational translocation of nascent proteins in mammals and bacteria, and 6) studies on the regulation of skeletal muscle contraction by thin filaments. The addition of a large format CCD to our existing TEM will allow us to remain at the forefront of our respective fields. The 1K x 1K CCD that is currently on theTF20 is an excellent tool for astigmatism correction, focus adjustment, and for checking a sample after a low dose image has been recorded on film. The requested large format CCD captures images of an area that is ten times larger than our current CCD. This increase in the amount of sample imaged is essential, as the limits imposed by our current CCD exclude the possibility of collecting the large datasets required for high resolution image reconstructions. However, the existing CCD will be a significant asset for routine work that is done on our Philips CM12 electron microscope Thus, an additional key benefit of this proposal will be the transfer of our existing TVIPS 1K x 1K CCD to our Philips CM12 electron microscope, for which no funding is requested. The CM12 has a large user base that currently includes the five major TF20 users and seven additional projects, six of which are NIH-funded. There is also a steady influx of new users. Having a CCD on this microscope will be extremely beneficial both as a training tool and for data collection on the CM12. Public Health Relevance: This proposal requests funding to upgrade our FEI Tecnai F20 electron microscope with a digital imaging system. Research by users of the electron microscope, including 10 NIH-funded projects, provide cutting edge, state-of-the-art advances in the fields of cardiovascular disease, bacterial pathogenesis, cell death (apoptosis), viral replication, muscle regulation, protein biosynthesis, and energy generation. Addition of this hardware will significantly improve the productivity of all these research projects, advancing research toward the prevention or elimination of human disease.

描述（由申请人提供）：该提案要求为我们的FEI Tecnai F20（TF 20）电子显微镜购买TVIPS 4K x 4K CCD相机提供资金，并配备1 K x 1 K Fastscan CCD配件。TF 20的主要用户的研究，包括六个NIH资助的项目，提供了细菌发病机制，细胞凋亡，心血管疾病，病毒复制，蛋白质生物合成和肌肉调节领域的尖端，最先进的进展。小用户通过NIH资助的四项关于糖酵解途径、细菌氮代谢和线粒体呼吸途径的研究进一步扩大了我们的研究。为TF 20购买新的CCD相机将使我们能够为以下主要项目高效准确地收集数据：1）关于致病性细菌粘附皮利在其人类宿主中的特定器官定殖的特化的比较研究，2）关于线粒体体（一种对线粒体凋亡途径至关重要的大分子组装体）的研究，3）关于“坏胆固醇”在心血管疾病中的作用的研究：LDL低密度脂蛋白单独和与其结合受体LDLR复合的结构和功能，4）研究蛋白质寡聚化对脊髓灰质炎病毒RNA复制的作用，包括对分离的3Dpol聚合酶和通过蛋白质3AB与膜连接的3Dpol聚合酶的研究，5）对哺乳动物和细菌中介导新生蛋白质共翻译易位的核糖体-Sec 61和核糖体-SecY复合物的比较研究，（6）细肌丝对骨骼肌收缩的调节研究。在我们现有的TEM上增加一个大幅面CCD将使我们能够保持在各自领域的最前沿。目前TF 20上的1 K x 1 K CCD是一款出色的工具，可用于像散校正、焦距调整以及在胶片上记录低剂量图像后检查样品。要求的大格式CCD捕获的图像面积是我们目前CCD的十倍。成像样本量的增加至关重要，因为我们当前的CCD所施加的限制排除了收集高分辨率图像重建所需的大型数据集的可能性。然而，现有的CCD将是在我们的Philips CM 12电子显微镜上完成的常规工作的重要资产。因此，该提案的另一个主要好处是将我们现有的TVIPS 1 K x 1 K CCD转移到我们的Philips CM 12电子显微镜，无需资金。CM 12拥有庞大的用户群，目前包括五个主要的TF 20用户和七个额外的项目，其中六个是NIH资助的。新用户也在不断涌入。在这台显微镜上安装一个CCD将是非常有益的，既可以作为培训工具，也可以用于CM 12上的数据收集。公共卫生相关性：该提案要求提供资金，以升级我们的FEI Tecnai F20电子显微镜，配备数字成像系统。电子显微镜用户的研究，包括10个NIH资助的项目，提供了心血管疾病，细菌发病机制，细胞死亡（凋亡），病毒复制，肌肉调节，蛋白质生物合成和能量产生领域的尖端，最先进的进展。这些硬件的增加将大大提高所有这些研究项目的生产力，推动研究朝着预防或消除人类疾病的方向发展。

项目成果

Antibody-mediated disruption of the mechanics of CS20 fimbriae of enterotoxigenic Escherichia coli.

抗体介导的产肠毒素大肠杆菌 CS20 菌毛力学破坏。

• DOI: 10.1038/srep13678
• 发表时间: 2015
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Singh,Bhupender;Mortezaei,Narges;Uhlin,BerntEric;Savarino,StephenJ;Bullitt,Esther;Andersson,Magnus
• 通讯作者: Andersson,Magnus

Biomechanical and structural features of CS2 fimbriae of enterotoxigenic Escherichia coli.

产肠毒素大肠杆菌CS2菌毛的生物力学和结构特征。

• DOI: 10.1016/j.bpj.2015.05.022
• 发表时间: 2015
• 期刊: Biophysical journal
• 影响因子: 3.4
• 作者: Mortezaei,Narges;Singh,Bhupender;Zakrisson,Johan;Bullitt,Esther;Andersson,Magnus
• 通讯作者: Andersson,Magnus

Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled.

抗体会损害菌毛的弹性，导致它们变得僵硬和缠结。

• DOI: 10.1128/jb.00665-16
• 发表时间: 2017
• 期刊: Journal of bacteriology
• 影响因子: 3.2
• 作者: Singh,Bhupender;Mortezaei,Narges;Savarino,StephenJ;Uhlin,BerntEric;Bullitt,Esther;Andersson,Magnus
• 通讯作者: Andersson,Magnus