STRUCTURE

结构

• 批准号: 8121484
• 负责人: IAIN D CAMPBELL
• 金额: $ 13.13万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至 2014-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8121484
• 关键词: Actinin; Actins; Adhesions; Binding; Binding Sites; Cells; Complex; Data; Docking; Electron Microscopy; Goals; Image; Immigration; Individual; Integrins; Life; Light; Maps; Mediating; Methods; Microscopy; Organelles; Phase; Proteins; Resolution; Roentgen Rays; Structure; Three-dimensional analysis; Vinculin; base; light microscopy; migration; three dimensional structure; tomography

The continued goal of this Initiative is to determine the high resolution structures of two major migration organelles, adhesions and protrusions. Our approach is to combine structural methods that range from correlative light and high resolution electron microscopy to X-ray and NMR. As part of this effort, we are developing a new imaging concept that allows us to capture by light microscopy the spatial-temporal aspects of migration in living cells and to visualize the same structures using high resolution electron microscopy - 3D correlative light and electron microscopy ("top down approaches"). High resolution structures of individual proteins and their detailed interactions with binding partners obtained by NMR, X-ray or tomography ("bottom up approaches") can then be computationally docked into the three-dimensional maps of the complexes. In the first phase of the Consortium, this paradigm was applied to the structure of the actin network in protrusions, providing 3D structures of Arp2/3 mediated actin branches and the structural basis for Arp2/3 activation. We also determined the ultrastructure of the lamellipodia of living cells using our new 3D correlative EM and speckle microscopy and determined the vinculin binding site on a-actinin in the presence of integrin. A unified framework for segmentation and analysis of 3-D data was developed to enable the identification of assemblies in 3-D structures.

该倡议的持续目标是确定两个主要的高分辨率结构， 迁移细胞器、粘连和突起。我们的方法是将联合收割机的结构方法， 从相关光学和高分辨率电子显微镜到X射线和NMR。作为这项努力的一部分，我们 开发一种新的成像概念，使我们能够通过光学显微镜捕捉的时空方面， 迁移活细胞和可视化相同的结构，使用高分辨率电子显微镜- 3D 相关光学和电子显微镜（“自上而下的方法”）。个体的高分辨率结构 通过NMR、X射线或层析成像获得的蛋白质及其与结合配偶体的详细相互作用（“底部 向上方法”）然后可以计算地对接到复合体的三维图中。在 在联盟的第一阶段，这种范例被应用于突起中肌动蛋白网络的结构， 提供Arp 2/3介导的肌动蛋白分支的3D结构和Arp 2/3激活的结构基础。我们 还用我们新的三维相关电镜测定了活细胞板状伪足的超微结构， 斑点显微术，并确定在整联蛋白存在下α-辅肌动蛋白上粘着斑蛋白结合位点。统一的 开发了一个用于分割和分析三维数据的框架，以识别装配体 在三维结构中。