Tools For High-Throughput High-Resolution Three Dimensional Electron Microscopy

总结 三维电子显微镜（3DEM）是一套用于确定结构的技术， 大蛋白质和蛋白质复合物。已经表明，低温EM（cryo-EM）具有以下能力： 以足够高的分辨率确定结构，以明确地将侧链放置在EM密度中。 虽然低温EM越来越接近原子分辨率，但在这方面存在许多障碍。 为了实现侧链水平的分辨率，结构测定低对称性和无对称性 配合物在这里，我们建议开发工具，以促进高通量高分辨率3DEM。在 第一个目标，我们建议开发工具，使自动化标本制备和筛选， 为直接电子探测提供了新的工具。在第二个目标中，我们建议开发工具， 断层摄影数据收集和处理，包括探索新的图像采集模式 以及用于开发用于处理断层摄影子体积的基础设施。这项研究将有利于 通过促进确定与人类疾病相关的蛋白质结构来促进人类健康。

Summary Three-dimensional electron microscopy (3DEM) is a set of techniques for determining the structures of large proteins and protein complexes. It has been shown that cryogenic EM (cryo-EM) has the ability to determine structures to high enough resolution to unambiguously place side chains in the EM density. Though cryo-EM is increasingly reaching near-atomic resolution, a number of hurdles are present in order to achieve side-chain level resolution structure determination of low-symmetry and no symmetry complexes. Here we propose to develop tool to facilitate high-throughput high-resolution 3DEM. In the first aim, we propose to develop tools to enable automated specimen preparation and screening, and enable a new tools for direct electron detection. In the second aim, we propose to develop tools for tomographic data collection and processing including exploring new modalities for image acquisition and for developing an infrastructure for processing tomographic subvolumes. This research will benefit human health by facilitating the determination of protein structures that are relevant for human disease.