Structural Studies of Togaviruses

项目摘要 晶体学是第一个开启了结构研究的主要结构工具， 近原子分辨率的病毒例如，第一个近原子分辨率的动物结构 病毒，即普通感冒病毒，是我们在20世纪80年代中期使用晶体学方法确定的。 方法（Nature 317：145-153，1985）。然而，具有脂质包膜的较大病毒变异性太大， 它们的结构，并且它们的表面中涉及晶格接触的比例太小， 有序的晶体冷冻电子显微镜现在已经成为推进 结构生物学低温电子显微镜图像的单粒子平均现在经常被 利用直接电子探测设备和技术接近原子分辨率， 补偿由于数据采集期间的随机运动而导致的图像模糊。然而，限制 这项技术在越来越复杂的研究中变得越来越明显。 多形集合体冷冻电子断层扫描现在已经成为一个宝贵的工具， 确定单个病毒颗粒的结构，而不依赖于相同病毒颗粒的可用性， 均匀粒子然而，层析重建的分辨率通常是不够的 来确定原子结构的细节。因此，这一赠款申请的一小部分是专门用于 扩展层析成像图的分辨率。 甲病毒属和风疹病毒属构成披膜病毒科，因为它们在基因上相似 秩序在甲病毒中，基孔肯雅病毒是一个主要的世界健康问题，因为它的再- 出现在亚洲和非洲。我们计划扩展我们以前对二十面体，脂质- 包含甲病毒，并解决更困难的多形性风疹结构分析， 病毒具体来说，我们计划获得各种甲病毒复合时的结构 与宿主受体分子和抗体以及阻断成熟的小分子抑制剂一起， 附着和融合宿主细胞，以研究结构中遇到的中间体， 病毒生命周期这些信息将提供病毒靶点，可用于设计特异性抗病毒药物。 抗病毒治疗。我们还在开发新技术， 单个cryoEM图像，使它们更加相似和均匀，以提高分辨率 多形性生物组装体如风疹病毒的断层图像。

PROJECT SUMMARY Crystallography was the primary structural tool that first opened up structural investigation of viruses at near-atomic resolution. For instance, the first near-atomic resolution structure of an animal virus, namely a common cold virus, was determined by us in the mid-1980s using crystallographic methods (Nature 317:145-153, 1985). However, larger viruses with lipid envelopes are too variable in their structures and have too small a proportion of their surface involved in lattice contacts to make well-ordered crystals. Cryo-electron microscopy has now emerged as the leading tool for advancing structural biology. Single particle averaging of cryo-electron microscopic images is now frequently approaching atomic resolution with the use of direct electron detection devices and technology to compensate for blurring of images due to random motion during data acquisition. However, the limits of this technology are becoming apparent in the investigations of progressively more complex pleomorphic assemblies. Cryo-electron tomography has now emerged as an invaluable tool for the determination of the structure of individual viral particles without reliance on the availability of identical homogeneous particles. However, the resolution of tomographic reconstructions is usually insufficient to determine structures in atomic detail. Thus, a small part of this grant application is dedicated towards extending the resolution of tomographic maps. Alphaviruses and rubella virus constitute the Togavirus family because of their similarity in gene order. Among alphaviruses, Chikungunya virus is a major world health concern because of its re- emergence in Asia and Africa. We plan to extend our previous studies of the icosahedral, lipid- containing alphaviruses, and tackle the more difficult structural analysis of the pleomorphic rubella virus. Specifically, we are planning to obtain the structures of various alphaviruses when complexed with host receptor molecules and with antibodies and small molecule inhibitors that block maturation, attachment and fusion with host cells in order to study the structural intermediates encountered in the viral life cycle. This information will provide viral targets that can be exploited for the design of specific antiviral therapeutics. We are also developing new technology aimed at stretching and pulling individual cryoEM images to make them more similar and homogenous to improve the resolution of tomographic images of pleomorphic biological assemblies such as rubella virus.