IIBR RoL: Applying innovative structural tools to highlight RNA's structural dynamics as RNA-protein complexes self-assemble

IIBR RoL：应用创新的结构工具来突出 RNA-蛋白质复合物自组装时的结构动力学

• 批准号: 1930046
• 负责人: Lois Pollack
• 金额: $ 77.32万
• 依托单位: Cornell University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1930046&HistoricalAwards=false
• 关键词: IIBR; RoL; Applying; innovative; structural

Life functions are supported by the interaction of large biological molecules, including proteins and RNA. Although the shapes or structures of some of these complexes have been visualized, very little is known about their dynamics of assembly. How do these differing components interact to create complexes whose function exceeds that of either alone? This is a challenging problem because of the intertwined motions of the components. Through this project, new tools will be applied to highlight the motions of just one macromolecular species during the assembly of a complex. Although both proteins and RNA are present, only the RNA is visible. Students will design, build and apply the new technology to study the self-assembly of a small plant virus, an intricate structure consisting of a protein shell surrounding a nucleic acid core. These studies will allow us to more clearly define the rules that govern the joining of nucleic acids and proteins into biological complexes. Many important biological assembles are comprised of distinct macromolecular components, such as proteins in conjunction with the nucleic acids. Despite their importance, less than 2% of the static structures in the protein data bank reflect RNA-protein complexes. Even less is known about the dynamic interactions between RNA and protein components. Although much effort has been expended in watching one type macromolecule self-assemble, e.g. protein folding, it is much more difficult to interpret dynamic structural information for multicomponent machines because multiple species are present. The inability to separate the signal from the distinct components precludes a simple interpretation. This project will exploit and innovate existing experimental infrastructure to create tools that highlight the signal of only one species on the background of the other, vastly simplifying measurements of complexes' dynamics. Mixers that rapidly combine the protein and RNA precursors will be constructed and will allow experimenters to track the changing structures of only the RNA component during complex assembly. A small plant virus will serve as a model system for these studies. The development and application of two complementary techniques (small angle x-ray scattering and single molecule fluorescence) will offer a unique perspective on this self-assembly problem. Once established, these methods can be applied other RNA-protein complexes, gathering data to elucidate the rules governing their assembly. Project results, sorted by topic, will be posted at: https://pollack.research.engineering.cornell.edu/all_publications/. This project is jointly funded by the Division of Biological Infrastructure program for Infrastructure Innovation for Biological Research, the Molecular Biophysics and Genetic Mechanism Clusters in the Division of Molecular and Cellular Biosciences, and the Rules of Life initiative in the Emerging Frontiers office of the Biology Directorate.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

生命功能得到了大型生物分子的相互作用，包括蛋白质和RNA。 尽管其中一些复合物的形状或结构已被可视化，但对它们的组装动态知之甚少。 这些不同的组件如何相互作用以创建其功能超过任何一个功能的复合物？这是一个具有挑战性的问题，因为组件的相互交织动作。通过该项目，将应用新工具来突出一个复合物组装过程中仅一个大分子物种的运动。 尽管存在蛋白质和RNA，但仅可见RNA。 学生将设计，构建和应用新技术来研究小型植物病毒的自组装，这是一种复杂的结构，该结构由围绕核酸核心的蛋白质壳组成。这些研究将使我们更清楚地定义了控制核酸和蛋白质加入生物复合物的规则。许多重要的生物组装由不同的大分子成分组成，例如与核酸结合的蛋白质。尽管它们的重要性，但蛋白质数据库中不到2％的静态结构反映了RNA蛋白质复合物。关于RNA和蛋白质成分之间的动态相互作用的知之甚少。尽管在观看一种大分子自组装时已经花费了很多努力，例如蛋白质折叠，由于存在多种物种，因此很难解释多组分机器的动态结构信息。 无法将信号与不同组件分开的简单解释。 该项目将利用和创新现有的实验基础架构，以创建工具，以突出另一个物种在另一个物种的信号，从而大大简化了复合物动力学的测量。 将构建快速结合蛋白质和RNA前体的混合器，并允许实验者在复杂组装过程中仅跟踪仅RNA成分的结构。 小型植物病毒将作为这些研究的模型系统。两种互补技术（小角度X射线散射和单分子荧光）的开发和应用将在此自组装问题上提供独特的视角。建立后，可以应用其他RNA-蛋白质复合物，收集数据以阐明管理其组装的规则。 通过主题排序的项目结果将发布在以下位置：https：//pollack.research.engineering.cornell.edu/all_publications/。该项目由生物学基础设施创新生物基础设施计划共同资助，用于生物学研究，分子生物物理学和遗传机制在分子和细胞生物科学的部门中的分子和遗传机制群，以及生物学领域新兴领域办公室的生命规则规则该奖项反映了NSF的法定任务，并通过使用基金会的智力优点和更广泛的影响审查标准来评估值得支持。

项目成果

RNA structures and dynamics with Å resolution revealed by x-ray free-electron lasers.

• DOI: 10.1126/sciadv.adj3509
• 发表时间: 2023-09-29
• 期刊: SCIENCE ADVANCES
• 影响因子: 13.6
• 作者: Zielinski, Kara A.;Sui, Shuo;Pabit, Suzette A.;Rivera, Daniel A.;Wang, Tong;Hu, Qingyue;Kashipathy, Maithri M.;Lisova, Stella;Schaffer, Chris B.;Mariani, Valerio;Hunter, Mark S.;Kupitz, Christopher;Moss III, Frank R.;Poitevin, Frederic P.;Grant, Thomas D.;Pollack, Lois
• 通讯作者: Pollack, Lois

Visualizing a viral genome with contrast variation small angle X-ray scattering

通过小角度 X 射线散射对比变化可视化病毒基因组

• DOI: 10.1074/jbc.ra120.013961
• 发表时间: 2020
• 期刊: Journal of Biological Chemistry
• 影响因子: 4.8
• 作者: San Emeterio, Josue;Pollack, Lois
• 通讯作者: Pollack, Lois