DNP approach for structural determination of large protein-RNA complexes by solid-state NMR

通过固态 NMR 确定大蛋白质-RNA 复合物结构的 DNP 方法

• 批准号: 399211197
• 负责人: Professor Dr.-Ing. Björn Corzilius
• 金额: --
• 依托单位: Institut für Molekulare Biowissenschaften
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/399211197?language=en
• 关键词: DNP; approach; structural; determination; large

In this research project we aim to investigate the feasibility of dynamic nuclear polarization (DNP) aided experiments for structural studies of large protein-RNA complexes by solid-state NMR. Solid-state NMR has been successfully applied for studies of amyloid fibrils, membrane proteins and large protein complexes; first ssNMR studies on protein-RNA complexes have been reported recently. While structural information on small protein-RNA complexes can be obtained in a reasonable time, studies of large complexes are much more time consuming - and often unfeasible - due to lack of sensitivity and severe signal overlap. DNP provides a significant boost in sensitivity for NMR experiments by transferring electron spin polarization of a paramagnetic polarizing agent to nuclear spins. NMR signal enhancement factors typically reach two orders of magnitude. This leads to time savings of the order of tens of thousands and opens the possibility to perform 3D and 4D experiments within few days. The unique collaboration of our research groups, being active in RNA structural biology by MAS NMR as well as DNP-enhanced MAS NMR, will allow us to develop methods for structural studies of large RNP complexes in close-to-native environment at atomic resolution. The methodology developed within this project will find broad applicability in several areas of structural biology.

在本研究项目中，我们旨在研究动态核极化 (DNP) 辅助实验通过固态 NMR 对大蛋白质-RNA 复合物进行结构研究的可行性。固态核磁共振已成功应用于淀粉样原纤维、膜蛋白和大蛋白复合物的研究；最近报道了第一个关于蛋白质-RNA 复合物的 ssNMR 研究。虽然可以在合理的时间内获得小蛋白质-RNA 复合物的结构信息，但由于缺乏灵敏度和严重的信号重叠，对大复合物的研究更加耗时，而且通常不可行。 DNP 通过将顺磁极化剂的电子自旋极化转移到核自旋，显着提高 NMR 实验的灵敏度。 NMR 信号增强因子通常达到两个数量级。这可以节省数万数量级的时间，并为在几天内执行 3D 和 4D 实验提供了可能性。我们的研究小组通过 MAS NMR 和 DNP 增强的 MAS NMR 活跃于 RNA 结构生物学领域，通过独特的合作，我们将能够开发在接近天然环境中以原子分辨率进行大型 RNP 复合物结构研究的方法。该项目开发的方法将在结构生物学的多个领域得到广泛的应用。