Single Molecule FRET Study of Ribosome Translocation

核糖体易位的单分子 FRET 研究

• 批准号: 7158670
• 负责人: YUHONG WANG
• 金额: $ 5.04万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-12-01 至 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7158670
• 关键词: binding proteins; biotin; conformation; fluorescence microscopy; fluorescence resonance energy transfer; fluorescent dye /probe; postdoctoral investigator; protein biosynthesis; protein structure; protein transport; ribosomal proteins; ribosomes; technology /technique development; transfer RNA

DESCRIPTION (provided by applicant): A. Using single molecule fluorescence resonance energy transfer (FRET) technology coupled with total internal reflection fluorescence microscopy (TIRFM) to directly probe the relative structure change between ribosome L11 protein and EFG protein during translocation as a function of time using Cy3-Cy5 as the donor-acceptor pair. The first goal is to directly probe the relative structure change between ribosome L11 protein and EFG protein during translocation as a function of time. The information is both structural and dynamic under a biological relative condition. B. we will study the translocation more directly by monitoring FRET between L11 and tRNAs. Combining this and the L11 -EFG time dependent FRET changes, we will reveal the order or coupling of the EFG conformational change and the actual tRNA movements. Coupling of EFG conformation change and the tRNAs movement will be a strong indication that EFG is a motor protein, which remains controversial. C. We will extend our research thrust to processes involving other G proteins, such as IF2 and EF-Tu. We are also interested in study conformation changes of other ribosome locations, for example L1 protein.

描述（申请人提供）：A.使用单分子荧光共振能量转移（FRET）技术结合全内反射荧光显微镜（TIRFM），使用Cy3-Cy5作为供体-受体对，直接探测易位过程中核糖体L11蛋白和EFG蛋白之间的相对结构变化作为时间的函数。第一个目标是直接探测易位过程中核糖体 L11 蛋白和 EFG 蛋白之间的相对结构变化作为时间的函数。在生物相对条件下，信息既是结构性的又是动态的。 B. 我们将通过监测 L11 和 tRNA 之间的 FRET 更直接地研究易位。结合此和 L11 -EFG 时间依赖性 FRET 变化，我们将揭示 EFG 构象变化和实际 tRNA 运动的顺序或耦合。 EFG 构象变化和 tRNA 运动的耦合将有力地表明 EFG 是一种运动蛋白，但这仍然存在争议。 C. 我们将把我们的研究重点扩展到涉及其他 G 蛋白的过程，例如 IF2 和 EF-Tu。我们还对研究其他核糖体位置的构象变化感兴趣，例如 L1 蛋白。