Biophysical Studies on the Additive Effects of Modification in tRNA and rRNA

tRNA 和 rRNA 修饰相加效应的生物物理学研究

• 批准号: 0078501
• 负责人: Edward Nikonowicz
• 金额: $ 45.24万
• 依托单位: William Marsh Rice University
• 依托单位国家: 美国
• 项目类别: Continuing grant
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0078501&HistoricalAwards=false
• 关键词: Biophysical; Studies; Additive; Effects; Modification

Nikonowicz, Edward P.MCB 0078501The goal of this project is to understand key aspects of the post-transcriptional modification process of tRNA at the molecular level, including the basis of molecule specific RNA recognition by modification enzymes and the functional basis of modifications within RNA molecules. This research focuses on modification sites within the anticodon stem-loop E. coli tRNAPhe and the rRNA target of the dual specificity modification enzyme pseudouridine synthase RluA. Specifically, the following topics will be addressed: (1) chemical and enzymatic protection of MiaA RNA ligands to identify positions on the RNA that contact MiaA protein and to monitor binding induced RNA structure changes, (2) structural studies of an RNA ligand in complex with MiaA, and (3) structure determination of Helix-35 from E. coli 23S rRNA, the "second" target of the RluA enzyme. The results of these studies are directly relevant to understanding the specificity and affinity of protein-RNA interactions and the impact of modification on RNA structure and dynamics and its relationship to RNA function (e.g. translation). These investigations also will complement mechanistic studies of the prenylation reaction catalyzed by MiaA. Many RNA molecules interact with specific protein molecules either to be modified by those proteins or to perform unique cellular functions as a protein-RNA complex. Thus, it is of fundamental biological importance to learn the molecular principles that encode the specificity of individual protein-RNA interactions. At present, the molecular details of protein-RNA recognition are known for only a small number of complexes and even fewer general principles for recognition have emerged. The overall goal of this research is to determine the rules of protein-RNA recognition for two model bacterial proteins and to determine the structural and stability effects introduced by their enzymatic action.

Nikonowicz, Edward P.MCB 0078501该项目的目标是在分子水平上了解 tRNA 转录后修饰过程的关键方面，包括修饰酶识别分子特异性 RNA 的基础以及 RNA 分子内修饰的功能基础。本研究重点关注反密码子茎环大肠杆菌 tRNAPhe 内的修饰位点和双特异性修饰酶假尿苷合酶 RluA 的 rRNA 靶标。 具体来说，将讨论以下主题：(1) MiaA RNA 配体的化学和酶保护，以确定 RNA 上接触 MiaA 蛋白的位置并监测结合诱导的 RNA 结构变化，(2) 复合物中 RNA 配体的结构研究(3) 确定来自大肠杆菌 23S rRNA（RluA 酶的“第二个”靶标）的 Helix-35 的结构。这些研究的结果与理解蛋白质-RNA 相互作用的特异性和亲和力、修饰对 RNA 结构和动力学的影响及其与 RNA 功能（例如翻译）的关系直接相关。这些研究还将补充 MiaA 催化的异戊二烯化反应的机制研究。许多 RNA 分子与特定的蛋白质分子相互作用，要么被这些蛋白质修饰，要么作为蛋白质-RNA 复合物发挥独特的细胞功能。因此，了解编码个体蛋白质-RNA 相互作用特异性的分子原理具有重要的生物学基础。目前，仅了解少量复合物的蛋白质-RNA 识别的分子细节，并且出现的识别通用原理甚至更少。 本研究的总体目标是确定两种模型细菌蛋白质的蛋白质-RNA 识别规则，并确定它们的酶促作用引入的结构和稳定性效应。