STRUCTURE/FUNCTION OF TRANSFER RIBONUCLEIC ACIDS

转移核糖核酸的结构/功能

• 批准号: 6385008
• 负责人: UTTAM L RAJBHANDARY
• 金额: $ 49.05万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-06-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6385008
• 关键词: X ray crystallography; chemical structure function; gene mutation; genetic translation; hydroxymethyltransferases; nuclear magnetic resonance spectroscopy; site directed mutagenesis; translation factor

DESCRIPTION (Adapted from applicant's abstract): The objectives of this proposal are to understand: (1) the relationship between the structure and the many distinctive properties of E. coli initiator tRNA and (2) the molecular mechanisms underlying its specific interactions with components of the translational machinery. An important question is the molecular basis of the highly specific recognition of the tRNA by Met-tRNA transformylase. A combination of structural, biochemical, and genetic approaches will be used to study this. These include (I) NMR spectroscopy to analyze the structure of the tRNA substrate, (ii) crystallography to determine the structure of the protein and the tRNA-protein complex, (iii) investigation of the topology of interaction of the protein with the tRNA using crosslinking experiments, protection experiments and by examining suppressor mutations in the protein that compensate for defects in formylation of mutant tRNAs, and (iv) site-specific mutagenesis to identify amino acids important in the protein for tRNA selection and function. Similar approaches will be used to study interactions of tRNA with other proteins, in particular, in vivo selection and analysis of suppressor mutations in IF2, IF3, or other chromosomal genes. Such work could provide information on interactions between the initiation factors and the initiator tRNA and could lead to identification of new genes involved in translational initiation. Work on identification of intermediates in translation initiation in vivo will continue. Questions of specific interest are: (I) is IF2 a carrier of fMet-tRNA to the ribosome? (ii) Does the 30S ribosome bind first to the initiator tRNA and then to the mRNA or vice versa? (iii) Are the requirements in an initiator tRNA for translational reinitiation the same as for de novo initiation? Finally, the role of the All:U24 base pair unique to eubacterial initiator tRNAs in initiation will be studied along with questions of why introduction of the base pair to an E. coli elongator methionine tRNA prevents accumulation of the tRNA in vivo.

描述（改编自申请人的摘要）：本项目的目标 建议要理解：（1）结构和之间的关系 大肠杆菌起始子 tRNA 的许多独特特性以及 (2) 其与组分的特定相互作用的分子机制 翻译机器。 一个重要的问题是高度特异性的分子基础 Met-tRNA 转化酶对 tRNA 的识别。 的组合 将使用结构、生物化学和遗传学方法来研究这一点。 其中包括 (I) NMR 光谱分析 tRNA 的结构 底物，(ii) 晶体学以确定蛋白质的结构 和 tRNA-蛋白质复合物，(iii) 拓扑结构的研究 使用交联实验研究蛋白质与 tRNA 的相互作用， 保护实验并通过检查蛋白质中的抑制突变 补偿突变 tRNA 的甲酰化缺陷，以及 (iv) 通过位点特异性突变来鉴定蛋白质中重要的氨基酸 用于 tRNA 选择和功能。 类似的方法将用于研究 tRNA 与其他物质的相互作用 蛋白质，特别是抑制因子的体内选择和分析 IF2、IF3 或其他染色体基因突变。 这样的工作可以提供 有关引发因素和引发剂之间相互作用的信息 tRNA 并可能导致鉴定参与翻译的新基因 引发。 翻译中间体识别工作 体内引发将继续。 特别感兴趣的问题是：（一） IF2 是核糖体 fMet-tRNA 的载体吗？ (ii) 30S 核糖体 首先与起始 tRNA 结合，然后与 mRNA 结合，或者反之亦然？ (三) 启动子 tRNA 中翻译重新启动的要求是否满足 与从头启动相同吗？ 最后来说一下All:U24碱基对的作用 真细菌启动子 tRNA 在启动过程中所特有的特性将被研究 问题是为什么要将碱基对引入大肠杆菌延伸器 蛋氨酸 tRNA 可防止 tRNA 在体内积累。