STRUCTURE AND FUNCTION OF EUKARYOTIC INITIATOR TRNA

本提案的长期目标是了解 真核启动子tRNAS的结构和功能之间的关系，包括 其分子机制的组成部分的具体识别 蛋白质合成机制因为他们在启蒙中的独特作用 蛋白质生物合成的起始剂tRNAS从原核和真核 来源具有生物化学特性，这些特性不同于 非起始剂tRNAS。 这些包括（i）特异性结合起始位点， 因子eIF-2，（ii）直接结合核糖体上的P位点，和（iii） 排除与核糖体上的A位点结合。 沿着这些 特定的性质，引发剂tRNAS具有独特的序列和/或 在非起始剂tRNAS中未发现的结构特征。 的 本建议的直接目标是了解 这些特征与真核生物起始物的上述性质之间的关系 tRNAS。 所采用的方法包括分离和分析 突变体人tRNAS的性质。 位点特异性突变将用于 产生不同类型的突变体tRNA：（a）来自 已经失去一个、两个或所有三个特征的起始tRNA 启动子tRNA所特有的，和（B）来源于非启动子tRNA的那些 甲硫氨酸tRNA现在具有一个，两个或所有三个结构， 启动子tRNA的独特功能。 的功能和详细性质 将在酵母S. 在酿酒酵母和哺乳动物细胞中以及在体外。

The long range objective of this proposal is to understand the relationship between structure and function of eukaryotic initiator tRNAS, including the molecular mechanisms underlying their specific recognition by components of the protein synthesis machinery. Because of their unique role in initiation of protein biosynthesis, initiator tRNAS from prokaryotic and eukaryotic sources possess biochemical properties which are distinct from those of non-initiator tRNAS. These include (i) specific binding to the initiation factor eIF-2, (ii) direct binding to the P site on the ribosome and (iii) exclusion from binding to the A site on the ribosome. Along with these specific properties, initiator tRNAS possess unique sequence and/or structural features which are not found in non-initiator tRNAS. The immediate objective of this proposal is to understand the relationship between these features and the above properties of eukaryotic initiator tRNAS. The approach to be used involves isolation and analysis of properties of mutant human tRNAS. Site specific mutations will be used to generate different classes of mutant tRNAs: (a) those derived from initiator tRNAs which have lost one, two or all three of the features unique to initiator tRNAs and (b) those derived from the non-initiator methionine tRNA which now possess one, two or all three of the structural features unique to initiator tRNA. The function and detailed properties of these and other mutant tRNAs will be studied in vivo, in the yeast S. cerevisiae and in mammalian cells, and in vitro.