STRUCTURE AND FUNCTION OF EUKARYOTIC INITIATOR TRNA

这项建议的长期目标是理解 真核启动子tRNAs的结构和功能之间的关系，包括 其成分对其特异性识别的分子机制 蛋白质合成机器。因为他们在入会过程中的独特作用 蛋白质生物合成的启动子原核生物和真核生物的tRNAs 来源具有与那些不同的生化特性 非启动子tRNA。这些包括(I)与起始物的特定结合 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。再加上这些 特异性，启动子tRNA具有独特的序列和/或 非启动子tRNA中没有的结构特征。这个 这项建议的直接目标是了解两国关系 这些特性与真核生物启动子的上述特性之间的关系 TRNAs。要使用的方法包括分离和分析 突变的人tRNA的性质。特定部位的突变将被用于 产生不同类别的突变tRNA：(A)源自 失去一个、两个或全部三个功能的启动器tRNA 启动子tRNA所特有的；和(B)源自非启动子的那些 蛋氨酸tRNA现在拥有一个、两个或全部三个结构 启动子tRNA所特有的功能。的功能和详细属性 这些和其他突变的tRNAs将在体内进行研究，在酵母S. 在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.