EUKARYOTIC TRNA MATURATION

真核生物的基因表达涉及一系列复杂的RNA成熟过程 事件。这些反应的大部分方面在 与原核生物中类似的过程相比，因为 相对缺乏复杂的遗传方法，也缺乏 用来研究酶反应的前体RNA。最近一段时间 利用高效体外转录系统进行克隆的转录 TRNA基因使合成野生型和 选择性诱变的tRNA初级转录本。这些RNA充当 体外各种tRNA加工反应的底物，包括 5‘末端的核溶解成熟。已经引起了相当大的关注 重点放在原核生物的最后一个反应上，因为催化 似乎主要是由酶的RNA成分(核糖核酸酶)执行的 p)。 在酿酒酵母中，这个反应是由内切酶催化的 这似乎也是一个与RNA络合的蛋白质部分。这 活性将从酵母核中提纯，酶组分(S) 对任何RNA部分进行物理表征并测定其序列。在……里面 建立识别和切割机制，结构性的 底物和酶RNA的分析将在#年进行。 结合对所选突变体的底物利用研究 前tRNA底物。这些结果将与 类似的原核生物和线粒体反应，以建立一个 RNA和核糖核蛋白结构参与的机制 在核溶解催化中。

Gene expression in eukaryotes involves a complex series of RNA maturation events. Most aspects of these reactions are poorly understood in comparison to the analogous processes in prokaryotes because of the relative lack of sophisticated genetic approaches and a paucity of precursor RNAs with which to study the enzymatic reactions. Recently the use of highly efficient in vitro systems for the transcription of cloned tRNA genes has made it possible to synthesize both wild type and selectively mutagenized tRNA primary transcripts. These RNAs act as substrates for a variety of tRNA processing reactions in vitro, including nucleolytic maturation of the 5' terminus. Considerable attention has been focused on this last reaction in prokaryotes due to the fact that catalysis appears to be performed primarily by the RNA component of the enzyme (RNase P). In Saccharomyces cerevisiae this reaction is catalyzed by an endonuclease which also appears to be a protein moiety complexed with RNA. This activity will be purified from yeast nuclei, the enzymatic component(s) physically characterized and the sequence of any RNA moiety determined. In order to establish mechanisms for recognition and cleavage, structural analysis of the substrate and enzymic RNAs will be undertaken in conjunction with substrate utilization studies of selected mutants in the pre-tRNA substrates. These results will be compared with models for the analogous prokaryotic and mitochondrial reactions in order to establish a mechanism for the involvement of the RNA and ribonucleoprotein structures in nucleolytic catalysis.