STRUCTURE AND FUNCTION OF NUCLEIC ACIDS

真核基因包含内含子，这些内含子中断了 遗传信息。内含子被从该基因的RNA转录本中移除 通过RNA剪接获得基因。本项目继续调查 酵母中两种RNA剪接的机制：tRNA和mRNA. TRNA前体在两步反应中被拼接。一种核酸内切酶 去除内含子和需要三磷酸腺苷的连接酶，将外显子连接在一起。 核酸内切酶将被提纯至均一。这一机制通过它 它识别和切割前体将被研究。连接酶有 已经被提纯到同质。进一步的研究将探索其 结构域结构和连接酶反应的机理。关于黄曲霉的研究 人工合成的tRNA-Phe前体将阐明次级和 底物识别中成熟结构域的三级结构。 信使核糖核酸前体被剪接在一个叫做剪接体的大颗粒上。 酵母突变株被分离出来，称为rna 2-11，它定义了 剪接体。RNA基因产物将使用体外纯化方法进行纯化 互补试验。这些产品一旦提纯，就可以用来 定义和表征其他的RNA和蛋白质组分 剪接体。剪接体将被提纯并鉴定为 电子显微镜和生化分级。它所经过的路径 是组装的还是拆卸的将被确定。一个可能的角色是 还将研究mRNA剪接中的hnRNPs。

Eukaryotic genes contain introns which interrupt the continuity of the genetic information. Introns are removed from an RNA transcript of the gene by RNA splicing. This project continues an investigation of the mechanism of splicing of two kinds of RNA splicing in yeast: tRNA and mRNA. tRNA precursors are spliced in a two-step reaction. An endonuclease removes the intron and a ligase, requiring ATP, joins the exons together. The endonuclease will be purified to homogeneity. The mechanism by which it recognizes and cleaves the precursor will be studied. The ligase has already been purified to homogeneity. Further studies will explore its domain structure and the mechanism of the ligase reaction. Studies on a synthetic tRNA-Phe precursor will elucidate the role of secondary and tertiary structure of the mature domain in substrate recognition. mRNA precursors are spliced on a large particle called the spliceosome. Yeast mutants have been isolated called rna2-11 which define components of the spliceosome. The RNA gene products will be purified using an in vitro complementation assay. These products, once purified, can be used to define and characterize other RNA and protein components of the spliceosome. The spliceosome will be purified and characterized by electron microscopy and biochemical fractionation. The pathway by which it is assembled and disassembled will be determined. A possible role of hnRNPs in mRNA splicing will also be investigated.