tRNA Processing

tRNA处理

• 批准号: 7153482
• 负责人: Eric M. Phizicky
• 金额: $ 28.37万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-05-01 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7153482
• 关键词: Accounting; Address; Binding; Biochemical; Biochemical Genetics; Biochemistry; Bioinformatics; Biological; Biology; Catalysis; Cells; Class; Classification; Environment; Enzymes; Eukaryota; Eukaryotic Cell; Failure; Family; Generations; Genes; Genetic; Genomics; Goals; Homologous Gene; Human; Left; Methods; Methyltransferase; Modification; Numbers; Organism; Pathway interactions; Phenotype; Polynucleotides; Positioning Attribute; Process; Proteins; Reaction; Ribosomal RNA; Role; Site; Specificity; Substrate Specificity; Time; Transfer RNA; Work; Yeasts; base; chemical reaction; guanylyltransferase; in vivo; insight; interest; mutant; novel; tRNA Methyltransferases; tRNA guanylyltransferase

The long term goal of this project is to define all the biochemical steps of tRNA processing and to examine their roles in the cell. Despite more than 40 years of work on tRNA, we still do not have a complete picture of this pathway. Current work is focused on tRNA modifications, which comprise the bulk of unknown biochemical steps. The number of tRNA molecules in the yeast cell dwarfs that of any other class of polynucleotide, accounting for 10 times more molecules than rRNA. There are a total of 22 different modifications in yeast tRNAs, catalyzed by multiple gene products. Many of these modifications are highly conserved, biochemically uncharacterized, unassigned to genes, and of unknown role. Recent efforts in the field have led to the identification of a number of gene products involved in modification, using a combination of genetic, biochemical, and bioinformatic methods. Use of a biochemical genomics approach led to the identification of five gene products involved in four yeast modification activities. This has led to the beginnings of an understanding of the specificity, mechanism and function of many of these gene products, but leaves many unanswered questions. This work on tRNA processing will be continued by studying novel and interesting aspects of several tRNA modification enzymes, and by identifying new genes encoding modification activities. Four specific aims are proposed. First, the biochemistry and biology of tRNA His guanylyltransferase will be studied. tRNA His species from all studied organisms are different from other tRNAs in having an extra G residue at the -1 position, which is added post-transcriptionally in eukaryotes. Two gene products that co-purify with tRNA His guanylyltransferase activity have been identified; one is essential and required for activity in vivo. The identities and activities of components of tRNA His guanylyltransferase will be determined, as well as the basis for tRNA recognition by the enzyme. Second, m1G9 methyltransferase will be examined to determine important catalytic and binding residues, because this enzyme is different from other known enzymes of this type. Third, several other modification enzymes and their corresponding genes will be identified. Some 17 modification enzymes have not yet been assigned to genes. Many of these modifications are biochemically interesting and are located in important regions of the tRNA. Finally, the function of some tRNA modification enzymes will be studied in vivo.

该项目的长期目标是确定tRNA加工的所有生化步骤，并检查它们在细胞中的作用。尽管对tRNA进行了40多年的研究，我们仍然没有对这一途径有一个完整的了解。目前的工作集中在tRNA修饰上，它包含了大部分未知的生化步骤。酵母细胞中tRNA分子的数量比其他任何种类的多核苷酸都要多，比rRNA分子多10倍。酵母trna共有22种不同的修饰，由多个基因产物催化。许多这些修饰是高度保守的，生物化学上没有特征，没有分配给基因，也没有未知的作用。最近在该领域的努力已经导致鉴定了一些基因产物参与修饰，使用遗传，生化和生物信息学的方法相结合。利用生物化学基因组学方法鉴定了涉及酵母四种修饰活动的五个基因产物。这使得人们开始了解这些基因产物的特异性、机制和功能，但也留下了许多未解之谜。在tRNA加工方面的工作将通过研究一些tRNA修饰酶的新颖和有趣的方面，并通过识别编码修饰活性的新基因来继续。提出了四个具体目标。首先，对tRNA - His鸟苷基转移酶的生物化学和生物学进行研究。与其他tRNA不同的是，在-1位置有一个额外的G残基，这是真核生物转录后添加的。两个与tRNA共纯化的基因产物已被确定；一种是体内活动所必需的。将确定tRNA - His鸟苷转移酶组分的特性和活性，以及该酶识别tRNA的基础。其次，将检测m1G9甲基转移酶，以确定重要的催化和结合残基，因为这种酶不同于其他已知的这种类型的酶。第三，鉴定其他几种修饰酶及其对应的基因。大约有17种修饰酶尚未被分配到基因上。许多这些修饰在生物化学上是有趣的，并且位于tRNA的重要区域。最后，对一些tRNA修饰酶的体内功能进行研究。