Mechanisms and biological functions of SPOUT methyltransferases

SPOUT甲基转移酶的机制和生物学功能

基本信息

批准号：
10736306
负责人：
Graeme L Conn
金额：
$ 31.01万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-14 至 2027-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10736306
关键词：
Address Adopted Amino Acid Sequence Archaea Aspartic Acid-Specific tRNA Award Biochemical Biological Biological Assay Biological Models Biological Process Biology Cell physiology Cells Chemicals Complement Complex Cryoelectron Microscopy Defect Disease Endocrine Ensure Enzymatic Biochemistry Enzymes Eukaryota Exhibits Family Functional disorder Generations Genetic Genetic Code Genetic Models Health Homologous Gene Human Individual Investigation Knowledge Life Ligation Maintenance Methylation Methyltransferase Modeling Modification Molecular Mutation Neurologic Nucleic Acids Nucleotides Organism Orthologous Gene Pathway interactions Pattern Phenotype Positioning Attribute Process Production Protein Biosynthesis Proteins Purine Nucleotides Quality Control RNA RNA Biochemistry Ribosomes Role S-Adenosylhomocysteine S-Adenosylmethionine Saccharomyces cerevisiae Specificity Structure Substrate Specificity Syndrome System Time Tissues Transfer RNA Translations Yeasts Zebrafish analog base biochemical tools biological adaptation to stress cofactor dimer disease phenotype drug sensitivity fitness genetic approach human disease in vitro Assay in vivo insight interdisciplinary approach loss of function member monomer mutant new therapeutic target posttranscriptional tRNA Methyltransferases yeast genetics

项目摘要

PROJECT SUMMARY/ ABSTRACT Transfer RNAs (tRNAs) are the universal adaptor molecules necessary to convert the nucleic acid-based genetic code into protein sequence during protein synthesis (translation) by the ribosome. This process is universally conserved and fundamental to all life, and, as such, defects in the molecular players of translation, including tRNAs, result in diverse human diseases. Specific chemical modifications such as methylation are common in tRNA, but a detailed understanding of the enzymes that incorporate them and their contributions to tRNA function (and disfunction in disease) have only recently emerged for a few select examples. Since the discovery of the tRNA methyltransferase (Trm10) in Saccharomyces cerevisiae, an accumulating body of evidence, including phenotypes in yeast and a multisymptomatic disease associated with human mutations, has established a significant role for Trm10 in tRNA biology. To better understand the implications of Trm10 modification, the mechanisms by which Trm10 family enzymes specifically recognize and act on their substrate tRNA, and the impact of tRNA modifications on important cellular processes need to be addressed. This project will determine the molecular basis for Trm10 mechanism and function using a multi-disciplinary approach. Genetic, biochemical and molecular enzymology approaches will be combined with structural analyses of enzyme-tRNA complexes using synthetic analogs of the native methyl donor, S-adenosyl-L-methionine, to uniquely identify the role of Trm10 in the maintenance of a high-quality pool of tRNA. A newly developed vertebrate model for Trm10 function will enable investigation of previously challenging questions on Trm10's role in the biological function of multicellular eukaryotes. The studies will be performed in three complementary but independent aims that will: 1) Determine how specific tRNA substrates are selected for modification by yeast and vertebrate Trm10 enzymes using structural, biochemical and genetic approaches; 2) Assess the molecular basis for and biological significance of the uniquely conserved vertebrate m1A9 modification exploiting a new vertebrate model for Trm10 function, and 3) Identify tRNA-specific functions for G9 modification in yeast and zebrafish using complementary genetic approaches in both model species. Collectively, the proposed studies will advance the fields of enzymology, RNA biochemistry, and tRNA biology by providing mechanistic and biological insight into a tRNA modification enzyme that is universally conserved among eukaryotes and is critically important for human health, yet whose molecular mechanism and biological functions are not at all understood. These results will also provide new insight into the dynamic landscape of tRNA modifications in multicellular eukaryotes.

项目摘要/摘要转移RNA（TRNA）是转化基于核酸的遗传所需的通用衔接子分子核糖体蛋白质合成期间的蛋白质序列代码。这个过程是普遍的对所有生活的保守和基础 TRNA，导致各种人类疾病。特定的化学修饰（例如甲基化）在 tRNA，但对将其纳入酶及其对tRNA功能的贡献的酶有详细的理解（疾病中的失功）直到最近才出现了一些精选的例子。由于发现酿酒酵母中的tRNA甲基转移酶（TRM10），一种积累的证据，包括酵母中的表型和与人类突变相关的多肿瘤疾病已经建立了 TRM10在TRNA生物学中的重要作用。为了更好地了解TRM10修改的含义 TRM10家族酶专门识别并作用于其底物tRNA的机制，以及需要解决tRNA修饰对重要细胞过程的影响。这个项目将确定使用多学科方法的TRM10机制和功能的分子基础。遗传，生化和分子酶学方法将与酶-TRNA复合物的结构分析相结合使用天然甲基供体S-腺苷-L-甲硫氨酸的合成类似物来唯一识别 TRM10在维持高质量的tRNA池中。 TRM10功能的新开发的脊椎动物模型将对TRM10在TRM10在生物学功能中的作用进行调查多细胞真核生物。这些研究将以三个互补但独立的目的进行： 1）确定如何选择特定的TRNA底物进行酵母和脊椎动物TRM10酶修饰使用结构，生化和遗传方法； 2）评估分子基础和生物学独特保守的脊椎动物M1A9修饰的意义，利用了TRM10的新脊椎动物模型 3）使用互补的酵母和斑马鱼中的G9修饰的tRNA特异性函数两种模型物种的遗传方法。拟议的研究集体将推进酶学，RNA生物化学和tRNA生物学通过提供机械和生物学洞察力在真核生物中普遍保守的修饰酶，对人类健康至关重要，然而，根本不了解其分子机制和生物学功能。这些结果也将提供对多细胞真核生物中tRNA修饰的动态景观的新见解。

项目成果

期刊论文数量（8）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Tied up in knots: Untangling substrate recognition by the SPOUT methyltransferases.

DOI：
10.1016/j.jbc.2022.102393
发表时间：
2022-10
期刊：
JOURNAL OF BIOLOGICAL CHEMISTRY
影响因子：
4.8
作者：
Strassler, Sarah E.;Bowles, Isobel E.;Dey, Debayan;Jackman, Jane E.;Conn, Graeme L.
通讯作者：
Conn, Graeme L.

Insights into Catalytic and tRNA Recognition Mechanism of the Dual-Specific tRNA Methyltransferase from Thermococcus kodakarensis.

深入了解柯达热球菌双特异性 tRNA 甲基转移酶的催化和 tRNA 识别机制。