MITOCHONDRIAL TRNA MUTATIONS AND HUMAN GENETIC DISEASES

线粒体 TRNA 突变与人类遗传疾病

• 批准号: 2640266
• 负责人: JAMES M BULLARD
• 金额: $ 2.14万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2640266
• 关键词: aminoacid tRNA ligase; genetic disorder; genetic translation; human genetic material tag; lactic acidosis; mitochondria; mitochondrial DNA; molecular cloning; myoclonus epilepsy; stroke; transfer RNA

Mitochondrial tRNA can be used in translation by mitochondrial EF-Tu (EF-Tumt), but not by E. coli EF-Tu. We hypothesize that the conformation of mitochondrial tRNA uniquely inhibits its use by E. coli EF-Tu, but that EF-Tumt alters the shape of the tRNA and is able to present it to the ribosome in the proper conformation. Chimeric tRNAPhe will be prepared in which different regions (acceptor stem/loop, D stem/loop, anticodon stem/loop and TpsiC stem/loop) of mitochondrial tRNAPhe will be replaced with E. coli tRNAPhe. Structural characteristics of the chimeric tRNAs will be compared to mitochondrial and E. coli tRNAPhe. The ability of each chimeric tRNA to function at each stage in translation will be assessed using both EF-Tumt and E. coli EF-Tu, to determine which region(s) of the mitochondrial tRNA inhibits use by E. coli EF-Tu. Physical methods including scanning force microscopy and fluoresence labeling will be used to identify changes in the conformation of tRNA when bound to EF-Tu. Point mutations in human tRNALeu lead to a variety of human genetic diseases. Very little is known about the biochemical processes affected by these mutated tRNAs. Transcripts of human tRNALeu containing these mutations will be prepared and structurally analyzed. The ability of the mutants to function at each step in mitochondrial translation will be analyzed to determine how the mutation may lead to a human genetic disease.

线粒体tRNA可被线粒体EF-Tu用于翻译 （EF-Tumt），而不是E. coli EF-Tu. 我们假设 线粒体tRNA的构象独特地抑制其被E.杆菌 EF-Tu，但是EF-Tumt改变了tRNA的形状， 将其以正确的构象呈现给核糖体。 嵌合tRNAPhe 将制备其中不同区域（受体茎/环，D 茎/环，反密码子茎/环和TpsiC茎/环）的线粒体 tRNAPhe将替换为E。coli tRNAPhe。 结构 将嵌合tRNA的特征与线粒体tRNA的特征进行比较。 和E. coli tRNAPhe。 每种嵌合tRNA的功能， 翻译中的每个阶段将使用EF-Tumt和E. coli EF-Tu，以确定线粒体tRNA的哪个区域（S） 抑制E. coli EF-Tu. 包括扫描在内的物理方法 力显微镜和荧光标记将用于识别 当与EF-Tu结合时，tRNA的构象发生变化。 人类tRNALeu的点突变导致多种人类遗传学改变， 疾病 人们对受影响的生化过程知之甚少 被这些突变的tRNA所控制。 含有这些的人tRNALeu的转录物 将制备突变并进行结构分析。 的能力 在线粒体翻译的每一步起作用的突变体将 分析以确定突变如何导致人类遗传 疾病