MECHANISM OF PROTEIN SYNTHESIS IN MAMMALIAN MITOCHONDRIA

哺乳动物线粒体蛋白质合成机制

• 批准号: 6018592
• 负责人: LINDA L SPREMULLI
• 金额: $ 17.82万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-05-01 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6018592
• 关键词: aminoacyl tRNA; animal genetic material tag; chemical kinetics; chimeric proteins; genetic translation; guanine nucleotides; mitochondria; molecular cloning; nucleic acid sequence; protein biosynthesis; site directed mutagenesis; translation factor

The overall objective of the proposed research is to investigate the mechanism of protein biosynthesis in mammalian mitochondria. The first major focus of our research will be to study the translational elongation factor complex (EF-Tu Tsmt) in detail. EF-TUmt is responsible for promoting the binding of aminoacyl- tRNA to the A-site of the ribosome. It is present in a tight complex with its recycling factor (EF-Tsmt). This complex is tightly associated and, unlike the corresponding complex in Escherichia coli, it is not readily dissociated by either GDP or GTP. The cDNAs for both EF-Tumt and EF-Tsmt have been cloned and expressed in E. Coli. The expressed factors will be used to develop a detailed model for the elongation cycle in mammalian mitochondria. The regions of EF-Tumt and EF-Tsmt that are involved in the interaction between these two proteins will be probed by examining the properties of mutated derivatives of EF-Tumt is able to promote the binding of the structurally unusual mitochondrial aminoacyl-tRNAs to the A-site of the ribosome. In contrast, although E. Coli EF-Tu can form a ternary complex with several mitochondrial aminoacyl-tRNAs, it is unable to promote their binding to the a-site of the ribosome. The regions of EF- Tumt that allow it to use mitochondrial aminoacyl-tRNAs will be examined using chimeric proteins and site-directed mutagenesis. The second major goal of this application is to purify and characterize a new mammalian mitochondrial initiation factor that promotes initiation complex formation in the presence of the mRNA for cytochrome oxidase subunit II. Our long-term goal is to develop an understanding of the mechanism of protein synthesis in mammalian mitochondria and its integration into the complex metabolism of the eukaryotic cell.

拟议研究的总体目的是调查 哺乳动物线粒体中蛋白质生物合成的机制。 我们研究的第一个重点是研究 详细介绍了翻译伸长因子复合物（EF-TU TSMT）。 ef-tumt负责促进氨基环境的结合 tRNA到核糖体的A位点。 它很紧张 复合及其回收因子（EF-TSMT）。 这个复合物是 紧密相关，与相应的复合物不同 大肠杆菌，它不容易被GDP或 GTP。 EF-TUMT和EF-TSMT的cDNA已经 克隆并在大肠杆菌中表达。 表达的因素将是 用于开发延伸周期的详细模型 哺乳动物线粒体。 EF-TUMT和EF-TSMT的区域 这两种蛋白质之间涉及的相互作用将 通过检查突变的衍生物的特性来探测 ef-tumt能够促进结构上不寻常的结合 线粒体氨基酰基 - 核糖体的A位点。 在 对比，尽管大肠杆菌EF-TU可以与 几个线粒体氨基酰基trnas，它无法促进 它们与核糖体的A位点结合。 EF-的地区 允许其使用线粒体氨基酰基-trnas的tumt是 使用嵌合蛋白和定分诱变进行检查。 该应用程序的第二个主要目标是净化和 表征一个新的哺乳动物线粒体起始因子 在存在的情况下促进起始复合物的形成 MRNA用于细胞色素氧化酶亚基II。 我们的长期目标是 为了理解蛋白质合成的机理 在哺乳动物的线粒体及其在复合物中的整合 真核细胞的代谢。