PROTEIN IMPORT INTO YEAST MITOCHONDRIA

描述（申请人描述）：本研究的长期目标 建议是了解线粒体内膜蛋白质和 膜间隙介导两个事件：蛋白质易位到膜间隙， 线粒体基质和蛋白质分选到内膜。线粒体 是必不可少的细胞器，其功能需要输入数百个 不同的蛋白质在细胞核中编码，在细胞质中合成， 并输入细胞器。蛋白质输入是一个多步骤的途径， 包括前体蛋白与表面受体的结合， 穿过一个或两个线粒体膜，以及线粒体膜的折叠和组装， 从蛋白质中输入的蛋白质。为了了解这些过程 发生，我们已经分离并正在分析酵母，酵母突变体 酿酒酵母，这是有缺陷的线粒体蛋白输入。 许多研究表明，酵母中的线粒体将蛋白质输入到它们的细胞中。 这一过程与哺乳动物线粒体 进口蛋白质。因此，我们对酵母的研究将产生有价值的信息 所有生物体中线粒体的生物发生。为了支持这一观点，一个人 一种神经退行性疾病 以耳聋、肌张力障碍、智力迟钝和失明为特征， 最近被证明是由线粒体蛋白质输入缺陷引起的。 人类突变是在一个基因编码的蛋白质称为DDP，同源 酵母Tim8蛋白我们预计，我们未来的研究将继续 提供了对线粒体正常功能及其 与人类疾病的关系。 我们最近发现，线粒体内膜含有两个 易位复合物：TIM23复合物，其是易位所需的 前体穿过内膜进入基质和TIM 22复合物， 这是将多位蛋白插入到内 膜的Tim54p、Tim22p和Tim18p包含TIM22的膜亚基 复杂. 在这个建议中，我们提出了几个问题，旨在了解机制 蛋白质被插入内膜。(1)什么其他蛋白质 包括TIM22复合物？(2)Tim18p是否在导入中发挥直接作用？（三） Tim54p在插入过程中是否发挥对接作用？(4)是什么 蛋白质在其输入和插入过程中穿过的精确途径 内膜？(5)内膜蛋白插入能被重构吗 使用纯化的成分？

DESCRIPTION (applicant's description): The long range goal of this research proposal is to understand how proteins of the mitochondrial inner membrane and intermembrane space mediate two events: translocation of proteins into the mitochondrial matrix, and protein sorting to the inner membrane. Mitochondria are essential organelles, whose function requires the import of hundreds of different proteins that are encoded in the nucleus, synthesized in the cytosol and imported into the organelle. Protein import is a multistep pathway which includes the binding of precursor proteins to surface receptors, translocation across one or both mitochondrial membranes, and folding and assembly of the imported protein inside the mitochondrion. To understand how these processes occur, we have isolated and are analyzing mutants in the yeast, Saccharomyces cerevisiae, that are defective in mitochondrial protein import. Many studies indicate that mitochondria in yeast import proteins into their mitochondria by a process virtually identical to the way mammalian mitochondria import proteins. Hence our studies with yeast will yield valuable information about mitochondria biogenesis in all organisms. Supporting this view, a human disease called Mohr-Tranebjaerg syndrome, a neurodegenerative disease characterized by deafness, dystonia , mental retardation and blindness, has recently been shown to result from a defect in mitochondrial protein import. The human mutation is in a gene encoding a protein called DDP, homologous to the yeast Tim8 protein. We anticipate that our future studies will continue to provide insights into the normal functions of mitochondria and their relationships to human disease. We have recently shown that the mitochondrial inner membrane contains two translocation complexes: the TIM23 complex, which is required to translocate precursors across the inner membrane into the matrix and the TIM22 complex, which is required for the insertion of polytopic proteins into the inner membrane. Tim54p, Tim22p and Tim18p comprise the membrane subunits of the TIM22 complex. In this proposal, we ask several questions aimed at understanding the mechanism by which proteins are inserted into the inner membrane. (1) What other proteins comprise the TIM22 complex? (2) Does Tim18p play a direct role in import? (3) Does Tim54p play a docking role in the insertion process? (4) What is the precise pathway that proteins traverse during their import and insertion into the inner membrane? (5) Can inner membrane protein insertion be reconstituted using purified components?