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首页> 外文期刊>Molecular and Cellular Biology >Proteomic Analysis of Mitochondrial Protein Turnover: Identification of Novel Substrate Proteins of the Matrix Protease Pim1
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Proteomic Analysis of Mitochondrial Protein Turnover: Identification of Novel Substrate Proteins of the Matrix Protease Pim1

机译：线粒体蛋白质转换的蛋白质组学分析：基质蛋白酶Pim1的新型底物蛋白的鉴定。

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ATP-dependent oligomeric proteases are major components of cellular protein quality control systems. To investigate the role of proteolytic processes in the maintenance of mitochondrial functions, we analyzed the dynamic behavior of the mitochondrial proteome of Saccharomyces cerevisiae by two-dimensional (2D) polyacrylamide gel electrophoresis. By a characterization of the influence of temperature on protein turnover in isolated mitochondria, we were able to define four groups of proteins showing a differential susceptibility to proteolysis. The protein Pim1/LON has been shown to be the main protease in the mitochondrial matrix responsible for the removal of damaged or nonnative proteins. To assess the substrate range of Pim1 under in vivo conditions, we performed a quantitative comparison of the 2D protein spot patterns between wild-type and pim1Δ mitochondria. We were able to identify a novel subset of mitochondrial proteins that are putative endogenous substrates of Pim1. Using an in organello degradation assay, we confirmed the Pim1-specific, ATP-dependent proteolysis of the newly identified substrate proteins. We could demonstrate that the functional integrity of the Pim1 substrate proteins, in particular, the presence of intact prosthetic groups, had a major influence on the susceptibility to proteolysis.
机译：ATP依赖性寡聚蛋白酶是细胞蛋白质质量控​​制系统的主要组成部分。为了研究蛋白水解过程在维持线粒体功能中的作用，我们通过二维（2D）聚丙烯酰胺凝胶电泳分析了酿酒酵母（Saccharomyces cerevisiae）线粒体蛋白质组的动力学行为。通过表征温度对分离的线粒体中蛋白质更新的影响，我们能够定义四组蛋白质，这些蛋白质对蛋白水解表现出不同的敏感性。 Pim1 / LON蛋白已被证明是线粒体基质中负责去除受损或非天然蛋白的主要蛋白酶。为了评估在体内条件下Pim1的底物范围，我们对野生型和 pim1 Δ线粒体之间的二维蛋白质斑点模式进行了定量比较。我们能够确定线粒体蛋白的一个新的子集，它们是Pim1的假定内源性底物。使用内脏降解试验，我们确认了新鉴定的底物蛋白的Pim1特异性，ATP依赖性蛋白水解。我们可以证明，Pim1底物蛋白的功能完整性，特别是完整的修复基团的存在，对蛋白水解的敏感性具有重大影响。

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