IMol Polish Academy of Sciences, Warsaw, Poland.
IMol Polish Academy of Sciences, Warsaw, Poland.
Methods Enzymol. 2024;706:125-158. doi: 10.1016/bs.mie.2024.07.039. Epub 2024 Sep 13.
Mitochondria contain numerous proteins that utilize the chemistry of cysteine residues, which can be reversibly oxidized. These proteins are involved in mitochondrial biogenesis, protection against oxidative stress, metabolism, energy transduction to adenosine triphosphate, signaling and cell death among other functions. Many proteins located in the mitochondrial intermembrane space are imported by the mitochondrial import and assembly pathway the activity of which is based on the reversible oxidation of cysteine residues and oxidative trapping of substrates. Oxidative modifications of cysteine residues are particularly difficult to study because of their labile character. Here we present techniques that allow for monitoring the oxidative state of mitochondrial proteins as well as to investigate the mitochondrial import and assembly pathway. This chapter conveys basic concepts on sample preparation and techniques to monitor the redox state of cysteine residues in mitochondrial proteins as well as the strategies to study mitochondrial import and assembly pathway.
线粒体含有许多利用半胱氨酸残基化学性质的蛋白质，这些残基可以被可逆氧化。这些蛋白质参与线粒体生物发生、抵抗氧化应激、代谢、能量向三磷酸腺苷的转导、信号传递和细胞死亡等功能。许多位于线粒体膜间空间的蛋白质通过线粒体输入和组装途径进行输入，该途径的活性基于半胱氨酸残基的可逆氧化和底物的氧化捕获。由于半胱氨酸残基的不稳定性质，其氧化修饰特别难以研究。在这里，我们介绍了一些可以监测线粒体蛋白质氧化状态以及研究线粒体输入和组装途径的技术。本章介绍了监测线粒体蛋白质中半胱氨酸残基氧化状态的基本概念和技术，以及研究线粒体输入和组装途径的策略。
