Quirós Pedro M
Centro de Investigación Biomédica en Red de Cáncer (CIBERONC); Departamento de Bioquímica y Biología Molecular, Universidad de Oviedo, Oviedo, Spain.
Methods Mol Biol. 2018;1731:49-56. doi: 10.1007/978-1-4939-7595-2_5.
Mitochondrial aconitase is a reversible enzyme that catalyzes the conversion of citrate to isocitrate in the tricarboxylic acid cycle. Mitochondrial aconitase is very sensitive to oxidative inactivation and can aggregate and accumulate in the mitochondrial matrix causing mitochondrial dysfunction. Lon protease, one of the major quality control proteases in mitochondria, degrades oxidized aconitase maintaining mitochondrial homeostasis. This chapter describes a step-by-step protocol for a simple and reliable measurement of mitochondrial aconitase, as well as citrate synthase activity, using isolated mitochondria from cells. The protocol is simple and fast, and it is optimized for a 96-well plate using a microplate reader.
线粒体乌头酸酶是一种可逆酶，它在三羧酸循环中催化柠檬酸转化为异柠檬酸。线粒体乌头酸酶对氧化失活非常敏感，可在线粒体基质中聚集和积累，导致线粒体功能障碍。Lon蛋白酶是线粒体中主要的质量控制蛋白酶之一，它降解氧化的乌头酸酶以维持线粒体稳态。本章介绍了一种使用从细胞中分离的线粒体，简单可靠地测量线粒体乌头酸酶以及柠檬酸合酶活性的分步方案。该方案简单快速，并且针对使用酶标仪的96孔板进行了优化。
