Université Paris-Saclay, CNRS, INRAE, Univ Evry, Institute of Plant Sciences Paris-Saclay (IPS2), Orsay, France.
Methods Mol Biol. 2022;2526:135-141. doi: 10.1007/978-1-0716-2469-2_10.
Protein carbonylation is an irreversible oxidation process leading to a loss of function of carbonylated proteins. Carbonylation is largely considered as a hallmark of oxidative stress, the level of protein carbonylation being an indicator of the oxidative cellular status. The method described herein represents an adaptation to the commonly used 2,4-dinitrophenylhydrazine (DNPH)-based spectrophotometric method to monitor protein carbonylation level. The classical final sample precipitation was replaced by a gel filtration step avoiding the tedious and repetitive washings of the protein pellet to remove free DNPH while allowing optimal protein recovery.This improved protocol here implemented to assay protein carbonylation in plant leaves can potentially be used with any cellular extract.
蛋白质羰基化是一种导致羰基化蛋白质功能丧失的不可逆氧化过程。羰基化在很大程度上被认为是氧化应激的标志，蛋白质羰基化水平是氧化细胞状态的一个指标。本文所述的方法是对常用的 2,4-二硝基苯肼（DNPH）基于分光光度法监测蛋白质羰基化水平的方法的改编。经典的最终样品沉淀被凝胶过滤步骤所取代，避免了繁琐和重复的洗涤蛋白质沉淀以去除游离的 DNPH，同时允许最佳的蛋白质回收。该改进的方案在这里被用于检测植物叶片中的蛋白质羰基化，可能可以用于任何细胞提取物。
