Jung Tobias, Engels Martina, Kaiser Barbara, Poppek Diana, Grune Tilman
Research Institute of Environmental Medicine, Heinrich Heine University, Duesseldorf, Germany.
Free Radic Biol Med. 2006 Apr 15;40(8):1303-12. doi: 10.1016/j.freeradbiomed.2005.11.023. Epub 2005 Dec 28.
The production of free radicals and the resulting oxidative damage of cellular structures are always connected with the formation of oxidized proteins. The 20S proteasome is responsible for recognition and degradation of oxidatively damaged proteins. No detailed studies on the intracellular distribution of oxidized proteins during oxidative stress and on the distribution of the proteasome have been performed until now. Therefore, we used immunocytochemical methods to measure protein carbonyls, a form of protein oxidation products, and proteasome distribution within cells. Both immunocytochemical methods of measurement are semiquantitative and the load of oxidized proteins is increased after various oxidative stresses explored, with the highest increase in the perinuclear region of the cell. Distribution of the proteasome and the total protein content revealed the highest concentration of both in the nucleus. No redistribution of the proteasome during oxidative stress occurs. The normalized ratio of protein carbonyls to protein content was formed, indicating the highest concentration of oxidized proteins in the cytosolic region near the cell membrane. By forming the protein oxidation-to-proteasome ratio it was concluded that the highest load of oxidized proteins to the proteasome takes place in the cytosol, independent of the oxidant explored.
自由基的产生以及由此导致的细胞结构氧化损伤总是与氧化蛋白的形成相关联。20S蛋白酶体负责识别和降解氧化损伤的蛋白质。到目前为止，尚未对氧化应激期间氧化蛋白的细胞内分布以及蛋白酶体的分布进行详细研究。因此，我们使用免疫细胞化学方法来测量蛋白质羰基（一种蛋白质氧化产物的形式）以及细胞内蛋白酶体的分布。这两种免疫细胞化学测量方法都是半定量的，在探索的各种氧化应激后，氧化蛋白的负荷增加，在细胞核周围区域增加最为明显。蛋白酶体的分布和总蛋白含量显示两者在细胞核中的浓度最高。在氧化应激期间蛋白酶体不会发生重新分布。形成了蛋白质羰基与蛋白质含量的标准化比率，表明在细胞膜附近的胞质区域中氧化蛋白的浓度最高。通过形成蛋白质氧化与蛋白酶体的比率得出结论，氧化蛋白相对于蛋白酶体的最高负荷发生在细胞质中，与所探索的氧化剂无关。
