人肝谷胱甘肽过氧化物酶纯化及性质研究

Purification and Charicterization of Glutathione Peroxidase from Human Liver 86315

经硫酸铵分级沉淀，离子交换层析和凝胶过滤等步骤，从人肝中获得了ＰＡＧＥ单一条带的谷胱甘肽过氧化物酶，比活提高１２０倍，得率为２５％。凝胶过滤法测得分子量为９０９８０，ＳＤＳ－ＰＡＧＥ测定亚基分子量为２２４２３．原子吸收法测得每分子酶含有四个硒原子。等电聚焦显示该酶等电点为５．０．酶活力的最适ｐＨ为８．５，最适温度为３７℃。动力学实验提示该酶作用机理属于乒乓机制型。

Human liver glutathione peroxidase (GSHPX) was purified to homogeneity by using ammonium sulfate precipitation, ion exchange chromatography and gel filtration chromatography.The specific activity of purified GSHPX was about 120 fold higher than that of raw liver extracts. The yield was about 25% . The molecular weight of the enzyme , determined by gel filtration, was 90 980 and that of the subunit determined by SDS-PAGE was 22 423. The enzyme therefore consisted four identical subunits. Selenium contents were determined by atomic absorption spectrophotometer. The enzyme contained approximately 4 mols of selenium per mole of protein. The isoelectric point of the enzyme was 5. 0. The optium pH was 8. 5. The optium temperature was 37℃ . Kinetic studies showed that the reaction catalyed by the enzyme was in accordance with a Ping-Pong mechanism.