【摘要】 以亚洲玉米螟（Ｏｓｔｒｉｎｉａｆｕｒｎａｃａｌｉｓ）为材料，经ＳｅｐｈａｄｅｘＧ７５、ＤＥＡＥＳｅｐｈａｒｏｓｅＦａｓｔＦｌｏｗ、还原型谷胱甘肽（ＧＳＨ）为配基的亲和层析，分离纯化得到玉米螟谷胱甘肽转硫酶（ＧＳＴ）。以２，４二硝基氯苯（ＣＤＮＢ）为底物测得其比活为１０７０Ｕ／ｍｇ蛋白，收率为２２９％，提纯１１３８倍。经ＳＤＳＰＡＧＥ和ＰＡＧＥ鉴定，得到的ＧＳＴ为一条带，亚基的分子量为２５ｋＤ，经凝胶过滤测得其全酶分子量为５０ｋＤ，说明该酶是由两个相同亚基组成的。ＩＥＦ测得主带等电点为８２。该酶最适温度为４１℃，最适ｐＨ范围为６３～６９。经动力学测定该酶双底物反应机制为序列机制，对ＣＤＮＢ的Ｋｍ值为０１５ｍｍｏｌ／Ｌ，对ＧＳＨ的Ｋｍ值为０３５ｍｍｏｌ／Ｌ，两种底物的ｋｃａｔ均为７８３ｓ－１。Ｃｌ－对底物ＣＤＮＢ有竞争性抑制作用，对ＧＳＨ是非竞争性抑制作用；３，５二硝基苯甲酸（ＤＮＢ）对ＣＤＮＢ有竞争性抑制作用，对ＧＳＨ是反竞争性抑制作用。更多还原

【Abstract】 Glutathione S transferase(GST) was purified from larvae of Asian corn borer( Ostrina furnacalis) by Sephadex G 75,DEAE Sepharose Fast Flow and reduced gl utathione(GSH) affinity chromatography.The specific activity determined by 2,4 Dinitrochlorobenzene(CDNB) and GSH as substrates is 1070 U/mg protein.The purifi cation fold is 1138 and the total recovered activity is 22.9%.SDS PAGE and PAGE showed one band and the molecular weight of the subunit is 25?kD and is 50?kD determined by gel exclusion, which indicates that the enzyme is composed of two identical subunits. The pI point of the enzyme is 8.2. The kinetics constants of the purified GST toward substrate were investigated. For CDNB, K m is 0.15 ?mmol/L, k cat is 783?s -1 ,and for GSH, K m is 0.35?mmol/L, k cat is 783?s -1 .The reaction mechanism of GST for bisubstrate i s sequential mechanism by kinetic research. The optimum temperature is 41℃ and the optimum pH range is 6.3～6.9.The enzyme is inhibited by Cl - and 3,5 Dinit robenzoic Acid. The kind of inhibition by Cl - is competitive for CDNB and non competitive for GSH,respectively.The kind of inhibition by 3,5 Dinitrobenzoi c Acid is competitive for CDNB and un competitive for GSH,respectively.更多还原