小麦尿卟啉原合酶的分离和纯化  

Isolation and purification of uroporphyrinogen Ⅲ synthase from wheat leaf

作　　者：范军[1] 郭蔼光[1] 

机构地区：[1]西北农业大学基础科学系

出　　处：《西北农业大学学报》2000年第1期11-15,共5页Journal of Northwest Sci-Tech University of Agriculture and Forestry（Natural Science Edition)

基　　金：国家自然科学基金!( 395 70 0 6 6 )

摘　　要：以40%～70%饱和度硫酸铵分级沉淀、DEAESepharoseiCL-6B、SephadexG-100、羟基磷灰石、PhenylSepharoseCL-4B和制备电泳分离纯化了小麦的尿卟啉原合酶(UROS)。酶的亚基分子质量为54ku,全酶分子质量为66ku,酶的等电点6.3,在pH8.2时比活为257μmol/(mg·min),最适反应温度40℃,5mmol/L的巯基乙醇和Mg2+促进酶活,纯化的小麦UROS在-20℃下0.1mol/L,pH8.5的Tris-HCl,内含质量分数为50%的甘油,5mmol/L的巯基乙醇和MgCl2中贮藏7d酶活损失90%。Uroporphyrinogen Ⅲ synthase (UROS) is isolated and purified from wheat leaves by 40%-70% ammonium sulphate fractionation and followed by chromatography DEAE Sepharose CL 6B,Sephadex G 100,hydroxylatite,Phenyl Sepharose CL 4B and finally by electrophoresis.The purified enzyme has a specific activity of 257 μmol/(mg·min) protein at pH of 8.5.It shows homogeneity and has a subunit Mr of 54 ku by SDS PAGE.The whole enzyme has about a relative Mr of 66 ku by Sephadex G 100 and thus appears to be monomeric.Its pI is about 6.3.It was maximally active at a temperature of 40℃ and activated by 2 mercaptothanol and Mg 2+ .The purified enzyme after concentrated is stored with a loss of 90% activity for a week at -20℃ in the dark in 0.1 mol/L Tris HCl buffer (pH 8.5),including 50% of glycerol,5 mmol/L 2 mercap toethanol and MgCl 2.

关 键 词：尿卟啉原Ⅲ合酶 小麦 纯化 分离 代谢 层析法 

分 类 号：S512.101[农业科学—作物学] Q555.6[生物学—生物化学]