人胎肝超氧化物歧化酶的研究

Purification and Characterization of Superoxide Dismutase From Human Fetal Liver

采用离子交换和凝胶过滤层析法,从正常人胎肝提取、纯化铜锌超氧化物歧化酶(Cu.Zn-SOD),并对其性质作了研究,结果测得人胎肝组织Cu·Zn-SOD平均含量为6.44unit/g湿重,并发现随着胎龄的增加人胎肝Cu.Zn-SOD含量上升;纯化后的Cu·Zn-SOD在聚丙烯酰胺凝胶电泳图谱上呈现单一区带;其活性受氰化钾抑制;以原子吸收光谱测得其铜、锌含量分别为0.39％和0.1％,采用SDS-聚丙烯酰胺凝胶电泳测得其亚基分子量为16kD;氨基酸自动分析仪测得其亚基的氨基酸残基数为151.5;在紫外吸收光谱上于265nm和258nm处分别出现两个吸收高峰。本研究结果表明,人胎肝Cu.Zn-SOD与成人肝及其他组织的Cu.Zn-SOD理化性质上基本一致。

Copper,zinc superoxide dismutase (Cu.Zn-SOD) has been purified from fresh human fetal livers by the methods of ion exchange chromatograph (DEAE-32) and gel filtration (Sephadex G-75).Its properties, such as molecular weight, composition of amino acids, ultraviolet spectrum, metal contents and sensitivity to heat and cyanide, were also studied.The average enzyme content of the fetal hepatic tissue was 156.44 unit/g wet weight,much lower than that from adult liver. As the fetus grew, however , it increased. Cu. Zn-SOD purified was proved to be homogeneous, giving a single band on polyacrylamide gel electrophoresis. When subjected to cyanide, the enzyme activity was inhibited, but it could tolerate 50℃ for 10 min without any loss of the enzyme activite. The subunit of the enzyme has a molecular weight of 16 000 dalton and contains 151 amino acid residues, as were determined by SDS-PAGE and amino acid automatic analyzer, respectively. As showed on ultraviolet spectrum, it has a maximum absorption at 265 nm with a smaller shoulder at 258 nm. The copper and zinc contents of the enzyme, measured by using atomic absorption spectraphotometry, were 0.39% and 0.37%, respectively. The results of the study show that Cu.Zn-SOD from human fetal liver and that from adult liver are identical in properties.