Studies On The Purification And Characterization Of Copper/Zinc Superoxide Dismutase From Hog Blood

The isolation of crude Cu, Zn-SOD can be started with organic solvents (ethanol-chloroform) in the classical manner. Then we take two different methods:the first one is that chromatography is carried out on Sephadex G100, G50 at room temperature with PH7. 6, 0. 02M phosphate buffer, respectively, Following this, ion-exchange chromatography is carried at pH7. 6 with 0. 02M and 0. 4M phosphate buffer;the second is that the crude enzyme runs on Sephadex G100-G50 column under the same experimental conditions(only with Sephadex G100, G50).At last polyacrylamide gel electrophoresis and SOD assay tell us both of products are highly purified SOD. These SOD activities are very high, too.Crude hog superoxide dismutase treated with different temperatures and for different time, the result.shows as following: The treatment is most appropriate at 55-65 C and for 15-26 minutes for thermal denatured extracting SOD from hog blood and specific activi is very high.In this thesis, we have done some research on the characterization of Cu, Zn-SOD from hog blood by polyacrylamide gel electrophoresis, electric focusing electrophoresis and dealing with some different reagents. The result shows as follows: one is that hog Cu, Zn-SOD has no isozymes and it contains the same two subunits , with molecular masses of 32112 daltons . the other is that the polymorphism of the Cu, Zn-SOD from hog blood is caused by polymerization and conformational isomers. So it has been found that this polymorphism of SOD was changed under different experimental conditions, which showed that the polymorphism was not caused by the difference of genes or the difference of length or side groups of the peptide chains which was caused during the processing after transcript or the purification. According to thereferences about the protein research, we estimated that the possible reasons of the polymorphism were polymerization, conformational isomers and charge isomers. There was some research on the SOD charge isomers, but not on this polymorphism.