An efficient method for purification of cuprozinc superoxide dismutase from bovine erythrocytes

Abstract

Cuprozinc superoxide dismutase (Cu,Zn-SOD) was isolated from bovine erythrocytes by pH-controlled ammonium sulfate-methanol extraction (ASME extraction). Adjustment of the pH of a suspension of the lysed red cells in the presence of ammonium sulfate (90% saturation) to pH 5.0, followed by partition with an equal amount of methanol, resulted in isolation of the enzyme with specific, activity of greater than 2000 units/mg of protein. Further purification using DEAE-cellulose column chromatography gave a highly purified Cu,Zn-SOD showing a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Using this procedure about 14 mg of pure Cu,Zn-SOD with a specific activity of 4728 units/mg of protein can be recovered from one liter of bovine blood. The enzyme was characterized and the results obtained were in agreement with earlier reports. This procedure appears, therefore, to be a convenient method for isolating the enzyme.