A rapid purification method for human erythrocyte pyruvate kinase

Abstract

Human erythrocyte pyruvate kinase (ATP: pyruvate phosphotransferase. E.C.2.7.1.40) is purified 30,000-fold, using a method which includes ammonium sulfate precipitation, Sephadex G-75 filtration, and Blue Dextran-Sepharose 4B chromatography. The enzyme is resolved into two peaks on Blue Dextran-Sepharose 4B. The first peak with sp act of 300 corresponds to the mature form (R 4) whereas the second peak with sp act of 180 corresponds to R 2R 2. Peaks I and II give one band on 10% polyacrylamide gel without SDS. Peak II gives two bands on 10% SDS gel with molecular weights 60,000 (R′) and 57,500 (R). On the other hand peak I gives only one band on 10% SDS gel having a molecular weight of 57,500. Both the R 4 and R 2R 2 forms of the enzyme have the same pH optimum of 7.2.