Purification and properties of enolase of human erythrocytes

Abstract

1. 1. Human erythrocyte enolase (2-phospho-D-glycerate hydrolyase, EC 4.2.1.11) was purified I000-fold.

2. 2. The pH-optimum was at pH 6.5. The molecular weight, estimated by gel filtration, was found to be 95,000 ± 5,000.

3. 3. Electrophoresis on agar-agarose at pH 8.5 and 6.4 showed only one band in hemolysate as well as in purified preparations.

4. 4. The enzyme was stabilized by magnesium and ammonium ions.

5. 5. The Km values for 2-phosphoglyceric acid at pH 6.5 and pH 7.2 were found to be the same: Km, (2-phosphoglyceric acid) = 28 μM. The optimal magnesium concentration was between 1 and 5 mM.

6. 6. Ca2+, at physiological concentrations, inhibited the enzyme. This inhibition depends on the Mg2+ concentration.

7. 7. The enzyme was inhibited by fluoride. This inhibition depends on magnesium, fluoride and phosphate concentration. Moreover the F− inhibition was dependent on 2-phosphoglyceric acid concentration in a competitive way.