MOLECULAR-FORMS OF CHICKEN LIVER CYTOPLASMIC MALATE-DEHYDROGENASE

Cytoplasmic malate dehydrogenase was purified 20-fold from the supernatant fraction of chicken liver homogenates in 0.25 M sucrose (differential centrifugation at 100,000 .times. g) through Ca phosphate gel absorption, ammonium sulfate fractionation (45-80% saturation) and Sephadex G-100 chromatography. Molecular forms, A and B, of the enzyme are unmodified during the purification process, observed by electrophoresis. Two additional bands that appear in the electrophoretic patterns are not attributable to molecular forms of the enzymes, but to the lack of non-dehydrogenase activity of the preparation used. Both forms differ in their thermal stability (form A is more thermolabile) and net charge; they have the same MW (67,000 .+-. 5,000 daltons), determined by Sephadex G-200 chromatography. The enzyme in the purified preparations exhibits 2 KM values in the double reciprocal plot v [velocity] vs. [L-malate], due to the presence of 2 forms with different affinity for the hydroxyacid.