Some properties of purified isocitric enzyme

Abstract

The chemical and physical properties of the isocitric enzyme are described. The inactivation of the enzyme caused by dialysis against distilled water or 0.05 [image] aminotrishydroxymethylmethane buffer (pH 7.3) can be prevented by addition of 0.1 [image] ammonium sulfate or sodium sulfate to the dialyzing medium. The absolute activity of the dehydrogenase reaction of the isocitric enzyme at pH 7.3 at 24[degree] is 350 molecules of D-isocitrate oxidized/molecule of enzyme/minute (in presence of 0.00042 [image] triphosphopyridine nucleotide), and for the carboxylase reaction atp H 5.6 at 14[degree] is 560 molecules of oxalosuccinate decarboxylated/ molecule of enzyme/minute (in presence of 0.0013 [image] manganous chloride) when calculated from the velocity constants (Vm) of these reactions. The dehydrogenase reaction does not require Mn2- ions but, being closely linked to the carboxylase reaction, it will not proceed in a forward direction unless the carboxylase reaction also occurs. The backward carboxylase reaction depends on dehydrogenase activity. The Michaelis constant for D-isocitrate is 2.6 x 10-6 [image] at pH 7.3 at 24[degree]; that for oxalosuccinate is 5.6 x 10-4 [image] at pH 7.3 at 24[degree] for the reduction reaction, and 2.5 x 10-2 [image] at pH 5.6 at 14[degree] for the decarboxyla-tion reaction. A hypothesis to explain the mechanism of action of the isocitric enzyme is described. A similar mechanism is proposed for the malic enzyme.