Mechanism of Pigeon Liver Malic Enzyme

Abstract

Abstract Crystalline pigeon liver malic enzyme (malate dehydrogenase (decarboxylating), EC 1.1.1.40) catalyzes the slow, magnesium-dependent reduction of oxalacetate and conversion of l-malate to l-lactate in the presence of TPNH and TPN, respectively, with TPN acting in a catalytic manner. Pyruvate reductase activity associated with this enzyme has a pH optimum of 6.5 to 6.8 and a Michaelis constant of 5.52 mm for pyruvate. The activity ratio of oxidative decarboxylation of l-malate, reduction of pyruvate, and reduction of oxalacetate catalyzed by the enzyme is approximately 7.3:0.31:1. Titration of 4 sulfhydryl groups by 5,5'-dithiobis(2-nitrobenzoic acid) induced a reversible loss of malic enzyme activity and an increase in pyruvate reductase activity, while the oxalacetate reductase activity remained constant. The modified enzyme exhibited a new activity ratio of 0.3:0.6:1. These observations are consistent with both reductases being auxiliary reactions that could be expected from a sequential, ordered mechanism with oxalacetate as the enzyme-bound intermediate in a central complex. It is postulated that alteration of thiol groups led to a reversible blockage of the CO2 step, which was responsible for the observed changes in the diverse activities of the enzyme.