On the mechanism of sodium ion translocation by oxaloacetate decarboxylase of Klebsiella pneumoniae.

Proteoliposomes reconstituted with purified oxaloacetate decarboxylase of Klebsiella pneumoniae catalyzed the uptake of Na+ ions upon oxaloacetate decarboxylation. The degree of coupling between the chemical and the vectorial reaction is dependent on the reconstitution conditions, and with the best preparations approaches a stoichiometry of two Na+ ions per decarboxylation of one oxaloacetate. This coupling ratio is observed only in the absence of a delta mu Na+, immediately after oxaloacetate addition. The ratio gradually declines during development of the electrochemical Na+ ion gradient and becomes zero in the steady state. The Na+ pump, however, continued to decarboxylate oxaloacetate and to catalyze Na+ influx at the apparent stoichiometry of two Na+ ions per decarboxylation event. During the steady state, this influx must be compensated by Na+ efflux of the same size. The efflux is catalyzed by the Na+ pump upon oxaloacetate decarboxylation, because in the absence of the substrate the efflux rate dropped to less than 10%. Proteoliposomes loaded with Na2SO4 catalyzed a bicarbonate-dependent uptake of 22Na+ that was completely abolished after incubation with avidin. These results suggest coupling of Na+ translocation to the carboxylation/decarboxylation of the biotin prosthetic group without the requirement for the oxaloacetate/pyruvate interconversion. The oxaloacetate-dependent transport of Na+ into proteoliposomes was inhibited by the additional presence of the beta + gamma subunits of oxaloacetate decarboxylase. A model of Na+ translocation by oxaloacetate decarboxylase based on these experimental results is proposed.