The purified and reconstituted ornithine/citrulline carrier from rat liver mitochondria: electrical nature and coupling of the exchange reaction with H+ translocation

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Abstract
The mechanism and the electrical nature of ornithine/citrulline exchange has been investigated in proteoliposomes reconstituted with the ornithine/citrulline carrier purified from rat liver mitochondria. The stoichiometry of the exchanging substrates was close to 1:1. The exchange was not affected by inducing electrogenic flux of K+ with valinomycin. In contrast, the pH gradient generated by the K+/H+ exchanger nigericin in the presence of an outwardly directed K+ gradient stimulated the ornithineout/citrullinein exchange, but not the ornithine/ornithine homoexchange. Experiments in which either the internal or the external pH was varied, while keeping constant the pH in the other compartment, indicated that maximal exchange rates are found at pH 6 in the compartment containing citrulline and at pH 8 in the compartment containing ornithine. Changes in fluorescence of the pH indicator pyranine, included inside the proteoliposomes, showed that the exchanges ornithineout/citrullinein and citrullineout/ornithinein are accompanied by translocation of H+ in the same direction as citrulline. It is concluded that the mitochondrial ornithine/citrulline carrier catalyses an electroneutral exchange of ornithine+ for citrulline plus an H+. A reasonable model is one in which ornithine binds to a deprotonated carrier and citrulline to a protonated carrier and both substrate-carrier complexes are neutral. The physiological implications of this transport process are discussed.