MEMBRANE POTENTIAL-GENERATING TRANSPORT OF CITRATE AND MALATE CATALYZED BY CITP OF LEUCONOSTOC-MESENTEROIDES

Citrate uptake in Leuconostoc mesenteroides subsp. mesenteroides 19D i s catalyzed by a secondary citrate carrier (CitP). The kinetics and me chanism of CitP were investigated in membrane vesicles of L. mesentero ides. The transporter is induced by the presence of citrate in the med ium and transports both citrate and malate. In spite of sequence homol ogy to the Na+-dependent citrate carrier of Klebsiella pneumoniae, Cit P is not Na+-dependent, nor is CitP Mg2+-dependent. The pH gradient (D elta pH) is a driving force for citrate and malate uptake into the mem brane vesicles, whereas the membrane potential (Delta psi) counteracts transport. An inverted membrane potential (inside positive) generated by thiocyanide diffusion can drive citrate and malate uptake in membr ane vesicles. Analysis of the forces involved showed that a single uni t of negative charge is translocated during transport. Kinetic analysi s of citrate counterflow at different pH values indicated that CitP tr ansports the dianionic form of citrate (Hcit(2-)) with an affinity con stant of similar to 20 mu M. It is concluded that CitP catalyzes Hcit( 2-)/H+ symport. Translocation of negative charge into the cell during citrate metabolism results in the generation of a membrane potential t hat contributes to the protonmotive force across the cytoplasmic membr ane, i.e. citrate metabolism in L. mesenteroides generates metabolic e nergy. Efficient exchange of citrate and D-lactate, a product of citra te/carbohydrate co-metabolism, is observed, suggesting that under phys iological conditions, CitP may function as an electrogenic precursor/p roduct exchanger rather than a symporter. The mechanism and energetic consequences of citrate uptake are similar to malate uptake in lactic acid bacteria.