Mechanisms of Citrate Transport and Exchange in Corn Mitochondria

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Abstract
Previous work (Birnberg, Jayroe, Hanson 1982 Plant Physiol 70: 511-516) demonstrated that corn mitochondria (Zea mays L.) can accumulate citrate by a malate- and phosphate-independent proton symporter. This uptake and symport of other ions were investigated. Passive swelling experiments indicated that corn mitochondria can accumulate several other anions by proton symport, but only isocitrate is taken up nearly as effectively as citrate. At the optimal pH (4.5), active uptake of carrier-free [14C]citrate in 50 micromolar mersalyl is inhibited by fourteen anions, but only the I50 (the concentration of inhibitor required to reduce uptake of carrier-free [14C]citrate by 50%) values of citrate (0.08 millimolar) and d-and l-isocitrate (0.5 millimolar) are less than 4 millimolar. Isocitrate is a competitive inhibitor of citrate uptake and [14C]isocitrate is accumulated with a Km similar similar to its I50. Valinomycin reduces net active citrate accumulation at pH 7.5, consistent with the relatively low Vmax for citrate uptake. At pH 4.5, mersalyl reduces the rate of citrate uptake without changing the affinity of the carrier for citrate. Thus, the corn mitochondria have a high-affinity, mersalyl-insensitive carrier selective for citrate that also transports isocitrate. The pH optimum for oxidation of both endogenous substrates and citrate is approximately pH 6.8, but citrate oxidation is low at all pH values and is poorly coupled to ATP synthesis. Under active conditions only, at pH 7.0, malate/citrate exchange occurs with 4 millimolar malate being sufficient to remove about half the matrix citrate. Therefore, in vivo both citrate uptake by proton symport and efflux by malate/citrate exchange should occur, with the net direction of citrate movement determined by the cytoplasmic pH, and citrate and malate concentrations; in most cases, net efflux is likely to be favored.