Structure, function, and genomic organization of human Na⁺-dependent high-affinity dicarboxylate transporter

We have cloned and functionally characterized the human Na⁺-dependent high-affinity dicarboxylate transporter (hNaDC3) from placenta. The hNaDC3 cDNA codes for a protein of 602 amino acids with 12 transmembrane domains. When expressed in mammalian cells, the cloned transporter mediates the transport of succinate in the presence of Na⁺ [concentration of substrate necessary for half-maximal transport (K(t)) for succinate = 20 ± 1 μM]. Dimethylsuccinate also interacts with hNaDC3. The Na⁺-to-succinate stoichiometry is 3:1 and concentration of Na⁺ necessary for half-maximal transport (K_0.5(Na+)) is 49 ± 1 mM as determined by uptake studies with radiolabeled succinate. When expressed in Xenopus laevis oocytes, hNaDC3 induces Na⁺-dependent inward currents in the presence of succinate and dimethylsuccinate. At a membrane potential of -50 mV, K_0.5(Suc) is 102 ± 20 μM and K_0.5(Na+) is 22 ± 4 mM as determined by the electrophysiological approach. Simultaneous measurements of succinate-evoked charge transfer and radiolabeled succinate uptake in hNaDC3-expressing oocytes indicate a charge-to-succinate ratio of 1:1 for the transport process, suggesting a Na⁺-to-succinate stoichiometry of 3:1. pH titration of citrate-induced currents shows that hNaDC3 accepts preferentially the divalent anionic form of citrate as a substrate. Li⁺ inhibits succinate- induced currents in the presence of Na⁺. Functional analysis of rat-human and human-rat NaDC3 chimeric transporters indicates that the catalytic domain of the transporter lies in the carboxy-terminal half of the protein. The human NaDC3 gene is located on chromosome 20q12-13.1, as evidenced by fluorescent in situ hybridization. The gene is >80 kbp long and consists of 13 exons and 12 introns.