PURIFICATION, CHARACTERIZATION AND NUCLEOTIDE-SEQUENCE OF THE PERIPLASMIC C4-DICARBOXYLATE-BINDING PROTEIN (DCTP) FROM RHODOBACTER-CAPSULATUS

A periplasmic binding protein essential for high-affinity transport of the C4-dicarboxylates malate, succinate and fumarate across the cytoplasmic membrane of the purple photosynthetic bacterium Rhodobacter capsulatus has been purified to homogeneity and some of its ligand-binding properties characterized. The protein was not produced in a Tn5 insertion mutant unable to transport C4-dicarboxylates under aerobic conditions in the dark. Wild-type DNA corresponding to the location of the transposon insertion site was subcloned and a 1.5 kb section sequenced. A complete open reading frame of 999bp was identified that encoded a 333-residue protein (DctP) with a molecular weight of 36128 with a 26-residue amino-terminal signal peptide. The identity of this protein with the purified dicarboxylate-binding protein and the position of the predicted signal peptide cleavage site was confirmed by N-terminal sequencing. No significant homology with other proteins was detected in database searches. A GC-rich region of dyad symmetry was located 7 bp downstream of the dctP translational stop codon. This structure may be of significance in regulating the relative abundance of DctP and other dct gene products which comprise the high-affinity dicarboxylate transport system in this bacterium.