Biochemical characterization of the C4-dicarboxylate transporter DctA from Bacillus subtilis.
Bacterial secondary transporters of the DctA family mediate ion-coupled uptake of C(4)-dicarboxylates. Here, we have expressed the DctA homologue from Bacillus subtilis in the Gram-positive bacterium Lactococcus lactis. Transport of dicarboxylates in vitro in isolated membrane vesicles was assayed. We determined the substrate specificity, the type of cotransported ions, the electrogenic nature of transport, and the pH and temperature dependence patterns. DctA was found to catalyze proton-coupled symport of the four C(4)-dicarboxylates from the Krebs cycle (succinate, fumurate, malate, and oxaloacetate) but not of other mono- and dicarboxylates. Because (i) succinate-proton symport was electrogenic (stimulated by an internal negative membrane potential) and (ii) the divalent anionic form of succinate was recognized by DctA, at least three protons must be cotransported with succinate. The results were interpreted in the light of the crystal structure of the homologous aspartate transporter Glt(Ph) from Pyrococcus horikoshii.
Groeneveld
Maarten
M
Department of Biochemistry, University of Groningen, Groningen Biomolecular Science and Biotechnology Institute, Nijenborgh 4, 9747 AG Groningen, Netherlands.
Weme
Ruud G J Detert Oude
RG
Duurkens
Ria H
RH
Slotboom
Dirk Jan
DJ
D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000235:genetics / Q000502:physiology*; D029382:Dicarboxylic Acid Transporters / Q000235:genetics / Q000378:metabolism*; D003998:Dicarboxylic Acids / Q000378:metabolism*; D005650:Fumarates / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D008293:Malates / Q000378:metabolism; D013379:Substrate Specificity; D013386:Succinates / Q000378:metabolism; D013696:Temperature
D001426:Bacterial Proteins; D029382:Dicarboxylic Acid Transporters; D003998:Dicarboxylic Acids; D005650:Fumarates; D008293:Malates; D013386:Succinates; C030298:malic acid