4-Aminobutyrate (GABA) transporters from the amine-polyamine-choline superfamily: substrate specificity and ligand recognition profile of the 4-aminobutyrate permease from Bacillus subtilis.
A previous study [Ferson, Wray and Fisher (1996) Mol. Microbiol. 22, 693-701] has shown that transposon-mediated disruption of a protein 47% identical to the Escherichia coli GABA (4-aminobutyrate) transporter abolishes the ability of nitrogen-limited culture conditions to induce expression of a GABA transport activity in Bacillus subtilis. Here it is demonstrated directly that the B. subtilis GABA permease (gabP) gene can complement the transport defect in the gabP-negative E. coli strain. Unexpectedly, the ligand-recognition profile of the B. subtilis GabP was found to differ substantially from that of the highly homologous E. coli GabP. Unlike the E. coli GabP, the B. subtilis GabP: (i) exhibits approx. equal preference for the 3-carbon (beta-alanine, Km=9.6 microM) and the 4-carbon (GABA, Km=37 microM) amino acids, and (ii) resists inhibition by bulky, conformationally constrained compounds (e.g. nipecotic acid, guvacine), which are active against GABA transporters from brain. The present study shows additionally that the B. subtilis GabP can translocate several open-chain GABA analogues (3-aminobutyrate, 3-aminopropanoate, cis-4-aminobutenoate) across the membrane via counterflow against [3H]GABA. Thus, consistent with the idea that the ligand-recognition domain of the B. subtilis GabP is less spacious than that of the close homologue from E. coli, the former exhibits more stringent requirements than the latter for substrate recognition and translocation. These distinct functional characteristics of the E. coli and B. subtilis GABA transporters provide a basis by which to identify ligand-recognition domains within the amine-polyamine-choline transporter superfamily.
Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston, Texas 77555-0641, USA.
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Brechtel
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King
D000588:Amines / Q000378:metabolism; D001412:Bacillus subtilis / Q000201:enzymology* / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002794:Choline / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism; D029968:Escherichia coli Proteins; D050485:GABA Plasma Membrane Transport Proteins; D007700:Kinetics; D008024:Ligands; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D027361:Organic Anion Transporters; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D000588:Amines; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D050485:GABA Plasma Membrane Transport Proteins; D008024:Ligands; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D027361:Organic Anion Transporters; C513601:gabP protein, E coli; D002794:Choline