title: Functional characterization of the Betaine/gamma-aminobutyric acid transporter BGT-1 expressed in Xenopus oocytes.
abstract: Betaine is an osmolyte accumulated in cells during osmotic cell shrinkage. The canine transporter mediating cellular accumulation of the osmolyte betaine and the neurotransmitter gamma-aminobutyric acid (BGT-1) was expressed in Xenopus oocytes and analyzed by two-electrode voltage clamp and tracer flux studies. Exposure of oocytes expressing BGT-1 to betaine or gamma-aminobutyric acid (GABA) depolarized the cell membrane in the current clamp mode and induced an inward current under voltage clamp conditions. At 1 mM substrate the induced currents decreased in the following order: betaine = GABA > diaminobutyric acid = beta-alanine > proline = quinidine > dimethylglycine > glycine > sarcosine. Both the Vmax and Km of GABA- and betaine-induced currents were voltage-dependent, and GABA- and betaine-induced currents and radioactive tracer uptake were strictly Na+-dependent but only partially dependent on the presence of Cl-. The apparent affinity of GABA decreased with decreasing Na+ concentrations. The Km of Na+ also depended on the GABA and Cl- concentration. A decrease of the Cl- concentration reduced the apparent affinity for Na+ and GABA, and a decrease of the Na+ concentration reduced the apparent affinity for Cl- and GABA. A comparison of 22Na+-, 36Cl--, and 14C-labeled GABA and 14C-labeled betaine fluxes and GABA- and betaine-induced currents yielded a coupling ratio of Na+/Cl-/organic substrate of 3:1:1 or 3:2:1. Based on the data, a transport model of ordered binding is proposed in which GABA binds first, Na+ second, and Cl- third. In conclusion, BGT-1 displays significant functional differences from the other members of the GABA transporter family.
authors.affiliation: Department of Physiology, University of Tübingen, Tübingen 72076, Germany.
authors.forename: I
authors.initials: I
authors.lastname: Matskevitch
authors.forename: C A
authors.initials: CA
authors.lastname: Wagner
authors.forename: C
authors.initials: C
authors.lastname: Stegen
authors.forename: S
authors.initials: S
authors.lastname: Bröer
authors.forename: B
authors.initials: B
authors.lastname: Noll
authors.forename: T
authors.initials: T
authors.lastname: Risler
authors.forename: H M
authors.initials: HM
authors.lastname: Kwon
authors.forename: J S
authors.initials: JS
authors.lastname: Handler
authors.forename: S
authors.initials: S
authors.lastname: Waldegger
authors.forename: A E
authors.initials: AE
authors.lastname: Busch
authors.forename: F
authors.initials: F
authors.lastname: Lang
mesh_terms: D000818:Animals; D001622:Betaine / Q000378:metabolism*; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002414:Cations, Monovalent / Q000494:pharmacology; D002712:Chlorides / Q000494:pharmacology; D004285:Dogs; D004553:Electric Conductivity; D050485:GABA Plasma Membrane Transport Proteins; D008954:Models, Biological; D009865:Oocytes; D018408:Patch-Clamp Techniques; D011994:Recombinant Proteins / Q000378:metabolism; D012964:Sodium / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D014981:Xenopus; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D002414:Cations, Monovalent; D002712:Chlorides; D050485:GABA Plasma Membrane Transport Proteins; D011994:Recombinant Proteins; C072207:betaine plasma membrane transport proteins; D001622:Betaine; D005680:gamma-Aminobutyric Acid; D012964:Sodium