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(Received for publication, February 6, 1997, and in revised form, April 8, 1997)
From the The
Volume 272, Number 26,
Issue of June 27, 1997
pp. 16096-16102
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
-Aminobutyric Acid Transporter GAT-1 Plays
a Critical Role in Neurotransmitter Recognition
,
Department of Biochemistry, Hadassah Medical
School, the Hebrew University, Jerusalem 91120, Israel and the
§ Vollum Institute, Oregon Health Science University,
Portland, Oregon 97201
-aminobutyric acid (GABA) transporter
GAT-1 is located in nerve terminals and catalyzes the electrogenic
reuptake of the neurotransmitter with two sodium ions and one chloride.
We now identify a single tyrosine residue that is critical for GABA recognition and transport. It is completely conserved throughout the
superfamily, and even substitution to the other aromatic amino acids,
phenylalanine (Y140F) and tryptophan (Y140W), results in completely
inactive transporters. Electrophysiological characterization reveals
that both mutant transporters exhibit the sodium-dependent transient currents associated with sodium binding as well as the chloride-dependent lithium leak currents characteristic of
GAT-1. On the other hand, in both mutants GABA is neither able to
induce a steady-state transport current nor to block their transient currents. The nontransportable analog SKF 100330A potently inhibits the
sodium-dependent transient in the wild type GAT-1 but not in the Y140W transporter. It partly blocks the transient of Y140F. Thus, although sodium and chloride binding are unimpaired in the tyrosine mutants, they have a specific defect in the binding of GABA.
The total conservation of the residue throughout the family suggests
that tyrosine 140 may be involved in the liganding of the amino group,
the moiety common to all of the neurotransmitters.
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