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Conserved tyrosine-147 plays a critical role in the ligand-gated current of the epithelial cation/amino acid transporter/channel CAATCH1
1 The Department of Physiology and Functional Genomics, University of
Florida College of Medicine, Gainesville, FL 32652, USA
2 The Whitney Laboratory, University of Florida, 9505 Ocean Shore Boulevard,
St Augustine, FL 32080, USA
* Present address: Shriners Hospital for Children of Northern California,
Sacramento, CA 98517, USA
e-mail:
wharvey{at}whitney.ufl.edu
Accepted 21 May 2002
CAATCH1 functions both as an amino-acid-gated cation channel and as a
cation-dependent, proline-preferring, nutrient amino acid transporter in which
the two functions are thermodynamically uncoupled. This study focuses on the
ionic channel aspect, in which a Tyr147 (wild type) to
Phe147 (Y147F) site-directed mutation was investigated by
steady-state electrophysiological measurements in the Xenopus laevis
oocyte expression system. This tyrosine residue is conserved within the third
transmembrane domain in members of the Na+:neurotransmitter
transporter family (SNF), where it plays a role in binding pharmacological
ligands such as cocaine to the serotonin (SERT), dopamine (DAT) and
norepinephrine (NET) transporters. Epithelial CAATCH1 is a member of the SNF
family. The results show that amino acid ligand-gating selectivity and current
magnitudes in Na+- and K+-containing media are
differentially altered in CAATCH1 Y147F compared with the wild type. In the
absence of amino acid ligands, the channel conductance of Na+,
K+ and Li+ that is observed in the wild type was reduced
to virtually zero in Y147F. In the wild type, proline binding increased
conductance strongly in Na+-containing medium and moderately in
K+-containing medium, whereas in Y147F proline failed to elicit any
cation currents beyond those of N-methyl-D-glucamine- or
water-injected oocytes. In the wild type, methionine binding strongly
inhibited inward Na+ currents, whereas in Y147F it strongly
stimulated inward currents in both Na+ and K+-containing
media. Indeed, in Na+-containing medium, the relative potency
ranking for inward current inhibition in the wild type
(Met>Leu>Gly>Phe>Thr) was similar to the ranking of
ligand-permissive gating of large inward currents in Y147F. In
Na+-containing medium, current/voltage relationships elicited by
ligands in the wild type were complex and reversing, whereas in Y147F they
were linear and inwardly rectifying. In K+-containing medium,
current/voltage relationships remained non-linear in Y147F. Both wild-type and
Y147F currents were Cl--independent. Together, these data
demonstrate a critical role for Tyr147 in ligand-binding
selectivity and modulation of the ionic channel conductance in CAATCH1. The
results support the argument that inhibition of the CAATCH1 conductance by
free methionine shares some properties in common with ligand inhibition of
DAT, SERT, NET and the 
-aminobutyric acid transporter (GAT1).
Key words: ion channel, potassium, sodium, amino acid, transporter, ionic current, Xenopus laevis, Manduca sexta
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  V. F. Sacchi, M. Castagna, S. A. Mari, C. Perego, E. Bossi, and A. Peres Glutamate 59 is critical for transport function of the amino acid cotransporter KAAT1 Am J Physiol Cell Physiol, September 1, 2003; 285(3): C623 - C632. [Abstract] [Full Text] [PDF]
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 Z. Liu, B. R. Stevens, D. H. Feldman, M. A. Hediger, and W. R. Harvey K+ amino acid transporter KAAT1 mutant Y147F has increased transport activity and altered substrate selectivity J. Exp. Biol., March 2, 2003; 206(2): 245 - 254. [Abstract] [Full Text] [PDF]
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