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(Received for publication, December 26, 1995, and in revised form, February 23, 1996)
From the A cDNA encoding for a weakly inward
rectifying K+ channel (sWIRK: salmon weakly inward
rectifying K+ channel) was isolated from the masu salmon
brain by expression cloning. The sWIRK channel exhibited the highest
similarity with members of the ROMK1 subfamily, BIR10/KAB-2
(70% amino acid identity) and ROMK1 (46%). An ATP binding motif which
is characteristic of this subfamily was also conserved. The sWIRK RNA
was detected in the brain, but not in the heart, kidney, skeletal
muscle, liver, testis, and ovary. In the brain, the expression was
observed in the ependymoglial cells on the surface of the ventricles as
well as in the small perineuronal glia-like cells in the midbrain and
the medulla. When compared with the strong inward rectifier IRK1
channel, the sWIRK channel showed a much weaker inward rectification
property, and the activation kinetics upon hyperpolarization was slower
and less voltage-dependent. The slope conductance of the
single channel inward current was 37 pS (140 mM
K+o), and outward current channel events were also
observed. The weak rectification of sWIRK is significant in that it has
a negatively charged residue (glutamate) in the M2 region which is
reported to cause strong inward rectification. By introducing a point
mutation to remove this negative charge (glutamine), the sWIRK E179Q
mutant channel lost its inward rectification property completely, and
the single channel property (45 pS; 140 mM
K+o) was ohmic up to highly depolarized potential,
even in the presence of the physiological cytoplasmic blockers such as
Mg2+ and polyamines.
Volume 271, Number 26,
Issue of June 28, 1996
pp. 15729-15735
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
GLUTAMATE 179 IN THE SECOND TRANSMEMBRANE DOMAIN IS INSUFFICIENT
FOR STRONG RECTIFICATION
,
and
Department of Neurophysiology, Tokyo
Metropolitan Institute for Neuroscience, Musashidai 2-6, Fuchu,
Tokyo 183, Japan and the ¶ Department of Molecular Biology, Ocean
Research Institute, University of Tokyo, Minamidai 1-15-1, Nakano,
Tokyo 164, Japan
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