RT Journal Article
SR Electronic
T1 Novel Site on Sodium Channel α-Subunit Responsible for the Differential Sensitivity of Grayanotoxin in Skeletal and Cardiac Muscle
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 865
OP 872
VO 60
IS 4
A1 Takahiro Kimura
A1 Kaoru Yamaoka
A1 Eiji Kinoshita
A1 Hiroshi Maejima
A1 Tsunetsugu Yuki
A1 Masuhide Yakehiro
A1 Issei Seyama
YR 2001
UL http://molpharm.aspetjournals.org/content/60/4/865.abstract
AB We searched for sites on the α-subunit of the fast Na+channel responsible for the difference in GTX (grayanotoxin) sensitivity of the skeletal- and cardiac-muscle Na+current. cDNA clones, encoding the skeletal or cardiac isoforms of the α-subunit, were inserted into a mammalian expression vector and transiently transfected into human embryonic kidney cells. The expressed channels were measured using whole-cell patch-clamp techniques and examined for GTX sensitivity. As a measure of GTX sensitivity, we used relative chord conductance (ratio of maximum chord conductance of noninactivating GTX-modified Na+ currents to that of unmodified peak currents). Wild-type channels from skeletal muscle (μ1) were more sensitive to GTX modification than wild-type cardiac channels (rH1) by a factor of 1.6. To facilitate exploration of α-subunit sites determining GTX sensitivity, we used SHHH, a chimera of skeletal muscle (S) domain D1 and heart muscle (H) domains D2D3D4 with supernormal sensitivity to GTX I (1.5-fold of wild-type μ1). Successive replacement of Ser-251 (D1S4–S5 intracellular loop) and Ile-433 (D1S6 transmembrane segment), with corresponding rH1 residues Ala and Val, reduced, in a stepwise manner, the GTX sensitivity of the chimera and related mutants to that of wild-type rHl. We concluded that, in addition to Ile-433, known as the GTX-binding site, Ser-251 represents a novel site for GTX modification. The American Society for Pharmacology and Experimental Therapeutics