RT Journal Article
SR Electronic
T1 Molecular Determinants of Inactivation and Dofetilide Block in<em>ether a-go-go</em> (EAG) Channels and EAG-Related K<sup>+</sup> Channels
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 1343
OP 1348
DO 10.1124/mol.60.6.1343
VO 60
IS 6
A1 Eckhard Ficker
A1 Wolfgang Jarolimek
A1 Arthur M. Brown
YR 2001
UL http://molpharm.aspetjournals.org/content/60/6/1343.abstract
AB The major subunit of the cardiac delayed rectifier current IKr is encoded by the human ether a-go-gorelated gene (HERG). HERG/IKr channels are blocked selectively by class III antiarrhythmic methanesulfonanilide drugs such as dofetilide. The binding site for methanesulfonanilides is believed to be similar for nonantiarrhythmic drugs such as antihistamines, antibiotics, and antipsychotics. To gain further insight into the binding site, we examined the minimal structural changes necessary to transform low-affinity binding of dofetilide by the related bovine ether a-go-go channel bEAG to high-affinity binding of HERG. Previously, it was shown that high-affinity binding in HERG required intact C-type inactivation; the bovine ether a-go-go K+ channel (bEAG), unlike HERG, is noninactivating. Therefore, we introduced C-type inactivation into noninactivating bEAG using site-directed mutagenesis. Two point mutations in the pore region, T432S and A443S, were sufficient to produce C-type inactivation. Low concentrations of dofetilide produced block of bEAG T432S/A443S; unlike HERG, block was almost irreversible. Substitution of an additional amino acid in transmembrane domain S6 made the block reversible. Dofetilide blocked the triply mutated bEAG T432S/A443S/A453S with an IC50value of 1.1 μM. The blocking potency was 30-fold greater than bEAG WT and about one third that of HERG WT. We conclude that high affinity methanesulfonanilide binding to HERG channels is strongly dependent on C-type inactivation.