Abstract

Dofetilide, a methanesulfonanilide derivative, is a potent class III antiarrhythmic drug. Like other members of this class of K+ channel blockers, the sites in the channel to which the drug binds are unknown, although high and low affinity binding has been reported in cardiomyocytes. The most sensitive K+ channel target for dofetilide seems to be IKr, the rapid component of the repolarizing delayed rectifier K+ current. However, block of other K+ channels occurs at higher concentrations and is of special interest in regard to toxicity. Recently, we have demonstrated that hIRK, a cloned inward rectifier K+ channel (IRK) isolated from human atrium and expressed heterologously in Xenopus oocytes, is blocked by dofetilide. We report the localization of a site that is critical for dofetilide block in hIRK. We used chimeric constructs between hIRK and ROMK1, a related inward rectifier that is drug resistant. Substitution of hIRK-M2, the second putative transmembrane spanning segment of IRKs, with ROMK1-M2 increased unblocking of dofetilide by 10-20-fold in hIRK. Site-directed mutagenesis further pinpointed the effects to a single hydrophobic residue (I177) in M2. A reduction in hydrophobicity by the point mutation I177C increased recovery from block > 10-fold (1.17 sec in wild-type to 0.112 sec at -80 mV at physiological K+ concentrations), leading us to suggest that hydrophobic interactions are essential for dofetilide block in hIRK. A similar mechanism may explain dofetilide block in other ion channels, including IKr.