Received for publication April 23, 2004.
Revised August 10, 2004.
Accepted for publication August 11, 2004.

The low potency, voltage-dependent HERG blocker propafenone - molecular determinants and drug trapping

Abstract

The molecular determinants of high-affinity HERG potassium channel blockade by methanesulfonanilides include two aromatic residues (F656 and Y652) on the inner helices (S6) and residues on the pore helices that face into the inner cavity, but determinants for lower-affinity HERG-blockers may be different. In this study, alanine-substituted HERG channel mutants of inner cavity residues were expressed in Xenopus oocytes, and were used to characterise the HERG channel binding-site of the antiarrhythmic propafenone. Propafenone's blockade of HERG was strongly dependent on residue F656, but was insensitive or weakly sensitive to mutation of Y652, T623, S624, V625, G648, or V659, nor did it require functional inactivation. Homology models of HERG based on KcsA and MthK crystal structures, representing the closed and open forms of the channel, respectively, suggest propafenone is trapped in the inner cavity and is unable to interact exclusively with F656 in the closed state (whereas exclusive interactions between propafenone and F656 are found in the open-channel model). These findings are supported by very slow recovery of wild-type HERG channels from block at -120 mV, but extremely rapid recovery of D540K channels that re-open at this potential. The experiments and modelling suggest the open-state propafenone binding-site may be formed by the F656 residues alone. The binding-site for propafenone (which may involve pi-stacking interactions with two or more F656 side-chains) is either perturbed or becomes less accessible due to closed-channel gating. This adds further evidence for the existence of gating-induced changes in the spatial location of F656 side-chains.

Key words: Potassium, Antiarrhythmic drugs, Structure-activity relationships and modeling, Func. analysis receptor/ion channel mutants