An allosteric mechanism for drug block of the human cardiac potassium channel KCNQ1

Mol Pharmacol. 2013 Feb;83(2):481-9. doi: 10.1124/mol.112.081513. Epub 2012 Nov 28.

Abstract

The intracellular aspect of the sixth transmembrane segment within the ion-permeating pore is a common binding site for many voltage-gated ion channel blockers. However, the exact site(s) at which drugs bind remain controversial. We used extensive site-directed mutagenesis coupled with molecular modeling to examine mechanisms in drug block of the human cardiac potassium channel KCNQ1. A total of 48 amino acid residues in the S6 segment, S4-S5 linker, and the proximal C-terminus of the KCNQ1 channel were mutated individually to alanine; alanines were mutated to cysteines. Residues modulating drug block were identified when mutant channels displayed <50% block on exposure to drug concentrations that inhibited wild-type current by ≥90%. Homology modeling of the KCNQ1 channel based on the Kv1.2 structure unexpectedly predicted that the key residue modulating drug block (F351) faces away from the permeating pore. In the open-state channel model, F351 lines a pocket that also includes residues L251 and V254 in S4-S5 linker. Docking calculations indicated that this pocket is large enough to accommodate quinidine. To test this hypothesis, L251A and V254A mutants were generated that display a reduced sensitivity to blockage with quinidine. Thus, our data support a model in which open state block of this channel occurs not via binding to a site directly in the pore but rather by a novel allosteric mechanism: drug access to a side pocket generated in the open-state channel configuration and lined by S6 and S4-S5 residues.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alanine / genetics
  • Alanine / metabolism
  • Allosteric Regulation / drug effects*
  • Animals
  • Binding Sites
  • CHO Cells
  • Cell Line
  • Cricetinae
  • Cysteine / genetics
  • Cysteine / metabolism
  • Heart / drug effects*
  • Humans
  • KCNQ1 Potassium Channel / antagonists & inhibitors*
  • KCNQ1 Potassium Channel / genetics
  • KCNQ1 Potassium Channel / metabolism*
  • Models, Molecular
  • Mutagenesis, Site-Directed / methods
  • Myocardium / metabolism*
  • Quinidine / metabolism

Substances

  • KCNQ1 Potassium Channel
  • KCNQ1 protein, human
  • Quinidine
  • Cysteine
  • Alanine