A molecular lid mechanism for K+ channel blockers revealed by a cone peptide

Many venomous organisms carry in their arsenal short peptides that block K+ channels in a highly selective manner. These toxins may compete with the permeating ions directly via a "plug" mechanism or indirectly via a "pore-collapse" mechanism. An alternative "lid" mechanism was proposed, but remains poorly defined. Here we study the block of the Drosophila Shaker channel by Conknunitzin-S1 and Conkunitzin-C3, two highly similar toxins derived from cone venom. Despite their similarity, the two peptides exhibited differences in their binding poses and in biophysical assays, implying discrete modes of action. We show that while Conknunitzin-S1 binds tightly to the channel turret and acts via a "pore-collapse" mechanism, Conkunitzin-C3 does not contact this region and use a non-conserved Arg to divert the permeant ions and trap them in off-axis cryptic sites above the SF, a mechanism we term a "molecular-lid". Our study provides an atomic description of the "lid" K+ blocking mode and offers valuable insights for the design of therapeutics based on venom peptides.