Variation of Two S3b Residues in K4.1-4.3 Channels Underlies Their Different Modulations by Spider Toxin κ-LhTx-1.

The naturally occurred peptide toxins from animal venoms are valuable pharmacological tools in exploring the structure-function relationships of ion channels. Herein we have identified the peptide toxin κ-LhTx-1 from the venom of spider (the Lichen huntsman spider) as a novel selective antagonist of the K4 family potassium channels. κ-LhTx-1 is a gating-modifier toxin impeded K4 channels' voltage sensor activation, and mutation analysis has confirmed its binding site on channels' S3b region. Interestingly, κ-LhTx-1 differently modulated the gating of K4 channels, as revealed by toxin inhibiting K4.2/4.3 with much more stronger voltage-dependence than that for K4.1. We proposed that κ-LhTx-1 trapped the voltage sensor of K4.1 in a much more stable resting state than that for K4.2/4.3 and further explored the underlying mechanism. Swapping the non-conserved S3b segments between K4.1(FVPK) and K4.3(VMTN) fully reversed their voltage-dependence phenotypes in inhibition by κ-LhTx-1, and intensive mutation analysis has identified P282 in K4.1, D281 in K4.2 and N278 in K4.3 being the key residues. Furthermore, the last two residues in this segment of each K4 channel (P282/K283 in K4.1, T280/D281 in K4.2 and T277/N278 in K4.3) likely worked synergistically as revealed by our combinatorial mutations analysis. The present study has clarified the molecular basis in K4 channels for their different modulations by κ-LhTx-1, which have advanced our understanding on K4 channels' structure features. Moreover, κ-LhTx-1 might be useful in developing anti-arrhythmic drugs given its high affinity, high selectivity and unique action mode in interacting with the K4.2/4.3 channels.