Increasing the molecular contacts between maurotoxin and Kv1.2 channel augments ligand affinity

Scorpion toxins interact with their target ion channels through multiple molecular contacts. Because a "gain of function" approach has never been described to evaluate the importance of the molecular contacts in defining toxin affinity, we experimentally examined whether increasing the molecular contacts between a toxin and an ion channel directly impacts toxin affinity. For this purpose, we focused on two scorpion peptides, the well-characterized maurotoxin with its variant Pi-like disulfide bridging (MTXPi1), used as a molecular template, and butantoxin (BuTX), used as an N-terminal domain provider. BuTX is found to be 60-fold less potent than MTXPi1 in blocking Kv1.2 (IC50 values of 165 nM for BuTX versus 2.8 nM for MTXPi1). Removal of its N-terminal domain (nine residues) further decreases BuTX affinity for Kv1.2 by 5.6-fold, which is in agreement with docking simulation data showing the importance of this domain in BuTX-Kv1.2 interaction. Transfer of the BuTX N-terminal domain to MTXPi1 results in a chimera with five disulfide bridges (BuTX-MTXPi1) that exhibits 22-fold greater affinity for Kv1.2 than MTXPi1 itself, in spite of the lower affinity of BuTX as compared to MTXPi1. Docking experiments performed with the 3-D structure of BuTX-MTXPi1 in solution, as solved by 'H-NMR, reveal that the N-terminal domain of BuTX participates in the increased affinity for Kv1.2 through additional molecular contacts. Altogether, the data indicate that acting on molecular contacts between a toxin and a channel is an efficient strategy to modulate toxin affinity.