Electrostatic interaction between charybdotoxin and a tetrameric mutant of Shaker K(+) channels.

The scorpion toxin, Charybdotoxin (CTX), blocks homotetrameric, voltage-gated K(+) channels by binding near the outer entrance to the pore in one of four indistinguishable orientations. We have determined the pH-dependence of CTX block of a tetrameric Shaker potassium channel with a single copy of a histidine replacing the wild-type phenylalanine at position 425. We compared this pH-dependence with that from homotetrameric channels with four copies of the mutation. We found that protonation of a single amino acid at position 425 had a large effect on the affinity of the channel for CTX-much larger than expected if only one of the four CTX binding orientations was disrupted. The pK(a) for the H(+)-ion induced protection from CTX block indicates that the electrostatic environment near position 425 is likely basic in nature, perhaps because of the proximity of lysine 427. We also examined the pH-dependence of block of channels with one and four copies of the histidine mutation by CTX containing neutralizing mutations of four basic residues on the active face of the toxin. The results suggested an orientation of CTX on the channel that places three of the positively charged CTX residues very near three of the four Shaker 425 positions.