Gene expression, mutation, and structure-function relationship of scorpion toxin BmP05 active on SKCa channels

Four peptide inhibitors of small-conductance Ca2+-activated, apamin-sensitive K+ channels (SKCa) have been isolated from the venom of the Chinese scorpion Buthus martensi, named BmP01, BmP02, BmP03, and BmP05, respectively [Romi-Lebrun, R. (1997) Eur. J. Biochem. 245, 457-464]. Among them BmP05 with 31 amino acid residues has been intensively studied due to its most potent toxicity. To investigate the structure-function relationship of BmP05, its wild type and seven mutants (their C-termini unamidated) were successfully expressed in the yeast secretion system and purified with a high yield over 8 mg/L. Their toxicity to mice and electrophysiological activity on the K+ currents (SKCa and Kv) in rat adrenal chromaffin cells were measured and compared. The results indicated the following: (1) As a selective antagonist against SKCa, 1 muM rBmP05 is equivalent to 0.2 muM apamin, and its IC50 is 0.92 muM. (2) The basic residues Lys and Arg located at positions 6 and 13 in the N-terminal alpha-helix region are essential and synergetic in the interaction of the toxin with SKCa. (3) Disruption of the alpha-helix by mutation of Gln at position 9 with Pro results in almost total loss of toxicity. (4) The C-terminal residue His31 plays an auxiliary role in the interaction of the toxin with SKCa. (5) The beta-turn connecting two beta-sheets near the C-terminal part is responsible for the specificity of the toxin to the different subtypes of K+ channels.