Blockade of a KCa channel with synthetic peptides from noxiustoxin: a K+ channel blocker.

Using the outside-out configuration of the patch-clamp method, we studied the effect of several synthetic peptides corresponding to various segments from the N-terminal region of noxiustoxin (NTX) on single Ca(2+)-activated K+ (KCa) channels of small conductance obtained from cultured bovine aortic endothelial cells. These peptides induced diverse degrees of fast blockade in the endothelial KCa channel. The most effective blockers were the peptides NTX1-39 (IC50 = 0.5 microM) and NTX1-20 comprising the first 20 amino acids from the native toxin (IC50 approximately 5 microM), while less effective was the hexapeptide NTX1-6, from the first six amino acid residues of NTX (IC50 = 500 microM). This was the minimum sequence required to block the channel. By testing overlapping sequences from the entire molecule, specially those corresponding to the N-terminal region of NTX, we have been able to determine their different apparent affinities for the KCa channel. Synthetic peptides from the C-terminal region produced no effect on the KCa channel at the concentrations tested (up to 1 mM). These results confirm that in the N-terminal region of the NTX is located part of the sequence that may recognize K+ channels, as we have suggested previously from in vivo experiments. The blockade induced by native NTX was poorly affected by changes in membrane potential; however, the blockage induced by synthetic peptides lacking the C-terminal region was partially released by depolarization.