A new cysteine framework in sodium channel blocking conotoxins

Two novel sodium channel blocking peptides from the venom of the molluscivorous snail Conus pennaceus, muPnIVA and muPnIVB, are described. Elucidation of their amino acid sequences was complicated by a previously undescribed anomalous product of reduction and pyridylethylation, which occurs on N-terminal cysteine residues and gives a PTH derivative eluting at the same position as PTH-Trp in reverse-phase chromatography. The amino acid sequences of the toxins were determined by a combination of Edman degradation and mass-spectrometric techniques as CCKYGWTCLLGCSPCGC (PnIVA) and CCKYGWTCWLGCSPCGC (PnIVB). These toxins block sodium channels in molluscan neurons, but have no effect on sodium currents in bovine chromaffin cells or in rat brain synaptosomes. Although there is only one amino acid difference in the two sequences, PnIVB is approximately 6 times more potent than PnIVA in blockade of the sodium current in Lymnaea neurons. The PnIV sequences reveal a new cysteine residue framework for conotoxins (CC-----C---C--C-C). Strikingly, the only charged residue in PnIVA/B is Lys3. Iodination reaction experiments on the adjacent Tyr4 suggest that this region of the peptide must be solvent exposed and essential for activity. These structurally novel mu-conotoxins target a sodium channel subtype with low affinity for tetrodotoxin and therefore provide new probes for functional studies on sodium channels.