Multiple, Distributed Interactions of μ-Conotoxin PIIIA Associated with Broad Targeting among Voltage-Gated Sodium Channels

The first mu-conotoxin studied, mu CTX GIIIA, preferentially blocked voltage-gated skeletal muscle sodium channels, Na(v)1.4, while mu CTX PIIIA was the first to show significant blocking action against neuronal voltage-gated sodium channels. PIIIA shares > 60% sequence identity with the well-studied GIIIA, and both toxins preferentially block the skeletal muscle sodium channel isoform. Two important features of blocking by wild-type GIIIA are the toxin's high binding affinity and the completeness of block of a single channel by a bound toxin molecule. With GIIIA, neutral replacement of the critical residue, Arg-13, allows a residual single-channel current (similar to 30% of the unblocked, unitary amplitude) when the mutant toxin is bound to the channel and reduces the binding affinity of the toxin for Na(v)1.4 (similar to 100-fold) [Becker, S., et al. (1992) Biochemistry 31, 8229-8238]. The homologous residue in PIIIA, Arg-14, is also essential for completeness of block but less important in the toxin's binding affinity (similar to 55% residual current and similar to 11-fold decrease in affinity when substituted with alanine or glutamine). The weakened dominance of this key arginine in PIIIA is also seen in the fact that there is not just one (R13 in GIIIA) but three basic residues (R12, R14, and K17) for which individual neutral replacement enables a substantial residual current through the bound channel. We suggest that, despite a high degree of sequence conservation between GIIIA and PIIIA, the weaker dependence of PIIIA's action on its key arginine and the presence of a nonconserved histidine near the C-terminus may contribute to the greater promiscuity of its interactions with different sodium channel isoforms.