Voltage Sensor of Sodium Channels Probed by a β-Scorpion Toxin

Voltage sensing by voltage-gated sodium channels determines the electrical excitability of cells, but the molecular mechanism is unknown. ß-scorpion toxins bind specifically to neurotoxin receptor site 4 and induce a negative shift in the voltage dependence of activation through a voltagesensor trapping mechanism. Kinetic analysis showed that β-scorpion toxin binds to the resting state and subsequently traps the voltage sensor in the activated state in a voltagedependent, but concentration-independent manner. Mutations of E779 in extracellular loop IIS1-S2 and both E837 and L840 in extracellular loop IIS3-S4 reduce the binding affinity of ß-scorpion toxin. Mutations of positively charged and hydrophobic amino acid residues in the IIS4 segment do not affect βscorpion toxin binding, but these mutations alter the voltage dependence of activation and enhance ß-scorpion toxin action. Molecular modeling with the Rosetta program and docking of β-scorpion toxin yielded a threedimensional model of the toxin-receptor complex with the IIS4 voltage sensor, which is formed at the extracellular surface of the protein. Our results define the position of the voltage sensor in the resting state of the sodium channel and favor voltage-sensing models in which the S4 segment spans the membrane in both resting and activated states.