Structure and Function of the Voltage Sensor of Sodium Channels Probed by a β-Scorpion Toxin

Sandrine Cestèle,Vladimir Yarov-Yarovoy,Yusheng Qu,François Sampieri,Todd Scheuer,William A Catterall

doi:10.1074/jbc.m603814200

Sandrine Cestèle, Vladimir Yarov-Yarovoy + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m603814200

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Abstract

Voltage sensing by voltage-gated sodium channels determines the electrical excitability of cells, but the molecular mechanism is unknown. beta-Scorpion toxins bind specifically to neurotoxin receptor site 4 and induce a negative shift in the voltage dependence of activation through a voltage sensor-trapping mechanism. Kinetic analysis showed that beta-scorpion toxin binds to the resting state, and subsequently the bound toxin traps the voltage sensor in the activated state in a voltage-dependent but concentration-independent manner. The rate of voltage sensor trapping can be fit by a two-step model, in which the first step is voltage-dependent and correlates with the outward gating movement of the IIS4 segment, whereas the second step is voltage-independent and results in shifted voltage dependence of activation of the channel. Mutations of Glu(779) in extracellular loop IIS1-S2 and both Glu(837) and Leu(840) in extracellular loop IIS3-S4 reduce the binding affinity of beta-scorpion toxin. Mutations of positively charged and hydrophobic amino acid residues in the IIS4 segment do not affect beta-scorpion toxin binding but alter voltage dependence of activation and enhance beta-scorpion toxin action. Structural modeling with the Rosetta algorithm yielded a three-dimensional model of the toxin-receptor complex with the IIS4 voltage sensor at the extracellular surface. Our results provide mechanistic and structural insight into the voltage sensor-trapping mode of scorpion toxin action, 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.

Highlights

Able cells [2] and are the molecular target for several groups of neurotoxins, which bind to different receptor sites and alter voltage-dependent activation, conductance, and inactivation [3, 4]
A Two-step Mechanism for Binding and Voltage Sensor Trapping by Css IV Toxin—Incubation of tsA-201 cells expressing sodium channels with Css IV toxin at a negative membrane potential has no effect on sodium channel function, but strong depolarization to fully activate sodium channels in the presence of Css IV toxin strongly enhances the activation process by shifting its voltage dependence negatively by 32 mV [34]
Based on the voltage sensor-trapping model [34], we hypothesized that this process involves a two-step mechanism of toxin action: first binding to sodium channels without functional effect on activation at the resting membrane potential followed by trapping the IIS4 voltage sensor in its activated conformation upon channel activation in the presence of bound toxin

Summary

Introduction

Able cells [2] and are the molecular target for several groups of neurotoxins, which bind to different receptor sites and alter voltage-dependent activation, conductance, and inactivation [3, 4]. These results suggest that Css IV binds to these two with the more rapid rate constant for IIS4 movement, because extracellular loops in domain II through a combination of electhe rate of voltage-dependent activation of voltage sensor trap- trostatic and hydrophobic interactions with the negatively ping in our experiments at 20 mV and 24 °C (7 msϪ1) correlates charged and hydrophobic side chains of these amino acid well with the rate of movement of the IIS4 segment at 20 mV residues.

Results

Conclusion