Crystallographic insights into sodium-channel modulation by the β4 subunit

Abstract

Voltage-gated sodium (Na v ) channels are embedded in a multicomponent membrane signaling complex that plays a crucial role in cellular excitability. Although the mechanism remains unclear, β-subunits modify Na v channel function and cause debilitating disorders when mutated. While investigating whether β-subunits also influence ligand interactions, we found that β4 dramatically alters toxin binding to Na v 1.2. To explore these observations further, we solved the crystal structure of the extracellular β4 domain and identified 58 Cys as an exposed residue that, when mutated, eliminates the influence of β4 on toxin pharmacology. Moreover, our results suggest the presence of a docking site that is maintained by a cysteine bridge buried within the hydrophobic core of β4. Disrupting this bridge by introducing a β1 mutation implicated in epilepsy repositions the 58 Cys-containing loop and disrupts β4 modulation of Na v 1.2. Overall, the principles emerging from this work (i) help explain tissuedependent variations in Na v channel pharmacology; (ii) enable the mechanistic interpretation of β-subunit-related disorders; and (iii) provide insights in designing molecules capable of correcting aberrant β-subunit behavior.voltage-gated sodium channel | beta4 subunit | ProTx-II | X-ray structure | disease mutations

Pharmacology of the Na v 1.1 domain IV voltage sensor reveals coupling between inactivation gating processes

“…To test whether shifts in SSI are a common feature of VSDIVbinding toxins, we examined the effects of LqqIV, a VSDIV-binding α-scorpion toxin (22,23) using this same protocol. Similar to Hm1a, LqqIV shifted the SSI curve to more positive potentials (V 1/2 = 67 ± 3 mV) (Fig.…”

“…Statistical analysis was performed as appropriate, and data are reported in the figure legends or text. To avoid functional interference of auxiliary proteins (23), Na v channel gating properties were investigated in oocytes without the presence of β subunits.…”

Voltage-Gated Sodium Channels

“…In this Chapter, we provide a comprehensive survey of the available X-ray crystal data and accompanying simulation studies that have been undertaken to discern the fundamental features regulating permeation and selectivity in Na + channels, with particular focus on voltage-gated sodium (Na V ) channels. The atomic structures of associated β-subunits have recently been determined (Gilchrist, Das, Van Petegem, & Bosmans, 2013;Namadurai et al, 2014), yet in this review, the focus is on the principal subunits (designated α or α1) which are responsible for the voltage-gated ion conductance which is the defining characteristic of these proteins.…”

Disease Caused by Mutations in NaV-β Subunit Genes

“…1). 1,2 Na V -b subunits interact with all known voltagegated sodium channel isoforms, altering their expression and function. [3][4][5][6] Recent studies have found that Na V -b subunits are also capable of shaping the channel pharmacology with crucial therapeutic implications.…”

“…[3][4][5][6] Recent studies have found that Na V -b subunits are also capable of shaping the channel pharmacology with crucial therapeutic implications. 2,7,8 Five Na V -b subunits have been identified in humans: b1, b1b, b2, b3, and b4, encoded by SCN1B, SCN2B, SCN3B, and SCN4B genes, respectively. All Na V -b subunits share similar topology (Fig.…”