The β3‐subunit modulates the effect of venom peptides ProTx‐II and OD1 on Na_V1.7 gating

Abstract

The voltage‐gated sodium channel NaV1.7 is involved in various pain phenotypes and is physiologically regulated by the NaV‐β3‐subunit. Venom toxins ProTx‐II and OD1 modulate NaV1.7 channel function and may be useful as therapeutic agents and/or research tools. Here, we use patch‐clamp recordings to investigate how the β3‐subunit can influence and modulate the toxin‐mediated effects on NaV1.7 function, and we propose a putative binding mode of OD1 on NaV1.7 to rationalise its activating effects. The inhibitor ProTx‐II slowed the rate of NaV1.7 activation, whilst the activator OD1 reduced the rate of fast inactivation and accelerated recovery from inactivation. The β3‐subunit partially abrogated these effects. OD1 induced a hyperpolarising shift in the V1/2 of steady‐state activation, which was not observed in the presence of β3. Consequently, OD1‐treated NaV1.7 exhibited an enhanced window current compared with OD1‐treated NaV1.7‐β3 complex. We identify candidate OD1 residues that are likely to prevent the upward movement of the DIV S4 helix and thus impede fast inactivation. The binding sites for each of the toxins and the predicted location of the β3‐subunit on the NaV1.7 channel are distinct. Therefore, we infer that the β3‐subunit influences the interaction of toxins with NaV1.7 via indirect allosteric mechanisms. The enhanced window current shown by OD1‐treated NaV1.7 compared with OD1‐treated NaV1.7‐β3 is discussed in the context of differing cellular expressions of NaV1.7 and the β3‐subunit in dorsal root ganglion (DRG) neurons. We propose that β3, as the native binding partner for NaV1.7 in DRG neurons, should be included during screening of molecules against NaV1.7 in relevant analgesic discovery campaigns.

Isoform-specific N-linked glycosylation of voltage-gated sodium channel alpha-subunits alters beta-subunit binding sites

“…Indeed, the DIV VSD of Nav1.7 has been shown to confer isoform-specific targeting of Nav1.7 and is also the binding site of venom toxins (e.g. OD1) specific to Nav1.7 (Kschonsak et al, 2023; Salvage et al, 2023c; Jalali et al, 2005).…”

“…Indeed, the DIV VSD of Nav1.7 has been shown to confer isoform-specific targeting of Nav1.7 and is also the binding site of venom toxins (e.g. OD1) specific to Nav1.7 (Kschonsak et al, 2023;Salvage et al, 2023c;Jalali et al, 2005). Therefore, alongside the electrostatic and structural effects, further investigation into the roles that glycans play in modulating interactions between Nav channels and other binding partners may reveal new insights into higher order complexes at the cell surface, coordination of intracellular signaling pathways, and site-specific drug target selection.…”

Sodium Channel β Subunits—An Additional Element in Animal Tetrodotoxin Resistance?

“…Interestingly, the activity of certain toxins from spiders (ProTx-II from the velvet tarantula Thrixopelma pruriens), scorpions (OD1 from the yellow Iranian scorpion Odontobuthus doriae), and cone snails (µ-and µO-conotoxins) against Na v 1.2, Na v 1.6, and/or Na v 1.7 appears to be reduced in the presence of certain β subunits but enhanced by others [31][32][33][34][35]. While ProTx-II, OD1, and µO-conotoxins are gating modifiers and therefore more likely to be affected by β subunits (which are themselves known to regulate channel gating) [34,36], µ-conotoxins are pore blockers like TTX [37], thus supporting the influence of β subunits on pore-blocking properties. However, the binding site of TTX within the channel pore differs from (although it overlaps with) that of µ-conotoxins [6], which likely explains why TTX itself and the nearly identical saxitoxin (STX) were seemingly unaffected by the presence of β subunits [14,31].…”