Abstract
Venoms from cone snails and arachnids are a rich source of peptide modulators of voltage-gated sodium (NaV) channels, however relatively few venom-derived peptides with activity at the mammalian NaV1.8 subtype have been isolated. Here, we describe the discovery and functional characterisation of β-theraphotoxin-Eo1a, a peptide from the venom of the Tanzanian black and olive baboon tarantula Encyocratella olivacea that modulates NaV1.8. Eo1a is a 37-residue peptide that increases NaV1.8 peak current (EC50 894 ± 146 nM) and causes a large hyperpolarising shift in both the voltage-dependence of activation (ΔV50–20.5 ± 1.2 mV) and steady-state fast inactivation (ΔV50–15.5 ± 1.8 mV). At a concentration of 10 μM, Eo1a has varying effects on the peak current and channel gating of NaV1.1–NaV1.7, although its activity is most pronounced at NaV1.8. Investigations into the binding site of Eo1a using NaV1.7/NaV1.8 chimeras revealed a critical contribution of the DII S3-S4 extracellular loop of NaV1.8 to toxin activity. Results from this work may form the basis for future studies that lead to the rational design of spider venom-derived peptides with improved potency and selectivity at NaV1.8.
Highlights
Voltage-gated sodium (NaV) channels are pore-forming transmembrane proteins that permit the influx of Na+ ions across cell membranes
The macroscopic current is a function of the total number of channels, the single channel current, and the open probability (Sigworth, 1980), and if we assume that the former two parameters remain constant, it is most likely that Eo1a increases the open probability of NaV1.8, this remains to be experimentally confirmed by singlechannel recordings
Eo1a is the most selective activator of NaV1.8 described to date; our data indicates that Eo1a has some off-target activity, which limits its use as a pharmacological tool to selectively activate NaV1.8 in native neurons
Summary
Voltage-gated sodium (NaV) channels are pore-forming transmembrane proteins that permit the influx of Na+ ions across cell membranes As such, they are responsible for the rising phase of action potentials and essential for regulating the excitability of neuronal, cardiac and skeletal muscle cells. While small molecules with NaV1.8 selectivity, such as A-803467 and PF-01247324, have been described (Jarvis et al, 2007; Payne et al, 2015), they are state-dependent inhibitors that bind to DIV preferentially in the inactivated state. As it is unclear how physiologically relevant this state is in vivo, more pharmacological tools are required to understand the contribution of NaV1.8 to sensory neuron function
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