Modulatory features of the novel spider toxin μ‐TRTX‐Df1a isolated from the venom of the spider Davus fasciatus

Background and Purpose Naturally occurring dysfunction of voltage‐gated sodium (Na V ) channels results in complex disorders such as chronic pain, making these channels an attractive target for new therapies. In the pursuit of novel Na V modulators, we investigated spider venoms for new inhibitors of Na V channels. Experimental Approach We used high‐throughput screens to identify a Na V modulator in venom of the spider Davus fasciatus . Further characterization of this venom peptide was undertaken using fluorescent and electrophysiological assays, molecular modelling and a rodent pain model. Key Results We identified a potent Na V inhibitor named μ‐TRTX‐Df1a. This 34‐residue peptide fully inhibited responses mediated by Na V 1.7 endogenously expressed in SH‐SY5Y cells. Df1a also inhibited voltage‐gated calcium (Ca V 3) currents but had no activity against the voltage‐gated potassium (K V 2) channel. The modelled structure of Df1a, which contains an inhibitor cystine knot motif, is reminiscent of the Na V channel toxin ProTx‐I. Electrophysiology revealed that Df1a inhibits all Na V subtypes tested (hNa V 1.1–1.7). Df1a also slowed fast inactivation of Na V 1.1, Na V 1.3 and Na V 1.5 and modified the voltage‐dependence of activation and inactivation of most of the Na V subtypes. Df1a preferentially binds to the domain II voltage‐sensor and has additional interactions with the voltage sensors domains III and IV, which probably explains its modulatory features. Df1a was analgesic in vivo, reversing the spontaneous pain behaviours induced by the Na V activator OD1. Conclusion and Implications μ‐TRTX‐Df1a shows potential as a new molecule for the development of drugs to treat pain disorders mediated by voltage‐gated ion channels.