Manipulation of a spider peptide toxin alters its affinity for lipid bilayers and potency and selectivity for voltage-gated sodium channel subtype


Journal of Biological Chemistry


Akello J. Agwa, Poanna Tran, Alexander Mueller, Hue N. T. Tran, Jennifer R. Deuis, Mathilde R. Israel, Kirsten L. McMahon, David J. Craik, Irina Vetter and Christina I. Schroeder



Huwentoxin-IV (HwTx-IV) is a gating modifier peptide toxin from spiders that has weak affinity for the lipid bilayer. As some gating modifier toxins have affinity for model lipid bilayers, a tripartite relationship among gating modifier toxins, voltage-gated ion channels and the lipid membrane surrounding the channels has been proposed. We previously designed an HwTx-IV analog (gHwTx-IV) with reduced negative charge and increased hydrophobic surface profile, which displays increased lipid bilayer affinity and in vitro activity at the voltage-gated sodium channel subtype 1.7 (NaV1.7), a channel targeted in pain management. Here, we show that replacements of the positively charged residues that contribute to the activity of the peptide can improve gHwTx-IV’s potency and selectivity for NaV1.7. Using HwTx-IV, gHwTx- IV,
[R26A]gHwTx-IV, [K27A]gHwTx-IV, and [R29A]gHwTx-IV variants, we examined their potency and selectivity at human NaV1.7 and their affinity for the lipid bilayer. [R26A]gHwTxIV consistently displayed the most improved potency and selectivity for NaV1.7, examined alongside off-target NaVs, compared with HwTxIV and gHwTx-IV. The lipid affinity of each of the three novel analogs was weaker than that of gHwTx-IV, but stronger than that of HwTx-IV, suggesting a possible relationship between in vitro potency at NaV1.7 and affinity for lipid bilayers. In a murine NaV1.7 engagement model, [R26A]gHwTx-IV exhibited an efficacy comparable to that of native HwTx- IV. In summary, the present study reports the development of an HwTx-IV analog with improved in vitro selectivity for the pain target NaV1.7 and with an in Vivo efficacy similar to that of native HwTx-IV.

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