Neuronally selective μ-conotoxins from Conus striatus utilize an α-helical motif to target mammalian sodium channels

Christina I. Schroeder, Jenny Ekberg, Katherine J. Nielsen, Denise Adams, Marion L. Loughnan, Linda Thomas, David J. Adams, Paul F. Alewood, Richard J. Lewis

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Abstract

μ-Conotoxins are small peptide inhibitors of muscle and neuronal tetrodotoxin (TTX)-sensitive voltage-gated sodium channels (VGSCs). Here we report the isolation of μ-conotoxins SIIIA and SIIIB by 125I- TIIIA-guided fractionation of milked Conus striatus venom. SIIIA and SIIIB potently displaced 125I-TIIIA from native rat brain Nav1.2 (IC50 values 10 and 5 nM, respectively) and muscle Nav1.4 (IC50 values 60 and 3 nM, respectively) VGSCs, and both inhibited current through Xenopus oocyte-expressed Nav1.2 and Na v1.4. An alanine scan of SIIIA-(2-20), a pyroglutamate-truncated analogue with enhanced neuronal activity, revealed residues important for affinity and selectivity. Alanine replacement of the solvent-exposed Trp-12, Arg-14, His-16, Arg-18 resulted in large reductions in SIIIA-(2-20) affinity, with His-16 replacement affecting structure. In contrast, [D15A]SIIIA-(2-20) had significantly enhanced neuronal affinity (IC50 0.65 nM), while the double mutant [D15A/H16R]SIIIA-(2-20) showed greatest Nav1.2 versus 1.4 selectivity (136-fold). 1H NMR studies revealed that SIIIA adopted a single conformation in solution comprising a series of turns and an α-helical motif across residues 11-16 that is not found in larger μ-conotoxins. The structure of SIIIA provides a new structural template for the development of neuronally selective inhibitors of TTX-sensitive VGSCs based on the smaller μ-conotoxin pharmacophore.

Original languageEnglish
Pages (from-to)21621-21628
Number of pages8
JournalJournal of Biological Chemistry
Volume283
Issue number31
DOIs
Publication statusPublished - 1 Aug 2008
Externally publishedYes

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    Schroeder, C. I., Ekberg, J., Nielsen, K. J., Adams, D., Loughnan, M. L., Thomas, L., Adams, D. J., Alewood, P. F., & Lewis, R. J. (2008). Neuronally selective μ-conotoxins from Conus striatus utilize an α-helical motif to target mammalian sodium channels. Journal of Biological Chemistry, 283(31), 21621-21628. https://doi.org/10.1074/jbc.M802852200