Title

Isolation and structure-activity of μ-conotoxin TIIIA, a potent inhibitor of tetrodotoxin-sensitive voltage-gated sodium channels

RIS ID

105695

Publication Details

Lewis, R. J., Schroeder, C. I., Ekberg, J., Nielsen, K. J., Loughnan, M. L., Thomas, L., Adams, D., Drinkwater, R., Adams, D. J. & Alewood, P. F. (2007). Isolation and structure-activity of μ-conotoxin TIIIA, a potent inhibitor of tetrodotoxin-sensitive voltage-gated sodium channels. Molecular Pharmacology, 71 (3), 676-685.

Abstract

μ-Conotoxins are three-loop peptides produced by cone snails to inhibit voltage-gated sodium channels during prey capture. Using polymerase chain reaction techniques, we identified a gene sequence from the venom duct of Conus tulipa encoding a new μ-conotoxin-TIIIA (TIIIA). A 125I-TIIIA binding assay was established to isolate native TIIIA from the crude venom of Conus striatus. The isolated peptide had three post-translational modifications, including two hydroxyproline residues and C-terminal amidation, and <35% homology to other μ-conotoxins. TIIIA potently displaced [ 3H]saxitoxin and 125I-TIIIA from rat brain (Na v1.2) and skeletal muscle (Nav1.4) membranes. Alanine and glutamine scans of TIIIA revealed several residues, including Arg14, that were critical for high-affinity binding to tetrodotoxin (TTX)-sensitive Na + channels. We were surprised to find that [E15A]TIIIA had a 10-fold higher affinity than TIIIA for TTX-sensitive sodium channels (IC50, 15 vs. 148 pM at rat brain membrane). TIIIA was selective for Nav1.2 and -1.4 over Nav1.3, -1.5, -1.7, and -1.8 expressed in Xenopus laevis oocytes and had no effect on rat dorsal root ganglion neuron Na + current. 1H NMR studies revealed that TIIIA adopted a single conformation in solution that was similar to the major conformation described previously for μ-conotoxin PIIIA. TIIIA and analogs provide new biochemical probes as well as insights into the structure-activity of μ-conotoxins.

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Link to publisher version (DOI)

http://dx.doi.org/10.1124/mol.106.028225