Interactions of disulfide-deficient selenocysteine analogs of u-conotoxin BuIIIB with the a-subunit of the voltage-gated sodium channel subtype 1.3

Brad Reed Green, Minmin Zhang, Sandeep Chhabra, Samuel Robinson, Michael John Wilson, Addison Redding, Baldomero Marquez Olivera, Doju Yoshikami, Grzegorz Bulaj, Raymond Stanley Norton

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Abstract

Inhibitors of the a-subunit of the voltage-gated sodium channel subtype 1.3 (NaV1.3) are of interest as pharmacological tools for the study of neuropathic pain associated with spinal cord injury and have potential therapeutic applications. The recently described ?-conotoxin BuIIIB (?-BuIIIB) from Conus bullatus was shown to block NaV1.3 with submicromolar potency (Kd = 0.2 ?m), making it one of the most potent peptidic inhibitors of this subtype described to date. However, oxidative folding of ?-BuIIIB results in numerous folding isoforms, making it difficult to obtain sufficient quantities of the active form of the peptide for detailed structure-activity studies. In the present study, we report the synthesis and characterization of ?-BuIIIB analogs incorporating a disulfide-deficient, diselenide-containing scaffold designed to simplify synthesis and facilitate structure-activity studies directed at identifying amino acid residues involved in NaV1.3 blockade. Our results indicate that, similar to other ?-conotoxins, the C-terminal residues (Trp16, Arg18 and His20) are most crucial for NaV1 blockade. At the N-terminus, replacement of Glu3 by Ala resulted in an analog with an increased potency for NaV1.3 (Kd = 0.07 ?m), implicating this position as a potential site for modification for increased potency and/or selectivity. Further examination of this position showed that increased negative charge, through ?-carboxyglutamate replacement, decreased potency (Kd = 0.33 ?m), whereas replacement with positively-charged 2,4-diamonobutyric acid increased potency (Kd = 0.036 ?m). These results provide a foundation for the design and synthesis of ?-BuIIIB-based analogs with increased potency against NaV1.3. The possibility of creating a molecule that latches only onto the channels in nerves that signal pain is of considerable interest for human health and well-being. Such a molecule could conceivably block pain neurons - therefore blocking the sensation of pain - without interfering with other nerves including those that signal muscles to move. A toxin extracted from a marine cone venom, ?-conotoxin BuIIIB, targets the voltage-gated sodium channel Nav1.3, an important mediator of neuropathic pain. In this issue, Green and colleagues provide a new approach for synthesizing disulfide-deficient and diselenide-containing analogs of this peptide that are functionally active. This approach offers new opportunities for the design of molecules targeting Nav 1.3 channels.
Original languageEnglish
Pages (from-to)2885 - 2898
Number of pages14
JournalFEBS Journal
Volume281
Issue number13
DOIs
Publication statusPublished - 2014

Cite this

Green, B. R., Zhang, M., Chhabra, S., Robinson, S., Wilson, M. J., Redding, A., Olivera, B. M., Yoshikami, D., Bulaj, G., & Norton, R. S. (2014). Interactions of disulfide-deficient selenocysteine analogs of u-conotoxin BuIIIB with the a-subunit of the voltage-gated sodium channel subtype 1.3. FEBS Journal, 281(13), 2885 - 2898. https://doi.org/10.1111/febs.12835