Cloning, synthesis, and characterization of αO-conotoxin GeXIVA, a potent α9α10 nicotinic acetylcholine receptor antagonist

Document Type

Article

Publication Title

Proceedings of the National Academy of Sciences of the United States of America

Abstract

We identified a previously unidentified conotoxin gene from Conus generalis whose precursor signal sequence has high similarity to the O1-gene conotoxin superfamily. The predicted mature peptide, αO-conotoxin GeXIVA (GeXIVA), has four Cys residues, and its three disulfide isomers were synthesized. Previously pharmacologically characterized O1-superfamily peptides, exemplified by the US Food and Drug Administration-approved pain medication, ziconotide, contain six Cys residues and are calcium, sodium, or potassium channel antagonists. However, GeXIVA did not inhibit calcium channels but antagonized nicotinic AChRs (nAChRs), most potently on the α9α10 nAChR subtype (IC50 = 4.6 nM). Toxin blockade was voltage-dependent, and kinetic analysis of toxin dissociation indicated that the binding site of GeXIVA does not overlap with the binding site of the competitive antagonist α-conotoxin RgIA. Surprisingly, the most active disulfide isomer of GeXIVA is the bead isomer, comprising, according to NMR analysis, two wellresolved but uncoupled disulfide-restrained loops. The ribbon isomer is almost as potent but has a more rigid structure built around a short 310-helix. In contrast to most α-conotoxins, the globular isomer is the least potent and has a flexible, multiconformational nature. GeXIVA reduced mechanical hyperalgesia in the rat chronic constriction injury model of neuropathic pain but had no effect on motor performance, warranting its further investigation as a possible therapeutic agent.

First Page

E4026

Last Page

E4035

DOI

10.1073/pnas.1503617112

Publication Date

7-28-2015

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