Solution structure of the sodium channel antagonist conotoxin GS: a new molecular caliper for probing sodium channel geometry

AbstractBackground: The venoms of Conus snails contain small, disulfide-rich inhibitors of voltage-dependent sodium channels. Conotoxin GS is a 34-residue polypeptide isolated from Conus geographus that interacts with the extracellular entrance of skeletal muscle sodium channels to prevent sodium ion conduction. Although conotoxin GS binds competitively with μ conotoxin GIIIA to the sodium channel surface, the two toxin types have little sequence identity with one another, and conotoxin GS has a four-loop structural framework rather than the characteristic three-loop μ-conotoxin framework. The structural study of conotoxin GS will form the basis for establishing a structure–activity relationship and understanding its interaction with the pore region of sodium channels.Results: The three-dimensional structure of conotoxin GS was determined using two-dimensional NMR spectroscopy. The protein exhibits a compact fold incorporating a β hairpin and several turns. An unusual feature of conotoxin GS is the exceptionally high proportion (100%) of cis-imide bond geometry for the three proline or hydroxyproline residues. The structure of conotoxin GS bears little resemblance to the three-loop μ conotoxins, consistent with the low sequence identity between the two toxin types and their different structural framework. However, the tertiary structure and cystine-knot motif formed by the three disulfide bonds is similar to that present in several other polypeptide ion channel inhibitors.Conclusions: This is the first three-dimensional structure of a ‘four-loop’ sodium channel inhibitor, and it represents a valuable new structural probe for the pore region of voltage-dependent sodium channels. The distribution of amino acid sidechains in the structure creates several polar and charged patches, and comparison with the μ conotoxins provides a basis for determining the binding surface of the conotoxin GS polypeptide