Three-Dimensional Structure of a γ-Carboxyglutamic Acid-Containing Conotoxin, Conantokin G, from the Marine Snail Conus geographus:  The Metal-Free Conformer

Conantokin G is a γ-carboxyglutamic acid-containing conotoxin from the venom of the marine cone snail Conus geographus. The 17-residue peptide, which contains five γ-carboxyglutamic acid (Gla) residues and an amidated C-terminal asparagine amide, was synthesized chemically in a form identical to the natural conantokin G. To gain insight into the role of γ-carboxyglutamic acid in the structure of this peptide, we determined the three-dimensional structure of conantokin G by 1H NMR and compared its structure to other conotoxins and to the γ-carboxyglutamic acid-containing regions of the vitamin K-dependent blood-clotting proteins. Complete resonance assignments were made by two-dimensional 1H NMR spectroscopy in the absence of metal ions. NOE cross-peaks d αN, d NN, and d βN provided interproton distance information, and vicinal spin−spin coupling constants 3 J HNα were used to calculate φ torsion angles. Distance geometry and simulated annealing methods were used to derive 20 convergent structures from a set of 227 interproton distance restraints and 13 torsion angle measurements. The backbone rmsd to the geometric average for 20 final structures is 0.8 ± 0.1 Å. Conantokin G consists of a structured region commencing at Gla 3 and extending through arginine 13. This structure includes a partial loop centered around Gla 3 and Gla 4, a distorted type I turn between glutamine 6 and glutamine 9, and two type I turns involving Gla 10, leucine 11, and isoleucine 12 and arginine 13. Together, these two turns define approximately 1.6 turns of a distorted 310 helix. The observed structure possesses structural elements similar to those seen in the disulfide-linked conotoxins.