Role of γ-Carboxyglutamic Acid in the Calcium-Induced Structural Transition of Conantokin G, a Conotoxin from the Marine Snail <i>Conus geographus</i>^,

Abstract

Conantokin G is a gamma-carboxyglutamic acid- (Gla-) containing conotoxin isolated from the venom of the marine cone snail Conus geographus. This 17-residue polypeptide, which contains five gamma-carboxyglutamic acid residues, is a N-methyl-d-aspartate- (NMDA-) type glutamate receptor antagonist. To investigate the role of gamma-carboxyglutamic acid in the calcium-induced structural transition of conantokin G, we determined the three-dimensional structure of the conantokin G/Ca2+ complex by two-dimensional 1H NMR spectroscopy and compared it to the high-resolution structure of conantokin G in the absence of metal ions [Rigby et al. (1997) Biochemistry 36, 6906]. Complete resonance assignments were made by two dimensional 1H NMR spectroscopy at pH 5.6 in the presence of saturating amounts of Ca2+. Distance geometry and simulated annealing methods were used to derive 23 convergent structures from a set of 302 interproton distance restraints and two torsion angle measurements. A high-resolution structure, with the backbone root mean square deviation to the geometric average of the 23 structures of 0.6 +/- 0.1 A, contains a linear alpha-helix from Gla 3 to Lys 15. Gla residues 3, 7, 10, and 14 are aligned in a linear array on one face of the helix. A genetic algorithm was applied to determine the calcium positions in conantokin G, and the conantokin G/Ca2+ complex refined by molecular simulation. Upon binding of Ca2+ to gamma-carboxyglutamic acid, conantokin G undergoes a conformational transition from a distorted curvilinear 310 helix to a linear alpha-helix. Occupancy of the metal binding sites, defined by gamma-carboxyglutamic acids, results in formation of a calcium-carboxylate network that linearizes the helix and exposes the hydrophobic amino acids on the opposite face of the helix.

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Affiliated Institutions

• Tufts University US
• National Institute of Environmental Health Sciences US
• Marine Biological Laboratory US

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