Tertiary structure of conotoxin GIIIA in aqueous solution

Abstract

The three-dimensional structure of conotoxin GIIIA, an important constituent of the venom from the marine hunting snail Conus geographus L., was determined in aqueous solution by two-dimensional proton nuclear magnetic resonance and simulated annealing based methods. On the basis of 162 assigned nuclear Overhauser effect (NOE) connectivities obtained at the medium field strength frequency of 400 MHz, 74 final distance constraints of sequential and tertiary ones were derived and used together with 18 torsion angle (phi, chi 1) constraints and 9 distance constraints derived from disulfide bridges. A total of 32 converged structures were obtained from 200 runs of calculations. The atomic root-mean-square (RMS) difference about the mean coordinate positions (excluding the terminal residues 1 and 22) is 0.8 A for backbone atoms (N, C alpha, C). Conotoxin GIIIA is characterized by a particular folding of the 22 amino acid peptidic chain, which is stabilized by three disulfide bridges arranged in cage at the center of a discoidal structure of approximately 20-A diameter. The seven cationic side chains of lysine and arginine residues project radially into the solvent and form potential sites of interaction with the skeletal muscle sodium channel for which the toxin is a strong inhibitor. The present results provide a molecular basis to elucidate the remarkable physiological properties of this neurotoxin.

Structure-Activity Relationships of Hainantoxin-IV and Structure Determination of Active and Inactive Sodium Channel Blockers

“…The potency reductions produced by Ala substitutions were primarily due to the direct interaction of the essential residues Lys 27 and Arg 29 with sodium channels rather than to a conformational change. After comparison of these structures and activities with correlated toxins, we hypothesized that residues Lys 27 clustered on one face of HNTX-IV were responsible for ligand binding.…”

“…Based on the structural comparison and sequence assignment with other related toxins, we hypothesized that the positively charged residues Arg 26 , Lys 27 , and Arg 29 in loop IV and vicinal polar residue Ser 12 clustered on one face of HNTX-IV may be the potential interaction site. Mutants S12A, R26A, K27A, and R29A were synthesized by solid-phase Fmoc chemistry followed by oxidative refolding of purified peptides under the optimal conditions.…”

“…1 H NMR analysis showed similar molecular conformations for native HNTX-IV and synthetic mutants. Pharmacological studies indicated that residues Ser 12 and Arg 26 were not important for the activities, while Lys 27 and Arg 29 were critical for the bioactivities. These results provide useful information for structure-activity relationships of toxins like HNTX-IV.…”

Characterization and 2D NMR study of the stable [9–21, 15–27] 2 disulfide intermediate in the folding of the 3 disulfide trypsin inhibitor EETI II

“…comm.). Some other proteins present the same connectivity, e.g., Escherichia coli enterotoxin (Garitpy et al, 1986) and conotoxin GIIIA (Lancelin et al, 1991); however, no loop penetration (Klapper & Klapper, 1980) can be observed in these structures.…”

Developmental Biology of Neoplastic Growth

“…5. The NMR structures of the muscle selective GIIIA (Lancelin et al 1991) may not reflect the structure that binds the VGSC based on comparisons with NMR structure of TIIIA ).…”