Tertiary structure of endothelin-1 in water by 1H NMR and molecular dynamics studies

The 3D structure in pure water of endothelin-1, a recently discovered potent vasoconstrictor peptide, has been determined at different pH and temperatures, using two-dimensional 1H NMR spectroscopy and constrained molecular dynamics (MD). 170 Inter- and intra-residue NOE interactions were quantified as volume integrals and translated into distances. All the coupling constants between the amidic and the α-protons have been measured and several dihedral angles, thus obtained, have been used as constraints for the MD. Some stereospecific assignments have also been performed. A family of eleven structures, satisfying the distance constraints to within 0.3 Å, was obtained and showed that the C-terminal, determinant for the binding with the receptor, has a well defined conformation. The correlation time measurements gave an average molecular volume consistent with monomeric species. The tertiary structure at neutral pH is that of a compact molecule, in which the C-terminal of the peptide folds back toward the α-helical segment (residues 9–16), in close proximity to the pro-R methyl group of Val12, as defined by important NOEs involving residues 17–21 and the α-helical core residues 9–14. The results are in agreement with the deuterium exchange experiments, which confirm the existence of a hydrophobic region also at the site of the C-terminal residues 19–21.