STRUCTURE AND DYNAMICS OF FERROCYTOCHROME C(553) FROM DESULFOVIBRIO-VULGARIS STUDIED BY NMR-SPECTROSCOPY AND RESTRAINED MOLECULAR-DYNAMICS

The solution structure of Desulfovibrio vulgaris Hildenborough (DvH) f errocytochrome c(553) has been determined by nuclear magnetic resonanc e spectroscopy and combined simulated annealing/high temperature restr ained molecular dynamics calculations. This three-stage protocol consi sts of an initial determination of overall fold from randomised co-ord inates, followed by a 20 picosecond exploratory stage, during which th e non-bonded terms are simplified to facilitate as broad a sampling of conformational space as possible, and a 26 picosecond refinement stag e, using the full AMBER force field. This latter stage systematically improved the energetic and convergence characteristics of the ensemble , while still satisfying the experimental restraints. Forty structures have been obtained from a total of 875 distance constraints for this protein of 79 amino acid residues. The root-mean-square deviation over all residues with respect to the mean is 0.70(+/-0.12)Angstrom for th e backbone (N, C(alpha)and C') atoms. Two conformations of the turn mo tif at the solvent/heme cleft interface have been identified, both ful filling the experimental data and having equally viable energetic char acteristics. The stability of the ensemble and the dynamic characteris tics have been further investigated by subjecting ten of the structure s to constraint-free molecular dynamics calculations (130 picoseconds) in vacuo. The structures were found to be stable to within 1.5 Angstr om of the initial backbone conformation. Comparison with the dynamic b ehaviour of the restrained molecular dynamics calculations has been us ed to identify regions of inherent flexibility in the molecule.