CHARACTERIZATION OF RESIDUAL STRUCTURE IN THE THERMALLY DENATURED STATE OF BARNASE BY SIMULATION AND EXPERIMENT - DESCRIPTION OF THE FOLDING PATHWAY

Residual structure in the denatured state of a protein may contain clu es about the early events in folding. We have simulated by molecular d ynamics the denatured state of barnase, which has been studied by NMR spectroscopy. An ensemble of 10(4) structures was generated after 2 ns of unfolding and following for a further 2 ns. The ensemble was heter ogeneous, but there was nonrandom, residual structure with persistent interactions. Helical structure in the C-terminal portion of helix alp ha 1 (residues 13-17) and in helix alpha 2 as well as a turn and nonna tive hydrophobic clustering between beta 3 and beta 4 were observed, c onsistent with NMR data. In addition, there were tertiary contacts bet ween residues in alpha 1 and the C-terminal portion of the beta-sheet. The simulated structures allow the rudimentary NMR data to be fleshed out. The consistency between simulation and experiment inspires confi dence in the methods. A description of the folding pathway of barnase from the denatured to the native state can be constructed by combining the simulation with experimental data from phi value analysis and NMR .