Direct and Indirect Models for Protein Chemical Denaturation are Characteristic of Opposite Dynamic Properties

Two major models, namely direct and indirect models, have been proposed for the protein chemical denaturation but it remains challenging to experimentally demonstrate and distinguish between them. Here, by use of CD and NMR spectroscopy, we succeeded in differentiating the effects on a small but well-folded protein WW4, of GdmCl and NaSCN at diluted concentrations ([≥]200 mM). Both denaturants up to 200 mM have no alternation of its average structure but do reduce its thermodynamic stability to different degrees. Despite acting as the stronger denaturant, GdmCl only weakly interacts with amide protons, while NaSCN shows extensive interactions with both hydrophobic side chains and amide pro