Line Shape Analysis of F NMR-Monitored Chemical Denaturation of a Fold-Switching Protein RfaH Reveals Its Slow Folding Dynamics.

The recent discovery of metamorphic proteins, which can switch between multiple conformations under native conditions, has challenged the well-established one sequence-one structure paradigm of protein folding. This is exemplified in the C-terminal domain of the multidomain transcription factor RfaH, which converts from an α-helical coiled-coil conformation in its autoinhibited state to a β-barrel conformation upon activation. Here, we use multisite line shape analysis of F NMR-monitored equilibrium chemical denaturation measurements of two F-labeled variants of full-length RfaH, to show that it folds/unfolds slowly on the NMR time scale, in an apparent all-or-none fashion at physiological pH and room temperature in the 3.