NMR and protein folding: Equilibrium and stopped‐flow studies

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Abstract
NMR studies are now unraveling the structure of intermediates of protein folding using hydrogen—deuterium exchange methodologies. These studies provide information about the time dependence of formation of secondary structure. They require the ability to assign specific resonances in the NMR spectra to specific amide protons of a protein followed by experiments involving competition between folding and exchange reactions. Another approach is to use 19F‐substituted amino acids to follow changes in side‐chain environment upon folding. Current techniques of molecular biology allow assignments of 19F resonances to specific amino acids by site‐directed mutagenesis. It is possible to follow changes and to analyze results from 19F spectra in real time using a stopped‐flow device incorporated into the NMR spectrometer.