Thioamides as Fluorescence Quenching Probes: Minimalist Chromophores to Monitor Protein Dynamics

One of the great challenges facing biochemists is to understand the rapid and complex structural dynamics of proteins.  Fluorescence measurements can be made on the ns timescale, and distance-dependent interactions such as Förster resonant energy transfer (FRET) can be used to determine the separation of chromophore labels to glean time-resolved structural information on protein motions.  However, the relatively large size of common fluorophores precludes assigning these motions at atomic resolution.  We have recently demonstrated that a thioamide, a single-atom substitution of the peptide backbone, can be used as a fluorescence quenching probe to monitor structural changes in proteins.  For a given protein, thioamide substitutions could be used to track motions with a much greater number of measurements than current fluorescence probes, providing a dense array of data with which to model conformational changes.