Title: Multiparameter Fluorescence Spectroscopy for Structural Dynamics of Biomolecules

Abstract: Single molecule Förster Resonance Energy Transfer experiments are rich in information of biomolecules. By time-stamping photon detection events with high temporal resolution, it is possible to construct various data representations that highlight the time evolution of multiple fluorescence parameters reflecting the structure and dynamics of molecules of interest. This can include the FRET efficiency, the fluorescence anisotropy, fluorescence decay, or the correlation of fluorescence fluctuations. Treating each representation independently provides only limited information. Nonetheless, the holistic nature of multiparameter fluorescence spectroscopy (MFS) allows for joint analysis of single molecules, thus allowing correction for potential artifacts, reducing uncertainty, and linking the structure and dynamics to the functions of biomolecules. Several examples of recent work are presented, highlighting the ample capability of MFS. A brief outlook of the remaining challenges will be presented.