Single-molecule spectroscopy of protein folding dynamics-expanding scope and timescales

Single-molecule spectroscopy has developed into an important method for probing protein structure and dynamics, especially in structurally heterogeneous systems. A broad range of questions in the diversifying field of protein folding have been addressed with single-molecule Forster resonance energy transfer (FRET) and photo-induced electron transfer (PET). Building on more than a decade of rapid method development, these techniques can now be used to investigate a wide span of timescales, an aspect that we focus on in this review. Important current topics range from the structure and dynamics of unfolded and intrinsically disordered proteins, including the coupling of folding and binding, to transition path times, the folding and misfolding of larger proteins, and their interactions with molecular chaperones.