(69g) Unfolded Peptide Structure and Conformation with FRET Dyes

Single molecule Forster resonance energy transfer (FRET) experiments are often used to study the properties of unfolded and intrinsically disordered proteins. Synthetic heteroaromatic chromophores covalently linked to the protein are often used as donor and acceptor fluorophores. A key issue in the interpretation of such experiments is the extent to which the properties of the unfolded chain may be affected by the presence of these chromophores. In this work, we look for an answer of this question using all-atom explicit solvent replica exchange molecular simulations of three different unfolded or intrinsically disordered proteins. We find that the secondary structure and long-range contacts are largely the same in the presence or absence of the fluorophores, and that the dimensions of the chain with and without chromophores are similar. This suggests that extrinsic fluorophores have little effect on the structural and conformational properties of unfolded or disordered proteins. We also find that the critical FRET orientational factor, kappa², has an average value and equilibrium distribution very close to that expected for isotropic orientations, which supports one of the assumptions frequently made when interpreting FRET efficiency in terms of distances.