Figure 3: Discrepancies between smFRET and time averaging results enable discovery of a novel lysozyme fold.

A) FRET efficiencies and (C) observed inter-dye distances for T4 Lysozyme labeled at position 44 and 150 with Alexa 488 and Alexa 647. Experimental traces are in black, calculations resulting from an MSM that only included crystal-like states in purple, and calculations resulting from an MSM including the alternate state in orange. Experimental donor only counts (E < 0.25) have been removed for ease of comparison. B) Example conformations of the crystal-like state of lysozyme (blue), or the alternate state (red). Residues 44 and 150 labeled for clarity. D) Qualitative comparison of smFRET distributions from experiment and simulation results including the alternate pose of lysozyme. E) Wasserstein distance between experimental and limited sampling dataset (purple) or extended sampling dataset (orange) for all labeled pairs.