Label-Free Analysis of Protein Aggregation and Phase Behaviour

Phase transitions of protein molecules are central to biological function and malfunction. One such transition commonly encountered in nature is the conversion of soluble monomeric states into solid phases, which include crystals and amyloid fibrils, the latter of which are associated with the onset and development of neurodegenerative diseases. Monitoring aggregate formation and protein phase behavior is essential in gaining mechanistic insights into these fundamental processes. Fluorescence techniques have proven invaluable in observing biological molecules; yet, most such approaches rely on the use of an extrinsic fluorophore that binds to the molecule of interest, the installation of which can perturb the molecular systems under study. However, most proteins also possess aromatic amino acids within their peptide sequence and therefore exhibit intrinsic fluorescence. Here, we show that by measuring in space and time tryptophan autofluorescence for three proteins, reconstituted silk fibroin, β-lactoglob...