We present a fluorogenic method to visualize misfolding and aggregation of a specific protein-of-interest in live cells using structurally modulated fluorescent protein chromophores. Combining photophysical analysis, X-ray crystallography, and theoretical calculation, we show that fluorescence is triggered by inhibition of twisted-intramolecular charge transfer of these fluorophores in the rigid microenvironment in viscous solvent or protein aggregates. Based on these molecules, we further developed the AggTag (aggregation tag) method that employs  bioorthogonal conjugation of the fluorophore to Halo-tag fused proteins. The AggTag method allows for fluorogenic detection of both misfolded oligomers and insoluble aggregates in live cells. Unlike other methods, the AggTag method is capable of detecting previously invisible misfolded soluble proteins. Taken together, this work provides the first application of fluorescent protein chromophores to detect the entire process of protein aggregation in live cells.

Detecting protein aggregation in live cells with turn-on fluorescence using chemically modulated fluorescent protein chromophores

2019 Spring National Meeting

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