A Solvatochromic Fluorescent Probe Reveals Polarity Heterogeneity upon Protein Aggregation in Cells.

Physicochemical details at the interphase of protein aggregation have been less explored especially in live cells. Herein, we reported a crystallization induced emission fluorophore to quantitatively interrogate the polarity of aggregated proteins. This solvatochromic probe, namely "AggRetina" probe, inherently binds to aggregated proteins and exhibits both polarity-dependent fluorescence emission wavelength shift and viscosity-dependent fluorescence intensity increase. Regulation of its polarity sensitivity was achieved by extending the conjugation length. Empowered by this probe, we observed that different proteins bear diverse polarity upon aggregation, leading to different resistance to proteolysis. We further showed that polarity primarily decreases during protein misfolding but viscosity mainly increases upon the formation of insoluble aggregates.  Finally, we quantified the polarity of aggregated protein-of-interest in live cells via HaloTag bioorthogonal labeling, revealing polarity heterogeneity within cellular aggregates. This work highlights the enriched micro-environment details inside misfolded and aggregated proteins that may correlate to their bio-chemical properties and pathogenicity.