Modulation of Fluorescent Protein Chromophores To Detect Protein Aggregation with Turn-On Fluorescence

Area of Science:

• Biochemistry
• Cell Biology
• Biophysics

Background:

• Protein misfolding and aggregation are implicated in various diseases.
• Current methods struggle to detect soluble misfolded proteins in live cells.

Purpose of the Study:

• To develop a novel fluorogenic method for visualizing specific protein misfolding and aggregation in live cells.
• To enable the detection of previously undetectable misfolded soluble proteins.

Main Methods:

• Utilized structurally modulated fluorescent protein chromophores.
• Combined photophysical analysis, X-ray crystallography, and theoretical calculations.
• Employed bioorthogonal conjugation of fluorophores to Halo-tag fused proteins.

Main Results:

• Demonstrated that fluorescence is triggered by the inhibition of twisted-intramolecular charge transfer in rigid environments.
• Successfully visualized both misfolded and aggregated protein species in live cells.
• Showcased the ability to detect previously invisible misfolded soluble proteins.

Conclusions:

• The developed fluorogenic method provides a powerful tool for studying protein conformational changes in real-time.
• This approach offers unprecedented sensitivity for detecting misfolded proteins, advancing our understanding of proteinopathies.
• Represents the first application of fluorescent protein chromophores for detecting protein conformational collapse in live cellular systems.