Photoactivatable BODIPY Dyes Conjugated to Self-Labeling Protein Tags.

Area of Science:

• Chemical Biology
• Molecular Imaging
• Biophysics

Background:

• Photoactivatable synthetic dyes offer advanced capabilities for time-lapse measurements and super-resolution imaging.
• Selective labeling of subcellular structures is crucial for investigating cellular processes using these dyes.
• Self-labeling protein tags provide structural control for attaching synthetic dyes to intracellular targets.

Purpose of the Study:

• To design and synthesize photoactivatable fluorescent dyes for selective labeling of HaloTag and SNAP-tag fusion proteins.
• To preserve the targeting ability and photochemical/photophysical properties of BODIPY dyes upon conjugation.
• To investigate the influence of protein environment on dye properties and demonstrate intracellular labeling in model cells.

Main Methods:

• Multistep synthesis of borondipyrromethene (BODIPY) dyes functionalized with HaloTag/SNAP-tag ligands.
• Spectroscopic measurements (ensemble and single-molecule) to characterize dye properties.
• Molecular dynamics simulations to assess protein-dye interactions.
• Cellular experiments demonstrating intracellular fluorescence photoactivation.

Main Results:

• Successfully synthesized BODIPY dyes with photoactivatable fluorescence and specific protein-tag ligands.
• Demonstrated preservation of ligand targeting and dye photophysical properties in the final constructs.
• Confirmed that the protein environment minimally impacts the dye's fluorescence characteristics.
• Achieved efficient intracellular labeling and photoactivation of fusion proteins in model cells.

Conclusions:

• The developed synthetic dyes are suitable for selective intracellular protein labeling.
• These photoresponsive dyes enable high-contrast fluorescence photoactivation within cells.
• The probes hold potential for investigating intracellular protein structures and dynamics with precise spatiotemporal control.