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Updated: Jun 6, 2026
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Identification of Small Molecule-binding Proteins in a Native Cellular Environment by Live-cell Photoaffinity Labeling
Published on: September 20, 2016
Construction of Coumarin-Conjugated Macromolecular Probe for Photoactivatable Bioimaging
Xuekun Zhang1, Qin Xu1, Ying Wang1
1Key Laboratory of Carbohydrate Vaccines and Drugs in Jiangsu Province, School of Life Sciences and Health Engineering, Jiangnan University, Wuxi, China.
Summary
Researchers developed new bioorthogonal fluorescent probes for protein imaging. Probe Y12 enables high-efficiency photoactivated fluorescence labeling of STING proteins, significantly reducing background noise for real-time dynamic studies.
Area of Science:
- Chemical Biology
- Molecular Imaging
- Biotechnology
Background:
- Fluorophore-conjugated imaging is crucial for visualizing protein localization and trafficking.
- High background noise frequently limits the performance of current protein imaging techniques.
Purpose of the Study:
- To design and synthesize novel coumarin-based bioorthogonal fluorescent probes for enhanced protein imaging.
- To evaluate the performance of these probes, particularly their photoactivated fluorescence and signal-to-noise ratio.
Main Methods:
- Rational design and synthesis of four coumarin-based bioorthogonal fluorescent probes.
- Site-specific conjugation of photoactivatable caging groups and HaloTag ligands.
- Fluorescence characterization, including aggregation-induced emission (AIE) and photoresponse.
- Intracellular fluorescence labeling of STING protein using developed probes.
Main Results:
- Probes Y8 and Y9 showed aggregation-induced emission (AIE).
- Probes Y11 and Y12 exhibited rapid photoresponse; Y12 demonstrated stable HaloTag conjugation and robust photoactivated fluorescence.
- Probe Y12 combined with STING-Halo achieved 16.5-fold fluorescence enhancement, enabling ultra-high signal-to-background ratio (S/N).
- Real-time monitoring of STING activation and translocation was achieved.
Conclusions:
- The developed fluorescent dye/protein hybrid system, particularly probe Y12, significantly improves photoactivated fluorescence labeling efficiency.
- This system facilitates high-S/N ratio imaging, enabling real-time tracking of protein dynamics.
- Holds promise for advancing research in cell homeostasis and medicine.
