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Resin-Assisted Capture Coupled with Isobaric Tandem Mass Tag Labeling for Multiplexed Quantification of Protein Thiol Oxidation
Published on: June 21, 2021
"IEDDA"-activated fluorescent sensing for protein cysteine redox modifications profiling.
Linfeng Chen1, Yifu Zheng1, Ziyi Wang1
1National Key Laboratory of Green Pesticide, College of Chemistry, International Joint Research Center for Intelligent Biosensing Technology and Health, Central China Normal University, Wuhan, 430079, China.
Researchers developed a new "turn-on" fluorescent probe strategy for tracking protein cysteine redox modifications. This method offers high-contrast imaging in cells and organisms, advancing redox biology research.
Area of Science:
- Redox Biology and Chemical Biology
- Bioorthogonal Chemistry and Imaging
Background:
- Protein cysteine sulfenylation is a critical reversible modification in redox signaling and disease.
- Existing fluorescent probes for sulfenylation have limitations, including high background signals and inability for real-time tracking.
Purpose of the Study:
- To develop a background-free, "turn-on" fluorescent sensing strategy for monitoring protein cysteine redox modifications.
- To enable dynamic, real-time visualization of cysteine redox events in living systems.
Main Methods:
- Designed selective probes (BCN-SH and BCN-SOH) utilizing a bicyclo[6.1.0]non-4-yne (BCN) handle for tagging protein thiols and sulfenic acids.
- Employed the inverse electron-demand Diels-Alder (IEDDA) reaction for bioorthogonal activation with a tetrazine-based fluorophore (BODIPY-tetrazine, BTZ).
- Validated probe selectivity, stability, and performance in live HeLa cells and Caenorhabditis elegans.
Main Results:
- Achieved a 248-fold fluorescence enhancement upon IEDDA reaction, enabling high-contrast detection.
- Successfully visualized endogenous cysteine redox dynamics in live HeLa cells under oxidative stress.
- Demonstrated the strategy's applicability for fluorescence imaging in Caenorhabditis elegans.
Conclusions:
- The developed platform integrates selective cysteine chemistry with fluorogenic bioorthogonal activation for background-free visualization of dynamic redox events.
- This strategy expands the chemical toolbox for redox biology, facilitating mechanistic studies of oxidative signaling pathways.
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