Proteome-Wide Profiling of Targets of Cysteine reactive Small Molecules by Using Ethynyl Benziodoxolone Reagents

In this study, we present a highly efficient method for proteomic profiling of cysteine residues in complex proteomes and in living cells. Our method is based on alkynylation of cysteines in complex proteomes using a “clickable” alkynyl benziodoxolone bearing an azide group. This reaction proceeds f...

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Bibliographic Details
Published in:Angewandte Chemie International Edition 2015-09, Vol.54 (37), p.10852-10857
Main Authors: Abegg, Daniel, Frei, Reto, Cerato, Luca, Prasad Hari, Durga, Wang, Chao, Waser, Jerome, Adibekian, Alexander
Format: Article
Language:English
Subjects:
activity-based protein profiling
Cellular
Chromatography, Liquid - methods
curcumin
Cysteine
Cysteine - metabolism
Enzymes
HeLa Cells
Humans
Indicators and Reagents - chemistry
Kinases
mass spectrometry
Natural products
Oxolinic Acid - chemistry
Pharmaceutical industry
Profiling
Proteome
Proteomics
Residues
Tandem Mass Spectrometry - methods
target identification
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Summary:In this study, we present a highly efficient method for proteomic profiling of cysteine residues in complex proteomes and in living cells. Our method is based on alkynylation of cysteines in complex proteomes using a “clickable” alkynyl benziodoxolone bearing an azide group. This reaction proceeds fast, under mild physiological conditions, and with a very high degree of chemoselectivity. The formed azide‐capped alkynyl–cysteine adducts are readily detectable by LC‐MS/MS, and can be further functionalized with TAMRA or biotin alkyne via CuAAC. We demonstrate the utility of alkynyl benziodoxolones for chemical proteomics applications by identifying the proteomic targets of curcumin, a diarylheptanoid natural product that was and still is part of multiple human clinical trials as anticancer agent. Our results demonstrate that curcumin covalently modifies several key players of cellular signaling and metabolism, most notably the enzyme casein kinase I gamma. We anticipate that this new method for cysteine profiling will find broad application in chemical proteomics and drug discovery. Good grip: Alkynyl benziodoxolones (EBX reagents) swiftly and highly selectively react with cysteine residues in cellular lysates and in living cells under physiological conditions. A “clickable” EBX probe allowed identification of the biological targets of natural product curcumin. This new method for cysteine labeling is particularly useful for various chemical proteomics applications.
ISSN:1433-7851
1521-3773
DOI:10.1002/anie.201505641