Broad-Spectrum Kinase Profiling in Live Cells with Lysine-Targeted Sulfonyl Fluoride Probes
Open accessAbstract
Protein kinases comprise a large family of structurally related enzymes. A major goal in kinase-inhibitor development is to selectively engage the desired kinase while avoiding myriad off-target kinases. However, quantifying inhibitor interactions with multiple endogenous kinases in live cells remains an unmet challenge. Here, we report the design of sulfonyl fluoride probes that covalently label a broad swath of the intracellular kinome with high efficiency. Protein crystallography and mass spectrometry confirmed a chemoselective reaction between the sulfonyl fluoride and a conserved lysine in the ATP binding site. Optimized probe 2 (XO44) covalently modified up to 133 endogenous kinases, efficiently competing with high intracellular concentrations of ATP. We employed probe 2 and label-free mass spectrometry to quantify intracellular kinase engagement by the approved drug, dasatinib. The data revealed saturable dasatinib binding to a small subset of kinase targets at clinically relevant concentrations, highlighting the utility of lysine-targeted sulfonyl fluoride probes in demanding chemoproteomic applications.
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A sulfonyl fluoride-bearing promiscuous kinase inhibitor that targets a conserved lysine in the ATP-binding site was used as a probe to profile kinase inhibitor selectivity in live cells.
“A sulfonyl fluoride-bearing promiscuous kinase inhibitor that targets a conserved lysine in the ATP-binding site was also used as a probe to profile kinase inhibitor selectivity in live cells208.”
From Advances in covalent drug discovery (2022) · cites this paper as [208]
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- Authors
- Zhao, Q., Ouyang, X., Wan, X., Gajiwala, K. S., Kath, J. C., Jones, L. H., Burlingame, A. L., & Taunton, J.
- Affiliations
- University of California, San Francisco (US)
- Pfizer (United States) (US)
- Corresponding author
- Taunton, Jack
- ORCID
- Venue
- Journal of the American Chemical Society
- Publisher
- American Chemical Society (ACS)
- Published
- 2017-01-04
- Volume
- 139
- Issue
- 2
- Pages
- 680-685
- ISSN
- 0002-7863 (ISSN-L)
- 1520-5126
- PMC
- PMC5858558
- OpenAlex
- W2569937202
- Cited by
- 388
- References
- 31
- Citations by year
- 2026: 36 · 2025: 50 · 2024: 58 · 2023: 38 · 2022: 37 · 2021: 45 · 2020: 32 · 2019: 48 · 2018: 25 · 2017: 18
- FWCI
- 14.28
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Journal of the American Chemical Society (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- eScholarship (California Digital Library) (submittedVersion)
- Funders
- Howard Hughes Medical Institute
- Pfizer
- Topics
- Physical Sciences › Chemistry › Organic Chemistry › Click Chemistry and Applications
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Keywords
- protein kinases, kinase profiling, chemoselective labeling, lysine modification, ATP binding site, dasatinib, kinase inhibitors, chemoproteomics, label-free mass spectrometry
- MeSH terms
- Dasatinib, Adenosine Triphosphate, Binding Sites, Cells, Cultured, Lysine, Models, Biological, Protein Kinases, Mass Spectrometry, Sulfinic Acids, Molecular Probes, Molecular Structure, Drug Delivery Systems
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