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Cited by 44Advanced Fluorescence Microscopy TechniquesBiochemical and Structural CharacterizationBiotin and Related Studies

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SNAP-tag2 for faster and brighter protein labeling

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Kühn, Stefanie, Nasufovic, Veselin, Wilhelm, Jonas, Kompa, Julian, de Lange, Eline M. F., Lin, Yin-Hsi, Egoldt, Cornelia, Fischer, Jonas, Lennoi, Artem, Tarnawski, Miroslaw, Reinstein, Jochen, Vlijm, Rifka, Hiblot, Julien, Johnsson, Kai·2025·Nature Chemical Biology
DOI 10.1038/s41589-025-01942-zPMC PMC12568630

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Abstract

SNAP-tag is a powerful tool for labeling proteins with synthetic fluorophores in bioimaging. However, its utility in live-cell applications can be constrained by its relatively slow labeling kinetics and the limited cell permeability of its substrates. Here, we introduce improved labeling substrates and an engineered SNAP-tag for faster labeling in vitro and in live cells. SNAP-tag2 presents a second-order rate constant with rhodamine substrates that approaches 10 7 s −1 M −1 , a 100-fold improvement over the corresponding SNAP-tag–substrate pairs. When labeled with highly fluorogenic dyes, SNAP-tag2 also shows a fivefold increase in fluorescence brightness relative to currently used SNAP-tag. The increased labeling kinetics and brightness of SNAP-tag2 translate into greatly improved performance in various live-cell (super-resolution) imaging applications.

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Metadata

Authors
Kühn, S., Nasufovic, V., Wilhelm, J., Kompa, J., de Lange, E. M. F., Lin, Y.-H., Egoldt, C., Fischer, J., Lennoi, A., Tarnawski, M., Reinstein, J., Vlijm, R., Hiblot, J., & Johnsson, K.
Affiliations
  • Max Planck Institute for Medical Research (DE)
  • University of Groningen (NL)
Corresponding author
  • Hiblot, Julien
  • Johnsson, Kai
ORCID
Venue
Nature Chemical Biology
Publisher
Springer Science and Business Media LLC
Published
2025-07-03
Volume
21
Issue
11
Pages
1754-1761
ISSN
  • 1552-4450 (ISSN-L)
  • 1552-4469
DOI
10.1038/s41589-025-01942-z
PMC
PMC12568630
OpenAlex
W4412008163
Cited by
44
References
2
Citations by year
2026: 26 · 2025: 18
FWCI
16.50
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Chemical Biology (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • University of Groningen research database (University of Groningen / Centre for Information Technology) (submittedVersion, cc-by)
Funders
  • Max-Planck-Gesellschaft
  • Deutsche Forschungsgemeinschaft (TRR 186)
  • École Polytechnique Fédérale de Lausanne
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Biotin and Related Studies
Keywords
SNAP-tag, protein labeling, fluorogenic dyes, fluorescence intensity, live-cell imaging, super-resolution imaging, membrane permeability
MeSH terms
Animals, Fluorescent Dyes, Humans, Kinetics, Proteins, Rhodamines, Staining and Labeling

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