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Journal Article

Kinetic and structural characterization of the self-labeling protein tags HaloTag7, SNAP-tag, and CLIP-tag

MPS-Authors
/persons/resource/persons264410

Wilhelm,  Jonas
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons264412

Kühn,  Stefanie
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons117897

Tarnawski,  Miroslaw
Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons264414

Tünnermann,  Jana
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons264416

Mertes,  Nicole
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons212625

Xue,  Lin
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons94928

Reinstein,  Jochen
Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons212621

Hiblot,  Julien
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

/persons/resource/persons203696

Johnsson,  Kai
Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society;

External Resource

https://doi.org/10.1021/acs.biochem.1c00258
(Any fulltext)

https://ndownloader.figstatic.com/files/29020878
(Supplementary material)

https://pubs.acs.org/doi/pdf/10.1021/acs.biochem.1c00258
(Any fulltext)

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Citation

Wilhelm, J., Kühn, S., Tarnawski, M., Gotthard, G., Tünnermann, J., Tänzer, T., et al. (2021). Kinetic and structural characterization of the self-labeling protein tags HaloTag7, SNAP-tag, and CLIP-tag. Biochemistry, 60(33), 2560-2575. doi:10.1021/acs.biochem.1c00258.


Cite as: https://hdl.handle.net/21.11116/0000-0008-FCD2-F
Abstract
The self-labeling protein tags (SLPs) HaloTag7, SNAP-tag, and CLIP-tag allow the covalent labeling of fusion proteins with synthetic molecules for applications in bioimaging and biotechnology. To guide the selection of an SLP-substrate pair and provide guidelines for the design of substrates, we report a systematic and comparative study of the labeling kinetics and substrate specificities of HaloTag7, SNAP-tag, and CLIP-tag. HaloTag7 reaches almost diffusion-limited labeling rate constants with certain rhodamine substrates, which are more than 2 orders of magnitude higher than those of SNAP-tag for the corresponding substrates. SNAP-tag labeling rate constants, however, are less affected by the structure of the label than those of HaloTag7, which vary over 6 orders of magnitude for commonly employed substrates. Determining the crystal structures of HaloTag7 and SNAP-tag labeled with fluorescent substrates allowed us to rationalize their substrate preferences. We also demonstrate how these insights can be exploited to design substrates with improved labeling kinetics.