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

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.

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 Creators:
Wilhelm, Jonas1, Author              
Kühn, Stefanie1, Author              
Tarnawski, Miroslaw2, Author              
Gotthard, Guillaume, Author
Tünnermann, Jana1, Author              
Tänzer, Timo, Author
Karpenko, Julie, Author
Mertes, Nicole1, Author              
Xue, Lin1, Author              
Uhrig, Ulrike, Author
Reinstein, Jochen2, Author              
Hiblot, Julien1, Author              
Johnsson, Kai1, Author              
Affiliations:
1Chemical Biology, Max Planck Institute for Medical Research, Max Planck Society, ou_2364732              
2Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Max Planck Society, ou_1497700              

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Free keywords: Kinetic modeling, Fluorescence, Labeling, Fluorophores, Kinetics
 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.

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Language(s): eng - English
 Dates: 2021-07-232021-05-032021-08-022021-08-24
 Publication Status: Published in print
 Pages: 16
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Degree: -

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Title: Biochemistry
Source Genre: Journal
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Publ. Info: Columbus, Ohio : American Chemical Society
Pages: - Volume / Issue: 60 (33) Sequence Number: - Start / End Page: 2560 - 2575 Identifier: ISSN: 0006-2960
CoNE: https://pure.mpg.de/cone/journals/resource/954925384103