Kinetic and Structural Characterization of the Self-Labeling Protein Tags HaloTag7, SNAP-tag, and CLIP-tag
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.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2021 |
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| Erschienen: |
2021 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:60 |
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| Enthalten in: |
Biochemistry - 60(2021), 33 vom: 24. Aug., Seite 2560-2575 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Wilhelm, Jonas [VerfasserIn] |
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| Links: |
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| Themen: |
Comparative Study |
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| Anmerkungen: |
Date Completed 19.11.2021 Date Revised 03.04.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1021/acs.biochem.1c00258 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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NLM328836982 |
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| 500 | |a Date Revised 03.04.2024 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a 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 | ||
| 650 | 4 | |a Comparative Study | |
| 650 | 4 | |a Journal Article | |
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| 650 | 7 | |a Fluorescent Dyes |2 NLM | |
| 650 | 7 | |a Recombinant Fusion Proteins |2 NLM | |
| 650 | 7 | |a Rhodamines |2 NLM | |
| 650 | 7 | |a O(6)-Methylguanine-DNA Methyltransferase |2 NLM | |
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| 700 | 1 | |a Kühn, Stefanie |e verfasserin |4 aut | |
| 700 | 1 | |a Tarnawski, Miroslaw |e verfasserin |4 aut | |
| 700 | 1 | |a Gotthard, Guillaume |e verfasserin |4 aut | |
| 700 | 1 | |a Tünnermann, Jana |e verfasserin |4 aut | |
| 700 | 1 | |a Tänzer, Timo |e verfasserin |4 aut | |
| 700 | 1 | |a Karpenko, Julie |e verfasserin |4 aut | |
| 700 | 1 | |a Mertes, Nicole |e verfasserin |4 aut | |
| 700 | 1 | |a Xue, Lin |e verfasserin |4 aut | |
| 700 | 1 | |a Uhrig, Ulrike |e verfasserin |4 aut | |
| 700 | 1 | |a Reinstein, Jochen |e verfasserin |4 aut | |
| 700 | 1 | |a Hiblot, Julien |e verfasserin |4 aut | |
| 700 | 1 | |a Johnsson, Kai |e verfasserin |4 aut | |
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