A designed fusion tag for soluble expression and selective separation of extracellular domains of fibroblast growth factor receptors
© 2021. The Author(s)..
Fibroblast growth factor receptors (FGFRs) generate various transduction signals by interaction with fibroblast growth factors (FGFs) and are involved in various biological functions such as cell proliferation, migration, and differentiation. Malfunction of these proteins may lead to the development of various diseases, including cancer. Accordingly, FGFRs are considered an alternative therapeutic target for protein and/or gene therapy. However, the screening of antagonists or agonists of FGFRs is challenging due to their complex structural features associated with protein expression. Herein, we conducted the development of a protease-free cleavable tag (PFCT) for enhancing the solubility of difficult-to express protein by combining maltose-binding protein (MBP) and the C-terminal region of Npu intein. To validate the availability of the resulting tag for the functional production of extracellular domains of FGFRs (Ec_FGFRs), we performed fusion of PFCT with the N-terminus of Ec_FGFRs and analyzed the expression patterns. Almost all PFCT-Ec_FGFR fusion proteins were mainly detected in the soluble fraction except for Ec_FGFR4. Upon addition of the N-terminal region of Npu intein, approximately 85% of the PFCT-Ec_FGFRs was separated into PFCT and Ec_FGFR via intein-mediated cleavage. Additionally, the structural integrity of Ec_FGFR was confirmed by affinity purification using heparin column. Taken together, our study demonstrated that the PFCT could be used for soluble expression and selective separation of Ec_FGFRs.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2021 |
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| Erschienen: |
2021 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:11 |
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| Enthalten in: |
Scientific reports - 11(2021), 1 vom: 02. Nov., Seite 21453 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Cheong, Dae-Eun [VerfasserIn] |
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| Links: |
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| Themen: |
Journal Article |
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| Anmerkungen: |
Date Completed 24.01.2022 Date Revised 24.01.2022 published: Electronic Citation Status MEDLINE |
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| doi: |
10.1038/s41598-021-01029-4 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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NLM332668789 |
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| 245 | 1 | 2 | |a A designed fusion tag for soluble expression and selective separation of extracellular domains of fibroblast growth factor receptors |
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| 520 | |a © 2021. The Author(s). | ||
| 520 | |a Fibroblast growth factor receptors (FGFRs) generate various transduction signals by interaction with fibroblast growth factors (FGFs) and are involved in various biological functions such as cell proliferation, migration, and differentiation. Malfunction of these proteins may lead to the development of various diseases, including cancer. Accordingly, FGFRs are considered an alternative therapeutic target for protein and/or gene therapy. However, the screening of antagonists or agonists of FGFRs is challenging due to their complex structural features associated with protein expression. Herein, we conducted the development of a protease-free cleavable tag (PFCT) for enhancing the solubility of difficult-to express protein by combining maltose-binding protein (MBP) and the C-terminal region of Npu intein. To validate the availability of the resulting tag for the functional production of extracellular domains of FGFRs (Ec_FGFRs), we performed fusion of PFCT with the N-terminus of Ec_FGFRs and analyzed the expression patterns. Almost all PFCT-Ec_FGFR fusion proteins were mainly detected in the soluble fraction except for Ec_FGFR4. Upon addition of the N-terminal region of Npu intein, approximately 85% of the PFCT-Ec_FGFRs was separated into PFCT and Ec_FGFR via intein-mediated cleavage. Additionally, the structural integrity of Ec_FGFR was confirmed by affinity purification using heparin column. Taken together, our study demonstrated that the PFCT could be used for soluble expression and selective separation of Ec_FGFRs | ||
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| 700 | 1 | |a Yoo, Su-Kyoung |e verfasserin |4 aut | |
| 700 | 1 | |a Lee, Hun-Dong |e verfasserin |4 aut | |
| 700 | 1 | |a Kim, Geun-Joong |e verfasserin |4 aut | |
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