Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron
Abstract Methods that allow for the manipulation of genes or their products have been highly fruitful for biomedical research. Here, we describe a method that allows the control of protein abundance by a genetically encoded regulatory system. We developed a dormant N‐degron that can be attached to t...
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| Format: | Article |
| Language: | English |
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Springer Nature
2009-04-01
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| Series: | Molecular Systems Biology |
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| Online Access: | https://doi.org/10.1038/msb.2009.25 |
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| _version_ | 1849225788241477632 |
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| author | Christof Taxis Gunter Stier Roberta Spadaccini Michael Knop |
| author_facet | Christof Taxis Gunter Stier Roberta Spadaccini Michael Knop |
| author_sort | Christof Taxis |
| collection | DOAJ |
| description | Abstract Methods that allow for the manipulation of genes or their products have been highly fruitful for biomedical research. Here, we describe a method that allows the control of protein abundance by a genetically encoded regulatory system. We developed a dormant N‐degron that can be attached to the N‐terminus of a protein of interest. Upon expression of a site‐specific protease, the dormant N‐degron becomes deprotected. The N‐degron then targets itself and the attached protein for rapid proteasomal degradation through the N‐end rule pathway. We use an optimized tobacco etch virus (TEV) protease variant combined with selective target binding to achieve complete and rapid deprotection of the N‐degron‐tagged proteins. This method, termed TEV protease induced protein inactivation (TIPI) of TIPI‐degron (TDeg) modified target proteins is fast, reversible, and applicable to a broad range of proteins. TIPI of yeast proteins essential for vegetative growth causes phenotypes that are close to deletion mutants. The features of the TIPI system make it a versatile tool to study protein function in eukaryotes and to create new modules for synthetic or systems biology. |
| format | Article |
| id | doaj-art-b7256db0f2af493a82fe9fe5f5f8f2b7 |
| institution | Kabale University |
| issn | 1744-4292 |
| language | English |
| publishDate | 2009-04-01 |
| publisher | Springer Nature |
| record_format | Article |
| series | Molecular Systems Biology |
| spelling | doaj-art-b7256db0f2af493a82fe9fe5f5f8f2b72025-08-24T11:59:22ZengSpringer NatureMolecular Systems Biology1744-42922009-04-01511710.1038/msb.2009.25Efficient protein depletion by genetically controlled deprotection of a dormant N‐degronChristof Taxis0Gunter Stier1Roberta Spadaccini2Michael Knop3EMBL, Cell Biology and Biophysics UnitEMBL, Cell Biology and Biophysics UnitEMBL, Cell Biology and Biophysics UnitEMBL, Cell Biology and Biophysics UnitAbstract Methods that allow for the manipulation of genes or their products have been highly fruitful for biomedical research. Here, we describe a method that allows the control of protein abundance by a genetically encoded regulatory system. We developed a dormant N‐degron that can be attached to the N‐terminus of a protein of interest. Upon expression of a site‐specific protease, the dormant N‐degron becomes deprotected. The N‐degron then targets itself and the attached protein for rapid proteasomal degradation through the N‐end rule pathway. We use an optimized tobacco etch virus (TEV) protease variant combined with selective target binding to achieve complete and rapid deprotection of the N‐degron‐tagged proteins. This method, termed TEV protease induced protein inactivation (TIPI) of TIPI‐degron (TDeg) modified target proteins is fast, reversible, and applicable to a broad range of proteins. TIPI of yeast proteins essential for vegetative growth causes phenotypes that are close to deletion mutants. The features of the TIPI system make it a versatile tool to study protein function in eukaryotes and to create new modules for synthetic or systems biology.https://doi.org/10.1038/msb.2009.25developmentprotein inactivationregulated protein inactivationtissue‐specific |
| spellingShingle | Christof Taxis Gunter Stier Roberta Spadaccini Michael Knop Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron Molecular Systems Biology development protein inactivation regulated protein inactivation tissue‐specific |
| title | Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron |
| title_full | Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron |
| title_fullStr | Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron |
| title_full_unstemmed | Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron |
| title_short | Efficient protein depletion by genetically controlled deprotection of a dormant N‐degron |
| title_sort | efficient protein depletion by genetically controlled deprotection of a dormant n degron |
| topic | development protein inactivation regulated protein inactivation tissue‐specific |
| url | https://doi.org/10.1038/msb.2009.25 |
| work_keys_str_mv | AT christoftaxis efficientproteindepletionbygeneticallycontrolleddeprotectionofadormantndegron AT gunterstier efficientproteindepletionbygeneticallycontrolleddeprotectionofadormantndegron AT robertaspadaccini efficientproteindepletionbygeneticallycontrolleddeprotectionofadormantndegron AT michaelknop efficientproteindepletionbygeneticallycontrolleddeprotectionofadormantndegron |