Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase
Abstract Background Setting of graded levels of a protein for in vivo studies by controlled gene expression has inconveniences, and we here explore the use of the t-degron technique instead. Results In a yeast t-degron (ubiquitin-argDHFRts)- phosphoglycerate mutase (GPM1) fusion strain, increasing p...
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BMC
2002-07-01
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| Series: | BMC Genetics |
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| Online Access: | https://doi.org/10.1186/1471-2156-3-39 |
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| author | Katja Heidrich Dan G Fraenkel |
| author_facet | Katja Heidrich Dan G Fraenkel |
| author_sort | Katja Heidrich |
| collection | DOAJ |
| description | Abstract Background Setting of graded levels of a protein for in vivo studies by controlled gene expression has inconveniences, and we here explore the use of the t-degron technique instead. Results In a yeast t-degron (ubiquitin-argDHFRts)- phosphoglycerate mutase (GPM1) fusion strain, increasing periods of exposure to the non-permissive temperature 37°C, even in the presence of cycloheximide, gave decreasing function, as assessed at 23°C in vivo by glucose metabolism and confirmed by immunoblot. Conclusion An ideal system would set a range of lower levels of a protein, do so without compensating protein synthesis, and give stable activity for in vitro comparisons. Although the first two aims appear obtainable, the third was not in this example of the application, limiting its uses for some but not all purposes. |
| format | Article |
| id | doaj-art-1dc57bcd580943bfb111b2cd59af1074 |
| institution | OA Journals |
| issn | 1471-2156 |
| language | English |
| publishDate | 2002-07-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Genetics |
| spelling | doaj-art-1dc57bcd580943bfb111b2cd59af10742025-08-20T02:17:46ZengBMCBMC Genetics1471-21562002-07-01311610.1186/1471-2156-3-39Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutaseKatja Heidrich0Dan G Fraenkel1Dept. of Microbiology and Molecular Genetics, Harvard Medical SchoolDept. of Microbiology and Molecular Genetics, Harvard Medical SchoolAbstract Background Setting of graded levels of a protein for in vivo studies by controlled gene expression has inconveniences, and we here explore the use of the t-degron technique instead. Results In a yeast t-degron (ubiquitin-argDHFRts)- phosphoglycerate mutase (GPM1) fusion strain, increasing periods of exposure to the non-permissive temperature 37°C, even in the presence of cycloheximide, gave decreasing function, as assessed at 23°C in vivo by glucose metabolism and confirmed by immunoblot. Conclusion An ideal system would set a range of lower levels of a protein, do so without compensating protein synthesis, and give stable activity for in vitro comparisons. Although the first two aims appear obtainable, the third was not in this example of the application, limiting its uses for some but not all purposes.https://doi.org/10.1186/1471-2156-3-39Phosphoglycerate MutaseNdeI SiteRequire Protein SynthesisDifco Yeast ExtractCUP1 Promoter |
| spellingShingle | Katja Heidrich Dan G Fraenkel Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase BMC Genetics Phosphoglycerate Mutase NdeI Site Require Protein Synthesis Difco Yeast Extract CUP1 Promoter |
| title | Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase |
| title_full | Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase |
| title_fullStr | Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase |
| title_full_unstemmed | Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase |
| title_short | Setting of graded levels of a protein in yeast by a t-degron technique as applied to phosphoglycerate mutase |
| title_sort | setting of graded levels of a protein in yeast by a t degron technique as applied to phosphoglycerate mutase |
| topic | Phosphoglycerate Mutase NdeI Site Require Protein Synthesis Difco Yeast Extract CUP1 Promoter |
| url | https://doi.org/10.1186/1471-2156-3-39 |
| work_keys_str_mv | AT katjaheidrich settingofgradedlevelsofaproteininyeastbyatdegrontechniqueasappliedtophosphoglyceratemutase AT dangfraenkel settingofgradedlevelsofaproteininyeastbyatdegrontechniqueasappliedtophosphoglyceratemutase |