Yeast membrane proteomics using leucine metabolic labelling: bioinformatic data processing and exemplary application to the ER-intramembrane protease Ypf1
We describe in detail the usage of leucine metabolic labelling in yeast in order to monitor quantitative proteome alterations, e.g. upon removal of a protease. Since laboratory yeast strains are typically leucine auxotroph, metabolic labelling with trideuterated leucine (d3-leucine) is a straightfor...
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| Main Authors: | , , , , , |
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| Format: | Article (Journal) |
| Language: | English |
| Published: |
15 July 2016
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| In: |
Biochimica et biophysica acta. Proteins and proteomics
Year: 2016, Volume: 1864, Issue: 10, Pages: 1363-1371 |
| ISSN: | 1878-1454 |
| DOI: | 10.1016/j.bbapap.2016.07.002 |
| Online Access: | Verlag, lizenzpflichtig, Volltext: https://doi.org/10.1016/j.bbapap.2016.07.002 Verlag, lizenzpflichtig, Volltext: http://www.sciencedirect.com/science/article/pii/S1570963916301388 
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| Author Notes: | Lars Nilse, Dönem Avci, Patrick Heisterkamp, Oliver Serang, Marius K. Lemberg, Oliver Schilling |
| Summary: | We describe in detail the usage of leucine metabolic labelling in yeast in order to monitor quantitative proteome alterations, e.g. upon removal of a protease. Since laboratory yeast strains are typically leucine auxotroph, metabolic labelling with trideuterated leucine (d3-leucine) is a straightforward, cost-effective, and ubiquitously applicable strategy for quantitative proteomic studies, similar to the widely used arginine/lysine metabolic labelling method for mammalian cells. We showcase the usage of advanced peptide quantification using the FeatureFinderMultiplex algorithm (part of the OpenMS software package) for robust and reliable quantification. Furthermore, we present an OpenMS bioinformatics data analysis workflow that combines accurate quantification with high proteome coverage. In order to enable visualization, peptide-mapping, and sharing of quantitative proteomic data, especially for membrane-spanning and cell-surface proteins, we further developed the web-application Proteator (http://proteator.appspot.com). Due to its simplicity and robustness, we expect metabolic leucine labelling in yeast to be of great interest to the research community. As an exemplary application, we show the identification of the copper transporter Ctr1 as a putative substrate of the ER-intramembrane protease Ypf1 by yeast membrane proteomics using d3-leucine isotopic labelling. |
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| Item Description: | Gesehen am 03.11.2020 |
| Physical Description: | Online Resource |
| ISSN: | 1878-1454 |
| DOI: | 10.1016/j.bbapap.2016.07.002 |