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Identification of phosphorylated proteins.
Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
2007 (English)In: Methods in Molecular Biology, ISSN 1064-3745, E-ISSN 1940-6029, Vol. 355, 305-316 p.Article in journal (Refereed) Published
Abstract [en]

Reversible protein phosphorylation is crucially involved in all aspects of plant cell physiology. The highly challenging task of revealing and characterizing the dynamic protein phosphorylation networks in plants has only recently begun to become feasible, owing to application of dedicated proteomics and mass spectrometry techniques. The experimental methodology that identified most of the presently known proteins phosphorylated in vivo is based on protein cleavage with trypsin, following chromatographic enrichment of phosphorylated peptides and mass spectrometric fragmentation and sequencing of these phosphopeptides. This procedure is most efficient when it is limited to the tryptic digestion of proteins in distinct isolated fractions or compartments of plant cells. Immobilized metal affinity chromatography (IMAC) is most useful for phosphopeptide enrichment after methylation of the peptides in the complex protein digests. The following tandem mass spectrometry of the isolated phosphopeptides results in both identification of phosphorylated proteins and mapping of the in vivo phosphorylation sites. The relative quantitation of the extent of phosphorylation at individual protein modification sites may be accomplished by either stable isotope labeling technique or dedicated liquid chromatography-mass spectrometry measurements.

Place, publisher, year, edition, pages
2007. Vol. 355, 305-316 p.
Keyword [en]
IMAC – mass spectrometry – protein phosphorylation – phosphoproteins – phosphopeptides – peptide
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:liu:diva-125936DOI: 10.1385/1-59745-227-0:305PubMedID: 17093319OAI: oai:DiVA.org:liu-125936DiVA: diva2:910444
Available from: 2016-03-09 Created: 2016-03-09 Last updated: 2016-03-21

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Turkina, Maria VVener, Alexander V
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Division of Cell BiologyFaculty of Medicine and Health Sciences
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Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)

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