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Analytical Strategies in Mass Spectrometry-Based Phosphoproteomics

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Gel-Free Proteomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 753))

Abstract

Phosphoproteomics, the systematic study of protein phosphorylation events and cell signaling networks in cells and tissues, is a rapidly evolving branch of functional proteomics. Current phosphoproteomics research provides a large toolbox of strategies and protocols that may assist researchers to reveal key regulatory events and phosphorylation-mediated processes in the cell and in whole organisms. We present an overview of sensitive and robust analytical methods for phosphopeptide analysis, including calcium phosphate precipitation and affinity enrichment methods such as IMAC and TiO2. We then discuss various tandem mass spectrometry approaches for phosphopeptide sequencing and quantification, and we consider aspects of phosphoproteome data analysis and interpretation. Efficient integration of these stages of phosphoproteome analysis is highly important to ensure a successful outcome of large-scale experiments for studies of phosphorylation-mediated protein regulation.

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Acknowledgments

Research in O.N.J.’s laboratory is funded by grants from the Danish Research Councils, The Danish National Research Foundation, and The Lundbeck Foundation. H.R. was funded by a Ph.D. fellowship from the German Research Council (DFG, grant number WI 2044/5-1).

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230, Odense, Denmark

    Juanying Ye & Ole N. Jensen

  2. Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow G4 0RE, Scotland, UK

    Heidi Rosenqvist

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  1. Heidi Rosenqvist
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  1. , VIB Department of Medical Protein Resear, VIB and Ghent University, Albert Baertsoenkaai 3, Gent, 9000, Belgium

    Kris Gevaert

  2. , VIB Department of Medical Protein Resear, VIB and Ghent University, Albert Baertsoenkaai 3, Gent, 9000, Belgium

    Joël Vandekerckhove

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Rosenqvist, H., Ye, J., Jensen, O.N. (2011). Analytical Strategies in Mass Spectrometry-Based Phosphoproteomics. In: Gevaert, K., Vandekerckhove, J. (eds) Gel-Free Proteomics. Methods in Molecular Biology, vol 753. Humana Press. https://doi.org/10.1007/978-1-61779-148-2_13

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