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Dimethyl-Labeling-Based Quantification of the Lysine Acetylome and Proteome of Plants

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Photorespiration

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1653))

Abstract

Photorespiratory enzymes in different cellular compartments have been reported to be posttranslational modified by phosphorylation, disulfide formation, S-nitrosylation, glutathionylation, and lysine acetylation. However, not much is known yet about the function of these modifications to regulate the activities, localizations, or interactions of the proteins in this metabolic pathway. Hence, it will be of great importance to study these modifications and their temporal and conditional occurrence in more detail. Here, we focus on the analysis of lysine acetylation as a relatively newly discovered modification on plant metabolic enzymes. The acetylation of lysine residues within proteins is a highly conserved and reversible posttranslational modification which occurs in all living organisms. First discovered on histones and implied in the regulation of gene expression, lysine acetylation also occurs on a diverse set of cellular proteins in different subcellular compartments and is particularly abundant on metabolic enzymes. Upon lysine acetylation, the function of proteins can be modulated due to the loss of the positive charge of the lysine residue. Lysine acetylation was also discovered on proteins involved in photosynthesis and novel tools are needed to study the regulation of this modification in dependence on the environmental conditions, tissues, or plant genotype. This chapter describes a method for the identification and relative quantification of lysine-acetylated proteins in plant tissues using a dimethyl labeling technique combined with an anti-acetyl lysine antibody enrichment strategy. Here, we describe the protein purification, labeling of trypsinated peptides, as well as immuno-enrichment of lysine-acetylated peptides followed by liquid chromatography tandem mass spectrometry (LC-MS/MS) data acquisition and analysis.

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Author information

Authors and Affiliations

  1. Plant Proteomics, Max-Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Köln, Germany

    Ines Lassowskat, Markus Hartl & Iris Finkemeier

  2. Mass Spectrometry Facility, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Dr. Bohr-Gasse 3, 1030, Vienna, Austria

    Markus Hartl

  3. Proteomics and Signal Transduction, Max-Planck Institute of Biochemistry, Am Klopferspitz 18, 82152, Martinsried, Germany

    Fabian Hosp, Paul J. Boersema & Matthias Mann

  4. Department of Biology, Institute of Biochemistry, ETH Zurich, Otto-Stern-Weg 3, 8093, Zurich, Switzerland

    Paul J. Boersema

  5. Institute of Plant Biology and Biotechnology, University of Münster, Schlossplatz 7, 48149, Münster, Germany

    Iris Finkemeier

Authors
  1. Ines Lassowskat
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  2. Markus Hartl
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  3. Fabian Hosp
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  4. Paul J. Boersema
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  5. Matthias Mann
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  6. Iris Finkemeier
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Corresponding author

Correspondence to Iris Finkemeier .

Editor information

Editors and Affiliations

  1. Max Planck Institute of Molecular Plant Physiology, Potsdam-Golm, Germany

    Alisdair R. Fernie

  2. Plant Physiology Department, University of Rostock, Rostock, Germany

    Hermann Bauwe

  3. Institute of Plant Biochemistry, Cluster of Excellence on Plant Sciences (CEPLAS), Heinrich-Heine-University, Düsseldorf, Germany

    Andreas P.M. Weber

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Lassowskat, I., Hartl, M., Hosp, F., Boersema, P.J., Mann, M., Finkemeier, I. (2017). Dimethyl-Labeling-Based Quantification of the Lysine Acetylome and Proteome of Plants. In: Fernie, A., Bauwe, H., Weber, A. (eds) Photorespiration. Methods in Molecular Biology, vol 1653. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7225-8_5

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  • DOI: https://doi.org/10.1007/978-1-4939-7225-8_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7224-1

  • Online ISBN: 978-1-4939-7225-8

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