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Preparation of Arabidopsis thaliana Seedling Proteomes for Identifying Metacaspase Substrates by N-terminal COFRADIC

  • Liana Tsiatsiani
  • Simon Stael
  • Petra Van Damme
  • Frank Van Breusegem
  • Kris Gevaert
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1133)

Abstract

Proteome-wide discovery of in vivo metacaspase substrates can be obtained by positional proteomics approaches such as N-terminal COFRADIC, for example by comparing the N-terminal proteomes (or N-terminomes) of wild-type plants to transgenic plants not expressing a given metacaspase. In this chapter we describe a protocol for the preparation of plant tissue proteomes, including differential isotopic labelling allowing for a comparison of in vivo N-terminomes that serves as the starting point for N-terminal COFRADIC studies.

Key words

Metacaspases Positional proteomics N-terminal COFRADIC Protease substrates Neo-N-termini Tissue samples Degradomics 

Notes

Acknowledgments

L.T. acknowledges support from the VIB International PhD Program and the Netherlands Proteomics Centre, a program embedded in The Netherlands Genomics Initiative. P.V.D. is a Postdoctoral Fellow of the Research Foundation Flanders (FWO-Vlaanderen) and S.S. is indebted to the Special Research Fund of Ghent University for a postdoctoral fellowship. F.V.B acknowledges support from grants of the Ghent University Multidisciplinary Research Partnership “Ghent BioEconomy” 27 (project no. 01MRB510W) and of the Belgian Science Policy Office (project IAP7/29). F.V.B. and K.G. acknowledge support from the Research Foundation Flanders (FWO-Vlaanderen), research project G.0038.09.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Liana Tsiatsiani
    • 1
    • 2
  • Simon Stael
    • 3
    • 4
  • Petra Van Damme
    • 5
    • 6
  • Frank Van Breusegem
    • 3
    • 4
  • Kris Gevaert
    • 5
    • 6
  1. 1.Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical SciencesUtrecht UniversityUtrechtThe Netherlands
  2. 2.Netherlands Proteomics CenterUtrechtthe Netherlands
  3. 3.Department of Plant Systems BiologyVIBGhentBelgium
  4. 4.Department of Plant Biotechnology and BioinformaticsGhent UniversityGhentBelgium
  5. 5.Department of Medical Protein ResearchVIBGhentBelgium
  6. 6.Department of BiochemistryGhent UniversityGhentBelgium

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