Isolation of Plant Cell Wall Proteins

  • Elisabeth Jamet
  • Georges Boudart
  • Giséle Borderies
  • Stephane Charmont
  • Claude Lafitte
  • Michel Rossignol
  • Herve Canut
  • Rafael Pont-Lezica
Part of the Methods in Molecular Biology™ book series (MIMB, volume 425)


The quality of a proteomic analysis of a cell compartment strongly depends on the reliability of the isolation procedure for the cell compartment of interest. Plant cell walls possess specific drawbacks: (1) the lack of a surrounding membrane may result in the loss of cell wall proteins (CWP) during the isolation procedure; (2) polysaccharide networks of cellulose, hemicelluloses, and pectins form potential traps for contaminants such as intracellular proteins; (3) the presence of proteins interacting in many different ways with the polysaccharide matrix require different procedures to elute them from the cell wall. Three categories of CWP are distinguished: labile proteins that have little or no interactions with cell wall components, weakly bound proteins extractable with salts, and strongly bound proteins. Two alternative protocols are decribed for cell wall proteomics: (1) nondestructive techniques allowing the extraction of labile or weakly bound CWP without damaging the plasma membrane; (2) destructive techniques to isolate cell walls from which weakly or strongly bound CWP can be extracted. These protocols give very low levels of contamination by intracellular proteins. Their application should lead to a realistic view of the cell wall proteome at least for labile and weakly bound CWP extractable by salts.

Key Words

Arabidopsis thaliana bioinformatics cell fractionation cell wall cell wall protein plant proteomics 



The authors are grateful to the Universit’e Paul Sabatier (Toulouse III, France) and the CNRS for support.


  1. 1.
    Hunter, T. C., Andon, N. L., Koller, A., Yates, J. R. and Haynes, P. A. (2002) The functional proteomics toolbox: methods and applications. J. Chromatogr. B 782, 161–181.CrossRefGoogle Scholar
  2. 2.
    Carpita, N. and Gibeaut, D. (1993) Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth. Plant J. 3, 1–30.CrossRefPubMedGoogle Scholar
  3. 3.
    Cosgrove, D. J. (2005) Growth of the plant cell wall. Nat. Rev. Mol. Cell. Biol. 6, 850–861.CrossRefPubMedGoogle Scholar
  4. 4.
    Jamet, E., Canut, H., Boudart, G. and Pont-Lezica, R. F. (2006) Cell wall proteins: a new insight through proteomics. Trends Plant Sci. 11, 33–9.CrossRefPubMedGoogle Scholar
  5. 5.
    Varner, J. E. and Lin, L.-S. (1989) Plant cell wall architecture. Cell 56, 231–39.CrossRefPubMedGoogle Scholar
  6. 6.
    Brady, J. D., Sadler, I. H., and Fry, S.C. (1996) Di-isodityrosine, a novel tetrameric derivative of tyrosine in plant cell wall proteins: a new potential cross-link. J. Biochem. 315, 323–27.Google Scholar
  7. 7.
    Schnabelrauch, L. S., Kieliszewski, M. J., Upham, B. L., Alizedeh, H. and Lamport, D. T. A. (1996) Isolation of pI 4.6 extensin peroxidase from tomato cell suspension cultures and identification of Val-Tyr-Lys as putative intermolecular cross-link site. Plant J. 9, 477–89.CrossRefPubMedGoogle Scholar
  8. 8.
    Shah, K., Penel, C., Gagnon, J., and Dunand, C. (2004) Purification and identification of a Ca+2-pectate binding peroxidase from Arabidopsis leaves. Phytochem. 65, 307–12.CrossRefGoogle Scholar
  9. 9.
    Boudart, G., Jamet, E., Rossignol, M., et al. (2005) Cell wall proteins in apoplastic fluids of Arabidopsis thaliana rosettes: Identification by mass spectrometry and bioinformatics. Proteomics 5, 212–21.CrossRefPubMedGoogle Scholar
  10. 10.
    Borderies, G., Jamet, E., Lafitte, C., et al. (2003) Proteomics of loosely bound cell wall proteins of Arabidopsis thaliana cell suspension cultures: a critical analysis. Electrophoresis 24, 3421–32.CrossRefPubMedGoogle Scholar
  11. 11.
    Charmont, S., Jamet, E., Pont-Lezica, R., and Canut, H. (2005) Proteomic analysis of secreted proteins from Arabidopsis thaliana seedlings: improved recovery following removal of phenolic compounds. Phytochem. 66, 453–61.CrossRefGoogle Scholar
  12. 12.
    Held, M. A., Tan, L., Kamyab, A., Hare, M., Shpak, E. and Kieliszewski, M. J. (2004) Di-isodityrosine is the intermolecular cross-link of isodityrosine-rich extensin analogs cross-linked in vitro. J. Biol. Chem. 279, 55474–82.CrossRefPubMedGoogle Scholar
  13. 13.
    Miller, J. G. and Fry, S. C. (1992) Production and harvesting of ionically wall-bound extensin from living cell suspension cultures. Plant Cell Tissue Organ Cult. 31, 61–66.Google Scholar
  14. 14.
    Murashige, T. and Skoog, F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol. Plant. 15, 473–97.CrossRefGoogle Scholar
  15. 15.
    Loomis, W. D. (1974) Overcoming problems of phenolics and quinones in the isolation of plant enzymes and organelles. Meth.Enzymol. 31, 528–45.CrossRefPubMedGoogle Scholar
  16. 16.
    Ramagli, L. S. and Rodriguez, L. V. (1985) Quantitation of microgram amounts of protein in two-dimensional polyacrylamide electrophoresis sample buffer. Electrophoresis 6, 559–63.CrossRefGoogle Scholar
  17. 17.
    Axelos, M., Curie, C., Mazzolini, L., Bardet, C. and Lescure, B. (1992) A protocol for transient gene expression in Arabidopsis thaliana protoplasts isolated from cell suspension cultures. Plant Physiol. Biochem. 30, 123–28.Google Scholar
  18. 18.
    Voigt, J. (1985) Extraction by lithium chloride of hydroxyproline-rich glycoproteins from intact cells of Chlamydomonas reinhardii. Planta 164, 379–89.CrossRefGoogle Scholar
  19. 19.
    Smith, J., Muldoon, E., and Lamport, D. (1984) Isolation of extensin precursors by direct elution of intact tomato cell suspension cultures. Phytochem. 23, 1233–39.CrossRefGoogle Scholar
  20. 20.
    Angyal, S. (1989) Complexes of metal cations with carbohydrates in solution. Adv. Carbohydr. Chem. Biochem. 47, 1–44.CrossRefGoogle Scholar
  21. 21.
    van Buren, J. (1991) Function of pectin in plant tissue structure and firmness in, The Chemistry and Technology of Pectin (Walter, R. H. ed.), Academic Press, New York, pp. 1–22.Google Scholar
  22. 22.
    Bardy, N., Carrasco, A., Galaud, J. P., Pont-Lezica, R. and Canut, H. (1998) Free-flow electrophoresis for fractionation of Arabidopsis thaliana membranes. Electrophoresis 19, 1145–53.CrossRefPubMedGoogle Scholar
  23. 23.
    Rabilloud, T. (2002) Two-dimensional gel electrophoresis in proteomics: old, old fashioned, but still climbs up the mountains. Proteomics 2, 3–10.CrossRefPubMedGoogle Scholar
  24. 24.
    Stasyk, T. and Huber, L. A. (2004) ,Zooming in fractionation strategies in proteomics. Proteomics 4, 3704–16.CrossRefPubMedGoogle Scholar
  25. 25.
    Lescuyer, P., Hochstrasser, D. F., Sanchez, J. C. (2004) Comprehensive proteome analysis by chromatographic protein prefractionation. Electrophoresis 25,1125–1135.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Elisabeth Jamet
    • 1
  • Georges Boudart
    • 1
  • Giséle Borderies
    • 1
  • Stephane Charmont
    • 2
  • Claude Lafitte
    • 1
  • Michel Rossignol
    • 1
  • Herve Canut
    • 1
  • Rafael Pont-Lezica
    • 1
  1. 1.UMR 5546 CNRS-Université Paul Sabatier-Toulouse IIICastanet-TolosanFrance
  2. 2.Novartis Pharma AGBaselSwitzerland

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