Abstract
Large-scale and high-throughput approaches play an essential role in the study of the highly complex biological systems. In plant biology, obtaining the complete genome sequence of the model plant Arabidopsis thaliana has definitively encouraged the incorporation of a completely new set of tools for the identification, characterization, and quantification of its proteome. Proteomics is a new branch of the protein sciences that makes use of these tools and the set of experimental methods specifically designed for them. In this chapter, we describe a complete method for the isolation, fractionation, and identification of a set of proteins obtained from Arabidopsis thaliana cells, stressing methodological aspects that could be of potential interest.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Anderson, N. G., Matheson, A., and Anderson, N. L. (2001) Back to the future: the human protein index (HPI) and the agenda for post-proteomic biology. Proteomics 1, 3–12.
Blackstock, W. P. and Weir, M. P. (1999) Proteomics: quantitative and physical mapping of cellular proteins. Trends Biotechnol. 17, 121–127.
Kersten, B., Bürkle, L., Eckehard, J. K., et al. (2002) Large-scale plant proteomics. Plant Mol. Biol. 48, 133–141.
Yamazaki, D., Motohashi, K., Kasama, T., Hara, Y., and Hisabori, T. (2004) Target proteins of the cytosolic thioredoxins in Arabidopsis thaliana. Plant Cell Physiol. 45, 18–27.
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, 3421–3432.
Zabrouskov, V., Giacomelli, L., Van Wijk, K. J., and McLafferty, F. W. (2003) A new approach for plant proteomics: characterization of chloroplast proteins of Arabidopsis thaliana by top-down mass spectrometry. Mol. Cell. Proteomics 2, 1253–1260.
Froehlich, J. E., Wilkerson, C. G., Ray, W. K., McAndrew, R. S., Osteryoung, K. W., Gage, D. A., and Phinney, B. S. (2003) Proteomic study of the Arabidopsis thaliana chloroplastic envelope membrane utilizing alternatives to traditional two-dimensional electrophoresis. J. Proteome Res. 2, 413–425.
Ferro, M., Salvi, D., Brugiere, S., et al. (2003) Proteomics of the chloroplast envelope membranes from Arabidopsis thaliana. Mol. Cell. Proteomics 2, 325–345.
Luche, S., Santoni, V., and Rabilloud, T. (2003) Evaluation of nonionic and zwitterionic detergents as membrane protein solubilizers in two-dimensional electrophoresis. Proteomics 3, 249–253.
Gallardo, K., Job, C., Groot, S. P., Puype, M., Demol, H., Vandekerckhove, J., and Job, D. (2002) Proteomics of Arabidpsis seed germination. A comparative study of wild-type and gibberellin-deficient seeds. Plant Physiol. 129, 823–837.
O’Farrell, P. H. (1975). High resolution two-dimensional electrophoresis of proteins. J. Biol. Chem. 250, 4007–4021.
Görg, A., Obermaier, C., Boguth, G., Harder, A., Scheibe, B., Wildgruber, R., and Weiss, W. (2000) The current state of two-dimensional electrophoresis with immobilized pH gradients. Electrophoresis 21, 1037–1053.
Henzel, W.J., Billeci, T. M., Stults, J. T., Wong, S. C., Grimley, C., and Watanabe, C. (1993) Identifying proteins from two-dimensional gels by molecular mass searching of peptide fragments in protein sequence databases. Proc. Natl. Acad. Sci. USA 90, 5011–5015.
Mann, M., Hojrup, P., and Roepstorff, P. (1993) Use of mass spectrometric molecular weight information to identify proteins in sequence databases. Biol. Mass Spectrom. 22, 338–345.
Pappin, D. J. C., Hojrup, P., and Bleasby, A. J. (1993) Rapid identification of proteins by peptide-mass fingerprinting. Curr. Biol. 3, 327–332.
Yates, J. R. III, Speicher, S., Griffin, P. R., and Hunkapiller, T. (1993) Peptide mass maps: a highly informative approach to protein identification. Anal. Biochem. 214, 397–408.
Link, A. J., Eng, J., Schieltz, D. M., et al. (1999). Nature Biotechnol. 17, 676–682.
Simpson, R. J. (2003) Proteins and Proteomics. A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
Pennington, S. R. and Dunn, M. J. (2001) Proteomics: From Protein Sequence to Function. BIOS Scientific Publishers, Oxford, UK.
Shevchenko, A., Wilm, M., Vorm, O., and Mann, M. (1996) Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal. Chem. 68, 850–858.
Robins, R. H. and Guido, J. E. (1997) Design, construction and application of a simple packed capillary high performance liquid chromatography/electrospray mass spectrometry system. Rapid Commun. Mass Spectrom. 11, 1661–1666.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2006 Humana Press Inc.
About this protocol
Cite this protocol
Paradela, A., Albar, J.P. (2006). Protein Microsequencing. In: Salinas, J., Sanchez-Serrano, J.J. (eds) Arabidopsis Protocols. Methods in Molecular Biology™, vol 323. Humana Press. https://doi.org/10.1385/1-59745-003-0:421
Download citation
DOI: https://doi.org/10.1385/1-59745-003-0:421
Publisher Name: Humana Press
Print ISBN: 978-1-58829-395-4
Online ISBN: 978-1-59745-003-4
eBook Packages: Springer Protocols