Abstract
Detailed knowledge of the proteome is crucial to advance the biological sciences. Low-abundant proteins are of particular interest to many biologists as they include, for example those proteins involved in signal transduction. Recent technological advances resulted in a tremendous increase in protein identification sensitivity by mass spectrometry (MS). However, the dynamic range in protein abundance still forms a fundamental problem that limits the detection of low-abundant proteins in complex proteomes. These proteins will typically escape detection in shotgun MS experiments due to the presence of other proteins at an abundance several-fold higher in order of magnitude. Therefore, specific enrichment strategies are required to overcome this technical limitation of MS-based protein discovery. We have searched for novel signal transduction proteins, more specifically kinases and calcium-binding proteins, and here we describe different approaches for enrichment of these low-abundant proteins from isolated chloroplasts from pea and Arabidopsis for subsequent proteomic analysis by MS. These approaches could be extended to include other signal transduction proteins and target different organelles.
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Acknowledgements
Work in the author’s lab is supported by grants from the Austrian Science Fund (FWF) to MT (P 23435-B12 and P 25359-B21) and the Marie Curie Initial training network (ITN) “CALIPSO” (GA ITN 2013-607607) from the European Union.
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Bayer, R.G., Stael, S., Teige, M. (2015). Chloroplast Isolation and Affinity Chromatography for Enrichment of Low-Abundant Proteins in Complex Proteomes. In: Posch, A. (eds) Proteomic Profiling. Methods in Molecular Biology, vol 1295. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2550-6_16
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DOI: https://doi.org/10.1007/978-1-4939-2550-6_16
Publisher Name: Humana Press, New York, NY
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