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  Metabolic labeling of plant cell cultures with K(15)NO3 as a tool for quantitative analysis of proteins and metabolites

Engelsberger, W. R., Erban, A., Kopka, J., & Schulze, W. X. (2006). Metabolic labeling of plant cell cultures with K(15)NO3 as a tool for quantitative analysis of proteins and metabolites. Plant Methods, 2, 14. doi:10.1186/1746-4811-2-14.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0014-2AA0-4 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0014-2AA1-2
Genre: Journal Article

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 Creators:
Engelsberger, W. R.1, Author              
Erban, A.2, Author              
Kopka, J.2, Author              
Schulze, W. X.1, Author              
Affiliations:
1Signalling Proteomics, Department Stitt, Max Planck Institute of Molecular Plant Physiology, Max Planck Society, ou_1753330              
2Applied Metabolome Analysis, Department Willmitzer, Max Planck Institute of Molecular Plant Physiology, Max Planck Society, ou_1753338              

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 Abstract: Strategies for robust quantitative comparison between different biological samples are of high importance in experiments that address biological questions beyond the establishment of protein lists. Here, we propose the use of N-15-KNO3 as the only nitrogen source in Arabidopsis cell cultures in order to achieve a metabolically fully labeled cell population. Proteins from such metabolically labeled culture are distinguishable from unlabeled protein populations by a characteristic mass shift that depends on the amino acid composition of the tryptic peptide analyzed. In addition, the metabolically labeled cell extracts are also suitable for comparative quantitative analysis of nitrogen-containing cellular metabolic complement. Protein extracts from unlabeled and from standardized N-15-labeled cells were combined into one sample for joined analytical processing. This has the advantage of (i) reduced experimental variability and (ii) immediate relative quantitation at the level of single extracted peptide and metabolite spectra. Together ease and accuracy of relative quantitation for profiling experiments is substantially improved. The metabolic labeling strategy has been validated by mixtures of protein extracts and metabolite extracts from the same cell cultures in known ratios of labeled to unlabeled extracts (1:1, 1:4, and 4:1). We conclude that saturating metabolic N-15-labeling provides a robust and affordable integrative strategy to answer questions in quantitative proteomics and nitrogen focused metabolomics.

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Language(s): eng - English
 Dates: 2006-09-052006
 Publication Status: Published in print
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 Identifiers: ISI: ISI:000208349600014
DOI: 10.1186/1746-4811-2-14
ISSN: 1746-4811 (Electronic) 1746-4811 (Linking)
URI: ://000208349600014 http://www.plantmethods.com/content/pdf/1746-4811-2-14.pdf
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Title: Plant Methods
Source Genre: Journal
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Pages: - Volume / Issue: 2 Sequence Number: - Start / End Page: 14 Identifier: -