Performance of isobaric and isotopic labeling in quantitative plant proteomics

Fábio C S Nogueira, Giuseppe Palmisano, Veit Schwämmle, Francisco A P Campos, Martin R. Larsen, Gilberto B. Domont, Peter Roepstorff

Research output: Contribution to journalArticle

Abstract

Mass spectrometry has become indispensable for peptide and protein quantification in proteomics studies. When proteomics technologies are applied to understand the biology of plants, two-dimensional gel electrophoresis is still the prevalent method for protein fractionation, identification, and quantitation. In the present work, we have used LC-MS to compare an isotopic (ICPL) and isobaric (iTRAQ) chemical labeling technique to quantify proteins in the endosperm of Ricinus communis seeds at three developmental stages (IV, VI, and X). Endosperm proteins of each stage were trypsin-digested in-solution, and the same amount of peptides was labeled with ICPL and iTRAQ tags in two orders (forward and reverse). Each sample was submitted to nanoLC coupled to an LTQ-Orbitrap high-resolution mass spectrometer. Comparing labeling performance, iTRAQ was able to label 99.8% of all identified unique peptides, while 94.1% were labeled by ICPL. After statistical analysis, it was possible to quantify 309 (ICPL) and 321 (iTRAQ) proteins, from which 95 are specific to ICPL, 107 to iTRAQ, and 214 common to both labeling strategies. We noted that the iTRAQ quantification could be influenced by the tag. Even though the efficiency of the iTRAQ and ICPL in protein quantification depends on several parameters, both labeling methods were able to successfully quantify proteins present in the endosperm of castor bean during seed development and, when combined, increase the number of quantified proteins.

Original languageEnglish
Pages (from-to)3046-3052
Number of pages7
JournalJournal of Proteome Research
Volume11
Issue number5
DOIs
Publication statusPublished - May 4 2012

Keywords

  • high-resolution mass spectrometry
  • ICPL
  • iTRAQ
  • plant proteomics
  • quantitative proteomics

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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  • Cite this

    Nogueira, F. C. S., Palmisano, G., Schwämmle, V., Campos, F. A. P., Larsen, M. R., Domont, G. B., & Roepstorff, P. (2012). Performance of isobaric and isotopic labeling in quantitative plant proteomics. Journal of Proteome Research, 11(5), 3046-3052. https://doi.org/10.1021/pr300192f