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Whole Proteome Analysis of the Protozoan Parasite Trypanosoma brucei Using Stable Isotope Labeling by Amino Acids in Cell Culture and Mass Spectrometry

  • Olivera Cirovic
  • Torsten Ochsenreiter
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1188)

Abstract

The single-celled protozoan Trypanosoma brucei spp. is the causative agent of human African trypanosomiasis and nagana in cattle. Quantitative proteomics for the first time has allowed for the characterization of the proteome from several different life stages of the parasite (Butter et al., Mol Cell Proteomics 12:172–179, 2013; Gunasekera et al., BMC Genomics 13:556, 2012; Urbaniak et al., PloS One 7(5):e36619, 2012). To achieve this, stable isotope labeling by amino acids in cell culture (SILAC) (Ong et al., Mol Cell Proteomics 1:376–386, 2002) was adapted to T. brucei spp. cultures. T. brucei cells grown in standard media with dialyzed fetal calf serum containing heavy isotope-labeled amino acids (arginine and lysine) show efficient incorporation of the labeled amino acids into the whole cell proteome (8–12 divisions) and no detectable amino acid conversions. The method can be applied to both of the major life stages of the parasite and in combination with RNAi or gene knockout approaches.

Key words

Trypanosoma brucei SILAC Mass spectrometry Cell culture Cell differentiation 

Notes

Acknowledgements

The authors would like to thank Kapila Gunasekera and Roman Trikin for critically reading the manuscript. The research was supported by the Swiss National Science Foundation.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Institute of Cell BiologyUniversity of BernBernSwitzerland
  2. 2.The Graduate School for Cellular and Biomedical SciencesUniversity of BernBernSwitzerland

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