Abstract
Classical quantitative proteomics studies focus on the relative or absolute concentration of proteins at a given time. In contrast, the investigation of protein turnover reveals the dynamics leading to these states. Analyzing the balance between synthesis and degradation of individual proteins provides insights into the regulation of protein concentration and helps understanding underlying biological processes. Comparing the half-lives of proteins allows detecting functional relationships and common regulation mechanisms. Moreover, comparing turnover of individual brain and plasma proteins between control- and treatment-groups indicates turnover changes induced by the treatment.
Here, we describe a procedure for determining turnover information of individual proteins in mice on a proteome-wide scale based on partial 15N metabolic labeling. We will outline the complete experimental workflow starting from 15N labeling the animals over sample preparation and mass spectrometric measurement up to the analysis of the data.
Stefan Reckow and Christian Webhofer contributed equally.
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Pratt JM, Petty J, Riba-Garcia I et al (2002) Dynamics of protein turnover, a missing dimension in proteomics. Mol Cell Proteomics 1:579–591
Schoenheimer R, Rittenberg D, Foster GL et al (1938) The application of the nitrogen isotope 15N for the study of protein metabolism. Science 88:599–600
Doherty MK, Beynon RJ (2006) Protein turnover on the scale of the proteome. Expert Rev Proteomics 3:97–110
Eng J, McCormack A, Yates J (1994) An approach to correlate tandem mass spectral data of peptides with amino acid sequences in a protein database. J Am Soc Mass Spectrom 5:976–989
Keller A, Nesvizhskii AI, Kolker E, Aebersold R (2002) Empirical statistical model to estimate the accuracy of peptide identifications made by MS/MS and database search. Anal Chem 74:5383–5392
Deutsch EW, Mendoza L, Shteynberg D et al (2010) A guided tour of the trans-proteomic pipeline. Proteomics 10:1150–1159
Nesvizhskii AI, Keller A, Kolker E, Aebersold R (2003) A statistical model for identifying proteins by tandem mass spectrometry. Anal Chem 75:4646–4658
Zhang Y, Reckow S, Webhofer C et al (2011) Proteome scale turnover analysis in live animals using stable isotope metabolic labeling. Anal Chem 83:1665–1672
Prince JT, Marcotte EM (2008) mspire: mass spectrometry proteomics in Ruby. Bioinformatics 24(23):2796–2797
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Reckow, S., Webhofer, C. (2014). Analysis of Individual Protein Turnover in Live Animals on a Proteome-Wide Scale. In: Martins-de-Souza, D. (eds) Shotgun Proteomics. Methods in Molecular Biology, vol 1156. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0685-7_9
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DOI: https://doi.org/10.1007/978-1-4939-0685-7_9
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-0685-7
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