Abstract
Protein labeling methods prior to separation and analysis have become indispensable approaches for proteomic profiling. Basically, three different types of tags are employed: stable isotopes, mass tags, and fluorophores. While proteins labeled with stable isotopes and mass tags are measured and differentiated by mass spectrometry, fluorescent labels are detected with fluorescence imagers. The major purposes for protein labeling are monitoring of biological processes, reliable quantification of compounds and specific detection of protein modifications and isoforms in multiplexed samples, enhancement of detection sensitivity, and simplification of detection workflows. Proteins can be labeled during cell growth by incorporation of amino acids containing different isotopes, or in biological fluids, cells or tissue samples by attaching specific groups to the ε-amino group of lysine, the N-terminus, or the cysteine residues. The principles and the modifications of the different labeling approaches on the protein level are described; benefits and shortcomings of the methods are discussed.
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References
Wöhlbrand L, Trautwein K, Rabus R (2013) Proteomic tools for environmental microbiology—a roadmap from sample preparation to protein identification and quantification. Proteomics 13:2700–2730
Miller I, Crawford J, Gianazza E (2006) Protein stains for proteomic applications: which, when, why? Proteomics 6:5385–5408
Moche M, Albrecht D, Maaß S et al (2013) The new horizon in 2D electrophoresis – new technology to increase resolution and sensitivity. Electrophoresis 34:1510–1518
Zhou S, Bailey MJ, Dunn MJ et al (2005) A quantitative investigation into the losses of proteins at different stages of a two-dimensional gel electrophoresis procedure. Proteomics 5:2739–2747
Go Y-M, Jones DP (2013) The redox proteome. J Biol Chem 288:26512–26520
Silva JC, Denny R, Dorschel CA et al (2005) Quantitative proteomic analysis by accurate mass retention time pairs. Anal Chem 77:2187–2200
Ünlü M, Morgan ME, Minden JS (1997) Difference gel electrophoresis: a single gel method for detecting changes in protein extracts. Electrophoresis 18:2071–2077
Westermeier R, Scheibe B (2008) Difference gel electrophoresis based on Lys/Cys tagging. In: Posch A (ed) Sample preparation and fractionation for 2-D PAGE/proteomics. Methods in molecular biology, vol 424. Humana, New York, pp 73–85
Berendt FJ, Fröhlich T, Bolbrinker P et al (2009) Highly sensitive saturation labeling reveals changes in abundance of cell cycle-associated proteins and redox enzyme variants during oocyte maturation in vitro. Proteomics 9:550–564
Personal communication from Dr. Rudolf Lichtenfels, Martin Luther University Halle-Wittenberg, Institute of Medical Immunology, Halle (Saale), Germany
Cramer R, Westermeier R (eds) (2012) Difference gel electrophoresis (DIGE): methods and protocols, vol 854. Springer GmbH, Heidelberg
Griebel A, Obermaier C, Westermeier R et al (2013) Simplification and Improvement of Protein Detection in two-dimensional Electrophoresis Gels with SERVA HPE™ LightningRed. Arch Physiol Biochem 119:94–99
Gygi SP, Rist B, Gerber SA et al (1999) Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. Nat Biotechnol 17:994–999
Hansen KC, Schmitt-Ulms G, Chalkley RJ et al (2003) Mass spectrometric analysis of protein mixtures at low levels using cleavable 13C-isotope-coded affinity tag and multidimensional chromatography. Mol Cell Proteomics 2:299–314
Li J, Steen H, Gygi SP (2003) Protein profiling with cleavable isotope-coded affinity tag (cICAT) reagents: the yeast salinity stress response. Mol Cell Proteomics 2:1198–1204
Ong SE, Blagoev B, Kratchmarova I et al (2002) Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol Cell Proteomics 1:376–386
Schwanhäusser B, Gossen M, Dittmar G et al (2009) Global analysis of cellular protein translation by pulsed SILAC. Proteomics 9:205–209
Schmidt A, Kellermann J, Lottspeich F (2005) A novel strategy for quantitative proteomics using isotope-coded protein labels. Proteomics 5:4–15
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Obermaier, C., Griebel, A., Westermeier, R. (2015). Principles of Protein Labeling Techniques. 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_13
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DOI: https://doi.org/10.1007/978-1-4939-2550-6_13
Publisher Name: Humana Press, New York, NY
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