Abstract
The small ubiquitin-like modifiers (SUMOs) alter the function of cellular proteins by covalent attachment to lysine side-chains. SUMOs can target themselves for modification so generating SUMO polymers, the functions of which are beginning to be unraveled.
The identification and quantitation of SUMO chains is essential for the functional investigation of SUMO polymerization. Classical techniques, such as site-directed mutagenesis and western blotting, are indirect and often inconclusive methods for the study of SUMO polymers. On the contrary, direct detection is possible with mass spectrometry (MS) by the identification of the SUMO–SUMO branched peptide remnant after proteolytic digestion. In this chapter, we describe a straightforward workflow that incorporates a modified database to efficiently detect SUMO polymers from simple and complex protein samples. In combination with stable isotope labeling by amino acids in cell culture (SILAC), this proteomic strategy allows accurate relative quantitation of SUMO polymers from different biological samples.
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Acknowledgments
The authors would like to thank Michael H. Tatham for comments on the manuscript. IM is a Sir Henry Wellcome Postdoctoral Fellow.
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Matic, I., Hay, R.T. (2012). Detection and Quantitation of SUMO Chains by Mass Spectrometry. In: Dohmen, R., Scheffner, M. (eds) Ubiquitin Family Modifiers and the Proteasome. Methods in Molecular Biology, vol 832. Humana Press. https://doi.org/10.1007/978-1-61779-474-2_17
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DOI: https://doi.org/10.1007/978-1-61779-474-2_17
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