Abstract
Studying postubiquitylation events has always been a difficult task due to the labile nature of these posttranslational modifications. When utilized in tandem, ubiquitin-binding entities (TUBEs) not only increase up to thousand times the affinity for poly-ubiquitin chains but also protect ubiquitylated proteins from the action of the proteasome and de-ubiquitylating enzymes.
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References
Reyes-Turcu FE, Ventii KH, Wilkinson KD (2009) Regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes. Annu Rev Biochem 78:363–397.
Schwartz AL, Ciechanover A (2009) Targeting proteins for destruction by the ubiquitin system: implications for human pathobiology. Annu Rev Pharmacol Toxicol 49:73–96.
Tagwerker C, Flick K, Cui M et al (2006) A tandem affinity tag for two-step purification under fully denaturing conditions: application in ubiquitin profiling and protein complex identification combined with in vivo cross-linking. Mol Cell Proteomics 5:737–748.
Xu P, Duong DM, Seyfried NT et al (2009) Quantitative proteomics reveals the function of unconventional ubiquitin chains in proteasomal degradation. Cell 137:133–145.
Ding Q, Dimayuga E, Markesbery WR et al (2006) Proteasome inhibition induces reversible impairments in protein synthesis. Faseb J 20:1055–1063.
Hjerpe R, Rodriguez MS (2008) Efficient approaches for characterizing ubiquitinated proteins. Biochem Soc Trans 36:823–827.
Hjerpe R, Aillet F, Lopitz-Otsoa F et al (2009) Efficient protection and isolation of ubiquitylated proteins using tandem ubiquitin-binding entities. EMBO Rep 10:1250–1258.
Lopitz-Otsoa F, Rodriguez MS, Aillet F (2010) Properties of natural and artificial proteins displaying multiple ubiquitin-binding domains. Biochem Soc Trans 38:40–45.
Hjerpe R, Aillet F, Lopitz-Otsoa F et al (2010) Oligomerization conditions Mdm2-mediated efficient p53 polyubiquitylation but not its proteasomal degradation. Int J Biochem Cell Biol 42:725–735.
Acknowledgments
We would like to thank Seth Goldenberg for the critical reading of this document. This work was funded by the Ramón y Cajal Program, Ministerio de Educación y Ciencia grant BFU2006-12991 and BFU2008-01108/BMC, Fondo de Investigaciones Sanitarias (FIS) CIBERhed, Government of the Autonomous Community of the Basque Country grant PI09-05, Department of Industry, Tourism and Trade of the Government of the Autonomous Community of the Basque Country (Etortek Research Programs 2008/2009), and from the Innovation Technology Department of the Bizkaia County.
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Aillet, F., Lopitz-Otsoa, F., Hjerpe, R., Torres-Ramos, M., Lang, V., Rodríguez, M.S. (2012). Isolation of Ubiquitylated Proteins Using Tandem Ubiquitin-Binding Entities. 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_12
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DOI: https://doi.org/10.1007/978-1-61779-474-2_12
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