Abstract
Expression of proteins in E. coli is often plagued by insolubility of the protein of interest. A solution to this problem is the expression of proteins as fusions to solubility tags such as the SUMO protein. SUMO fusion proteins can be cleaved to remove the SUMO moiety using SUMO-specific proteases such as Ulp1. Here, we describe the use of vectors for the expression of recombinant proteins in E. coli as fusions to the Drosophila SUMO protein. This includes a vector that encodes not only the SUMO tagged protein of interest but also SUMO-tagged Ulp1. Coexpression of these two proteins results in the in vivo cleavage of the protein of interest from the SUMO tag, while still leaving the protein of interest in a form that can be purified from a soluble cell lysate by nickel affinity chromatography.
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Burgess RR (2009) Refolding solubilized inclusion body proteins. Methods Enzymol 463:259–282. doi:10.1016/S0076-6879(09)63017-2
Malhotra A (2009) Tagging for protein expression. Methods Enzymol 463:239–258. doi:10.1016/S0076-6879(09)63016-0
Butt TR, Edavettal SC, Hall JP, Mattern MR (2005) SUMO fusion technology for difficult-to-express proteins. Protein Expr Purif 43(1):1–9. doi:10.1016/j.pep.2005.03.016
Malakhov MP, Mattern MR, Malakhova OA, Drinker M, Weeks SD, Butt TR (2004) SUMO fusions and SUMO-specific protease for efficient expression and purification of proteins. J Struct Funct Genomics 5(1–2):75–86. doi:10.1023/B:JSFG.0000029237.70316.52
Marblestone JG, Edavettal SC, Lim Y, Lim P, Zuo X, Butt TR (2006) Comparison of SUMO fusion technology with traditional gene fusion systems: enhanced expression and solubility with SUMO. Protein Sci 15(1):182–189. doi:10.1110/ps.051812706
Panavas T, Sanders C, Butt TR (2009) SUMO fusion technology for enhanced protein production in prokaryotic and eukaryotic expression systems. Methods Mol Biol 497:303–317. doi:10.1007/978-1-59745-566-4_20
Zuo X, Mattern MR, Tan R, Li S, Hall J, Sterner DE, Shoo J, Tran H, Lim P, Sarafianos SG, Kazi L, Navas-Martin S, Weiss SR, Butt TR (2005) Expression and purification of SARS coronavirus proteins using SUMO-fusions. Protein Expr Purif 42(1):100–110. doi:10.1016/j.pep.2005.02.004
Kuo D, Nie M, De Hoff P, Chambers M, Phillips M, Hirsch AM, Courey AJ (2011) A SUMO-Groucho Q domain fusion protein: characterization and in vivo Ulp1-mediated cleavage. Protein Expr Purif 76(1):65–71. doi:10.1016/j.pep.2010.08.008
Smith M, Turki-Judeh W, Courey AJ (2012) SUMOylation in Drosophila development. Biomolecules 2(3):331–349. doi:10.3390/biom2030331
Studier FW (2005) Protein production by auto-induction in high density shaking cultures. Protein Expr Purif 41(1):207–234
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Kuo, D., Nie, M., Courey, A.J. (2014). SUMO as a Solubility Tag and In Vivo Cleavage of SUMO Fusion Proteins with Ulp1. In: Giannone, R., Dykstra, A. (eds) Protein Affinity Tags. Methods in Molecular Biology, vol 1177. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1034-2_6
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DOI: https://doi.org/10.1007/978-1-4939-1034-2_6
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Publisher Name: Humana Press, New York, NY
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