Improved isolation of proteins tagged with glutathione S-transferase |
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| Authors: | Nicholas K. Vinckier Arkadiusz Chworos Stanley M. Parsons |
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| Affiliation: | 1. Laboratory of Environmental Toxicology and Aquatic Ecology, Department Applied Ecology and Environmental Biology, Faculty of Bioscience Engineering, Ghent University (UGent), Jozef Plateaustraat 22, 9000 Gent, Belgium;2. Laboratory Aquatic Ecology, Evolution and Conservation, University of Leuven, Charles Deberiotstraat 32, 3000 Leuven, Belgium;1. Departamento de Bioquímica Clínica, Centro de Investigaciones en Bioquímica Clínica e Inmunología (CIBICI-CONICET), Facultad de Ciencias Químicas-Universidad Nacional de Córdoba, Córdoba, CP 5000, Argentina;2. Graduate Program in Cellular and Molecular Biology, Center of Biotechnology, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil;3. Department of Biophysics, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil;3. Graduate Institute of Biomedical Sciences,;4. School of Medicine,;5. Department of Cell and Molecular Biology, and;6. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan;;1. Department of Internal Medicine, University Hospital Brussels, Brussels, Belgium;2. Department of Radiology, University Hospital Brussels, Brussels, Belgium;3. Department of Emergency Medicine, University Hospital Brussels, Brussels, Belgium;1. Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA;2. Center for Systems and Synthetic Biology, The University of Texas at Austin, Austin, TX, 78712, USA;1. Departments of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA;2. Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA;3. Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA |
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| Abstract: | ![]()
A common affinity tag used to express and purify fusion proteins is glutathione S-transferase. However, many researchers have reported difficulty eluting GST-tagged proteins from the affinity matrix. This report demonstrates that the problem likely is due to the propensity of glutathione S-transferase to dimerize combined with a propensity of the tagged protein to oligomerize, which results in formation of large oligomers of fusion protein that are chelated by the affinity matrix. The solution to the problem is to use S-butylglutathione instead of glutathione to elute, as S-butylglutathione binds more tightly to glutathione S-transferase and overcomes the chelate effect. Moreover, in contrast to glutathione, S-butylglutathione has no reducing capability that might inactivate a tagged protein. |
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