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We established a purification system for glutathione-S-transferase (GST) fusion protein using glutathione coupled magnetic particle. Glutathione was coupled covalently to the surface of magnetic particles with isothiocyanate functional groups. Cell lysate, containing the fusion protein, was then incubated with these glutathione coupled magnetic particles at room temperature. Unbound and non-specifically bound proteins were removed by wash steps. Subsequently, the GST-fusion protein was eluted from the magnetic particles by the addition of reduced glutathione. The resulting fusion protein was tested for purity using SDS-PAGE and demonstrated by Western blotting. The concentration of the fusion protein was measured by Bradford method. Both the conditions for incubation and washing were optimized. The results showed that 150 microg glutathione could be bound on 1 mg of particle surface and 10 mg of the glutatione-coupled magnetic particles was suitable for 100 microL lysate, the optimal incubation time for reaction between particles and lysate was 40 min. The magnetic particles could help purify efficiently GST-fusion protein with a yield of around 516 microg fusion protein per 10 mg particles. Magnetic particles can be successfully used in a simple, rapid and reliable method for the purification of GST-fusion proteins.
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
October 2020,
Nano letters,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
May 2001,
Current protocols in molecular biology,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
January 2002,
Methods in enzymology,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
August 1995,
FEMS microbiology letters,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
April 1992,
Biotechnology and bioengineering,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
February 1997,
Frontiers in bioscience : a journal and virtual library,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
December 1995,
Protein expression and purification,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
January 1982,
Acta chemica Scandinavica. Series B: Organic chemistry and biochemistry,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
October 2006,
Zhongguo yi xue ke xue yuan xue bao. Acta Academiae Medicinae Sinicae,
Jingjing Zhu, and
Liu Yang, and
Lei Yang, and
Chao Chen, and
Yali Cui
April 1993,
Analytical biochemistry,
