期刊
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
卷 166, 期 5, 页码 1264-1274出版社
HUMANA PRESS INC
DOI: 10.1007/s12010-011-9512-8
关键词
Recombinant human G-CSF; Inclusion bodies; High pH solubilization; Protein refolding; Protein purification; Liquid chromatography
资金
- National Natural Science Foundation in China [20705028]
- National Fund for Talent Training in Basic Science [J0830417]
- State Key Laboratory of Supramolecular Structure and Materials [SKLSSM201133]
- Open Foundation of Key Laboratory of Synthetic and Natural Functional Molecule Chemistry of Ministry of Education
- Open Foundation of Shaanxi Provincial Key Laboratory of Electroanalytical Chemistry
Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a very efficient therapeutic protein drug which has been widely used in human clinics to treat cancer patients suffering from chemotherapy-induced neutropenia. In this study, rhG-CSF was solubilized from inclusion bodies by using a high-pH solution containing low concentration of urea. It was found that solubilization of the rhG-CSF inclusion bodies greatly depended on the buffer pH employed; alkalic pH significantly favored the solubilization. In addition, when small amount of urea was added to the solution at high pH, the solubilization was further enhanced. After solubilization, the rhG-CSF was renatured with simultaneous purification by using weak anion exchange, strong anion exchange, and hydrophobic interaction chromatography, separately. The results indicated that the rhG-CSF solubilized by the high-pH solution containing low concentration of urea had much higher mass recovery than the one solubilized by 8 M urea when using anyone of the three refolding methods employed in this work. In the case of weak anion exchange chromatography, the high pH solubilized rhG-CSF could get a mass recovery of 73%. The strategy of combining solubilization of inclusion bodies at high pH with refolding of protein using liquid chromatography may become a routine method for protein production from inclusion bodies.
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