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Solubilization and refolding with simultaneous Purification of Recombinant Human Granulocyte Colony-stimulating Factor expressed by Escherichia coli as Inclusion Bodies
Wang Chaozhan *,Wang Lili #,Geng Xindu
Key Laboratory of Synthetic and Natural Functional Molecule Chemistry of Ministry of Education, Institute of Modern Separation Science, Key Laboratory of Separation Science in Shaanxi Province, Department of Chemistry, Northwest University
*Correspondence author
#Submitted by
Subject:
Funding: 教育部博士点基金,陕西省教育厅重点实验室重点科研计划项目（No.20040697002,05JS61）
Opened online: 6 January 2008
Accepted by: none
Citation: Wang Chaozhan ,Wang Lili ,Geng Xindu .Solubilization and refolding with simultaneous Purification of Recombinant Human Granulocyte Colony-stimulating Factor expressed by Escherichia coli as Inclusion Bodies[OL]. [ 6 January 2008] http://en.paper.edu.cn/en_releasepaper/content/17777
 
 
Recombinant human granulocyte colony-stimulating factor (rhG-CSF) was solubilized and purified from inclusion bodies expressed in Escherichia coli. Effect of both pH and urea on the solubilization of rhG-CSF inclusion bodies were investigated, the results indicate that the existence both ionic and hydrophobic interactions in the inclusion bodies. Various solvents with reduced/oxidized glutathione (GSH/GSSG) were used to solubilize rhG-CSF inclusion bodies，the extent of solubilization was compared with each other and the behaviors of rhG-CSF in high performance hydrophobic interaction chromatography (HPHIC) were investigated. Solubilization of rhG-CSF inclusion bodies by 2% Sarkosyl in the presence of GSH/GSSG probably helps oxidizing rhG-CSF to form disulfide bonds, but Sarkosyl in the solution interferes seriously with the chromatographic process. To solubilize rhG-CSF inclusion bodies by combination of 2% Sarkosyl and 8.0 molL-1 urea can overcome this problem very well. And the so solubilized rhG-CSF was simultaneously refolded and purified by one step of HPHIC, with a specific activity of 1.9 × 108 IUmg-1 and a purity of 97%, the mass recovery was 41%.
Keywords:Recombinant human G-CSF; Inclusion bodies; Solubilization; Protein refolding; Purification.
 
 
 

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