Solubilization and refolding with simultaneous purification of recombinant human stem cell factor.
Recombinant human stem cell factor (rhSCF) was solubilized and renatured from inclusion bodies expressed in Escherichia coli. The effect of both pH and urea on the solubilization of rhSCF inclusion bodies was investigated; the results indicate that the solubilization of rhSCF inclusion bodies was significantly influenced by the pH of the solution employed, and low concentration of urea can drastically improve the solubilization of rhSCF when solubilized by high pH solution. The solubilized rhSCF can be easily refolded with simultaneous purification by ion exchange chromatography (IEC), with a specific activity of 7.8 x 10(5) IU x mg(-1), a purity of 96.3%, and a mass recovery of 43.0%. The presented experimental results show that rhSCF solubilized by high pH solution containing low concentration of urea is easier to be renatured than that solubilized by high concentration of urea, and the IEC refolding method was more efficient than dilution refolding and dialysis refolding for rhSCF. It may have a great potential for large-scale production of rhSCF.
Wang
Chaozhan
C
Institute of Modern Separation Science, Department of Chemistry, Northwest University, 229 Tai Bai North Road, Xi'an 710069, People's Republic of China. czwang@nwu.edu.cn
Liu
Jiahua
J
Wang
Lili
L
Geng
Xindu
X
D002021:Buffers; D002852:Chromatography, Ion Exchange; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry* / Q000187:drug effects / Q000302:isolation & purification*; D012995:Solubility; D019089:Stem Cell Factor / Q000737:chemistry* / Q000187:drug effects / Q000302:isolation & purification*; D014508:Urea / Q000737:chemistry
D002021:Buffers; D011994:Recombinant Proteins; D019089:Stem Cell Factor; D014508:Urea