Expression, purification and refolding of recombinant human bone morphogenetic protein- 2 in Escherichia coli
- VernacularTitle:重组人骨形成蛋白-2在大肠杆菌中的表达、纯化和复性
- Author:
Fenyong SUN
;
Ju WANG
;
Jinhua SUN
;
Yun DAI
;
Chuiyuan QIU
;
An HONG
- Publication Type:Journal Article
- Keywords:
Bone morphogenetic proteins;
Escherichia coli;
Human
- From:
Chinese Journal of Pathophysiology
2005;21(8):1480-1485
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To get high biological activity of recombinant human bone morphogenetic protein -2 (rhBMP-2) expressed in Escherichia coli by the methods of refolding. METHODS: The rhBMP-2,expressed in Escherichia coli, was washed by Triton X- 100 and further purified by DEAE chromatography.The inclusion bodies were resolved in 8 mol/L urea, and were refolded and dimerized in the redox systems (reduced and oxidized glutathione). Finally, a one - step purification procedure based on the heparin affinity chromatography was implemented. The biological activity of purified rhBMP - 2 were tested by induction of the alkaline phosphatase activity in C2C12 cells. RESULTS: The rhBMP - 2 was expressed in Escherichia coli in a non - active aggregated form of inclusion bodies using a temperature - inducible expression system. The high - purified rhBMP - 2 was obtained in the form of inclusion bodies by several purification courses. The rhBMP - 2 was refolded and dimerized in the redox systems (reduced and oxidized glutathione) and a one - step purification procedure based on the heparin affinity chromatography was implemented to isolate the rhBMP - 2 dimers and monomers. The purified rhBMP - 2 dimers showed the higher biological activity than the commercial rhBMP - 2. CONCLUSIONS: The method achieved the refolding of rhBMP - 2 would be applied to the whole TGF - β superfamily because the BMP - 2 belongs to the superfamily. Meanwhile, the inexpensive,high yield rhBMP - 2 is suitable for clinic application.
