Expression and purification of recombinant bone morphogenetic protein-2 in E.coli.
OBJECTIVE
To explore the method for producing human bone morphogenetic protein-2 (hBMP-2) by gene engineering techniques.


METHODS
E.coli BL21 was transformed with recombinant plasmid pYR (pBV220-hBMP-2) under different conditions, and SDS-PAGE analysis was conducted to observe the effects of the activation status and induction time of the bacterium on the target protein expression. The inclusion bodies obtained from E.coli were purified by anion exchange chromatography DEAE and molecular sieve S-300, and the recombinant protein was renatured by dialyse.


RESULTS
SDS-PAGE analysis showed a conspicuous band after induction signifying a new foreign protein with relative molecular mass of approximately 13 000. After activation of the bacteria when D600 was about 0.45, most efficient expression of rhBMP-2 was achieved which reached the peak 4 h after induction with heat. Implantation of the purified recombinant hBMP-2 resulted in proliferation of mesenchymal cells and new cartilage and bone formation, as shown by histological analysis 4 weeks after implantation.


CONCLUSIONS
hBMP-2 produced by gene engineering techniques possesses the biological capacity of ectopic bone formation.
Xiong
Shao-Hu
SH
Institute of Clinical Anatomy, First Military Medical University, Guangzhou 510515, China. ln-xsh@sohu.com
Yu
Lei
L
Tan
Hai-Yan
HY
Huang
Tao
T
Wang
Chao-Yang
CY
Liu
Da-Yong
DY
Zhong
Shi-Zhen
SZ
D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D006801:Humans; D008297:Male; D051379:Mice; D010012:Osteogenesis / Q000187:drug effects; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016212:Transforming Growth Factor beta
C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; C499469:recombinant human bone morphogenetic protein-2