| The member of fibroblast growth factor familiy,fibroblast growth factor-8, isoform a(FGF8a)is accepted as a crucial organizing signal during the development of vertebrate embryos. Furthermore, the FGF8a is also efficient in healing organic wound, culturing the neural stem cells and induction of dopaminergic neurons. So it is suggested that the potential application is huge for FGF8a to benefit the cliniacal treatment .This study is based on the expressed vector FGF-pET28a-Rosetta. RhFGF8a occurs both in the cytoplasm of soluble and insoluble components of broken bacteria by IPTG induction. It is very diffculty to obtain the aim protein in soluble through various purification means . And we can purify rhFGF8a easily from insoluble components only through nickel affinity chromatography under denaturalization. We also do several parallel experiments to get the optimal conditions for the resumption of rhFGF8a.The results show that FGF8a which is diluted to 50ug/ ml refold best in pH 8.0 refolding buffer with the oxidation / reduction of glutathione. Cultivating cell with rhFGF8a and mensured by MTT method, which shows that FGF8a plays a certain role in promoting the growth of fibroblasts(ED50 is about 50ng/ml).RhFGF8, a strong basic protein,whose isoelectric point is about 10.5. According to previous experience, proteins could be refolded better by negative charge.As rhFGF8a has high isoelectric poin , it is not easy to make rhFGF8a attach negative charge in general buffer. By groping the experimental conditions, the protein having certain biological activity are successfully gained ultimately.The purity of rhFGF8a can get to 95% tested by SDS-PAGE. The study will play a certain reference for other strong basic protein purification.
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