The Study of Pancreas-targeted Epidermal Growth Factor Biosynthesis and Targeting
OBJECTIVE To design and biosynthesis a pancreas-targeted epidermal growth factor(EGF)fusion proteinusing Escherichiacoli as a host cell to improve the stability and targeting ability of EGF.METHODS The recombinant plasmids containing the pancreatic targeting peptide TRC(CHVLWSTRC)and the recombinant polypeptide(Pastag 200)modified EGF gene were introduced into E.coli for EGF fusion protein expression,purified by affinity chromatography and identified by Western blot.The biological functions were verified using mouse embryonic fibroblasts(NIH/3T3)and pancreatic islet endothelial cells(MSl).The in vivo targeting effects of the EGF fusion protein were investigated using KM mouse.RESULTS The results of agarose gel electrophoresis and sequencing indicated that the recombinant vector was successfully constructed.Western blot technique verified the specificity of the EGF fusion protein.EGF-Pastag200-TRC fusion protein had certain pro-proliferative ability on NIH/3T3 cells(P＜0.01)and showed higher biological activity and structural stability.Besides,10 pg-mL-1 EGF-Pastag200-TRC had a significant pro-proliferative effect on MS1 cells.After intraperitoneal injection,the distribution of EGF-Pastag200-TRC was significantly higher in pancreatic tissues than in other tissues(P＜0.01),demonstrating its targeting ability to pancreatic tissues.CONCLUSION EGF modified with pancreatic targeting peptides and Pastag200 shows higher biological activity and enhanced stability.It also shows good pancreatic targeting ability in this study,and providesa new idea for the use of EGF in the treatment of diabetes.
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