Study on the Expression of Exendin-4 and Its activity to Decrease Plasma Glucose

For the sake of a high expression of Exendin-4 in Escherichia coli,the original Exendin-4 gene was modified in light of its amino acid sequence(GeneBank:AAB22006) and the code bias of E.coli.Not only 70% of the codes were changed,but also the sites of KpnI,NcoI and enterokinase were added to the two terminals of Exendin-4 if the gene respectively.The recombinant expression vector of pET-32a(+)-Exendin-4 was constructed and transformed into E.coli BL21(DE3).The fusion protein was 32 percent of total protein in E.coli after induction with 1 mmol/L IPTG for 6 hours at 30℃.With Ni-NTA chromatography,6.44 mg exendin-4 fusion protein from one gram wet bacteria was obtained and the purity was up to 98%.After the fusion protein was digested by enterokinase,affinity chromatography was used to bind the N terminals of 6×His tag and to purify Exendin-4.The yield of Exendin-4 was up to 1.59 mg per gram wet bacteria with the purity of 99%.The experiment in BKS.Cg-m+/+ Lepr db/J mice revealed that intraperitoneal injection of Exendin-4 had prominent glucose-lowering effects.The optimal dose was 7μg/kg and the maximal decrease in plasma glucose was up to 51.93% at 4 h after intraperitoneal injection.