重组人胰岛素样生长因子1的原核可溶性表达及活性鉴定

摘要

目的:原核可溶性表达人胰岛素样生长因子1(hlGF-1)并进行生物活性测定.方法:PCR扩增hlGF-1核酸序列,克隆至融合表达载体pET-DsbC中,转入大肠杆菌BL21(DE3)进行诱导、表达和纯化,对融合蛋白DsbC -hlGF-1分别进行酶切、Western印迹和生物活性测定.结果:融合蛋白DsbC-hlGF-1在原核系统内经IPTG诱导,主要以可溶性形式表达,其表达量占菌体总蛋白量的30％～35％,Western印迹和MTT法测定结果证明重组DsbC-hlGF-1蛋白具有较强的抗原活性和明显的促NIH/3T3细胞增殖作用.结论:融合蛋白DsbC-hlGF在大肠杆菌中以可溶性表达形式存在并具有类似天然hIGF-1的生物活性,这对进一步研究hIGF-1的结构和功能,开发相应的基因工程产品具有重要意义.%Objective: To obtain human insulin like growth factor 1 (hIGF-1) expressed in solubility in prokary-otic system and detect its biological activity. Methods: hIGF-1 gene was obtained by PCR and was cloned into pET-DsbC fusion vector. After induction, expression and purification, the antigenicity and biological activity of the product were examined by Western blot and MTT assay. Results: DsbC-hlGF-1 was expressed in solubility in Es-cherichia coli and its expression quantity was about 30%~35% to bacterial total protein with IPTG induction, which also showed a strong antigenic ability and stimulate proliferation of NIH/3T3 cells evidently. Conclusion: The results suggest that the DsbC- hIGF-1 protein has the similar biological activity to wide type hIGF-1 and could express in solubility in E.coli It provides a futher investigation of the IGF-1 structure and function and build the foundation for the large scale production of IGF-1.