【摘要】 目的:构建干扰素α1b突变体IFNα1b/31K,以期获得高效低毒的新型药物分子。方法:根据合理药物设计,采用定点突变技术,将干扰素α1b第31位氨基酸残基突变为K,并构建表达IFNα1b/31K重组蛋白。纯化后,对其抗病毒活性、抗肿瘤细胞增殖活性和动物体内急性毒性进行考察。结果:IFNα1b/31K表达量占菌体总蛋白的30%以上。纯化后的IFNα1b/31K纯度大于95%,比活性约为IFNα1b的1.7倍,抗肿瘤增殖活性比IFNα1b降低,未见对实验动物的急性毒性作用。结论:成功设计构建并表达了高效低毒的IFNα1b突变蛋白分子。更多还原

【Abstract】 Interferon alpha are used clinically to treat a group of viral diseases and cancer because of its broad anti-viral,anti-tumor and immune regulation activities.Nevertheless,the clinical usage of interferon-α can still be improved in many ways,such as increasing activity and so on.In order to obtain a new type of molecule with higher potency,human interferon α1b mutant IFNα1b/31K was constructed.ATG was substituted by AAG at 31 of IFN α1b through PCR site-directed mutagenesis in vitro.The amplified fragments were inserted into pET-23b expression vector,and the recombinant plasmid was transformed into Escherichia coli BL21(DE3).After being cultured in complex auto-inducing media and purification process,IFNα1b/31K was expressed as soluble protein with the yield more than 30% of total bacterial protein.The purity was more than 95%,the anti-viral activity was 1.7 times comparing with IFNα1b,anti-tumor activity was lower than IFNα1b,and the p-value of cytotoxicity & acute toxicity was more than 0.05.更多还原