目的 采用合成生物学的原理与方法，将自行构建的融合蛋白重组人IFNα2b（recombinant human IFNα2b，rhIFNα2b）-TNFα刺激基因6（TNFα stimulated gene 6，TSG6）在BL21大肠埃希工程菌中高效表达，并检测其抗炎、抗病毒活性。方法 采用Swiss-model软件同源建模法在线预测该融合蛋白的三级结构等；将经密码子优化的人IFNα2b与TSG6基因片段通过连接肽连接，构建成含rhIFNα2b-TSG6融合基因的pET-32a重组质粒，转化至E. coli BL21(DE3)菌株，经发酵、包涵体变复性及纯化后获得融合蛋白rhIFNα2b-TSG6。采用SDS-PAGE对其纯度和蛋白含量进行分析，并通过蛋白质印迹检测蛋白特异性。采用WISH/水疱性口炎病毒活性检测系统，以微量细胞病变抑制法测定融合蛋白的抗病毒活性。在小鼠巨细胞病毒感染肝炎模型中观察该融合蛋白对肝细胞的保护作用及对IL-1β mRNA水平的抑制。测试rhIFNα2b-TSG6在家兔体内的药代动力学。结果 生物信息学分析结果显示，rhIFNα2b-TSG6符合设计预期。经鉴定，该基因成功以包涵体形式表达出目的蛋白，蛋白质表观相对分子量约为68 000，表达量达30%~40%。纯化后的rhIFNα2b-TSG6纯度达90%，能分别与抗人IFNα和抗人TSG6单克隆抗体特异性结合；其抗病毒比活性为（1.80±0.16）×10⁶ 国际单位/mg，约为单独rhIFNα2b的3.5倍。在小鼠病毒感染肝炎模型中起到明显抑制炎症、保护肝脏的作用。在家兔中血药浓度于24 h达到峰值。结论 在大肠埃希工程菌中高效表达了有明显抗病毒抑炎活性的rhIFNα2b-TSG6，为融合蛋白的规模化生产及临床治疗病毒性疾病打下基础。

Objective To efficiently express the self-constructed recombinant human IFNα2b (rhIFNα2b)-TNFα stimulated gene 6 (TSG6) fusion protein in Escherichia coli BL21 using principles and methods of synthetic biology, and to assess its anti-inflammatory and anti-viral activities.Methods The tertiary structure of the fusion protein was predicted using Swiss-model software online. The codon-optimized human IFNα2b gene was linked to a TSG6 gene fragment via a linker peptide. The pET-32a recombinant plasmid containing rhIFNα2b-TSG6 fusion gene was constructed and transformed into E. coli BL21(DE3) strain. Fermentation, inclusion body denaturation-renaturation, and purification processes were undertaken to obtain the fusion protein. Purity assessment along with protein content analysis were conducted through SDS-PAGE while Western blotting was applied to confirm specificity. The antiviral activity of rhIFNα2b-TSG6 was determined by WISH/vesicular stomatitis virus activity detection system with micro-cytopathic effect inhibition assay. Protective effects on hepatocytes as well as IL-β mRNA expression suppression were observed in mouse hepatitis models infected with murine cytomegalovirus (MCMV). The rhIFNα2b-TSG6 in vivo pharmacokinetic study was performed in rabbits.Results The results of bioinformatics analysis showed that rhIFNα2b-TSG6 met the design expectation. The target protein was successfully expressed in the form of inclusion bodies. The apparent relative molecular weight of rhIFNα2b-TSG6 was ~68 000, and the expression rate was 30%-40%. The purity of purified rhIFNα2b-TSG6 was up to 90%. The fusion protein could specifically bind to anti-IFNα and anti-TSG6 monoclonal antibodies. The specific antiviral activity of rhIFNα2b-TSG6 was (1.80±0.16)×10⁶ international unit/mg, 3.5 times of rhIFNα2b alone. It also protected the liver and inhibited inflammation in the MCMV mouse infected hepatitis model.The blood drug concentration reached peak at 24 h in rabbits.Conclusion Efficient prokaryotic expression of rhIFNα2b-TSG6 is achieved, accompanied by pronounced antiviral activities, laying foundational ground work for large-scale production aimed at clinical applications addressing acute chronic viral diseases.