抗胶质瘤单链抗体—人肿瘤坏死因子α融合基因的构建及表达

目的 制备单链抗体-人肿瘤坏死因子α（hTNF-α)融合蛋白。方法 将hTNF-α的成熟肽cDNA以重组PCR技术融合于胶质瘤单链抗体基因的3′端，构建39ScFv-hTNF-α融合基因，克隆入大肠杆菌分泌型表达pET-20b( ),并诱导表达，结果 融合蛋白的表达量占菌体总蛋白的20%，还原SDS-PAGE和Western blot显示，其相对分子质量（Mr)为44000，具有识别胶质瘤相关抗原的活发表主TNF-α的抑瘤活性。结论 完成了39ScFv-hTNF-α融合基因的构建，并于大肠杆菌中表达了双功能融合蛋白，为胶质瘤的临床治疗提供了新的高效免疫导向药物。

Construction and expression of anti-glioma single-chain Fv/human tumor necrosis factor

Aim To construct and express the fused gene of anti-glioma single chain Fv(39ScFv)/ human tumor necrosis factor α (TNF-α ). Methods The mature peptide cDNA of TNF-α was fused to 3′ -terminus of ScFv gene. The 39ScFv-TNFα fusion gene was cloned into expression vector pET20b(＋ ) and induced to express in E.coli. Results The fusion protein expressed in E.coli account for 20％ of the total bacterial protein. It displayed a single band of Mr 44 000 by reducing SDS-PAGE and Western blot, and retained both of its anti-glioma immunoreactivity and biological activity of TNF. Conclusion The fusion gene was constructed and expressed successfully in E.coli. The fusion protein possessed bifunctional activities, it may prove useful in targeting immunotherapy against glioma.