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目的:评价原核表达、纯化的6×His-硫氧还蛋白(TRX)-人肿瘤坏死因子α(TNFα)抑制肽-C端抗炎酸性尾巴融合蛋白的生物学功能。方法:在大肠杆菌中分别表达带His标签的TRX对照蛋白及TRX蛋白融合的人TNFα抑制肽-抗炎酸性尾巴融合蛋白,并对2种蛋白进行N2+金属螯合层析纯化,采用MTT法检测纯化后的蛋白及化学合成多肽抑制TNFα标准品对L929细胞的细胞毒活性。结果:与对照蛋白相比,融合蛋白人TNFα抑制肽-C端抗炎酸性尾巴及合成多肽均能拮抗TNFα标准品对L929细胞的细胞毒作用。结论:融合蛋白人TNFα抑制肽-C端抗炎酸性尾巴及合成肽均能有效拮抗TNFα的生物学作用,为今后发展抑制TNFα为主的抗炎生物药物奠定了基础。
OBJECTIVE: To evaluate the biological function of prokaryotic expressed and purified 6 × His-thioredoxin (TRX) -citing human tumor necrosis factor α (TNFα) inhibitory peptide-C-terminal anti-inflammatory acid tail fusion protein. METHODS: The His-tagged TRX control protein and TRX fusion protein human TNFα inhibitor peptide-anti-inflammatory acid tail fusion protein were expressed respectively in E. coli. The two proteins were purified by N2 + metal chelate chromatography and detected by MTT assay Purified proteins and chemically synthesized polypeptides inhibit the cytotoxic activity of TNFα standards on L929 cells. Results: Compared with the control protein, the fusion protein human TNFα inhibitory peptide C-terminal anti-inflammatory acid tail and synthetic peptides could antagonize the cytotoxic effect of TNFα standard on L929 cells. CONCLUSION: The fusion protein human TNFα inhibitory peptide C-terminal anti-inflammatory acid tail and synthetic peptide can both effectively antagonize the biological effects of TNFα and lay a foundation for the future development of anti-inflammatory biological drugs that inhibit TNFα.