含硒抗病毒多肽的半胱氨酸缺陷型表达及鉴定

doi:10.13523/j.cb.20140403

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中国生物工程杂志 ›› 2014, Vol. 34 ›› Issue (4) : 16-20. DOI: 10.13523/j.cb.20140403

研究报告

含硒抗病毒多肽的半胱氨酸缺陷型表达及鉴定

王程^1,2, 隋春红¹, 闫岗林², 吕绍武², 牟颖²

作者信息 +

Cysteine Auxotrophic Expression and Identification of Antiviral Selenium-containing Peptide

WANG Cheng^1,2, SUI Chun-hong¹, YAN Gang-lin², LÜ Shao-wu², MU Ying²

Author information +

文章历史 +

摘要

目的：在大肠杆菌中进行含硒抗病毒多肽的可溶性半胱氨酸缺陷型融合蛋白的表达，并对其纯化蛋白进行生物学活性的初步鉴定。方法：将含硒抗病毒多肽Se-GBVA10基因克隆到GST融合表达载体pGEX-2T中，转化至半胱氨酸缺陷型大肠杆菌BL21 cysE51 后进行半胱氨酸缺陷型表达，通过谷胱甘肽Sepharose 4B 亲和柱纯化目的蛋白质，并对其含硒量、抗病毒活力等性质进行初步鉴定。结果：利用半胱氨酸缺陷型表达法成功地可溶性表达了sjGST-Se-GBVA10和sjGST-GBVA10融合蛋白，并经凝血酶切后获得了纯化的Se-GBVA10和GBVA10抗病毒多肽。测定Se-GBVA10的含硒量为0.974 mol/mol peptide，GPX活力为47.52 U/μmol，其EC₅₀为21.73μmol/L，CC₅₀为849.41μmol/L。结论：Se-GBVA10具有与GBVA10相同的抗病毒活性，并具有一定的抗氧化活力。

Abstract

Objective: To express the soluble antiviral peptide, the cysteine auxotrophic expression system was used in Escherichia coli (E. coli). And the biological activities of the purified antiviral peptide were initially identified. Method: The gene sequence of antiviral peptide Se-GBVA10 was chemically synthesized according to the optimized codons of E.coli, and the gene was cloned into GST fusion expression vector pGEX-2T. Then, the plasmid pGEX-2T-GBVA10 was converted to cysteine auxotrophic expression strains (E. coli BL21 cysE51). Expressed proteins were purified by glutathione Sepharose 4B affinity column. Finally, the content of selenium, the antiviral activity and the antioxidant activity (the vitality of glutathione peroxidase) of the antiviral peptides were detected. Results: The fusion protein, sjGST-Se-GBVA10 and sjGST-GBVA10, were successfully expressed by means of cysteine auxotrophic expression system. After cutting with thrombin, the purified antiviral peptides, Se-GBVA10 and GBVA10 were obtained. The content of selenium of the Se-GBVA10 was 0.974 mol/mol peptide, the glutathione peroxidase vitality of Se-GBVA10 was 47.52 U/μmol, the concentration for 50% of maximal effect (EC₅₀) of Se-GBVA10 was 21.73μmol/L, and the median cytotoxic concentration (CC₅₀) of Se-GBVA10 was 849.41μmol/L. Conclusion: The antiviral peptide Se-GBVA10 has the same antiviral activity as the GBVA10, but also has some antioxidant activity.

导出引用

王程, 隋春红, 闫岗林, 吕绍武, 牟颖. 含硒抗病毒多肽的半胱氨酸缺陷型表达及鉴定[J]. 中国生物工程杂志, 2014, 34(4): 16-20 https://doi.org/10.13523/j.cb.20140403

WANG Cheng, SUI Chun-hong, YAN Gang-lin, LÜ Shao-wu, MU Ying. Cysteine Auxotrophic Expression and Identification of Antiviral Selenium-containing Peptide[J]. China Biotechnology, 2014, 34(4): 16-20 https://doi.org/10.13523/j.cb.20140403

中图分类号： O627.6

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