A Method for Rapid Screening, Expression, and Purification of Antimicrobial Peptides

文献类型： 外文期刊

第一作者： Zhang, Yingli

作者： Zhang, Yingli;Li, Zhongchen;Li, Li;Ma, Lixin;Wang, Yaping;Rao, Ben

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关键词： antimicrobial peptides expression; CL7 tag; Escherichia coli; Pichia pastoris

期刊名称：MICROORGANISMS （ 影响因子：4.128； ）

ISSN：

年卷期： 2021 年 9 卷 9 期

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收录情况： SCI

摘要： In this study, a method for the rapid screening, expression and purification of antimicrobial peptides (AMPs) was developed. AMP genes were fused to a heat-resistant CL7 tag using the SLOPE method, and cloned into Escherichia coli and Pichia pastoris expression vectors. Twenty E. coli and ten P. pastoris expression vectors were constructed. Expression supernatants were heated, heteroproteins were removed, and fusion proteins were purified by nickel affinity (Ni-NTA) chromatography. Fusion proteins were digested on the column using human rhinovirus (HRV) 3C protease, and AMPs were released and further purified. Five AMPs (1, 2, 6, 13, 16) were purified using the E. coli expression system, and one AMP (13) was purified using the P. pastoris expression system. Inhibition zone and minimum inhibitory concentration (MIC) tests confirmed that one P. pastoris not sign -derived and two E. coli-derived AMPs have the inhibition activity. The MIC of AMP 13 and 16 from E. coli was 24.2 mu M, and the MIC of AMP 13 from P. pastoris was 8.1 mu M. The combination of prokaryotic and eukaryotic expression systems expands the universality of the developed method, facilitating screening of a large number of biologically active AMPs, establishing an AMP library, and producing AMPs by industrialised biological methods.

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