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

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 and expression vectors. Twenty and ten 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 expression system, and one AMP (13) was purified using the expression system. Inhibition zone and minimum inhibitory concentration (MIC) tests confirmed that one ⌐-derived and two -derived AMPs have the inhibition activity. The MIC of AMP 13 and 16 from was 24.2 μM, and the MIC of AMP 13 from was 8.1 μM. 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.