Effect of Amino Acid Substitutions on Biological Activity of Antimicrobial Peptide: Design, Recombinant Production, and Biological Activity

Recently, antimicrobial peptides have been introduced as potent antibiotics with a wide rangeof antimicrobial activities. They have also exhibited other biological activities, including antiinflammatory,growth stimulating, and anti-cancer activities. In this study, an analog of MagaininII was designed and produced as a recombinant fusion protein. The designed sequence contained24 amino acid residues (P24), in which Lys, His, Ser residues were substituted with Arg and also,hydrophobic Phe was replaced with Trp. Recombinant production of P24 in Escherichia coli (E.coli) BL21 using pTYB21, containing chitin binding domain and intein sequence at the N-terminusof the peptide gene, resulted in 1 μg mL-1 product from culture. Chitin column chromatography,followed by online peptide cleavage with thiol reducing agent was applied to purify the peptide.Antimicrobial activity was evaluated using five bacteria strains including Staphylococcus aureus,Enterococcus faecalis, Klebsiella pneumonia, E. coli, and Pseudomonas aeruginosa. DesignedAMP exhibited promising antimicrobial activities with low minimum inhibitory concentration, inthe range of 64-256 μg/mL. P24 showed potent antimicrobial activity preferably against Grampositivebacteria, and more potent than pexiganan as a successful Magainin II analog for topicalinfections. In general, further modification can be applied to improve its therapeutic index.