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NZ17074, a member of the arenicin family from Arenicola marina , exhibited potent antimicrobial activity against bacteria and fungi, but had some toxicity to human erythrocytes and porcine intestinal epithelial cells. To resolve this problem, one novel peptide N2 was designed based on NZ17074 by replacing Gly in position 1, 12 with Ala. N2 exhibited a typical β-sheet conformation in aqueous and almost had no hemolysis. N2 was expressed in Pichia pastoris using the small ubiquitin-like modifier fusion technology. The yield of N2 was approximately 6.5 mg/l, and its molecular weight was 2665.0 Da. The minimum inhibitory concentrations of N2 against Gram-negative bacteria ranged from 0.125 to 4 μg/ml. The antibacterial activity of N2 against Escherichia coli CVCC 195 and Salmonella enteritidis CVCC 3377 was enhanced up to 1–3-fold over parent NZ17074. This result provides some information that assists in the rational design of toxic antimicrobial peptides. [ABSTRACT FROM AUTHOR]

Currently, more antimicrobial drug candidates are urgently needed to combat the rise in drug-resistance among pathogenic microbes. A new antimicrobial peptide, MP1102, a variant of NZ2114, was designed, evaluated, and overexpressed in Pichia pastoris. The total secreted protein in cultures reached 695 mg/l, and the concentration of the recombinant MP1102 (rMP1102) was 292 mg/l. rMP1102 was purified from the fermentation supernatant by one-step cation exchange chromatography to obtain a yield of 197.1 mg/l with 96.4 % purity. rMP1102 exhibited potent activity against Gram-positive bacteria, and its minimum inhibitory concentrations (MICs) for four Staphyloccocus aureus ( S. aureus) strains ranged from 0.028 to 0.11 μM, and it had stronger activity (MIC = 0.04 to 0.23 μM) to 20 clinical isolates of MRSA (cMRSA) than rNZ2114 (MIC = 0.11 to 0.90 μM). rMP1102 was shown to kill over 99.9 % of tested S. aureus cells within 6 h when treated at one, two, and four times its MIC and over 90 % of S. aureus cells within 12 h at concentrations of 5, 10, and 20 mg/kg in a mouse thigh infection model. The higher sensitivity of MRSA to MP1102 than to its parental peptide, NZ2114, indicated by this initial pharmacodynamic analysis suggests a possible difference in the killing mechanism of these two molecules. rMP1102 caused less than 0.05 % hemolytic activity at 128 μg/ml and exhibited good thermostability from 20 to 80 °C, with its highest activity being observed at pH 8.0. These results suggest that this yeast expression system is feasible for large-scale production, and rMP1102 exerted stronger activity against S. aureus than NZ2114 via a different mechanism and exhibited potential as a new antimicrobial agent for S. aureus, especially MRSA infections. [ABSTRACT FROM AUTHOR]