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NZ2114 is a promising candidate for therapeutic application owing to its potent activity to Staphylococcus aureus. Our objective was to identify NZ2114 derivatives with improved activity through substitution of His16 and His18 with residues Arginine and Lysine. Eight mutants were designed and expressed in Pichia pastoris X-33 via pPICZαA. Five of them exhibited strong antimicrobial activity against S. aureus at low minimal inhibitory concentrations (MICs) of 0.057-0.454 μM. Among them, H1, H2, and H3 showed ideal pharmacodynamic effects on methicillin-resistant S. aureus ATCC43300. The total protein level of H1, H2, and H3 reached 1.70, 1.77 and 1.54 g/l at 120 h of induction in the 5-l fermenter, respectively. They killed over 99.9% of pathogens within 1.5 h at 2× and 4× MIC. The post antibiotic effect of H1, H2 and H3 to S. aureus ATCC43300 was 2.94, 1.75 and 1.55 h at 2× MIC, which was similar with their original peptide NZ2114 (1.43 h) and vancomycin (1.72 h). The fractional inhibitory concentration index (FICI) indicated indifferent effects between H1, H2, H3 and vancomycin, ampicillin, rifampicin. Additionally, they had low hemolysis and high stability in different environments (temperature, pH, proteases, and saline ions). All results indicate that H1, H2, and H3 can be produced in large-scale and have potential as therapeutic drugs against MRSA. [ABSTRACT FROM AUTHOR]

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]

MP1106 is a novel variant of plectasin with potent activity against some Gram-positive pathogenic bacteria, especially for Streptococcus suis and Staphylococcus aureus . MP1106 was successfully expressed in Pichia pastoris X-33 using the glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter. The total protein concentration and antimicrobial activity were 998.3 mg/L and 51,200 AU/mL, respectively. After purification by cation exchange chromatography, recombinant MP1106 (rMP1106) with a concentration of 100.13 mg/L and a purity of 95.2% was obtained from fermentation culture. Its antimicrobial spectrum was similar to that of plectasin, and the antibacterial activity against S. suis CVCC3309 was enhanced up to 29-fold over that of plectasin. In addition, 97.9% of the S. suis CVCC606 bacteria were killed by 16× minimal inhibitory concentration (MIC) rMP1106 within 6 h. The percentage hemolysis by rMP1106 against mouse erythrocytes was 3.8% at a concentration of 512 μg/mL. Exposure of porcine intestinal epithelial cells to 16–128 μg/mL rMP1106 caused a 9.5% decrease in cell viability. The rMP1106 was stable over broad ranges of temperature, pH and NaCl concentration. In addition, rMP1106 was highly resistant to papain, proteinase K and snailase digestion but susceptible to trypsin and slightly susceptible to pepsin. These results suggested that rMP1106 has potential as a promising therapeutic drug against S. suis infections. [ABSTRACT FROM AUTHOR]