Aurein-1.2

Background: With the emergence of drug-resistant fungi and the increased population prone to fungal infections, more effective antifungal drugs are needed. Aurein 1.2 is a potent antimicrobial peptide. Here, we designed a novel derivative of Aurein 1.2, called Aurein N3, which is a modified form of Aurein N2 (another Aurein 1.2 derivative), in which Lys 8 residue was replaced with Leu 13, and was also modified by creating two other mutations. Methods: Aurein N3 was designed using several algorithms and docking studies. All peptides were synthesized and some of their bio-activity indices such as antifungal properties on 11 fungi, cytotoxicity, hemolysis, and time of the killing were investigated. Electron microscopy, lived/dead staining, and ergosterol binding assay were performed to study their mechanism of action. Results: In comparison to Aurein 1.2 and N2, the docking studies showed that Aurein N3 has reduced binding energy toward ergosterol. The antifungal assessments showed that both Aurein N2 and N3 had strong activity against many fungi. Aurein N3 had lower cytotoxicity and higher binding capability to ergosterol. The hemolytic activity of Aurein N2 and N3 was less than parental Aurein 1.2. All peptides were able to attack the cell wall/membrane and enter the fungi cells. Conclusion: Here we introduced a novel derivative of Aurein 1.2 which has lower cytotoxicity, higher ergosterol-binding capability, and comparable antifungal activity compared to the original peptides. It can bind to ergosterol and can also attack the cell wall/membrane of fungi, although more studies are required to find its accurate mechanism of action.

Antibacterial peptides have potential as novel therapeutic agents for bacterial infections. Aurein 1.2 is one of the smallest antibacterial peptides extracted from an anuran. LLAA is a more active analogue of aurein 1.2. Antibacterial peptides usually accomplish their function by interacting with bacterial membrane selectively. In this study, we tried to find the reasons for the stronger antibacterial activity of LLAA compared with aurein 1.2. For this purpose, the interaction of aurein 1.2 and LLAA with dipalmitoylphosphatidylcholine (DPPC) was investigated by molecular dynamics (MD) simulation. In addition, the structure of peptides and their antibacterial activity were investigated by circular dichroism (CD) and dilution test method, respectively. MD results showed that LLAA is more flexible compared with aurein 1.2. Furthermore, LLAA loses its structure more than aurein 1.2 in the DPPC bilayer. A higher amount of water molecules penetrate into bilayer in the presence of LLAA relative to aurein 1.2. According to the antibacterial result that indicated LLAA is remarkably more active than aurein 1.2, it can be concluded that flexibility of the peptide is a determining factor in antibacterial activity. Probably, flexibility of the peptides facilitates formation of effective pores in the lipid bilayer.

Antimicrobial peptides (AMPs) are promising alternative agents for treating multidrug-resistant bacterial infections. Aurein 1.2 is a natural 13-amino acid AMP with antibacterial activity against Gram-positive bacteria. In this study, we designed three novel AMPs: aurein M1 (A10W), aurein M2 (D4K, E11K), and aurein M3 (A10W, D4K, E11K) to analyze the effect of Trp substitution and enhancement of positive charge on the activity of aurein 1.2. The AMP probability, physicochemical properties, secondary and tertiary structures, and amphipathic structure were predicted by various bioinformatics tools. After the synthesis of the peptides, their antibacterial activity, hemolysis, cytotoxicity, and structural analysis were assayed. Compared to the selectivity of aurein 1.2, the selectivity of aurein M2 and M3 with a net positive charge of +5 was improved 11.30- and 8.00-fold against Gram-positive and -negative bacteria, respectively. The hemolytic activity of aurein M2 was lower than that of aurein 1.2 and M3, while the higher percentage of human fibroblast cells were alive in the presence of aurein M3. Also, the MICs of aurein M3 toward Staphylococcus aureus and Escherichia coli at the physiologic salt were ≤16, which is recommended as a promising candidate for clinical investigation. Circular dichroism analysis indicated an alpha-helical structure in the peptide analogs that is similar to aurein 1.2 in the presence of 10 mM SDS. Therefore, increasing positive charge can be used successfully as an approach for improving the potency and selectivity of AMPs. Moreover, the beneficial effect of Trp substitution depends on its position and the sequence of peptides.