RESUMO
The low stability and nonspecific toxicity are the main limiting factors for the clinical applications of melittin (MLT). This study aimed to design and synthesize new analogs of MLT to increase stability, reduce toxicity, and retain their antimicrobial properties against bacterial pathogens. At first, peptide analogs were designed computationally by inducing single mutations in MLT peptides and evaluating their physicochemical properties. The stability of the analogs with the highest scores was determined by Gromacs software. In vitro assays were performed to examine the antimicrobial activity and toxicity of the selected analogs. Two peptide analogs, M1 and M2, were selected based on the SVM score in cell PPD. The M1 analog was created by replacing alanine with leucine on the 15th. The M2 analog was designed by substituting alanine with leucine and isoleucine with arginine at the 15th and 17th positions. According to the Gromacs results, the M2 peptide indicated more stability. RMSD and RMSF results showed no undesirable fluctuations during the 200 ns MD simulation. The MIC and MBC values for the M1 peptide were calculated in a range of 8-128 µg/ml, while the M2 peptide limited the bacterial growth to 32-128 µg/mL. Both peptides indicated less toxicity than natural MLT, based on MTT assay results. The hemolytic activity of the M1 analog was more than M2 at 16 µg/mL concentration. M1 peptide displayed the highest selectivity index against S. aureus and A. baumannii, which were approximately 5.27-fold improvements compared to MLT. In conclusion, we introduced two analogs of MLT with low toxicity, low hemolytic activity, and higher stability, along with retaining antimicrobial properties against gram-negative and positive bacteria.
