SYNTHESIS, PURIFICATION, CHARACTERIZATION AND BIOPHYSICAL STUDIES OF NATURAL PEPTIDES EXTRACTED FROM THE WASP Polybia paulista

Antimicrobial resistance is a growing problem, resulting from the indiscriminate use of antibiotics, which compromises the effective treatment of bacterial and fungal infections. In this way, this study aims to optimize the antimicrobial activities of peptides derived from the Polybia paulista wasp, focusing on increasing efficacy against pathogens without causing hemolytic effects, ensuring greater safety for animal cells. To achieve this, peptide analogues were designed replacing specific amino acids by His, Lys, and non-standard amino acids (Dap) at specific positions in the MP2 peptide sequence and them synthesized using the standard solid-phase peptide synthesis method. These modifications aim to enhance selectivity, reduce toxicity, and increase the stability of the peptides against proteolytic degradation. Additionally, the secondary structure and membrane permeabilization in LUVs membrane mimetics containing POPC:POPG (70:30, mol%) were evaluated at pH 7,4 and room temperature. The results showed that the peptide analogues [His⁵]MP2, [Dap^5,8]MP2, and [His^5,8]MP2 were successfully obtained by SPPS and purified and characterized by mass spectrometry, respectively, with purity degree higher than 95%. The CD studies indicated that the analogue peptides possess a disordered structure in aqueous solution but acquire an alpha-helical conformation when the LUVs were added to the system (helical fraction approximately 50%). Also, the K_p values were calculated and the peptide [His^5,8]MP2 presented the highest value, indicating that the affinity of the peptides is not exclusively related to their ability to form an α-helical structure, but rather to a set of physicochemical factors, such as net charge along the peptide sequence, presence of hydrophobic residues, and the ability to establish electrostatic interactions with the anionic membrane (due to the presence of POPG). In summary, this study can contribute to the search for therapeutic alternatives in the face of the growing challenge of antibiotic resistance.