Discovery, development and optimisation of a novel frog antimicrobial peptide with combined mode of action against drug-resistant bacteria

Antimicrobial peptides (AMP) have emerged as promising candidates for addressing the clinical challenges posed by the rapid evolution of antibiotic-resistant microorganisms. Brevinins, a representative frog-derived AMP family, exhibited broad-spectrum antimicrobial activities, attacking great attentions in previous studies. However, their strong haemolytic activity and cytotoxicity, greatly limit their further development. In this work, we identified and characterised a novel brevinin-1 peptide, brevinin-1pl, from the skin secretions of the northern leopard frog, <i>Rana pipiens</i>. Like many brevinins, brevinin-1pl also displayed strong haemolytic activity, resulting in a lower therapeutic index. We employed several bioinformatics tools to analyse the structure and potential membrane interactions of brevinin-1pl, leading to a series of modifications. Among these analogues, des-Ala¹⁶-[Lys⁴]brevinin-1pl exhibited great enhanced therapeutic efficacy in both <i>in vitro</i> and <i>in vivo</i> tests, particularly against some antibiotics-resistant <i>Escherichia coli</i> strains. Mechanistic studies suggest that des-Ala¹⁶-[Lys⁴]brevinin-1pl may exert bactericidal effects through multiple mechanisms, including membrane disruption and DNA binding. Consequently, des-Ala¹⁶-[Lys⁴]brevinin-1pl holds promise as a candidate for the treatment of drug-resistant <i>Escherichia coli</i> infections.