Mechanistic Divergence and Differential Antibacterial Potency of the Proline-Rich Antimicrobial Peptide B7-005 Across ESKAPE + E Pathogens

The urgent need for new antimicrobials is driving the optimization of proline-rich antimicrobial peptides (PrAMPs) as a basis for novel antibiotics to combat multidrug-resistant pathogens. The PrAMP B7-005 has emerged from this process, displaying a broader spectrum of activity compared to similar native PrAMPs and reduced reliance on the bacterial transporter SbmA for its action. While the compatibility and interactions of B7-005 with various mammalian cell types have been investigated, most information on its molecular mechanism of antibacterial action has so far been limited to Escherichia coli. In this study, we investigated the antimicrobial potency and mechanisms of action of B7-005 across the full panel of ESKAPE pathogens, with E. coli included for comparison (ESKAPE + E). The potential of B7-005 to eradicate these pathogens was evaluated in both planktonic and biofilm forms, revealing distinct bactericidal and anti-biofilm effects across the ESKAPE + E pathogens. B7-005’s mechanism of action also varied depending on the target microorganism, ranging from intracellular inhibition of protein synthesis without membrane damage to varying levels of membrane permeabilization. Notably, B7-005 consistently inhibited protein synthesis across all ESKAPE + E pathogens, suggesting a possible combination of lytic and non-lytic mechanisms. Furthermore, biochemical analysis of its inhibitory effect on protein synthesis demonstrated that, despite acquiring membrane-destabilizing properties, B7-005 still blocks ribosome progression into the elongation phase, consistent with Class I PrAMPs. B7-005 thus retains the essential characteristics of native PrAMPs while offering a broadened spectrum of activity, highlighting its potential as a lead compound in the development of new antibiotics.