Selective Serine Substitutions of Antimicrobial Peptides Reveal Different Mechanistic Actions Toward Gram-Negative Bacteria

Antimicrobial resistance (AMR) is fast becoming a global healthcare burden, aggravated by the overuse and misuse of antibiotics. To reduce AMR-associated mobility, mortality and cost, we must actively seek alternative treatments, not primarily relying on antibiotics. Gram-negative bacteria, particularly <i>E. coli</i> and <i>P. aeruginosa</i>, are responsible for many AMR casualties worldwide. Rationally designed antimicrobial peptides (AMPs) are promising agents which destroy bacteria by direct and fast membrane disruption, impairing the ability of microbial pathogens to develop AMR. Here, we have explored how AMP's actions change based on selective substitutions of Lysine with Serine in the amino acid sequence of a designed AMP G(IIKK)₃I-NH₂. Through antimicrobial assays, neutron reflection and molecular dynamics (MD) simulations, we examined how alternating charges and amphiphilicity can impose different structural disruptions to the inner and outer membranes of Gram-negative bacteria, linking intramembrane aggregations to the improved antimicrobial actions of Serine-rich AMPs. The combined experiments and simulations have demonstrated the great potential of Serine-containing AMPs for further development into medical treatment against AMR infections.