ABSTRACT
We have designed and synthesized a series of cationic α-helical AMPs with improved antibacterial activity and selectivity against a broad spectrum of G(+) and G(-) bacteria. In the current study, we intended to gain further insight into the mechanisms of action between AMPs and cellular membranes using model liposomes of various phospholipid compositions. Circular dichroism measurements showed that AMPs adopted amphipathic α-helical conformation in the presence of negatively charged vesicles (DOPC/DOPG=1:3), while they were largely unstructured when incubated with neutral vesicles (DOPC). The interaction of AMPs with phospholipid vesicles were further analyzed by calcein leakage experiments. AMPs exhibited weak dye-leakage activity for DOPC (neutral) vesicles, while they effectively induced calcein leakage when interacted with DOPC/DOPG-entrapped vesicles. These results indicated that our newly designed cationic AMPs did show preferences for bacteria-mimicking anionic membranes. All of them exert their cytolytic activity by folding into an amphipathic helix upon selectively binding and insertion into the target membrane, leading to breakdown of the membrane structure, thus causing leakage of cell contents, resulting finally in cell death. Elucidating the mechanism of the membranolytic activity of AMPs may facilitate the development of more effective antimicrobial agents.
Subject(s)
Anti-Bacterial Agents , Antimicrobial Cationic Peptides , Bacteria/drug effects , Liposomes/metabolism , Phospholipids/metabolism , Amino Acid Sequence , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/metabolism , Anti-Bacterial Agents/pharmacology , Antimicrobial Cationic Peptides/chemical synthesis , Antimicrobial Cationic Peptides/metabolism , Antimicrobial Cationic Peptides/pharmacology , Fluoresceins/metabolism , Fluorescent Dyes/metabolism , Hemolysis/drug effects , Humans , Liposomes/chemistry , Molecular Sequence Data , Phospholipids/chemistryABSTRACT
Extensive use of classical antibiotics has led to the growing emergence of many resistant strains of pathogenic bacteria. Evidence has suggested that cationic antimicrobial peptides (AMPs) are of greatest potential to represent a new class of antibiotics. The largest group of AMPs comprises peptides that fold into an amphipathic alpha-helical conformation when interacting with the target microorganism. In the current study, a series of cationic AMPs of 20 amino acids was designed and synthesised based on four structural parameters, including charge, polar angle, hydrophobicity and hydrophobic moment. The effect of these parameters on antimicrobial activity and selectivity was assessed by structural and biological analyses. Our results indicated that high hydrophobicity and amphipathicity (hydrophobic moment) were correlated with increased haemolytic activity, whilst antimicrobial activity was found to be less dependent on these factors. Three of the synthetic AMPs (GW-Q4, GW-Q6 and GW-H1) showed higher antimicrobial activity and selectivity against a broad spectrum of Gram-positive and Gram-negative bacteria compared with the naturally occurring AMPs magainin 2a and pleurocidin. This study also demonstrates that these rationally designed cationic and amphipathic helical AMPs exhibited high selectivity against several Vibrio spp. and are potential agents for future use in the treatment of these marine pathogens.
