Myricetin protects Galleria mellonella against Staphylococcus aureus infection and inhibits multiple virulence factors.

Staphylococcus aureus is an opportunistic pathogen related to a variety of life-threatening infections but for which antimicrobial resistance is liming the treatment options. We report here that myricetin, but not its glycosylated form, can remarkably decrease the production of several S. aureus virulence factors, including adhesion, biofilm formation, hemolysis and staphyloxanthin production, without interfering with growth. Myricetin affects both surface proteins and secreted proteins which indicate that its action is unrelated to inhibition of the agr quorum sensing system. Analysis of virulence related gene expression and computational simulations of pivotal proteins involved in pathogenesis demonstrate that myricetin downregulates the saeR global regulator and interacts with sortase A and α-hemolysin. Furthermore, Myr confers a significant degree of protection against staphylococcal infection in the Galleria mellonella model. The present findings reveal the potential of Myr as an alternative multi-target antivirulence candidate to control S. aureus pathogenicity.

Membrane negative curvature induced by a hybrid peptide from pediocin PA-1 and plantaricin 149 as revealed by atomistic molecular dynamics simulations.

Antimicrobial peptides (AMPs) are cationic peptides that kill bacteria with a broad spectrum of action, low toxicity to mammalian cells and exceptionally low rates of bacterial resistance. These features have led to considerable efforts in developing AMPs as an alternative antibacterial therapy. In vitro studies have shown that AMPs interfere with membrane bilayer integrity via several possible mechanisms, which are not entirely understood. We have performed the synthesis, membrane lysis measurements, and biophysical characterization of a novel hybrid peptide. These measurements show that PA-Pln149 does not form nanopores, but instead promotes membrane rupture. It causes fast rupture of the bacterial model membrane (POPG-rich) at concentrations 100-fold lower than that required for the disruption of mammalian model membranes (POPC-rich). Atomistic molecular dynamics (MD) simulations were performed for single and multiple copies of PA-Pln149 in the presence of mixed and pure POPC/POPG bilayers to investigate the concentration-dependent membrane disruption by the hybrid peptide. These simulations reproduced the experimental trend and provided a potential mechanism of action for PA-Pln149. It shows that the PA-Pln149 does not form nanopores, but instead promotes membrane destabilization through peptide aggregation and induction of membrane negative curvature with the collapse of the lamellar arrangement. The sequence of events depicted for PA-Pln149 may offer insights into the mechanism of action of AMPs previously shown to induce negative deformation of membrane curvature and often associated with peptide translocation via non-bilayer intermediate structures.

Effect of dimerization on the mechanism of action of aurein 1.2.

The mechanism of action of antimicrobial peptides depends on physicochemical properties such as structure, concentration, and oligomerization. Here, we focused on the effect of dimerization on the mechanism of action of aurein 1.2 (AU). We designed a lysine-linked AU dimer, (AU)2K, and its interaction with membrane mimetics was studied using four biophysical techniques and molecular dynamics simulations. Circular dichroism and molecular dynamics studies showed that AU displayed a typical spectrum for disordered structures in aqueous solution whereas (AU)2K exhibited the typical spectrum of α-helices in a coiled-coil conformation, wherein helices are wrapped around each other. With the addition of large unilamellar vesicles (LUVs), AU adopted an α-helix structure whereas the coiled-coil structure of (AU)2K assumed an extended conformation. Carboxyfluorescein release experiments with LUVs showed that both peptides were able to permeabilize vesicles although the leakage response to increases in peptide concentration differed. Optical microscopy experiments showed that both peptides induced pore opening and the dimer eventually caused the vesicles to burst. Finally, calorimetric traces determined by isothermal titration calorimetry on the LUVs also showed significant differences in peptide-membrane interactions. Together, the results of our study demonstrated that dimerization changes the mechanism of action of AU.