A molecular dynamics study of antimicrobial peptide translocation across the outer membrane of Gram-negative bacteria

Abstract

With rising bacterial resistance, antimicrobial peptides (AMPs) have been widely investigated as potential antibacterial molecules to replace conventional antibiotics. Our understanding of the molecular mechanism for membrane disruption are largely based on AMP interactions with the inner phospholipid bilayers of both Gram-negative and Gram-positive bacteria. Mechanisms for AMP translocation across the outer membrane of Gram-negative bacteria composed of lipopolysaccharides and the asymmetric lipid bilayer are incompletely understood. In the current study, we have employed atomistic molecular dynamics and umbrella sampling simulations with an aggregate duration of ~ 8 microseconds to understand the free energy landscape of CM15 peptide within the OM of Gram-negative bacteria, E. coli. The peptide has a favourable binding free energy (-130 kJ mol-1) in the O-antigen region with a large barrier (150 kJ mol-1) at the interface between the anionic core-saccharides and upper bilayer leaflet made up of lipid A molecules. We have analyzed the peptide and membrane properties at each of the 100 ns duration umbrella sampling windows to study variations in the membrane and the peptide structure during the translocation through the OM. Interestingly the peptide is seen to elongate, adopting a membrane perpendicular orientation in the phospholipid region resulting in the formation of a transient water channel during it's translocation through the bilayer. The presence of the peptide at the lipid A and core-saccharide interface results in a 11% increase in the membrane area with the peptide adopting a predominantly membrane parallel orientation in this cation rich region. Additionally, the lateral displacement of the peptide is significantly reduced in this region, and increases toward the inner phospholipid leaflet and the outer O-antigen regions of the membrane. The peptide is found to be sufficiently hydrated across both the hydrophilic as well as hydrophobic regions of the membrane and remains unstructured without any gain in helical content. Our study unravels the complex free energy landscape for the translocation of the AMP CM15 across the outer membrane of Gram-negative bacteria and we discuss the implications of our findings with the broader question of how AMPs overcome this barrier during antimicrobial activity.

Differentiating interactions of antimicrobials with Gram-negative and Gram-positive bacterial cell walls using molecular dynamics simulations

“…58 Our recent study reports the free energy landscapes for CM15 peptide (cecropin A and melittin residues) for its translocation across the complex outer membrane of E. coli. 59 Although the interactions of melittin with lipid membranes have been extensively studied and several molecular insights have emerged [60][61][62][63][64] only few studies have reported the inter- The number of contacts made by melittin with individual units of peptidoglycans for S. aureus and E. coli models. The contacts are calculated using a distance cutoff of 0.5 nm, and averaged over 500 ns long trajectories as well as averaged over the melittin peptides.…”

“…58 Our recent study reports the free energy landscapes for CM15 peptide (cecropin A and melittin residues) for its translocation across the complex outer membrane of E. coli . 59…”