Farfield Group Press Releases

05 Sep 2008 - Abstract for ASB 2008 Mechanistic Analysis of AMP bind to SLB

Mechanistic Analysis of Antimicrobial Peptides Binding to Planar Supported Lipid Membranes

Tzong-Hsien Lee1, Christine Heng1, Kristopher Hall1, John D. Gehman2, Marcus Swann3, Gerry Ronan3, Frances Separovic2, Marie-Isabel Aguilar1

1. Department of Biochemistry and Molecular Biology, Monash University, Clayton Vic 3800, Australia
2. School of Chemistry, University of Melbourne, Melbourne VIC, 3010, Australia
3. Farfield Group, Farfield House, Southmere Court, Electra Way, Crewe Business Park, Crewe CW1 6GU, UK

Effective antimicrobial peptides (AMPs) distinguish between the host and microbial cells, show broad antimicrobial spectra and exhibit a fast killing mechanism. Although the precise cytotoxicity mechanism of AMPs is still not clearly understood, electrostatic and hydrophobic interactions are involved to different degrees during the binding process. The membrane architecture and lipid composition are also critical in understanding the molecular mechanism and specificity of membrane lysis.

In this study, planar supported lipid bilayers formed via in situ liposome adsorption were used for the mechanistic analysis of AMP action. A stable bilayer with a thickness of 4.5±0.1 nm, an area of 49.7±0.8Å/molecule were obtained at 20°C for DMPC and a thickness 4.7±0.2 nm and an area of 47.4±1.6Å/molecule for DMPC/DMPG. Peptide-induced structural changes to the lipid bilayer were examined in real time using dual polarisation interferometry. Parameters including lipid layer thickness, density, mass and optical birefringence (molecular ordering) were examined for Australian frog peptides, aurein 1.2, citropin 1.1, maculatin 1.1 and caerin 1.1 binding to DMPC and DMPC/DMPG (4:1) bilayers.

The overall process of peptide induced membrane destabilisation was examined by the changes in mass as a function of optical birefringence. Among all 4 peptides, only aurein 1.2 induced destabilization of the neutral membrane (DMPC), while all 4 peptides induced destabilisation of the negatively charged membrane (DMPC/DMPG). Aurein 1.2 adsorbed to the membrane surface of both lipid membranes. Citropin 1.1 bound to the DMPC bilayer surface but inserted into the DMPC/DMPG bilayer. In addition, no significant changes in thickness were observed for low peptide concentrations (5-10uM) on both lipid bilayers. In contrast, the increased density for maculatin1.1 and caerin 1.1 binding to both lipids reflects the insertion of these longer peptides (21-25 a.a.) into the membrane.

Overall, the geometrical structural changes of the planar lipid bilayer induced by AMP binding can be elucidated by the DPI analysis. Furthermore, the stoichiometry of peptide to lipid causing membrane lysis and the kinetics of different stages of peptide adsorption, insertion and membrane lysis can provide a more detailed understanding of the molecular mechanism of peptide-lipid interactions than other sensor technologies.