Final Activity Report Summary - BIOSAMP (Mechanism of interaction of antimicrobial peptides with the cell membrane - a study using bio-sensing technologies)
The goal of this work was to use a combination of novel and standard biosensing technologies to elucidate the molecular mechanisms that facilitate AMP interaction with microbial target cell membranes without lysing the host cell. Our approach in studying antimicrobial peptide/membrane interactions was to form simple membrane models that would retain the physiologically relevant membrane-fluidity but would luck other features that would complicate our study.
Specifically, work performed was focused on the detection of the mechanism of interaction of alpha-defensins, a class of antimicrobial peptides, with model membranes. Biosensors based mainly on acoustic but also on optical transducers were applied for the real time detection of the peptide/membrane interaction. Model membranes were formed on the surface of the biosensor device by using the vesicle-fusion technique.
Various combinations of surfaces and vesicles were tested in order to establish the optimum conditions for the bilayer formation. The effect of the membrane nature was assessed by forming supported lipid bilayers of various chemical compositions and/or surface charges; in addition, E.coli supported bilayers were formed based on E.coli lipids extract.
The effect of the membrane charge was clearly shown in the alpha-defensin binding; increase of the membrane-charge increased the amount of peptide binding, indicating a strong electrostatic interaction between the two. In addition, it was shown that in all case, binding did not result in any type of major membrane disruption (including solubilisation), as indicated by both biosensing and fluorescent measurements. In general, real time acoustic data was proven a valuable technique for monitoring membrane interactions and distinguishing between pore formation (for example, during melittin binding) and membrane surface-binding events.
Specifically, work performed was focused on the detection of the mechanism of interaction of alpha-defensins, a class of antimicrobial peptides, with model membranes. Biosensors based mainly on acoustic but also on optical transducers were applied for the real time detection of the peptide/membrane interaction. Model membranes were formed on the surface of the biosensor device by using the vesicle-fusion technique.
Various combinations of surfaces and vesicles were tested in order to establish the optimum conditions for the bilayer formation. The effect of the membrane nature was assessed by forming supported lipid bilayers of various chemical compositions and/or surface charges; in addition, E.coli supported bilayers were formed based on E.coli lipids extract.
The effect of the membrane charge was clearly shown in the alpha-defensin binding; increase of the membrane-charge increased the amount of peptide binding, indicating a strong electrostatic interaction between the two. In addition, it was shown that in all case, binding did not result in any type of major membrane disruption (including solubilisation), as indicated by both biosensing and fluorescent measurements. In general, real time acoustic data was proven a valuable technique for monitoring membrane interactions and distinguishing between pore formation (for example, during melittin binding) and membrane surface-binding events.