The electrical response of bilayers to the bee venom toxin melittin: Evidence for transient bilayer permeabilization

Abstract

Melittin is a 26-residue bee venom peptide that folds into amphipathic α-helix and causes membrane permeabilization via a mechanism that is still disputed. While an equilibrium transmembrane pore model has been a central part of the mechanistic dialogue for decades, there is growing evidence that a transmembrane pore is not required for melittin’s activity. In part, the controversy is due to limited experimental tools to probe the bilayer’s response to melittin. Electrochemical impedance spectroscopy (EIS) is a technique that can reveal details of molecular mechanism of peptide activity, as it yields direct, real-time measurements of membrane resistance and capacitance of supported bilayers. In this work, EIS was used in conjunction with vesicle leakage studies to characterize the response of bilayers of different lipid compositions to melittin. Experiments were carried out at low peptide to lipid ratios between 1:5000 and 1:100. The results directly demonstrate that the response of the bilayer to melittin at these concentrations cannot be explained by an equilibrium transmembrane pore model.

Multistep Molecular Dynamics Simulations Identify the Highly Cooperative Activity of Melittin in Recognizing and Stabilizing Membrane Pores

“…57 Wiedman et al reported a similar phenomenon for melittin. 58 High concentrations of melittin induced a rapid burst of leakage of vesicle-entrapped fluorescent dyes.…”

“…58 Here, we use coarse-grained MARTINI model simulations to elucidate the behaviour of ruptured pores in the presence of melittin peptides. The simulated systems are some ten times larger than the CHARMM 29 all-atom simulations presented above, albeit with a more crude representation of molecular interactions.…”

Current Understanding of the Mechanisms by which Membrane-Active Peptides Permeate and Disrupt Model Lipid Membranes

“…A melittin concentration of P: L=1:100 was shown to induce an initial and rapid leakage of vesicle-entrapped fluorescent dyes, which then slowed to zero within ~10 minutes [85]. This result suggests a dynamic evolution of the membrane pore from an initially large size upon rupture, shrinking within minutes to a final state of either no pore, or one with a radius too small to allow passage of the dye.…”

“…verified that melittin can generate pores by suddenly rupturing the lipid membrane [85]. A melittin concentration of P: L=1:100 was shown to induce an initial and rapid leakage of vesicle-entrapped fluorescent dyes, which then slowed to zero within ~10 minutes [85].…”

“…This result suggests a dynamic evolution of the membrane pore from an initially large size upon rupture, shrinking within minutes to a final state of either no pore, or one with a radius too small to allow passage of the dye. This behaviour was concentration dependent with dye efflux less than complete for concentrations less than P: L≈1:100 [85]. We have performed coarse-grained MD simulations on a range of amphipathic membrane active peptides, including melittin.…”