Gramicidin A disassembles large conductive clusters of its lysine-substituted derivatives in lipid membranes

Abstract

N-terminally substituted lysine derivatives of gramicidin A (gA), [Lys1]gA and [Lys3]gA, but not glutamate- or aspartate-substituted peptides have been previously shown to cause the leakage of carboxyfluorescein from liposomes. Here, the leakage induction was also observed for [Arg1]gA and [Arg3]gA, while [His1]gA and [His3]gA were inactive at neutral pH. The Lys3-containing analogue with all tryptophans replaced by isoleucines did not induce liposome leakage, similar to gA. This suggests that the presence of both tryptophans and N-terminal cationic residues is critical for pore formation. Remarkably, the addition of gA blocked the leakage induced by [Lys3]gA. By examining with fluorescence correlation spectroscopy the peptide-induced leakage of fluorescent markers from liposomes, we estimated the diameter of pores responsible for the leakage to be about 1.6 nm. Transmission electron cryo-microscopy imaging of liposomes with [Lys3]gA showed that the liposomal membranes contained high electron density particles with a size of about 40 Å, suggesting the formation of peptide clusters. No such clusterization was observed in liposomes incorporating gA or a mixture of gA with [Lys3]gA. Three-dimensional reconstruction of the clusters was compatible with their pentameric arrangement. Based on experimental data and computational modeling, we suggest that the large pore formed by [Lys3]gA represents a barrel-stave oligomeric cluster formed by antiparallel double-stranded helical dimers (DH). In a tentative model, the pentamer of dimers may be stabilized by aromatic Trp-Trp and cation-π Trp-Lys interactions between the neighboring DHs. The inhibiting effect of gA on the [Lys3]gA-induced leakage can be attributed to breaking of cation-π interactions, which prevents peptide clusterization and pore formation.

The anticancer peptide LL-III alters the physico-chemical properties of a model tumor membrane promoting lipid bilayer permeabilization

“…At a high local concentration, the CF emission is self-quenched. 36 This allows its release in solution to be monitored by following the fluorescence enhancement as a function of time both in the absence and in the presence of an increasing peptide concentration. The kinetic curves and the leakage percentage at 1500 s are reported in Fig.…”

The Integrative Approach to Study of the Structure and Functions of Cardiac Voltage-Dependent Ion Channels

“…Computer simulations have shown that this method can theoretically improve the reconstruction of membrane proteins. We applied it to study the clustering of gramicidin in liposomes [83]; however, the method did not allow achieving high resolution.…”

Characterization and Antimicrobial Activity of Amphiphilic Peptide AP3 and Derivative Sequences

“…There are several physical models that were proposed for the membrane disruptive activity of AMPs. The barrel stave model involves the insertion of the peptide into the membrane in forming a purely proteinaceous pore [22], the toroidal pore model causes bending of the lipid membrane leading to the formation protein and lipid headgroup lined pore [23], and the carpet model which causes destabilization of the cell membrane through a mass action effect similar micellization type mechanism [24].…”