Interaction of wheat α‐thionin with large unilamellar vesicles

Abstract

The interaction of the wheat antibacterial peptide a-thionin with large unilamellar vesicles has been investigated by means of fluorescence spectroscopy. Binding of the peptide to the vesicles is followed by the release of vesicle contents, vesicle aggregation, and lipid mixing. Vesicle fusion, i.e., mixing of the aqueous contents, was not observed. Peptide binding is governed by electrostatic interactions and shows no cooperativity. The amphipatic nature of wheat a-thionin seems to destabilize the membrane bilayer and trigger the aggregation of the vesicles and lipid mixing. The presence of distearoylphosphatidylethanolamine-poly(ethy1ene glycol 2000) (PEG-PE) within the membrane provides a steric barrier that inhibits vesicle aggregation and lipid mixing but does not prevent leakage. Vesicle leakage through discrete membrane channels is unlikely, because the release of encapsulated large fluorescent dextrans is very similar to that of 8-arninonaphthalene-l,3,6,trisulfonic acid (ANTS). A minimum number of 700 peptide molecules must bind to each vesicle to produce complete leakage, which suggests a mechanism in which the overall destabilization of the membrane is due to the formation of transient pores rather than discrete channels.

The Cytotoxic Plant Protein, β-Purothionin, Forms Ion Channels in Lipid Membranes

“…study determined that alterations in conductance were due to protein binding (11), and another assumed that large fluorescent cations would mimic the channels natural substrate (19). In this study, we show by direct measurement that purothionins form cation-selective channels in lipid membranes.…”

TrwD, the Hexameric Traffic ATPase Encoded by Plasmid R388, Induces Membrane Destabilization and Hemifusion of Lipid Vesicles

“…Glyceraldehyde-3-phosphate dehydrogenase, a soluble protein containing hydrophobic patches on the molecular surface, has been reported to destabilize and fuse SUV under certain conditions (26). A variety of amphipathic peptides act in a similar way (5,9,23,30). The ability of TrwD to interact with lipid bilayers in the presence of millimolar concentrations of calcium ions resembles the properties of annexins (7,11), although there is no similarity in the conserved residues of the two protein families (our unpublished observations).…”

Structural changes induced in thionins by chloride anions as determined by molecular dynamics simulations

“…Interactions of β-purothionin (βPTH) [6,17,20] with the model membranes were also consistent with protein channel formation, rather than membrane disruption [5,9]. In contrast, fluorescent probes, which could not move through ion channel pores, were released from lipid vesicles in the presence of α-purothionin suggesting destabilization and disruption of the membrane [18]. Another proposed model for membrane permeabilization by thionins is based on the ability to solubilize negatively charged phospholipids [3].…”

“…These data support the glycerol-binding site as found in the α1-purothionin crystal structure. Indeed, this site where all residues belong to one monomer is more probable because thionins are bound to the membrane bilayer in a monomeric form [3,5,18]. In contrast, K45 in the glycerol-binding site of the βPTH crystal structure belonged to a second monomer.…”

“…the negatively charged phospholipid phosphatidyl serine, which is present in bacterial, fungal, mammalian, and plant membranes. In model unilamellar vesicle studies, thionins permeabilize membranes only when the vesicles contain negatively charged phospholipids [18]. A thionin from Pyrularia pubera (PpTH) binds to phosphatidyl serine and phosphatidic acid, and effectively solubilizes these negatively charged phospholipids into the aqueous phase from the organic Biophysical Chemistry 147 (2010) [42][43][44][45][46][47][48][49][50][51][52] phase, suggesting the formation of a proteolipid complex [3].…”