The Membrane-Permeabilizing Effect of Avenacin A-1 Involves the Reorganization of Bilayer Cholesterol

Abstract

Avenacin A-1 is a member of a group of naturally occurring compounds called saponins. It is found in oat plants, where it protects against fungal pathogens. A combined electrical and optical chamber was used to determine the interaction of avenacin A-1 with Montal-Mueller planar lipid bilayers. This system allowed simultaneous measurement of the effect of avenacin A-1 on the fluorescence and lateral diffusion of a fluorescent lipid probe and permeability of the planar lipid bilayer. As expected, cholesterol was required for avenacin A-1-induced bilayer permeabilization. The planar lipid bilayers were also challenged with monodeglucosyl, bis-deglucosyl, and aglycone derivatives of avenacin A-1. The results show that the permeabilizing activity of the native avenacin A-1 was completely abolished after one, two, or all three sugar residues are hydrolyzed (monodeglucosyl, bis-deglucosyl, and aglycone derivatives, respectively). Fluorescence recovery after photobleaching (FRAP) measurements on cholesterol-containing planar lipid bilayers revealed that avenacin A-1 caused a small but significant reduction in the lateral diffusion of the phospholipid probe N-(7-nitrobenzoyl-2-oxa-1,3-diazol-4-yl)-1, 2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (NBD-PE). Similarly, with the sterol probe (22-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl)amino)-23,24-bisnor-5-cholen-3beta-ol (NBD-Chol), avenacin A-1, but not its derivatives, caused a more pronounced reduction in the lateral diffusion than that observed with the phospholipid probe. The data indicate that an intact sugar moiety of avenacin A-1 is required to reorganize membrane cholesterol into pores.

Induction of Highly Curved Structures in Relation to Membrane Permeabilization and Budding by the Triterpenoid Saponins, α- and δ-Hederin

“…It appears that a certain cholesterol density in the liposomes is required to allow ␣-hederin to induce pores. The importance of this threshold for pore formation by other saponins agrees with observations reported by others (9,10,75,76). On cells, THP-1 macrophages depleted in cholesterol were protected against cytotoxicity induced by ␣-hederin.…”

“…In particular, the interaction with cholesterol has been suggested to be involved in the membrane permeabilization by monodesmosidic saponins with a sugar moiety at C3 (9,10).…”

Exploiting Natural Variation of Secondary Metabolism Identifies a Gene Controlling the Glycosylation Diversity of Dihydroxybenzoic Acids in Arabidopsis thaliana

“…There are known cases where the sugar moiety of a glycoside has been shown to be important for the biological activity of the molecule. For example, an intact sugar moiety was found to be essential for the membrane disruption and antifungal activity of oat saponins, a group of triterpene glycosides (Armah et al 1999). It has also been reported that the 3-b-glucoside of resveratrol, piceid (Waterhouse and LamuelaRaventos 1994), has significant higher inhibitory effect than the aglycone on spore germination and penetration of Venturia inaequalis, the causal agent for apple scab (Schulze et al 2005).…”

Induced plant secondary metabolites for phytopatogenic fungi control: a review

“…Electron microscopic analysis and electrical conductivity measurements suggest the formation of transmembrane pores (Seeman et al, 1973;Armah et al, 1999), although steroidal glycoalkaloids have been proposed to interfere with membrane integrity by extracting sterols from membranes (Keukens et al, 1992). Aggregation of the saponin-sterol complexes in the membrane may be mediated by interactions between the sugar residues of the saponin molecules (Keukens et al, 1995;Armah et al, 1999).…”

“…Electron microscopic analysis and electrical conductivity measurements suggest the formation of transmembrane pores (Seeman et al, 1973;Armah et al, 1999), although steroidal glycoalkaloids have been proposed to interfere with membrane integrity by extracting sterols from membranes (Keukens et al, 1992). Aggregation of the saponin-sterol complexes in the membrane may be mediated by interactions between the sugar residues of the saponin molecules (Keukens et al, 1995;Armah et al, 1999). The sugar chain attached to C-3 is usually critical for both the membrane-permeabilizing and antifungal properties of saponins, and removal of these sugar residues often results in loss of biological activity (Keukens et al, 1995;Armah et al, 1999).…”