La3+-induced fusion of phosphatidylserine liposomes. Close approach, intermembrane intermediates, and the electrostatic surface potential

Abstract

The fusion of large unilamellar phosphatidylserine liposomes (PS LUV) induced by La3+ has been monitored using the 1-aminoapthalene-3,6,8-trisulfonic acid/p-xylenebis(pyridinium bromide) (ANTS/DPX) fluorescence assay for the mixing of aqueous contents. The fusion event is extensive and nonleaky, with up to 95% mixing of contents in the fused liposomes. However, addition of excess EDTA leads to disruption of the fusion products in a way that implies the existence of metastable intermembrane contact sites. The maximal fusion activity occurs between 10 and 100 microM La3+ and fusion can be terminated rapidly, without loss of contents, by the addition of excess La3+, e.g., 1 mM La3+ at pH 7.4. This observation is explained by the very large intrinsic binding constant (approximately 10(5) M-1) of La3+ to the PS headgroup, as measured by microelectrophoresis. Addition of 1 mM La3+ causes charge reversal of the membrane and a large positive surface potential. La3+ binding to PS causes the release of a proton. These data can be explained if La3+ can chelate to PS at two sites, with one of the sites being the primary amino group. This binding model successfully predicts that at pH 4.5 fusion occurs up to 2 mM La3+, due to reduced La3+ binding at low pH. We conclude that the general mechanism of membrane fusion includes three kinetic steps. In addition to (a) aggregation, there is (b) the close approach of the surfaces, or thinning of the hydration layer, and (c) the formation of intermembrane intermediates which determine the extent to which membrane destabilization leads to fusion (mixing of aqueous contents), as opposed to lysis. The lifetime of these intermembrane intermediates appears to depend upon La3+ binding to both PS sites.

Evaluation of Red CdTe and Near Infrared CdHgTe Quantum Dots by Fluorescent Imaging

“…An extremely active area of research that has constantly utilized liposomes has been the ubiquitous cellular process of membrane fusion. [5][6][7][8][9][10][11][12] For the last two decades, renewed interest has appeared in application of liposomes for drug delivery purposes. 3 More recently, liposomes have gained some more importance due to their significance in the field of bio-nanotechnology.…”

Selective Complexation and Transport of Europium Ions at the Interface of Vesicles

“…[22] Since initially the EPC vesicles were slightly negatively charged, [23] Eu III binding resulted in electrical neutralization of the vesicle surface and thus increased the attractive van der Waals forces between vesicles, which tended to aggregate. [7] These results were confirmed by freeze-fracture electron microscopy ( Figure 3). In the following, the ratio [Eu]/[lipid] was always less than 0.1, under which conditions the addition of lanthanide ions to EPC vesicles does not induce aggregation.…”

“…Indeed vesicles composed of an anionic phosphate-functionalized lipid effect the permeability to lanthanide ions by forming lipophilic complexes, [6] but the ion selectivity of these systems remains very poor because the interaction is essentially electrostatic. [7] To improve the ion selectivity, amphiphilic derivatives of b-diketone ligands were chosen in view of their ability to bind lanthanides and to form stable luminescent complexes with Eu III ions. [8] We describe here the preparation of functionalized vesicles bearing at the interface b-diketone groups that can selectively bind lanthanide ions.…”

Photoinduced Fusion of Liposomes

“…anionic liposomes in the presence of multivalent ions Ohki et al, 1982;Bentz & Duzgunes, 1985;CarmonaRibeiro et al, 1985;Bentz et al, 1988); liposomes containing polymorphic lipids, such as phosphatidylethanolamine (PE) 1, at temperatures near TH 82 (Ellens et al, 1985;Ellens et al, 1986a;Ellens et al, 1986b;Ellens et aI., 1987;Ellens et al, 1989;; neutralliposomes in the presence of dehydrating agents such as PEG (Parente & Lentz, 1986;Lentz et al, 1992;Massenburg & Lentz, 1993); so-called pH-sensitive liposomes at low pH or in the presence of multivalent cations (Connor et al, 1984;Ellens et al, 1984;Duzgunes et al, 1985;Nayar & Schroit, 1985;Straubinger et al, 1985;Leventis et al, 1987;Collins et al, 1989); formation of diacylglycerol (DAG) by reaction of phospholipase C with PC-containing liposomes (Nieva et aI., 1989;Luk et aI., 1993); and cationic liposomes in the presence of anionic liposomes (FeIgner et al, 1987;Stamatatos et al, 1988;Leventis & Silvius, 1990 In all cases, membrane destabilization most likely occurs only after interbilayer contact. The prerequisite of membrane contact to the destabilization of anionic membranes in the presence of divalent cations was demonstrated by a number of workers (Liao & Pestegard, 1980b;Liao & Pestegard, 1980a;Wilschut et al, 1980;Nir et al, 1983a).…”

“…In the model for Ca 2 + induced fusion of PS LUV, adhesion of the membranes promotes surface tension on the membranes, while overcoming the hydration forces, and resulting rupture of the membranes in the region of contact (Kachar et a1., 1986;Bentz et al, 1988). An intermembrane intermediate may mediate the fusion event, but their short lifetimes have in most cases precluded their experimental observation.…”