Fusion of dipalmitoylphosphatidylcholine vesicles at 4.degree.C

Wong, Martin; Anthony, F.; Tillack, Thomas W.; Thompson, T. E.

doi:10.1021/bi00260a032

Cited by 157 publications

(81 citation statements)

References 33 publications

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“…Unilamellar vesicles were prepared from the vortex suspension by ultrasonic irradiation in a cup-horn of a Branson Sonifier W185 (Danbury, Conn.I at above the phase-transition temperature as previously reported (Kamaya et al, 1984). The preparation was aged at 4~ for one week to obtain relatively homogeneous size distribution by fusing vesicles, as described by Wong et al (1982). When observed by electron microscopy after negative staining by 0.5% ammonium molybdate, the aged preparation showed an average diameter of 0,7/xm.…”

Section: Methodsmentioning

confidence: 99%

Miscibility of phosphatidylcholine binary mixtures in unilamellar vesicles: Phase equilibria

et al. 1986

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Miscibility among phospholipids with different lipid chain-lengths or with different head groups has attracted a number of research efforts because of its significance in biological membrane structure and function. The general consensus about the miscibility of phosphatidylcholines with varying lipid chain-lengths appears to be that binary mixtures of phospholipids with a difference of two carbon atoms in the lipid chain mix well at the main phase transition. Miscibility between phosphatidylcholines with differences of four carbon atoms appears to be inconclusive. Previous reports on the phase transition of binary phospholipid mixtures are concerned mainly with multilamellar vesicles and are mostly limited to the main transition. In the present study, unilamellar vesicles were used and miscibility in binary systems between dimyristoyl-, dipalmitoyl- and distearoyl-phosphatidylcholines at pretransition as well as main transition temperatures was evaluated by constructing phase diagrams. Two methods were used to monitor the phase transitions: differential scanning microcalorimetry and optical absorbance methods. The optical method has the advantage that unilamellar vesicles of dilute phospholipid concentrations can be used. The liquidus and solidus phase boundaries were determined by the onset temperature of heating and cooling scans, respectively, because the completion temperature of a phase transition has no meaning in binary solutions. Dimyristoyl- and distearoyl-phosphatidylcholines, where the difference in the lipid chain-length is four carbon atoms, mixed well even at pretransition temperature.

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Section: Methodsmentioning

confidence: 99%

Miscibility of phosphatidylcholine binary mixtures in unilamellar vesicles: Phase equilibria

et al. 1986

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“…The preparation was aged at 4~ for two weeks to obtain homogeneous and relatively large vesicle suspension (Wong et al, 1982). Long-chain alkanols were mixed with the vesicle suspension by sonication under the flow of nitrogen gas.…”

Section: Methodsmentioning

confidence: 99%

Dose-dependent nonlinear response of the main phase-transition temperature of phospholipid membranes to alcohols

Kamaya

Lin

1986

J. Membrain Biol.

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The effect of 1-alkanols upon the main phase-transition temperature of phospholipid vesicle membranes between gel and liquid-crystalline phases was not a simple monotonic function of alkanol concentration. For instance, 1-decanol decreased the transition temperature at low concentrations, but increased it at high concentrations, displaying a minimal temperature. This concentration-induced biphasic effect cannot be explained by the van't Hoff model on the effect of impurities upon the freezing point. To explain this nonlinear response, a theory is presented which treats the effect of 1-alkanols (or any additives) on the transition temperature of phospholipid membranes in a three-component mixture. By fitting the experimental data to the theory, the enthalpy of the phase transition delta H* and the interaction energy, epsilon*AB between the additive and phospholipid molecules may be estimated. The theory predicts that when epsilon*AB greater than 2 (where epsilon*AB = epsilon AB/RT0, T0 being the transition temperature of phospholipid), both minimum and maximum transition temperatures should exist. When epsilon*AB = 2, only one inflection point exists. When epsilon*AB less than 2, neither maximum nor minimum exists. The alkanol concentration at which the transition temperature is minimum (Xmin) depends on the epsilon*AB value: the larger the epsilon*AB values, the smaller the Xmin. When epsilon*AB is large enough, Xmin values become so small that the plot delta T vs. X shows positive delta T in almost all alkanol concentrations. The interaction energy between 1-alkanols and phospholipid molecules increased with the increase in the carbon chain-length of 1-alkanols. In the case of the dipalmitoylphosphatidylcholine vesicle membrane, the carbon chain-length of 1-alkanols that caused predominantly positive delta T was about 12.

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“…Fujii et al (1 993) used sonicated vesicles, which may have been composed of phase-separated domains of gel-phase DPPC and fluidphase DOPS. Such vesicles are probably unstable, and will have a greater tendency to undergo fusion and lysis (Nicolussi et al, 1982;Wong et al, 1982). As Fujii et al (1993) pointed out, the cationic and amphipathic nature of both native and reduced defensins may promote fusion and lysis in this system.…”

Section: Mechanism Of All-or-none Leakagementioning

confidence: 99%

Interactions between human defensins and lipid bilayers: Evidence for formation of multimeric pores

1994

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Defensins comprise a family of broad-spectrum antimicrobial peptides that are stored in the cytoplasmic granules of mammalian neutrophils and Paneth cells of the small intestine. Neutrophil defensins are known to permeabilize cell membranes of susceptible microorganisms, but the mechanism of permeabilization is uncertain. We report here the results of an investigation of the mechanism by which HNP-2, one of 4 human neutrophil defensins, permeabilizes large unilamellar vesicles formed from the anionic lipid palmitoyloleoylphosphatidylglycerol (POPG). As observed by others, we find that HNP-2 (net charge = +3) cannot bind to vesicles formed from neutral lipids. The binding of HNP-2 to vesicles containing varying amounts of POPG and neutral (zwitterionic) palmitoyloleoylphosphatidylcholine (POPC) demonstrates that binding is initiated through electrostatic interactions.Because vesicle aggregation and fusion can confound studies of the interaction of HNP-2 with vesicles, those processes were explored systematically by varying the concentrations of vesicles and HNP-2, and the P0PG:POPC ratio. Vesicles (300 pM POPG) readily aggregated at HNP-2 concentrations above 1 pM, but no mixing of vesicle contents could be detected for concentrations as high as 2 pM despite the fact that intervesicular lipid mixing could be demonstrated. This indicates that if fusion of vesicles occurs, it is hemi-fusion, in which only the outer monolayers mix at bilayer contact sites. Under conditions of limited aggregation and intervesicular lipid mixing, the fractional leakage of small solutes is a sigmoidal function of peptide concentration. For 300 pM POPG vesicles, 50% of entrapped solute is released by 0.7 p M HNP-2. We introduce a simple method for determining whether leakage from vesicles is graded or all-or-none. We show by means of this fluorescence "requenching" method that native HNP-2 induces vesicle leakage in an all-or-none manner, whereas reduced HNP-2 induces partial, or graded, leakage of vesicle contents. At HNP-2 concentrations that release 100% of small (-400 Da) markers, a fluorescent dextran of 4,400 Da is partially retained in the vesicles, and a 18,900-Da dextran is mostly retained. These results suggest that HNP-2 can form pores that have a maximum diameter of approximately 25 A. A speculative multimeric model of the pore is presented based on these results and on the crystal structure of a human defensin.

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Fusion of dipalmitoylphosphatidylcholine vesicles at 4.degree.C

Cited by 157 publications

References 33 publications

Miscibility of phosphatidylcholine binary mixtures in unilamellar vesicles: Phase equilibria

Miscibility of phosphatidylcholine binary mixtures in unilamellar vesicles: Phase equilibria

Dose-dependent nonlinear response of the main phase-transition temperature of phospholipid membranes to alcohols

Interactions between human defensins and lipid bilayers: Evidence for formation of multimeric pores

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