Phase Behavior Of Lipid Bilayers Research Articles

Probing Slipids Force Field for Phase Transitions in SOPC Lipid Bilayers with Various Cholesterol Concentrations

We explore the phase behavior of lipid bilayers containing SOPC (1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine) with various molar concentrations (0 mol%, 10 mol% and 30 mol%) of cholesterol. To this end, we performed extensive atomistic molecular dynamics simulations in conjunction with the Slipids force field with optimized parameters for the headgroups of phospholipids. We computed thermodynamic and structural quantities describing the ordering of the tails, the mobility of the heads and the arrangement of the lipids in the bilayers. We analyzed the behavior of the named quantities over the temperature range between 271 K and 283 K, where the experimentally determined melting temperature, Tm=279 K, lies, as well as at 400 K, which is used as a reference temperature. The obtained results are compared to available experimental data along with the outcome from molecular dynamics simulations of similar phospholipids containing different amounts of cholesterol. In the temperature interval of interest, we found evidence of the occurrence of a thermal-driven phase transition (melting) in both the pure system and the one with the lower concentration of cholesterol, while in the remaining system, the higher amount of cholesterol in the bilayer smears out the transitional behavior. Thus, we demonstrate the ability of the Slipids force field to predict the phase behavior of bilayers of SOPC and SOPC mixed with cholesterol.

Recapitulating the Lateral Organization of Membrane Receptors at the Nanoscale.

Many cell membrane functions emerge from the lateral presentation of membrane receptors. The link between the nanoscale organization of the receptors and ligand binding remains, however, mostly unclear. In this work, we applied surface molecular imprinting and utilized the phase behavior of lipid bilayers to create platforms that recapitulate the lateral organization of membrane receptors at the nanoscale. We used liposomes decorated with amphiphilic boronic acids that commonly serve as synthetic saccharide receptors and generated three lateral modes of receptor presentation─random distribution, nanoclustering, and receptor crowding─and studied their interaction with saccharides. In comparison to liposomes with randomly dispersed receptors, surface-imprinted liposomes resulted in more than a 5-fold increase in avidity. Quantifying the binding affinity and cooperativity proved that the boost was mediated by the formation of the nanoclusters rather than a local increase in the receptor concentration. In contrast, receptor crowding, despite the presence of increased local receptor concentrations, prevented multivalent oligosaccharide binding due to steric effects. The findings demonstrate the significance of nanometric aspects of receptor presentation and generation of multivalent ligands including artificial lectins for the sensitive and specific detection of glycans.

Cholesterol Spatial Distribution in Asymmetric Lipid Bilayers

While the effect of cholesterol on the phase behavior of lipid bilayers is well-known, the spatial distribution of cholesterol molecules in inner and outer leaflets is not fully understood. We have performed coarse grained molecular dynamics simulations to study spatial distribution and orientation of cholesterol molecules within asymmetric lipid bilayers at room and biological temperatures. From our studied systems we observed (i) there is a semi-linear relationship between cholesterol Z-position and Z-angle, and (ii) cholesterol molecules prefer to be located more in outer leaflet where ordered lipids are major components.

Spreading and Imbibition of Vesicle Dispersion Droplets on Porous Substrates

Vesicles have recently found widespread use in applications such as conditioning of textiles, paper and hair, as well as transdermal drug delivery. The mode of treatment in several such cases involves the application of droplets of aqueous dispersions of vesicles onto dry porous substrates like paper and textiles. One of the factors which affects the performance of such treatments is the rate at which the droplets spread and imbibe on the porous substrate. Depending upon the specific purpose of the treatment either a fast or slow droplet spreading kinetics could be desired. Therefore, it is important to have a good understanding of the droplet spreading process and the factors which influence it. In this work, an experimental investigation of the simultaneous spreading and imbibition of vesicle dispersion droplets on cellulose filter papers is carried out. Two different types of vesicles which are composed of similar lipid molecules but exhibit contrasting lipid bilayer phase behavior are used. Two different grades of filter papers with comparable porosities but different thicknesses are used as porous substrate. It is found that the droplet spreading behavior is of the “complete wetting” type on the thicker porous substrate, whereas it is of the “partial wetting” type on the thinner substrate. Furthermore, it is observed that the spreading of droplets containing vesicles with liquid-crystalline phase bilayers occurs faster than that of vesicles with solid-gel phase bilayers. The secondary radial penetration which commences after the initial droplet spreading is complete is also investigated and discussed.

Entropic forces drive clustering and spatial localization of influenza A M2 during viral budding

The influenza A matrix 2 (M2) transmembrane protein facilitates virion release from the infected host cell. In particular, M2 plays a role in the induction of membrane curvature and/or in the scission process whereby the envelope is cut upon virion release. Here we show using coarse-grained computer simulations that various M2 assembly geometries emerge due to an entropic driving force, resulting in compact clusters or linearly extended aggregates as a direct consequence of the lateral membrane stresses. Conditions under which these protein assemblies will cause the lipid membrane to curve are explored, and we predict that a critical cluster size is required for this to happen. We go on to demonstrate that under the stress conditions taking place in the cellular membrane as it undergoes large-scale membrane remodeling, the M2 protein will, in principle, be able to both contribute to curvature induction and sense curvature to line up in manifolds where local membrane line tension is high. M2 is found to exhibit linactant behavior in liquid-disordered-liquid-ordered phase-separated lipid mixtures and to be excluded from the liquid-ordered phase, in near-quantitative agreement with experimental observations. Our findings support a role for M2 in membrane remodeling during influenza viral budding both as an inducer and a sensor of membrane curvature, and they suggest a mechanism by which localization of M2 can occur as the virion assembles and releases from the host cell, independent of how the membrane curvature is produced.

Electrokinetic investigation of deposition of cationic fabric softener vesicles on anionic porous cotton fabrics

HypothesisColloidal deposition on porous substrates is a complex process influenced by both, (i) characteristics of colloidal permeation into porous substrates, and (ii) mechanism of colloidal deposition on solid surfaces. Such processes are quintessential to action of products such as hair conditioners and fabric softeners where the substrates to be treated are porous. The performance of these formulations is linked with the distribution of deposited colloids across porous substrates i.e. whether deposition is localized near substrate periphery, or deposition is homogeneously distributed. ExperimentsIn this work, we investigate the deposition of cationic vesicles, commonly used in fabric softeners, on anionic porous cotton yarns via spectrophotometric measurement of adsorption density of vesicles on yarns and electrokinetic measurement of cotton yarn apparent zeta potentials. Under the employed conditions, cotton yarn apparent zeta potentials are sensitive predominantly to external yarn surfaces. Therefore, these measurements can distinguish between deposition on external and internal yarn surfaces. FindingsThe phase behavior of lipid bilayers constituting the vesicles is identified as an important governing factor with solid-gel vesicles depositing more near yarn periphery, and liquid-crystalline vesicles depositing more uniformly throughout the yarns. Bulk electrical conductivity also influences the distribution of deposited vesicles. The results are explained with the help of a newly proposed theory.

Influence of lipid bilayer phase behavior and substrate roughness on the pathways of intact vesicle deposition: A streaming potential study

Deposition of vesicles on solid surfaces can in some cases result in the formation of assemblies of intact surface-bound vesicles known as supported vesicular layers (SVLs). Understanding the mechanism of SVL formation is thus important for effective utilization of vesicles in applications involving vesicle-substrate interactions such as drug delivery and fabric softening. In this work, we investigate SVL formation of two different types of cationic vesicles made up of lipid bilayers existing in the solid-gel and liquid-crystalline phases, respectively, on both smooth and rough anionic cellulose fibers. Deposition is studied quantitatively via spectrophotometric determination of bulk lipid concentrations, and characterization of fiber apparent zeta potentials via streaming potential measurements. Equilibrium deposition results, and profiles of variation of substrate apparent zeta potentials with deposition reveal that the deposition mechanisms of the two vesicle types are similar on rough surfaces (cotton) but significantly different on smooth surfaces (viscose). It is concluded that vesicle deposition mechanism is governed by the surface mobility of deposited vesicles which is dependent on substrate roughness and lipid bilayer phase behavior controlled vesicle properties. Results indicate that while both the vesicle types are immobile on rough surfaces, on smooth surfaces liquid-crystalline vesicles are preferentially mobile over solid-gel vesicles. Possible reasons underlying the vesicle surface mobility characteristics are discussed.

Contrasting Effects of a Rigid Core and an Alkyl Chain in nCB on the Phase Behavior of Lipid Bilayers.

Molecules incorporated into biomembranes often bear both a core and an alkyl chain in a single molecule (e.g., sterols). To clarify the effects of these two parts of a molecule, the phase behavior of a 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayer containing 4-n-alkyl-4'-cyanobiphenyl (nCB) (n = 0-8) was investigated. The trends of the main transition temperature (Tm) with respect to n and of the pretransition temperature (Tp) with respect to nCB content changed at n = 3. It was therefore suggested that the two parts of the molecule had opposing effects on the phase behavior of DPPC bilayers. The core appears to perturb molecular ordering in the gel phase and lowers Tm (like cholesterol), while alkyl chains appear to order the lipids in the gel phase and raise Tm (like n-alkanes). In addition, Tm exhibits the so-called odd-even effect based on the alkyl chain length of the minor component, nCB. Depending on the value of n, the variation in Tp was dependent on the additive content, although the pretransition was rarely observed at high contents.

Morphological Effects Induced In Vitro by Propranolol on Human Erythrocytes.

Despite the extended use and well-documented information, there are insufficient reports concerning the effects of propranolol on the structure and functions of cell membranes, particularly those of human erythrocytes. Aimed to better understand the molecular mechanisms of its interactions with cell membranes, human erythrocyte and molecular models of the red cell membrane were utilized. The latter consisted of bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representative of phospholipid classes located in the outer and inner monolayers of the human erythrocyte membrane, respectively. The capacity of propranolol to perturb the multibilayer structures of DMPC and DMPE was evaluated by X-ray diffraction. Moreover, we took advantage of the capability of differential scanning calorimetry to detect the changes in the thermotropic phase behavior of lipid bilayers resulting from propranolol interaction with DMPC and DMPE multilamellar vesicles. In an attempt to further elucidate their effects on cell membranes, the present work also examined their influence on the morphology of intact human erythrocytes by means of defocusing and scanning electron microscopy. Results indicated that propranolol induced morphological changes to human erythrocytes and interacted in a concentration-dependent manner with phospholipid bilayer.

Pressure Effects on Artificial and Cellular Membranes.

We review the combined effect of temperature and pressure on the structure, dynamics and phase behavior of lipid bilayers, differing in chain length, headgroup structure and composition as revealed by thermodynamic, spectroscopic and scattering experiments. The effect of additives, such as ions, cholesterol, and anaesthetics is discussed as well. Our data include also reports on the effect of pressure on the lateral organization of heterogeneous lipid membranes and lipid extracts from cellular membranes, as well as the influence of peptide and protein incorporation on the pressure-dependent structure and phase behavior of lipid membranes. Moreover, the effects of pressure on membrane protein function are summarized. Finally, we introduce pressure as a kinetic variable for studying the kinetics of various lipid phase transformations.

Adaptation of the Wine Bacterium Oenococcus oeni to Ethanol Stress: Role of the Small Heat Shock Protein Lo18 in Membrane Integrity

Malolactic fermentation in wine is often carried out by Oenococcus oeni. Wine is a stressful environment for bacteria because ethanol is a toxic compound that impairs the integrity of bacterial membranes. The small heat shock protein (sHsp) Lo18 is an essential actor of the stress response in O. oeni. Lo18 prevents the thermal aggregation of proteins and plays a crucial role in membrane quality control. Here, we investigated the interaction between Lo18 and four types of liposomes: one was prepared from O. oeni grown under optimal growth conditions (here, control liposomes), one was prepared from O. oeni grown in the presence of 8% ethanol (here, ethanol liposomes), one was prepared from synthetic phospholipids, and one was prepared from phospholipids from Bacillus subtilis or Lactococcus lactis. We observed the strongest interaction between Lo18 and control liposomes. The lipid binding activity of Lo18 required the dissociation of oligomeric structures into dimers. Protein protection experiments carried out in the presence of the liposomes from O. oeni suggested that Lo18 had a higher affinity for control liposomes than for a model protein. In anisotropy experiments, we mimicked ethanol action by temperature-dependent fluidization of the liposomes. Results suggest that the principal determinant of Lo18-membrane interaction is lipid bilayer phase behavior rather than phospholipid composition. We suggest a model to describe the ethanol adaptation of O. oeni. This model highlights the dual role of Lo18 in the protection of proteins from aggregation and membrane stabilization and suggests how modifications of phospholipid content may be a key factor determining the balance between these two functions.

SETUP FOR SIMULTANEOUS MICROWAVE HEATING AND REAL-TIME SPECTROFLUOROMETRIC MEASUREMENTS IN BIOLOGICAL SYSTEMS

In this paper, a delivery system allowing simultaneous microwave heating and real-time spectro∞uorometric measurements in biological systems is proposed and characterized. This system is used to investigate the phase behavior of lipid bilayers from about 15 - C to 45 - C. The delivery system is based on an open transverse electromagnetic (TEM) cell combined with a spectro∞uorometer via an optical cable system. A numerical and experimental dosimetry of the delivery system is conducted. The Speciflc Absorption Rate (SAR) e-ciency of the system is 26:1§2:1W/kg/W. Spectro∞uorometric measurements on Laurdan labeled small unilamellar vesicles (SUVs) are carried out. Generalized polarization (GP) of the SUV's membrane is obtained from the ∞uorescence intensities measured at two emission wavelengths.

Docosahexaenoic Acid Affects Gel Phase by Increasing Tilt Angle

The physical properties of docosahexasenoic acid (DHA) have dramatic effects on lipid bilayer phase behavior which could be a link to beneficial health. Its highly unsaturated structure makes fatty acid highly flexible and with a low transition temperature (Tm) of −80°C. In this investigation, we explore a binary system composed of the saturated lipid, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1-dipalmitoyl-2-docohaxeanoyl-sn-glycero-3-phosphoethanolamine (DHA-PE, 22:6) using small angle x-ray scattering (SAXS) and atomic force microscopy (AFM). Solution SAXS is a powerful tool for structural characterization and was used to identify the different phases (Lβ' and Lα) co-existing within the membrane. In a complementary study, AFM was used to investigate the morphology of coexisting domains. In a surprising result, we observed that the DHA-PE lipid used in this study induced phase separation at concentrations as low as 0.25mol% into a DHA-PE rich liquid crystalline (Lα) phase and a DHA-PE poor gel (Lβ') phase. Also unexpectedly the thickness of the gel phase is thinner than the fluid phase as shown in the d-spacing and electron density profile. We anticipated that DHA-PE would remove or decrease the angle of tilted Lβ' phase that is composed of high concentration of DPPC, but instead it does the opposite and increase the tilt angle. However, DHA-PE still maintain its thermal property and does lower the Tm of the Lβ' phase.

Acetylsalicylic acid (aspirin) and salicylic acid interaction with the human erythrocyte membrane bilayer induce in vitro changes in the morphology of erythrocytes

Despite the well-documented information, there are insufficient reports concerning the effects of salicylate compounds on the structure and functions of cell membranes, particularly those of human erythrocytes. With the aim to better understand the molecular mechanisms of the interaction of acetylsalicylic acid (ASA) and salicylic acid (SA) with cell membranes, human erythrocyte membranes and molecular models were utilized. These consisted of bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representative of phospholipid classes located in the outer and inner monolayers of the human erythrocyte membrane, respectively. The capacity of ASA and SA to perturb the multibilayer structures of DMPC and DMPE was evaluated by X-ray diffraction while DMPC unilamellar vesicles (LUV) were studied by fluorescence spectroscopy. Moreover, we took advantage of the capability of differential scanning calorimetry (DSC) to detect the changes in the thermotropic phase behavior of lipid bilayers resulting from ASA and SA interaction with PC and PE molecules. In an attempt to further elucidate their effects on cell membranes, the present work also examined their influence on the morphology of intact human erythrocytes by means of defocusing and scanning electron microscopy, while isolated unsealed human erythrocyte membranes (IUM) were studied by fluorescence spectroscopy. Results indicated that both salicylates interact with human erythrocytes and their molecular models in a concentration-dependent manner perturbing their bilayer structures.

Phase Behavior of Lipid Bilayers under Tension

Given the proposed importance of membrane tension in regulating cellular functions, we explore the effects of a finite surface tension on phase equilibrium using a molecular theory that captures the quantitative structure of the phase diagram of the tensionless DPPC/DOPC/Cholesterol lipid bilayer. We find that an increase in the surface tension decreases the temperature of the transition from liquid to gel in a pure DPPC system by ∼1.0 K/(mN/m), and decreases the liquid-disordered to liquid-ordered transition at constant chemical potentials by approximately the same amount. Our results quantitatively isolate the role of tension in comparison to other thermodynamic factors, such as pressure, in determining the phase behavior of lipid bilayers.

Effect of Mechanical Stresses on Lipid Membrane Phase Properties

We currently investigate how membrane tension modifies the miscibility properties of model lipid bilayers, namely giant unilamellar vesicles. We use fluorescence microscopy to observe coexisting liquid phases within temperature-controlled vesicles under surface tension induced by micropipette aspiration. We measure shifts in liquid/liquid miscibility temperatures as a function of applied tension.We compare our experimental data to the theoretical prediction of [1], by which a surface tension of ∼1mN/m should cause a decrease in miscibility temperature of ∼1K. This prediction may explain why Giant Plasma Membrane Vesicles (GPMVs) show large-scale liquid/liquid phase coexistence [2,3] whereas undisturbed cell membranes do not. GPMV's, which are detached from the cytoskeleton, are hypothesized to have lower surface tensions than cell membranes.[1] Uline, Schick and Szleifer. Changes in phase behavior of lipid bilayers under tension. Submitted.[2] Baumgart, Hammond, Sengupta, Hess, Holowka, Baird and Webb. Large-scale fluid/fluid phase separation of proteins and lipids in giant plasma membrane vesicles. Proc. Natl. Acad. Sci. USA, 104:3165-3170, 2007.[3] Veatch, Cicuta, Sengupta, Honerkamp-Smith, Holowka and Baird. Critical fluctuations in plasma membrane vesicles. ACS Chem Biol., 3:287-293, 2008.

High pressure effect on phase transition behavior of lipid bilayers

Phase behavior of lipid bilayers at high pressure is critical to biological processes. Using coarse grained molecular dynamic simulations, we report critical characteristics of dipalmitoylphosphatidylcholine bilayers with applied high pressure, and also show their phase transition by cooling bilayer patches. Our results indicate that the phase transition temperature of dipalmitoylphosphatidylcholine bilayers obviously shifts with pressure increasing in the rate of 37 °C kbar(-1), which are in agreement with experimental data. Moreover, the main phase transition is revealed to be strongly dependent on lipid area. A critical lipid area of ~0.57 nm(2) is found on the main phase transition boundary. Similar structures of acyl chains lead to the same sensitivity of phase transition temperature of different lipids to the pressure. Based on the lateral density and pressure profiles, we also discuss the different effects on bilayer structure induced by high temperature and high pressure, e.g., increasing temperature induces higher degree of interdigitation of lipid tails and thinner bilayers, and increasing pressure maintains the degree of interdigitation and bilayer thickness.

Dissipative particle dynamic simulation study of lipid membrane

The lipid membrane plays crucial roles in countless biologic processes, ranging from cell motility, endo- and exocytosis, and cell division to protein aggregation and trafficking. To gain a molecular insight in these biologic processes, the recently developed mesoscale simulation technique, dissipative particle dynamics (DPD) simulation, has become an invaluable tool. By providing a brief survey of existing atomistic and popular coarse-grained models used today in studying the dynamics (including vesicle formation and (protein-mediated) vesicle fusion) and phase behavior of lipid bilayers, this review illustrates how mesoscopic DPD models can be used to obtain a better understanding of these biologic processes currently inaccessible to atomistic and most coarse-grained models.

Bilayer disruption and liposome restructuring by a homologous series of small Arg-rich synthetic peptides

The effects of a series of low molecular weight water-soluble cationic linear peptide analogs (LPAs, <1000 MW) with increasing hydrophobic/hydrophilic balance on lipid bilayer phase behavior and permeability were examined using liposomes composed of zwitterionic dipalmitoylphosphatidylcholine (DPPC) and mixed zwitterionic/anionic DPPC/dipalmitoylphosphatidylglycerol (DPPG) lipid bilayers. LPAs were synthesized using a previously reported alkyl linkage strategy as Arg-C n -Arg-C n -Lys, where C n represents the saturated alkyl linkage separating the cationic residues ( n = 4, 7, or 11) (Ye et al., 2007 [1]). Differential scanning calorimetry results show that the cationic LPAs bound to and disrupted DPPC and, to a greater extent, DPPC/DPPG phase behavior. When added to preformed unilamellar liposomes, the LPAs led to significant structural changes based on cryogenic transmission electron microscopy (cryo-TEM). Coupling cryo-TEM with carboxyfluorescein leakage studies indicate that the LPAs induced permeabilization through bilayer expansion, which caused membrane thinning. The effects were inconsistent with increasing LPA hydrophobicity, which suggests that a cooperative effect between electrostatic binding and hydrophobic insertion determined the location of LPAs within the bilayer and their membrane activity. Our results for LPA-induced membrane disruption correlate with previous breast cancer cell uptake studies that showed minimal LPA-C 4 uptake, but high LPA-C 11 uptake through a non-endocytic mechanism.

Effect of pressure on membranes

Besides temperature, hydrostatic pressure has been used as a physical-chemical parameter for studying the energetics and phase behavior of membrane systems. First we review some theoretical aspects of lipid self-assembly. Then, the temperature and pressure dependent structure and phase behavior of lipid bilayers, differing in chain configuration, headgroup structure and composition as revealed by using thermodynamic, spectroscopic and scattering experiments is discussed. We also report on the lateral organization of phase-separated lipid membranes and model raft mixtures as well as the influence of peptide and protein incorporation on membrane structure and dynamics upon pressurization. Also the effect of other additives, such as ions, cholesterol, and anaesthetics is discussed. Furthermore, we introduce pressure as a kinetic variable. Applying the pressure-jump relaxation technique in combination with time-resolved synchrotron X-ray diffraction, the kinetics of various lipid phase transformations was investigated. Finally, also new data on pressure effects on membrane mimetics, such as surfactants and microemulsions, are presented.