Combined monte carlo and molecular dynamics simulation of fully hydrated dioleyl and palmitoyl-oleyl phosphatidylcholine lipid bilayers

We have applied a new equilibration procedure for the atomic level simulation of a hydrated lipid bilayer to hydrated bilayers of dioleyl-phosphatidylcholine (DOPC) and palmitoyl-oleyl phosphatidylcholine (POPC). The procedure consists of alternating molecular dynamics trajectory calculations in a constant surface tension and temperature ensemble with configurational bias Monte Carlo moves to different regions of the configuration space of the bilayer in a constant volume and temperature ensemble. The procedure is applied to bilayers of 128 molecules of POPC with 4628 water molecules, and 128 molecules of DOPC with 4825 water molecules. Progress toward equilibration is almost three times as fast in central processing unit (CPU) time compared with a purely molecular dynamics (MD) simulation. Equilibration is complete, as judged by the lack of energy drift in 200-ps runs of continuous MD. After the equilibrium state was reached, as determined by agreement between the simulation volume per lipid molecule with experiment, continuous MD was run in an ensemble in which the lateral area was restrained to fluctuate about a mean value and a pressure of 1 atm applied normal to the bilayer surface. Three separate continuous MD runs, 200 ps in duration each, separated by 10,000 CBMC steps, were carried out for each system. Properties of the systems were calculated and averaged over the three separate runs. Results of the simulations are presented and compared with experimental data and with other recent simulations of POPC and DOPC. Analysis of the hydration environment in the headgroups supports a mechanism by which unsaturation contributes to reduced transition temperatures. In this view, the relatively horizontal orientation of the unsaturated bond increases the area per lipid, resulting in increased water penetration between the headgroups. As a result the headgroup-headgroup interactions are attenuated and shielded, and this contributes to the lowered transition temperature.     

Effect of cholesterol on the structure and dynamic properties of unsaturated phospholipid bilayers

Molecular dynamics (MD) computer simulations of five different hydrated unsaturated phosphatidylcholine lipid bilayers built up by 18:0/18:1(n-9)cis PC, 18:0/18:2(n-6)cis PC, 18:0/18:3(n-3)cis PC, 18:0/20:4(n-6)cis PC, and 18:0/22:6(n-3)cis PC molecules with 40 mol% cholesterol, and the same five pure phosphatidylcholine bilayers have been performed at 303 K. The simulation box of a lipid bilayer contained 96 phosphatidylcholines, 64 cholesterols, and 3840 water molecules (48 phosphatidylcholine molecules and 32 cholesterols per layer and 24 water molecules per phospholipid or cholesterol in each case). The lateral self-diffusion coefficients of the lipids in these systems and mass density profiles with respect to the bilayer normal have been analyzed. It has been found that the lateral diffusion coefficients of phosphatidylcholine molecules increase with increasing number of double bonds in one of the lipid chains, both in pure bilayers and in bilayers with cholesterol. It has been found as well that the lateral diffusion coefficient of phosphatidylcholine molecules of a lipid bilayer with 40 mol% cholesterol is smaller than that for the corresponding pure phosphatidylcholine bilayer.     

Cholesterol surrogates: a comparison of cholesterol and 16:0 ceramide in POPC bilayers

Experimental evidence indicates that, under some circumstances, "surrogate" molecules may play the same role as cholesterol in ordering membrane lipids. The simplest molecule in this class is Ceramide. In this article, we describe atomic-level molecular dynamics simulations designed to shed light on this phenomenon. We run simulations of hydrated phosphoryl-oleoyl phosphatidylcholine (POPC) bilayers containing cholesterol, and containing ceramide, in concentrations ranging from 5% to 33%. We also perform a simulation of a pure POPC bilayer to verify the simulation force fields against experimental structural data for POPC. Our simulation data are in good agreement with experimental data for the partial molecular volumes, areas, form factors, and order parameters. These simulations suggest that ceramide and cholesterol have a very similar effect on the POPC bilayer, although ceramide is less effective in inducing order in the bilayer compared with cholesterol at the same concentrations.     

Lipid-mediated interactions between intrinsic membrane proteins: dependence on protein size and lipid composition.

The present study is an application of an approach recently developed by the authors for describing the structure of the hydrocarbon chains of lipid-bilayer membranes (LBMs) around embedded protein inclusions ( Biophys. J. 79:2867-2879). The approach is based on statistical mechanical integral equation theories developed for the study of dense liquids. First, the configurations extracted from molecular dynamics simulations of pure LBMs are used to extract the lateral density-density response function. Different pure LBMs composed of different lipid molecules were considered: dioleoyl phosphatidylcholine (DOPC), palmitoyl-oleoyl phosphatidylcholine (POPC), dipalmitoyl phosphatidylcholine (DPPC), and dimyristoyl phosphatidylcholine (DMPC). The results for the lateral density-density response function was then used as input in the integral equation theory. Numerical calculations were performed for protein inclusions of three different sizes. For the sake of simplicity, protein inclusions are represented as hard smooth cylinders excluding the lipid hydrocarbon core from a small cylinder of 2.5 A radius, corresponding roughly to one aliphatic chain, a medium cylinder of 5 A radius, corresponding to one alpha-helix, and a larger cylinder of 9 A radius, representing a small protein such as the gramicidin channel. The lipid-mediated interaction between protein inclusions was calculated using a closed-form expression for the configuration-dependent free energy. This interaction was found to be repulsive at intermediate range and attractive at short range for two small cylinders in POPC, DPPC, and DMPC bilayers, whereas it oscillates between attractive and repulsive values in DOPC bilayers. For medium size cylinders, it is again repulsive at intermediate range and attractive at short range, but for every model LBM considered here. In the case of a large cylinder, the lipid-mediated interaction was shown to be repulsive for both short and long ranges for the DOPC, POPC, and DPPC bilayers, whereas it is again repulsive and attractive for DMPC bilayers. The results indicate that the packing of the hydrocarbon chains around protein inclusions in LBMs gives rise to a generic (i.e., nonspecific) lipid-mediated interaction which favors the association of two alpha-helices and depends on the lipid composition of the membrane.     

Lateral pressure profiles in cholesterol–DPPC bilayers

By means of atomistic molecular dynamics simulations, we study cholesterol–DPPC (dipalmitoyl phosphatidylcholine) bilayers of different composition, from pure DPPC bilayers to a 1:1 mixture of DPPC and cholesterol. The lateral pressure profiles through the bilayers are computed and separated into contributions from the different components. We find that the pressure inside the bilayer changes qualitatively for cholesterol concentrations of about 20% or higher. The pressure profile in the inside of the bilayer then turns from a rather flat shape into an alternating sequence of regions with large positive and negative lateral pressure. The changes in the lateral pressure profile are so characteristic that specific interaction between cholesterol and molecules such as membrane proteins mediated solely via the lateral pressure profile might become possible.     

Structure of Sphingomyelin Bilayers: A Simulation Study

We have carried out a molecular dynamics simulation of a hydrated 18:0 sphingomyelin lipid bilayer. The bilayer contained 1600 sphingomyelin (SM) molecules, and 50,592 water molecules. After construction and initial equilibration, the simulation was run for 3.8 ns at a constant temperature of 50°C and a constant pressure of 1 atm. We present properties of the bilayer calculated from the simulation, and compare with experimental data and with properties of dipalmitoyl phosphatidylcholine (DPPC) bilayers. The SM bilayers are significantly more ordered and compact than DPPC bilayers at the same temperature. SM bilayers also exhibit significant intramolecular hydrogen bonding between phosphate ester oxygen and hydroxyl hydrogen atoms. This results in a decreased hydration in the polar region of the SM bilayer compared with DPPC. Since our simulation system is very large we have calculated the power spectrum of bilayer undulation and peristaltic modes, and we compare these data with similar calculations for DPPC bilayers. We find that the SM bilayer has significantly larger bending modulus and area compressibility compared to DPPC.     

The effect of cholesterol on the solubilization of phosphatidylcholine bilayers by the non-ionic surfactant Triton X-100

Most of the studies on the solubilization of model membranes by Triton X-100 (TR) involve one lipid. The aim of the present study was to evaluate the effect of the addition of cholesterol on the solubilization of bilayers made of palmitoyloleoylphosphatidylcholine (POPC) or dipalmitoylphosphatidylcholine (DPPC). Detailed investigation of the kinetics of solubilization of the cholesterol-containing bilayers by TR at different temperatures reveals that: (i) At 4 degrees C, solubilization of both systems is relatively slow. Hence, in order to prevent misleading conclusions from turbidity measurements it is important to monitor the solubilization after steady-state values of optical density (OD) are reached. (ii) Studies of the temperature-induced changes of the aggregates present in mixtures of TR, POPC and cholesterol indicate that the state of aggregation at all temperatures (including 4 degrees C) represents equilibrium. By contrast, for DPPC/cholesterol/TR mixtures "kinetic traps" may occur not only at 4 degrees C but at higher temperatures as well (e.g. 37 degrees C). (iii) The presence of cholesterol in POPC bilayers makes the bilayers more resistant to solubilization at low temperatures, especially at 4 degrees C. As a consequence, the temperature dependence of the TR concentration required for complete solubilization (Dt(sol)) is no longer a monotonically increasing function (as for POPC bilayers) but a bell-shaped function, with a minimum at about 25 degrees C. Inclusion of cholesterol in DPPC bilayers makes the bilayers more resistant to solubilization at all temperatures except 4 degrees C. In this system, we observe a bell-shaped dependence of Dt(sol) on temperature, with a minimum at 37 degrees C. (iv) Both the rate of vesicle size growth and the rate of the solubilization of POPC vesicles are not affected by the inclusion of cholesterol in the bilayers. Similarly, cholesterol did not affect significantly the rate of size growth of DPPC bilayers at all temperatures, but reduced the rate of solubilization at 4 degrees C.     

Interaction of ceramides with phosphatidylcholine, sphingomyelin and sphingomyelin/cholesterol bilayers

Ceramides (Cers) may exert their biological activity through changes in membrane structure and organization. To understand this mechanism, the effect of Cer on the biophysical properties of phosphatidylcholine, sphingomyelin (SM) and SM/cholesterol bilayers was determined using fluorescence probe techniques. The Cers were bovine brain Cer and synthetic Cers that contained a single acyl chain species. The phospholipids were 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1,2-dipalmitoyl-sn-glyero-3-phosphocholine (DPPC) and bovine brain, egg yolk and bovine erythrocyte SM. The addition of Cer to POPC and DPPC bilayers that were in the liquid-crystalline phase resulted in a linear increase in acyl chain order and decrease in membrane polarity. The addition of Cer to DPPC and SM bilayers also resulted in a linear increase in the gel to liquid-crystalline phase transition temperature (T(M)). The magnitude of the change was dependent upon Cer lipid composition and was much higher in SM bilayers than DPPC bilayers. The addition of 33 mol% cholesterol essentially eliminated the thermal transition of SM and SM/Cer bilayers. However, there is still a linear increase in acyl chain order induced by the addition of Cer. The results are interpreted as the formation of DPPC/Cer and SM/Cer lipid complexes. SM/Cer lipid complexes have higher T(M)s than the corresponding SM because the addition of Cer reduces the repulsion between the bulky headgroup and allows closer packing of the acyl chains. The biophysical properties of a SM/Cer-rich bilayer are dependent upon the amount of cholesterol present. In a cholesterol-poor membrane, a sphingomyelinase could catalyze the isothermal conversion of a liquid-crystalline SM bilayer to a gel phase SM/Cer complex at physiological temperature.     

Effects of a carane derivative local anesthetic on a phospholipid bilayer studied by molecular dynamics simulation

Molecular dynamics (MD) simulations of two hydrated palmitoyloleoylphosphatidylcholine (POPC) bilayers each containing eight carane derivative (KP-23) local anesthetic (LA) molecules in neutral (POPC-LA) or protonated (POPC-LAH) forms were carried out to investigate the effect of KP-23 and its protonation on the bilayer. 3-ns trajectories were used for analyses. A pure POPC bilayer was employed as a reference system. In both POPC-LA and POPC-LAH systems a few KP-23 molecules intercalated into the bilayer and moved near the bilayer/water interface. They were located on the hydrophobic core side of the interface in the POPC-LA bilayer, but on the water phase side in the POPC-LAH bilayer. The order of the POPC chains was higher in the POPC-LA bilayer than in the pure POPC bilayer and was lower in the POPC-LAH bilayer. Interactions between polar groups of KP-23 and POPC or water were responsible for a lower hydration of POPC headgroups in POPC bilayers containing KP-23 than in the pure POPC bilayer. KP-23 molecules were found to form aggregates both in POPC-LA and POPC-LAH bilayers. Due to higher amphiphilicity of LAH, the LAH aggregate was more micelle-like and larger than the LA one. The results demonstrate the rapid timescales of the initial processes that take place at and near the bilayer interface as well as details of the atomic level interactions between local anesthetic and the lipid matrix of a cell membrane.     

Phospholipid packing and hydration in pulmonary surfactant membranes and films as sensed by LAURDAN

The efficiency of pulmonary surfactant to stabilize the respiratory surface depends critically on the ability of surfactant to form highly packed films at the air-liquid interface. In the present study we have compared the packing and hydration properties of lipids in native pulmonary surfactant and in several surfactant models by analyzing the pressure and temperature dependence of the fluorescence emission of the LAURDAN (1-[6-(dimethylamino)-2-naphthyl]dodecan-1-one) probe incorporated into surfactant interfacial films or free-standing membranes. In interfacial films, compression-driven changes in the fluorescence of LAURDAN, evaluated from the generalized polarization function (GPF), correlated with changes in packing monitored by surface pressure. Compression isotherms and GPF profiles of films formed by native surfactant or its organic extract were compared at 25 or 37 °C to those of films made of dipalmitoylphosphatidylcholine (DPPC), palmitoyloleoylphosphatidylcholine (POPC), DPPC/phosphatidylglycerol (PG) (7:3, w/w), or the mixture DPPC/POPC/palmitoyloleoylphosphatidylglycerol (POPG)/cholesterol (Chol) (50:25:15.10), which simulates the lipid composition of surfactant. In general terms, compression of surfactant films at 25 °C leads to LAURDAN GPF values close to those obtained from pure DPPC monolayers, suggesting that compressed surfactant films reach a dehydrated state of the lipid surface, which is similar to that achieved in DPPC monolayers. However, at 37 °C, the highest GPF values were achieved in films made of full surfactant organic extract or the mixture DPPC/POPC/POPG/Chol, suggesting a potentially important role of cholesterol to ensure maximal packing/dehydration under physiological constraints. Native surfactant films reached high pressures at 37 °C while maintaining relatively low GPF, suggesting that the complex three-dimensional structures formed by whole surfactant might withstand the highest pressures without necessarily achieving full dehydration of the lipid environments sensed by LAURDAN. Finally, comparison of the thermotropic profiles of LAURDAN GPF in surfactant model bilayers and monolayers of analogous composition shows that the fluorophore probes an environment that is in average intrinsically more hydrated at the interface than inserted into free-standing bilayers, particularly at 37 °C. This effect suggests that the dependence of membrane and surfactant events on the balance of polar/non-polar interactions could differ in bilayer and monolayer models, and might be affected differently by the access of water molecules to confined or free-standing lipid structures.     

Phase separation is induced by phenothiazine derivatives in phospholipid/sphingomyelin/cholesterol mixtures containing low levels of cholesterol and sphingomyelin

Lipid rafts are membrane structures enriched in cholesterol, sphingomyelin and glycolipids. In majority raft-mimicking model systems high contents of cholesterol and sphingomyelin (approximately 30 mol%) are used. Existence of raft-like structures was, however, reported also in model and natural membranes containing low levels of cholesterol and sphingomyelin. In the present work differential scanning calorimetry and fluorescence spectroscopy with the use of Laurdan probe was employed to demonstrate the existence of phase separation in model systems containing DPPC with addition of 5 mol% or 10 mol% of both cholesterol and sphingomyelin. Additionally, the influence of three phenothiazine derivatives on phase separation in mixed DPPC/cholesterol/sphingomyelin bilayers was investigated. Chlorpromazine, thioridazine and trifluoperazine were able to induce phase separation in DPPC and DPPC/cholesterol/sphingomyelin bilayers in temperatures below lipid main phase transition. However, only trifluoperazine induced phase separation in temperatures close to or above main phase transition. Trifluoperazine also induced phase separation in bilayers composed of egg yolk PC or DOPC mixed with cholesterol and sphingomyelin. We concluded that presence of lipid domains can be observed in model membranes containing low levels of cholesterol and sphingomyelin. Among three phenothiazine derivatives studied, only trifluoperazine was able to induce a permanent phase separation in phosphatidylcholine/cholesterol/sphingomyelin systems.     

Effect of hydrostatic pressure on water penetration and rotational dynamics in phospholipid-cholesterol bilayers.

The effect of high hydrostatic pressure on the lipid bilayer hydration, the mean order parameter, and rotational dynamics of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) cholesterol vesicles has been studied by time-resolved fluorescence spectroscopy up to 1500 bar. Whereas the degree of hydration in the lipid headgroup and interfacial region was assessed from fluorescence lifetime data using the probe 1-(4-trimethylammonium-phenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), the corresponding information in the upper acyl chain region was estimated from its effect on the fluorescence lifetime of and 3-(diphenylhexatrienyl)propyl-trimethylammonium (TMAP-DPH). The lifetime data indicate a greater level of interfacial hydration for DPPC bilayers than for POPC bilayers, but there is no marked difference in interchain hydration of the two bilayer systems. The addition of cholesterol at levels from 30 to 50 mol% to DPPC has a greater effect on the increase of hydrophobicity in the interfacial region of the bilayer than the application of hydrostatic pressure of several hundred to 1000 bar. Although the same trend is observed in the corresponding system, POPC/30 mol% cholesterol, the observed effects are markedly less pronounced. Whereas the rotational correlation times of the fluorophores decrease in passing the pressure-induced liquid-crystalline to gel phase transition of DPPC, the wobbling diffusion coefficient remains essentially unchanged. The wobbling diffusion constant of the two fluorophores changes markedly upon incorporation of 30 mol% cholesterol, and increases at higher pressures, also in the case of POPC/30 mol% cholesterol. The observed effects are discussed in terms of changes in the rotational characteristics of the fluorophores and the phase-state of the lipid mixture. The results demonstrate the ability of cholesterol to adjust the structural and dynamic properties of membranes composed of different phospholipid components, and to efficiently regulate the motional freedom and hydrophobicity of membranes, so that they can withstand even drastic changes in environmental conditions, such as high external hydrostatic pressure.     

Phospholipase A2 hydrolysis of supported phospholipid bilayers: a neutron reflectivity and ellipsometry study

We have investigated the phospholipase A(2) catalyzed hydrolysis of supported phospholipid bilayers using neutron reflection and ellipsometry. At the hydrophilic silica-water interface, hydrolysis of phosphatidylcholine bilayers by phospholipase A(2) from Naja mossambica mossambica venom is accompanied by destruction of the bilayer at an initial rate, which is comparable for DOPC and DPPC but is doubled for POPC. The extent of bilayer destruction at 25 degrees C decreases from DOPC to POPC and is dramatically reduced for DPPC. Neutron reflectivity measurements indicate that the enzyme penetrates into the bilayers in increasing order for DOPC, POPC, and DPPC, while the amount of enzyme adsorbed at the interface is smallest for DPPC and exhibits a maximum for POPC. Penetration into the hydrophobic chain region in the bilayer is further supported by the fact that the enzyme adsorbs strongly and irreversibly to a hydrophobic monolayer of octadecyltrichlorosilane. These results are rationalized in terms of the properties of the reaction products and the effect of their accumulation in the membrane on the kinetics of enzyme catalysis.     

Interfacial behavior of phospholipid monolayers revealed by mesoscopic simulation

A mesoscopic model with molecular resolution is presented for dipalmitoyl phosphatidylcholine (DPPC) and palmitoyl oleoyl phosphatidylcholine (POPC) monolayer simulations at the air-water interface using many-body dissipative particle dynamics (MDPD). The parameterization scheme is rigorously based on reproducing the physical properties of water and alkane and the interfacial property of the phospholipid monolayer by comparison with experimental results. Using much less computing cost, these MDPD simulations yield a similar surface pressure-area isotherm as well as similar pressure-related morphologies as all-atom simulations and experiments. Moreover, the compressibility modulus, order parameter of lipid tails, and thickness of the phospholipid monolayer are quantitatively in line with the all-atom simulations and experiments. This model also captures the sensitive changes in the pressure-area isotherms of mixed DPPC/POPC monolayers with altered mixing ratios, indicating that the model is promising for applications with complex natural phospholipid monolayers. These results demonstrate a significant improvement of quantitative phospholipid monolayer simulations over previous coarse-grained models.     

The effect of cholesterol on measured interaction and compressibility of dipalmitoylphosphatidylcholine bilayers

We have examined the phase diagram of dipalmitoylphosphatidylcholine (DPPC)--cholesterol-water mixtures at low cholesterol content, and report phase separation between 3 and 10 mol% cholesterol. The two lamellar phases at equilibrium in this region appear to be pure DPPC and 11 mol% cholesterol in DPPC. For these two lamellar phases, which are made up of alternating layers of water and bimolecular lipid leaflets, we have measured the forces of interaction between leaflets and the lateral pressure and compressibility of the leaflets. Both bilayers experience a strong repulsive force when forced together only a few ?ngstr?ms (1 A = 0.1 nm) closer than their maximum separation in excess water. However, the presence of 11 mol% cholesterol causes the bilayers to move apart of 35-A separation from the 19-A characteristic of pure DPPC in excess water. This swelling may result from a decrease in van der Waals attraction between bilayers or from an increase in bilayer repulsion. Differences in bilayer interaction can be a cause for phase separation. More importantly these differences can cause changes in the composition of regions of membranes approaching contact. At 11 mol%, cholesterol substantially increases the lateral compressibility of DPPC bilayers leading to higher lateral density fluctuations and potentially higher bilayer permeability.     

Differential scanning calorimetric studies of the thermotropic phase behavior of membranes composed of dipalmitoyllecithin and mixed-acid unsaturated lecithins

The lecithins 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC) have been synthesized by reacylation of the appropriate lysolecithins with fatty acid anhydrides. These lecithins have been used to make model membranes in mixtures with dipalmitoyllecithin (DPPC), and phase diagrams of the two bilayer systems have been constructed. These diagrams show that there is essentially no gel-state miscibility in the POPC-DPPC bilayers at any composition, and that SOPC-DPPC bilayers show gel-state immiscibility at DPPC concentrations of less than 50 mol%, and partial miscibility above 50 mol% DPPC. Analysis of the POPC-DPPC phase diagram on the assumption of athermal solution in the liquid-crystalline phase shows that the two lipids mix nearly randomly above the phase transition. The liquidus curve of SOPC-DPPC bilayers showed deviations from calculated ideal behaviour, which indicated that there is a small excess tendency for the formation of pairs of like molecules in SOPC-DPPC bilayers in the liquid-crystalline phase. Thus, in the liquid-crystalline phase, SOPC and DPPC do not pack quite as well as do POPC and DPPC.     

Effect of fenitrothion on dipalmitoyl and 1-palmitoyl-2-oleoylphosphatidylcholine bilayers

The effects of the organophosphorous insecticide fenitrothion (phosphorothioic acid, O,O-dimethyl O-(3-methyl-4-nitrophenyl) ester; FS) on the physical state of pure dipalmitoyl (DPPC) and 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) membranes were investigated. FS lowers the phase transition temperature of DPPC. It has no large effects on the DPPC gel phase, but it increases the order of the liquid-crystalline state of DPPC and POPC. FS also decreases 1,6-diphenyl-1,3,5-hexatriene (DPH) lifetime (tau) in the DPPC and POPC liquid-crystalline states. Since a direct quenching of DPH emission by FS was ruled out, tau shortening is assigned to an increased water penetration in the bilayer. The effect of FS is different from most perturbing agents for which an increased order is accompanied by a higher tau. Furthermore, quenching of DPH by KI was increased by FS in POPC liposomes indicating an increased accessibility of the quencher to the hydrophobic core where DPH distributes. The effect of FS on dipole relaxation at the hydrophilic-hydrophobic interface of POPC bilayers was studied with 2-dimethylamino-6-lauroylnaphthalene (Laurdan). FS produces a decrease in Laurdan tau and a narrowing of its emission band. FS significantly increases the generalized polarization values at both emission band ends. These results indicate that FS may allow the coexistence of microdomains that have different physical properties.     

Membrane properties of plant sterols in phospholipid bilayers as determined by differential scanning calorimetry, resonance energy transfer and detergent-induced solubilization

The increased use of plant sterols as cholesterol-lowering agents warrants further research on the possible effects of plant sterols in membranes. In this study, the effects of the incorporation of cholesterol, campesterol, beta-sitosterol and stigmasterol in phospholipid bilayers were investigated by differential scanning calorimetry (DSC), resonance energy transfer (RET) between trans parinaric acid (tPA) and 2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexadecanoyl-sn-glycero-3-phosphocholine (NBD-PC), and Triton X-100-induced solubilization. The phospholipids used were 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), D-erythro-N-palmitoyl-sphingomyelin (PSM), and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). In DSC experiments, it was demonstrated that the sterols differed in their effect on the melting temperatures of both the sterol-poor and the sterol-rich domains in DPPC and PSM bilayers. The plant sterols gave rise to lower temperatures of both transitions, when compared with cholesterol. The plant sterols also resulted in lower transition temperatures, in comparison with cholesterol, when sterol-containing DPPC and PSM bilayers were investigated by RET. In the detergent solubilization experiments, the total molar ratio between Triton X-100 and POPC at the onset of solubilization (R(t,sat)) was higher for bilayers containing plant sterols, in comparison with membranes containing cholesterol. Taken together, the observations presented in this study indicate that campesterol, beta-sitosterol and stigmasterol interacted less favorably than cholesterol with the phospholipids, leading to measurable differences in their domain properties.     

Bilayer polarity and its thermal dependency in the l(o) and l(d) phases of binary phosphatidylcholine/cholesterol mixtures

Diverse variations in membrane properties are observed in binary phosphatidylcholine/cholesterol mixtures. These mixtures are nonideal, displaying single or phase coexistence, depending on chemical composition and other thermodynamic parameters. When compared with pure phospholipid bilayers, there are changes in water permeability, bilayer thickness and thermomechanical properties, molecular packing and conformational freedom of phospholipid acyl chains, in internal dipolar potential and in lipid lateral diffusion. Based on the phase diagrams for DMPC/cholesterol and DPPC/cholesterol, we compare the equivalent polarity of pure bilayers with specific compositions of these mixtures, by using the Py empirical scale of polarity. Besides the contrast between pure and mixed lipid bilayers, we find that liquid-ordered (l(o)) and liquid-disordered (l(d)) phases display significantly different polarities. Moreover, in the l(o) phase, the polarities of bilayers and their thermal dependences vary with the chemical composition, showing noteworthy differences for cholesterol proportions at 35, 40, and 45 mol%. At 20 degrees C, for DMPC/cholesterol at 35 and 45 mol%, the equivalent dielectric constants are 21.8 and 23.8, respectively. Additionally, we illustrate potential implications of polarity in various membrane-based processes and reactions, proposing that for cholesterol containing bilayers, it may also go along with the occurrence of lateral heterogeneity in biological membranes.     

Pressure effects on the physical properties of lipid bilayers detected by trans-parinaric acid fluorescence decay.

The effects of hydrostatic pressure on the physical properties of large unilamellar vesicles of single lipids dipalmitoyl phosphatidylcholine (DPPC) and dimyristoyl phosphatidylcholine (DMPC) and lipid mixtures of DMPC/DPPC have been studied from time-resolved fluorescence of trans-parinaric acid. Additional experiments were carried out using diphenylhexatriene to compare the results extracted from both probes. Fluorescence decays were analyzed by the maximum entropy method. Pressure does not influence the fluorescence lifetime distribution of trans-parinaric acid in isotropic solvents. However, in pressurized lipid bilayers an abrupt change was observed in the lifetime distribution which was associated with the isothermal pressure-induced phase transition. The pressure to temperature equivalence values, dT/dP, determined from the midpoint of the phase transitions, were 24 and 14.5 degrees C kbar-1 for DMPC and POPC, respectively. Relatively moderate pressures of about 500 bar shifted the DMPC/DPPC phase diagram 11.5 degrees C to higher temperatures. The effects of pressure on the structural properties of these lipid vesicles were investigated from the anisotropy decays of both probes. Order parameters for all systems increased with pressure. In the gel phase of POPC the order parameter was smaller than that obtained in the same phase of saturated phospholipids, suggesting that an efficient packing of the POPC hydrocarbon chains is hindered.