1-2-oleoylphosphatidylcholine and 1-2-distearoyllecithin

Biological membranes are two-dimensional mixtures of an enormous number of different components. Modeling cell membranes as simple bilayer mixtures reveals rich phase behavior, but how can we use the observed phase behavior to understand the real membranes?

Topics: Animals; Cell Membrane; Cholesterol; Humans; Lipid Bilayers; Lipids; Models, Chemical; Molecular Structure; Phase Transition; Phosphatidylcholines; Surface Properties

The plasma membrane (PM) is asymmetric in lipid composition. The distinct and characteristic lipid compositions of the exoplasmic and cytoplasmic leaflets lead to different lipid-lipid interactions and physical-chemical properties in each leaflet. The exoplasmic leaflet possesses an intrinsic ability to form coexisting ordered and disordered fluid domains, whereas the cytoplasmic leaflet seems to form a single fluid phase. To better understand the interleaflet interactions that influence domains, we compared asymmetric model membranes that capture salient properties of the PM with simpler symmetric membranes. Using asymmetric giant unilamellar vesicles (aGUVs) prepared by hemifusion with a supported lipid bilayer, we investigate the domain line tension that characterizes the behavior of coexisting ordered + disordered domains. The line tension can be related to the contact perimeter of the different phases. Compared to macroscopic phase separation, the appearance of modulated phases was found to be a robust indicator of a decrease in domain line tension. Symmetric GUVs of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)/1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)/1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC)/cholesterol (chol) were formed into aGUVs by replacing the GUV outer leaflet with DOPC/chol = 0.8/0.2 in order to create a cytoplasmic leaflet model. These aGUVs revealed lower line tension for the ordered + disordered domains of the exoplasmic model leaflet.

Topics: Cholesterol; Phosphatidylcholines; Surface Tension; Unilamellar Liposomes

Post-translational lipid modification of integral membrane proteins is recognized as a key mechanism to modulate protein-protein and membrane-protein associations. Despite numerous reports of lipid-modified proteins, molecular-level understanding of the influence of lipid-modification of key membrane proteins remains elusive. This study focuses on the lipid modification of one such protein-claudin-5, a critical component of the blood-brain barrier tight junctions. Claudin-5 proteins are responsible for regulating the size and charge-selective permeability at the blood-brain interface. Palmitoylation of the claudin family of proteins is implicated in influencing the tight junction permeability in prior experimental studies. Here, we investigate the impact of palmitoylation on claudin-5 self-assembly using multiscale molecular simulations. To elucidate protein-membrane interactions, we used three model membrane compositions (endoplasmic reticulum, cholesterol-enriched endoplasmic reticulum, and plasma membrane) that mimic the complexity of cell organelles encountered by a typical membrane protein in its secretion pathway. The results show that palmitoylation enhances protein's affinity for cholesterol-rich domains in a membrane, and it can elicit a site-specific response based on the location of the palmitoyl chain on the protein. Also, in claudin-5 self-assembly, palmitoylation restricts specific protein-protein conformations. Overall, this study demonstrates the significance of post-translational lipid modification of proteins in cellular and subcellular membranes, and the impact palmitoylation can have on critical cellular functions of the protein.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Blood-Brain Barrier; Cholesterol; Claudin-5; Lipoylation; Membrane Microdomains; Molecular Dynamics Simulation; Phosphatidylcholines; Protein Binding; Protein Domains; Protein Processing, Post-Translational; Tight Junctions

Preparation of lipid-based drug delivery systems by microfluidics has been increasingly popular, due to the reproducible, continuous and scalable nature of the microfluidic process. Despite exciting development in the field, versatility and superiority of microfluidics over conventional methods still need further evidence, since preparing clinically-relevant sterically stabilised liposomes has been lacking. The present study describes the optimisation of PEGylated liposomal formulations of various rigidity using staggered herringbone micromixer (SHM). The effect of both processing parameters (total flow rate (TFR) and aqueous-to-ethanol flow rate ratio (FRR)) and formulation parameters (lipid components and composition, initial lipid concentration and aqueous media) was investigated and discussed. Liposomal formulations consist of 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) or 1,2-distearoyl-sn-glycero-3-phosphatidylcholine (DSPC), with cholesterol and PEGylated lipid (DSPE-PEG

Topics: 1,2-Dipalmitoylphosphatidylcholine; Doxorubicin; Microfluidics; Particle Size; Phosphatidylcholines; Phosphatidylethanolamines; Polyethylene Glycols

We have studied the contributions of stored elastic energies in liquid-ordered (Lo) and liquid-disordered (Ld) domains to transmembrane proteins using the lateral pressure concept. In particular we applied previously reported experimental data for the membrane thickness, intrinsic curvature and bending elasticities of coexisting Lo/Ld domains to calculate whether proteins of simple geometric shapes would preferentially diffuse into Lo or Ld domains and form oligomers of a certain size. For the studied lipid mixture we generally found that proteins with convex shapes prefer sorting to Ld phases and the formation of large clusters. Lo domains in turn would be enriched in monomers of concave shaped proteins. We further observed that proteins which are symmetric with respect to the bilayer center prefer symmetric Lo or Ld domains, while asymmetric proteins favor a location in domains with Lo/Ld asymmetry. In the latter case we additionally retrieved a strong dependence on protein directionality, thus providing a mechanism for transmembrane protein orientation.

Topics: Cholesterol; Lipid Bilayers; Membrane Proteins; Molecular Dynamics Simulation; Phosphatidylcholines; Protein Multimerization; X-Ray Diffraction

Aluminum phthalocyanine chloride (AlClPc) is a second-generation photodynamic therapy (PDT) photosensitizer characterized for its high hydrophobicity and self-aggregation tendency in aqueous media, which hamper its potential application. Aiming at AlClPc solubilization we proposed here the use of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) at different proportions to form mixed lipid vesicles (LVs) as a drug delivery system. LVs were prepared by ethanol injection method and formed nano-sized vesicles (about 100nm) with suitable polydispersity index, negative zeta potential, and stable in aqueous medium for at least 50days. AlClPc strongly interacts with LV (high binding constant values), especially due to aluminum-phosphate specific interactions, which gives a surface localization to AlClPc molecules as demonstrated by fluorescence quenching data. Anisotropy, static and time-resolved fluorescence measurements corroborated with these results and demonstrated that AlClPc self-aggregation occurred even in the liposomes. However, formulation uptake by oral squamous cell carcinoma (OSCC) the AlClPc was distributed in cellular organelles and suffered a disaggregation process demonstrated by fluorescence life-time imaging microscopy. This amazing behavior is new and increases the scientific knowledge about the intracellular mechanism of action of PDT photosensitizers. In addition, these results open a new perspective to the potential use of AlClPc-LV formulations for photodynamic treatment.

Topics: Cell Line, Tumor; Ethanol; Fluorescence Polarization; Glycerylphosphorylcholine; Humans; Hydrophobic and Hydrophilic Interactions; Indoles; Liposomes; Microscopy, Fluorescence; Organometallic Compounds; Phosphatidylcholines; Photosensitizing Agents

In the present chapter we discuss the complex mixing behaviour of plasma membrane lipids. To do so, we first introduce the plasma membrane and membrane mixtures often used to model its complexity. We then discuss the nature of lipid phase behaviour in bilayers and the distinction between these phases and other manifestations of non-random mixing found in one-phase mixtures, such as clusters, micelles and microemulsions. Finally, we demonstrate the applicability of Gibbs phase diagrams to the study of increasingly complex model membrane systems, with a focus on phase coexistence, morphology and their implications for the cell plasma membrane.

Topics: Cholesterol; Emulsions; Kinetics; Lipid Bilayers; Membrane Microdomains; Membrane Proteins; Micelles; Models, Chemical; Monte Carlo Method; Phase Transition; Phosphatidylcholines; Thermodynamics

The study of the ability of drug molecules to enter cells through the membrane is of vital importance in the field of drug delivery. In cases where the transport of the drug molecules through the membrane is not easily accomplishable, other carrier molecules are used. Spherical fullerene molecules have been postulated as potential carriers of highly hydrophilic drugs across the plasma membrane. Here, we report the coarse-grain molecular dynamics study of the translocation of C60 fullerene and its derivatives across a cell membrane modeled as a 1,2-distearoyl-sn-glycero-3-phosphocholine bilayer. Simulation results indicate that pristine fullerene molecules enter the bilayer quickly and reside within it. The addition of polar functionalized groups makes the fullerenes less likely to reside within the bilayer but increases their residence time in bulk water. Addition of polar functional groups to one half of the fullerene surface, in effect creating a Janus particle, offers the most promise in developing fullerene models that can achieve complete translocation through the membrane bilayer.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Cell Membrane; Fullerenes; Lipid Bilayers; Molecular Dynamics Simulation; Phosphatidylcholines; Temperature

We report the first 4-component phase diagram for the lipid bilayer mixture, DSPC/DOPC/POPC/chol (distearoylphosphatidylcholine/dioleoylphosphatidylcholine/1-palmitoyl, 2-oleoylphosphatidylcholine/cholesterol). This phase diagram, which has macroscopic Ld+Lo phase domains, clearly shows that all phase boundaries determined for the 3-component mixture containing DOPC transition smoothly into the boundaries for the 3-component mixture containing POPC, which has nanoscopic phase domains of Ld+Lo. Our studies start from two published ternary phase diagrams, and show how these can be combined into a quaternary phase diagram by study of a few hundred samples of intermediate compositions.

Topics: Cholesterol; Fluorescence Resonance Energy Transfer; Phase Transition; Phosphatidylcholines

Cholesterol is a major constituent of biological membranes in mammalian cells. Experiments have shown that cholesterol influences the physical properties of the plasma membrane, such as lateral diffusion and phase equilibrium. In addition to controlling the 2-dimensional phase behaviour and mobility of lipids in membranes, cholesterol has also been implicated in the transbilayer diffusion of lipids across the bilayer. Sum-frequency vibrational spectroscopy (SFVS) is used to measure the intrinsic rate of lipid flip-flop for 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) in the presence of cholesterol using planar supported lipid bilayer (PSLB) model membranes. Asymmetric PSLBs were prepared using the Langmuir-Blodgett (LB) method by placing a perdeuterated lipid analogue in one leaflet of the PSLB. SFVS was used to directly measure the asymmetric distribution of DSPC within the membrane by measuring the decay in the CH3 vs intensity at 2875 cm(-1) with time and as a function of temperature. A complete kinetic analysis of DSPC flip-flop and the effect of cholesterol on the DSPC dynamics are presented. An analysis of the kinetic data in the framework of Eyring theory provides important insight into the transition state enthalpy (deltaH(double dagger)), entropy (deltaS(double dagger)) and free energy (deltaG(double dagger)) for this important biological process. In addition, the transmembrane migration of cholesterol molecules was also explored by SFVS. These combined studies are aimed at providing new insight in to the transbilayer migration of phospholipids and cholesterol in biological membranes and the effects cholesterol plays in membrane dynamics.

Topics: Cell Membrane; Cholesterol; Lipid Bilayers; Membrane Lipids; Models, Chemical; Phosphatidylcholines; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis; Thermodynamics; Vibration

The studies on the condensing and ordering effect of cholesterol by application of the Langmuir monolayer technique are usually performed on binary lipid/cholesterol systems. The results concerning a quantitative analysis of these effects in multicomponent monolayers are very limited. In this work the condensing and ordering effect of cholesterol in ternary (SM/DSPC/Chol and SM/DOPC/Chol) and quaternary (SM/DSPC/DOPC/Chol) films was investigated. It was evidenced that the systems containing saturated PC (both SM/DSPC and SM/DSPC/Chol) are always more condensed and chain-ordered than the systems containing unsaturated PC (SM/DOPC and SM/DSPC/DOPC and their mixtures with cholesterol). However, the magnitude of condensation provoked by cholesterol at higher surface pressures is stronger on the monolayers containing unsaturated PC. The addition of cholesterol into SM/PC films induces the increase of chain-ordering however, the effectiveness of cholesterol as an ordering agent is determined by the presence/absence of unsaturated phospholipid. The magnitude of the effect of cholesterol on the investigated mixed monolayer was analyzed in the context of the influence of sterol on lipid chains (ordering, straightening and reorientation of chains) as well as the reorientation of polar heads.

Topics: Animals; Chickens; Cholesterol; Eggs; Lipids; Materials Testing; Models, Chemical; Phosphatidylcholines; Phospholipids; Protein Conformation; Sphingomyelins; Surface Properties

Recently, a number of ternary phase diagrams of lipid mixtures have been constructed using various experimental techniques with a common goal of understanding the nature of lipid domains. An accurate experimental phase diagram can provide rich thermodynamic information and can also be used to extract molecular interactions using computer simulation. In this study, the liquid-ordered and liquid-disordered (l(o)-l(d)) phase boundary of DSPC/DOPC/Cholesterol ternary mixtures is simulated in a lattice model using pairwise interactions. The block composition distribution (BCD) technique was used to locate accurately the compositions of coexisting phases and thermodynamics tie-lines in the two-phase region, and the Binder ratio method was used to determine the phase boundary in the critical region. In simulations performed along a thermodynamic tie-line, the BCD method correctly samples the compositions as well as the relative amounts of coexisting phases, which is in excellent agreement with the lever rule. A "best-fit" phase boundary was obtained that has a top boundary closely resembling the experimental boundary. However, the width of the simulated two-phase region is significantly wider than the experimental one. The results show that pairwise interactions alone are not sufficient to describe the complexity of molecular interactions in the ternary lipid mixtures; more complex forms of interactions, possibly multibody interaction or domain interfacial energy, should be included in the simulation.

Topics: Cholesterol; Computer Simulation; Phosphatidylcholines; Thermodynamics

Molecular dynamics simulations are used to study the influence of chloroform in two different lipid membranes: one representative of a liquid-disordered phase and another one mixed with cholesterol and representative of a liquid-ordered phase. When chloroform is added to the cholesterol-containing membrane, a strong chain disordering is induced. In both cases, chloroform laterally disorganizes the membranes. The analysis of the main structural and dynamical membrane properties reveals that the interaction with cholesterol is the main factor to explain the strong disordering effect of chloroform in liquid-ordered phases. The results support and provide a molecular explanation to the observations of Regen et al. ( J. Am. Chem. Soc. 2009 , 131 , 5068 ) that suggest that chloroform loosens cholesterol-containing bilayers, thus changing their lateral lipid organization. This lipid-mediated mechanism is conjectured by Regen et al. to be responsible for the anesthetic effect of chloroform and other small volatile anesthetic compounds. This proposal is also discussed.

Topics: Anesthetics; Carbon Tetrachloride; Chloroform; Cholesterol; Hydrophobic and Hydrophilic Interactions; Lipid Bilayers; Membrane Fluidity; Molecular Dynamics Simulation; Phosphatidylcholines; Static Electricity; Water

We have found modulated phase morphology in a particular region of composition within the liquid-ordered + liquid-disordered coexistence region in the four-component lipid bilayer mixture DSPC/DOPC/POPC/Chol. By controlling lipid composition, we could see distinct types of modulated liquid-liquid phase morphologies, including linear, irregular, and angular features in giant unilamellar vesicles. We used a combination of confocal, two-photon, wide-field fluorescence, and differential interference contrast microscopies, and used stringent controls to minimize light-induced artifacts. These studies establish that both the size and morphology of membrane rafts can be controlled by the concentration and the type of low-melting lipid in mixtures with cholesterol and a high-melting lipid.

Topics: Cholesterol; Microscopy, Fluorescence; Nanoparticles; Phase Transition; Phosphatidylcholines; Temperature; Unilamellar Liposomes

Phase diagrams of ternary lipid mixtures containing cholesterol have provided valuable insight into cell membrane behaviors, especially by describing regions of coexisting liquid-disordered (Ld) and liquid-ordered (Lo) phases. Fluorescence microscopy imaging of giant unilamellar vesicles has greatly assisted the determination of phase behavior in these systems. However, the requirement for optically resolved Ld + Lo domains can lead to the incorrect inference that in lipid-only mixtures, Ld + Lo domain coexistence generally shows macroscopic domains. Here we show this inference is incorrect for the low melting temperature phosphatidylcholines abundant in mammalian plasma membranes. By use of high compositional resolution Förster resonance energy transfer measurements, together with electron spin resonance data and spectral simulation, we find that ternary mixtures of DSPC and cholesterol together with either POPC or SOPC, do indeed have regions of Ld + Lo coexistence. However, phase domains are much smaller than the optical resolution limit, likely on the order of the Förster distance for energy transfer (R(0), ∼2-8 nm).

Topics: Cholesterol; Electron Spin Resonance Spectroscopy; Ergosterol; Fluorescence Resonance Energy Transfer; Lipid Bilayers; Membrane Microdomains; Phase Transition; Phosphatidylcholines; Porphobilinogen; Surface Properties

Cell membrane lipids and proteins are heterogeneously distributed in the membrane plane. In recent years, much attention has been paid to the heterogeneous distribution of the lipid components, particularly the formation of cholesterol-rich domains that are thought to be important in signaling processes. This has led to renewed interest in the phase diagrams of complex lipid mixtures, such as three-component mixtures containing phospholipids and cholesterol. We report here a novel fluorescent probe (NBD-R595) that is useful for exploring the phase behaviors of one-, two-, and three-component large unilamellar vesicles. In one-component fluid-phase membranes, the probe has the expected spectral characteristic of monomeric 7-nitrobenzo-2-oxa-1,3-diazol, with a fluorescence maximum of 540 nm when excited at 470 nm. But below the gel-to-liquid crystalline phase transition temperature, an additional emission peak appears at approximately 610 nm, because of Förster resonance energy transfer from NBD-R595 monomers to NBD-R595 Jelley aggregates of limited size formed by the association of 7-nitrobenzo-2-oxa-1,3-diazol moieties. This may be the first report of Förster resonance energy transfer from a single fluorophore in two different physical states. In a test of the probe, we found NBD-R595 to be remarkably sensitive to the molar composition of large unilamellar vesicles formed from cholesterol, distearoylphosphatidylcholine, and dioleoylphosphatidylcholine.

Topics: Bacterial Proteins; Cholesterol; Fluorescence; Lipid Bilayers; Lipopolysaccharides; Phase Transition; Phosphatidylcholines; Salmonella; Spectrometry, Fluorescence; Temperature; Unilamellar Liposomes

Giant vesicles generated from synthetic and natural lipids such as phosphatidylcholines are useful models for understanding mechanical properties of cell membranes. Line tension is the one-dimensional force enabling the closing of transient pores on cell membranes. Transient pores were repeatedly and reproducibly formed on the membrane edge of giant vesicles generated from synthetic and natural phosphatidylcholines employing a nitrogen-pumped coumarin dye laser (440 nm). Line tension was determined at room temperature from closing of these pores that occurred over several seconds when the radius of the vesicle could be considered to be constant. The value of line tension depends on the nature of the lipid for single lipid systems, which, at room temperature, yielded a vesicle bilayer region in the gel, fluid, or mixed gel and fluid phases. The line tension for vesicles generated from phosphatidylcholines with saturated acyl chains of lengths of 12-18 carbon atoms ranges from 1 to 12 pN, exhibiting an increase with chain length. Vesicles generated from the natural Egg-PC, which is a mixture of lipids, are devoid of phase transition and exhibited the largest value of line tension (32 pN). This value is much larger than that estimated from the line tensions of vesicles obtained from lipids with homologous acyl chains. This study, to our knowledge, is the first to employ laser ablation to generate transient pores and determine line tension from the rate of pore closure and demonstrate a relationship between line tension and acyl chain length.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Algorithms; Chemical Phenomena; Chemistry, Physical; Dimyristoylphosphatidylcholine; Elasticity; Kinetics; Lipid Bilayers; Phosphatidylcholines; Surface Tension; Unilamellar Liposomes; Viscosity

This work presents a novel method for experimentally quantifying interfacial line tension, which can be readily applied to study a wide variety of different lipid mixtures exhibiting phase coexistence. The method combines AFM imaging of lipid domain nucleation with classical nucleation theories. The results, using symmetric and asymmetric domains, permit the prediction of key physical parameters (critical nuclei size and nucleation rate) in multicomponent bilayer systems with implications toward understanding the dynamic nature of submicrometer domains (i.e., lipid rafts) in cell membranes.

Topics: Galactosylceramides; In Vitro Techniques; Lipid Bilayers; Membrane Microdomains; Microscopy, Atomic Force; Phosphatidylcholines; Surface Tension; Thermodynamics

We have undertaken a series of experiments to examine the behavior of individual components of cell membranes. Here we report an initial stage of these experiments, in which the properties of a chemically simple lipid mixture are carefully mapped onto a phase diagram. Four different experimental methods were used to establish the phase behavior of the 3-component mixture DSPC/DOPC/chol: (1) confocal fluorescence microscopy observation of giant unilamellar vesicles, GUVs; (2) FRET from perylene to C20:0-DiI; (3) fluorescence of dilute dyes C18:2-DiO and C20:0-DiI; and (4) wide angle X-ray diffraction. This particular 3-component mixture was chosen, in part, for a high level of immiscibility of the components in order to facilitate solving the phase behavior at all compositions. At 23 degrees C, a large fraction of the possible compositions for this mixture give rise to a solid phase. A region of 3-phase coexistence of {Lalpha+Lbeta+Lo} was detected and defined based on a combination of fluorescence microscopy of GUVs, FRET, and dilute C20:0-DiI fluorescence. At very low cholesterol concentrations, the solid phase is the tilted-chain phase Lbeta'. Most of the phase boundaries have been determined to be within a few percent of the composition. Measurements of the perturbations of the boundaries of this accurate phase diagram could serve as a means to understand the behaviors of a range of added lipids and proteins.

Topics: Cholesterol; Fluorescence Resonance Energy Transfer; Lipid Bilayers; Phase Transition; Phosphatidylcholines; Spectrometry, Fluorescence; X-Ray Diffraction

Topics: Fatty Acids, Monounsaturated; Lipid Bilayers; Microscopy, Atomic Force; Nanoparticles; Phosphatidylcholines; Quaternary Ammonium Compounds

Topics: Animals; Biosensing Techniques; Cattle; Cholera Toxin; G(M1) Ganglioside; In Vitro Techniques; Kinetics; Membranes, Artificial; Models, Biological; Phosphatidylcholines

In order to understand the effect of cis unsaturation on the thermotropic and barotropic phase behavior of phospholipid bilayer membranes, the phase transitions of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) bilayer membranes were observed by high-pressure optical method. With respect to DOPC bilayer membrane, the so-called main transition between the liquid crystalline (Lalpha) and the lamellar gel (Lbeta) phases was observed in water at above 0 degrees C under high pressure, in addition to the transition between the Lalpha and the lamellar crystalline (L(C)) phases in 50% aqueous ethylene glycol. The pressure of main transition increased linearly with an increase in temperature. Extrapolation of temperature (T)-pressure (P) phase boundary to ambient pressure suggests the temperature of the main transition to be -40.3 degrees C, which has never been found by the DSC method. On the other hand, the temperature of L(C)/Lalpha phase transition in 50% aqueous ethylene glycol was found to be -12.0 degrees C at ambient pressure. The main transition temperatures for DSPC, SOPC and DOPC are 55.6, 6.7 and -40.3 degrees C, respectively, at ambient pressure. The substitution of cis unsaturated chain for saturated chains of DSPC brings about the depression of the main transition temperature by about 48 (+/-1) degrees C for each chain. The volume changes (deltaV) associated with the transitions were calculated from the transition enthalpy (deltaH) and the slope of T-P diagram (dT/dP) by means of the Clapeyron-Clausius equation. The value of deltaV for the main transition of SOPC bilayer membranes was reduced to half the volume change for DSPC bilayers, which means the introduction of the cis double bond in the acyl chain of lipids brings about the reduction of deltaV because of the disordered packing of unsaturated chains in the gel phase of lipid bilayer membranes.

Topics: Biophysical Phenomena; Biophysics; Ethylene Glycol; Gels; Lipid Bilayers; Phosphatidylcholines; Pressure; Stereoisomerism; Temperature; Thermodynamics; Water

Alpha-Hemolysin is an extracellular protein toxin (107 kDa) produced by some pathogenic strains of Escherichia coli. Although stable in aqueous medium, it can bind to lipid bilayers and produce membrane disruption in model and cell membranes. Previous studies had shown that toxin binding to the bilayer did not always lead to membrane lysis. In this paper, we find that alpha-hemolysin may bind the membranes in at least two ways, a reversible adsorption and an irreversible insertion. Reversibility is detected by the ability of liposome-bound toxin to induce hemolysis of added horse erythrocytes; insertion is accompanied by an increase in the protein intrinsic fluorescence. Toxin insertion does not necessarily lead to membrane lysis. Studies of alpha-hemolysin insertion into bilayers formed from a variety of single phospholipids, or binary mixtures of phospholipids, or of phospholipid and cholesterol, reveal that irreversible insertion is favored by fluid over gel states, by low over high cholesterol concentrations, by disordered liquid phases over gel or ordered liquid phases, and by gel over ordered liquid phases. These results are relevant to the mechanism of action of alpha-hemolysin and provide new insights into the membrane insertion of large proteins.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Adsorption; Bacterial Proteins; Bacterial Toxins; Cholesterol; Dimyristoylphosphatidylcholine; Escherichia coli; Escherichia coli Proteins; Fluorescent Dyes; Hemolysin Proteins; Kinetics; Lipid Bilayers; Phosphatidylcholines; Protein Binding; Spectrometry, Fluorescence

Partition coefficients, Kp, of four dopamine antagonists (pimozide, fluspirilene, haloperidol and domperidone) between the aqueous phase and lipid bilayer vesicles were determined as a function of lipid chain length, unsaturation and temperature encompassing the range of the lipid phase transition. Model membranes of egg phosphatidylcholine (PC), dimyristoyl (DMPC)-, dipalmitoyl (DPPC)-, distearoyl (DSPC)- and dioleoyl (DOPC)-phosphatidylcholines were studied. Kp values of the drugs are different in the various membranes under study and depend on temperature, aliphatic carbon chain-length and on the presence of unsaturation in the aliphatic lipid chain. First-order transition of membrane lipids from the gel to the liquid crystalline state is accompanied by a sharp increase of the partition coefficient of pimozide and fluspirilene in DMPC, DPPC and DSPC bilayers. For domperidone, Kp values are maximal within the mid-point of phase transition of DMPC and DPPC, while for DSPC Kp values increase progressively with increasing temperature. Haloperidol Kp values display a maximum at the mid-point of phase transition of DMPC, while a progressive increase of Kp is observed in DPPC and DSPC. The four drugs are easily accommodated in bilayers of short aliphatic chain lipids (DMPC), the partition coefficients being 17,137 for pimozide, 18,700 for fluspirilene, 686 for domperidone and 722 for haloperidol, at temperatures 10 degrees C below the mid-point of the lipid phase transition. Except for haloperidol, the partition of the drugs in DOPC (18:1) is higher than that in DSPC (18:0) bilayers at a temperature above the phase transition temperature of both lipids.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 1,2-Dipalmitoylphosphatidylcholine; Dimyristoylphosphatidylcholine; Domperidone; Dopamine Antagonists; Fluspirilene; Haloperidol; Lipid Bilayers; Phosphatidylcholines; Pimozide

Four fluorescent diphenylhexatriene derivatives were considered as membrane probes, namely two ammonium compounds, 3-(diphenylhexatrienyl)propyltrimethylammonium (TMAP-DPH) and 22-(diphenylhexatrienyl)docosyltrimethylammonium (LcTMA-DPH), and two phospholipids, 1-palmitoyl-2-[3-(diphenylhexatrienyl)propanoyl]-sn-glyc ero-3-phosphocholine (DPHpPC) and 1-palmitoyl-2-[21-(diphenylhexatrienyl)henicosanoyl]-sn-phos phocholine (LcDPHpPC). For each pair, the molecules differ by the length of the polymethylenic spacer between the fluorescent moiety and the polar head, so one pair comprises two short chain molecules (C3 spacer) and the other two long chain molecules (C21 or C22 spacer). The partitioning of these probes between gel and liquid crystalline phases of multilamellar vesicles with binary composition (DEPC and DSPC) was measured by a method based on fluorescence anisotropy. The partitioning was shown to depend strongly on the length of the spacer. Short chain probes preferably partition into fluid phases (Kf/s = 1.7 +/- 0.3 for TMAP-DPH; 2.6 +/- 0.11 for DPHpPC), whereas long chain probes show a strong preferential partitioning for gel phases of the vesicles (Kf/s = 0.12 +/- 0.06 for LcTMA-DPH; 0.22 +/- 0.11 for LcDPHpPC). This strong partitioning may be explained by the interdigitation of the long polymethylenic chains across the mid-point of the lipid bilayer (I.E. Mehlhorn et al. (1988) Biochim. Biophys. Acta 939, 151-159), which is enhanced by the better packing provided by a gel phase.

Topics: Diphenylhexatriene; Fluorescence Polarization; Fluorescent Dyes; Liposomes; Membrane Fluidity; Models, Biological; Molecular Structure; Phosphatidylcholines; Structure-Activity Relationship