Curvature effect on the structure of phospholipid bilayers

High-resolution small-angle X-ray scattering (SAXS), complemented by small-angle neutron scattering (SANS) and dynamic light scattering (DLS) experiments, was used to study the effect of curvature on the bilayer structure of dioleoyl-phosphatidylcholine (DOPC) and dioleoyl-phosphatidylserine (DOPS) unilamellar vesicles (ULVs). Bilayer curvature, as a result of finite vesicle size, was varied as a function of vesicle radius and determined by DLS and SANS measurements. Unilamellarity of large DOPC ULVs was achieved by the addition of small amounts (up to 4 mol %) of the charged lipid, DOPS. A comparison of SANS data over the range of 0.02 < q <0.2 A-1 indicated no change in the overall bilayer thickness as a function of ULV diameter (620 to 1840 A). SANS data were corroborated by high-resolution (0.06 < q <0.6 A-1) SAXS data for the same diameter ULVs and data obtained from planar samples of aligned bilayers. Both the inner and outer leaflets of the bilayer were found to be indistinguishable. This observation agrees well with simple geometric models describing the effect of vesicle curvature. However, 1220-A-diameter pure DOPS ULVs form asymmetric bilayers whose structure can most likely be rationalized in terms of geometrical constraints coupled with electrostatic interactions, rather than curvature alone.     

Two-component membrane material properties and domain formation from dissipative particle dynamics

The material parameters (area stretch modulus and bending rigidity) of two-component amphiphilic membranes are determined from dissipative particle dynamics simulations. The preferred area per molecule for each species is varied so as to produce homogeneous mixtures or nonhomogeneous mixtures that form domains. If the latter mixtures are composed of amphiphiles with the same tail length, but different preferred areas per molecule, their material parameters increase monotonically as a function of composition. By contrast, mixtures of amphiphiles that differ in both tail length and preferred area per molecule form both homogeneous and nonhomogeneous mixtures that both exhibit smaller values of their material properties compared to the corresponding pure systems. When the same nonhomogeneous mixtures of amphiphiles are assembled into planar membrane patches and vesicles, the resulting domain shapes are different when the bending rigidities of the domains are sufficiently different. Additionally, both bilayer and monolayer domains are observed in vesicles. We conclude that the evolution of the domain shapes is influenced by the high curvature of the vesicles in the simulation, a result that may be relevant for biological vesicle membranes.     

Influence of spontaneous curvature and microtubules on the conformations of lipid vesicles

The conformations of vesicles deformed by microtubules are studied within the framework of the curvature energy. The phenomenon in which the destruction of a microtubule is followed by the formation of peristaltic shapes on a protrusion created by the microtubule is investigated. The influence of the spontaneous curvature on the conformations of vesicles is examined, and the results are compared to existing experiments. The elastic properties of a vesicle deformed by the microtubule are studied.     

Properties of giant vesicles

We have discussed the specific properties of giant vesicles and their use as model systems for fluid interfaces and biomembranes. Recent advances in giant vesicle research include systematic measurements of visco-elastic parameters as a function of membrane composition, experiments with water-soluble amphiphiles and active membranes, as well as the investigation of hydrodynamic interactions. Notably, it has finally been possible to measure spontaneous curvatures of membranes for a variety of different systems. Experimentally, spontaneous curvature has been a somewhat obscure quantity so far. Furthermore, vesicles have been used to construct bioelectronic devices and new classes of vesicles made of polymers were introduced.     

Aqueous-mixed ionic liquid system: phase transitions and synthesis of gold nanocrystals

Micelle-vesicle-micelle (MVM) transitions are observed in the aqueous-mixed ionic liquid (1-butyl-3-methylimidazolium octyl sulfate and 3-methyl-1-octylimidazolium chloride) system. The surface activity of mixed ILs, phase behavior, and solution structures in the system have been thoroughly characterized using conductometry, tensiometry, fluorimetry, dynamic light scattering (DLS), viscometry, turbidity, atomic force microscopy (AFM), transmission electron microscopy (TEM), and (1)H NMR techniques. Synergetic interactions between the two ILs in monolayers at the air/water interface and in micelles/vesicles have been determined using the regular solution approach, and the origins of spontaneous vesicle formation in this novel system are discussed. Using a photoreduction method, the formation of stable gold nanoparticles (GNPs) and microscale nanosheets of different shapes and sizes in the micellar and vesicle solutions has been reported. The studies show the potential of a mixed IL system in constructing stable micelles/supramolecular assemblies, such as bilayer vesicles, which are effective in the preparation of the desired nanomaterials.     

Lipid-based mechanisms for vesicle fission

Shape transformations and topological changes of lipid vesicles, such as fusion, budding, and fission, have important chemical physical and biological significance. In this paper, we study the fission process of lipid vesicles. Two distinct routes are considered that are both based on an asymmetry of the lipid distribution within the membrane. This asymmetry consists of a nonuniform distribution of two types of lipids. In the first mechanism, the two types of lipids are equally distributed over both leaflets of the membrane. Phase separation of the lipids within both leaflets, however, results in the formation of rafts, which form buds that can split off. In the second mechanism, the asymmetry consists of a difference in composition between the two monolayers of the membrane. This difference in composition yields a spontaneous curvature, reshaping the vesicle into a dumbbell such that it can split. Both pathways are studied with molecular dynamics simulations using a coarse-grained lipid model. For each of the pathways, the conditions required to obtain complete fission are investigated, and it is shown that for the second pathway, much smaller differences between the lipids are needed to obtain fission than for the first pathway. Furthermore, the lipid composition of the resulting split vesicles is shown to be completely different for both pathways, and essential differences between the fission pathway and the pathway of the inverse process, i.e., fusion, are shown to exist.     

Size-dependent properties of small unilamellar vesicles formed by model lipids

The size-dependent behavior of small unilamellar vesicles is explored by dissipative particle dynamics, including the membrane characteristics and mechanical properties. The spontaneously formed vesicles are in the metastable state and the vesicle size is controlled by the concentration of model lipids. As the vesicle size decreases, the bilayer gets thinner and the area density of heads declines. Nonetheless, the area density in the inner leaflet is higher than that in the outer. The packing parameters are calculated for both leaflets. The result indicates that the shape of lipid in the outer leaflet is like a truncated cone but that in the inner leaflet resembles an inverted truncated cone. Based on a local order parameter, our simulations indication that the orientation order of lipid molecules decreases as the size of the vesicle reduces and this fact reveals that the bilayer becoming thinner for smaller vesicle is mainly attributed to the orientation disorder of the lipids. The membrane tension can be obtained through the Young-Laplace equation. The tension is found to grow with reducing vesicle size. Therefore, small vesicles are less stable against fusion. Using the inflation method, the area stretching and bending moduli can be determined and those moduli are found to grow with reducing size. Nonetheless, a general equation with a single numerical constant can relate bending modulus, area stretching modulus, and bilayer thickness irrespective of the vesicle size. Finally, a simple metastable model is proposed to explain the size-dependent behavior of bilayer thickness, orientation, and tension.     

Interleaflet interaction and asymmetry in phase separated lipid bilayers: molecular dynamics simulations

In order to investigate experimentally inaccessible, molecular-level detail regarding interleaflet interaction in membranes, we have run an extensive series of coarse-grained molecular dynamics simulations of phase separated lipid bilayers. The simulations are motivated by differences in lipid and cholesterol composition in the inner and outer leaflets of biological membranes. Over the past several years, this phenomenon has inspired a series of experiments in model membrane systems which have explored the effects of lipid compositional asymmetry in the two leaflets. The simulations are directed at understanding one potential consequence of compositional asymmetry, that being regions of bilayers where liquid-ordered (L(o)) domains in one leaflet are opposite liquid-disordered (L(d)) domains in the other leaflet (phase asymmetry). The simulated bilayers are of two sorts: 1) Compositionally symmetric leaflets where each of the two leaflets contains an identical, phase separated (L(o)/L(d)) mixture of cholesterol, saturated and unsaturated phospholipid; and 2) Compositionally asymmetric leaflets, where one leaflet contains a phase separated (L(o)/L(d)) mixture while the other contains only unsaturated lipid, which on its own would be in the L(d) phase. In addition, we have run simulations where the lengths of the saturated lipid chains as well as the mole ratios of the three lipid components are varied. Collectively, we report on three types of interleaflet coupling within a bilayer. First, we show the effects of compositional asymmetry on acyl chain tilt and order, lipid rotational dynamics, and lateral diffusion in regions of leaflets that are opposite L(o) domains. Second, we show substantial effects of compositional asymmetry on local bilayer curvature, with the conclusion that phase separated leaflets resist curvature, while inducing large degrees of curvature in an opposing L(d) leaflet. Finally, in compositionally symmetric, phase separated bilayers, we find phase asymmetry (domain antiregistration) between the two leaflets occurs as a consequence of mismatched acyl chain-lengths in the saturated and unsaturated lipids.     

Composition and asymmetry in supported membranes formed by vesicle fusion

The structure and formation of supported membranes at silica surfaces by vesicle fusion was investigated by neutron reflectivity and quartz crystal microbalance (QCM-D) measurements. The structure of equimolar phospholipid mixtures of DLPC-DPPC, DMPC-DPPC, and DOPC-DPPC depends intricately on the vesicle deposition conditions. The supported bilayer membranes exhibit varying degrees of compositional asymmetry between the monolayer leaflets, which can be modified by the deposition temperature as well as the salt concentration of the vesicle solution. The total lipid composition of the supported bilayers differs from the composition of the vesicles in solution, and the monolayer proximal to the silica surface is always enriched in DPPC compared to the distal monolayer. The results, which show unambiguougsly that some exchange and rearrangement of lipids occur during vesicle deposition, can be rationalized by considering the effects of salt screening and temperature on the rates of lipid exchange, rearrangement, and vesicle adsorption, but there is also an intricate dependence on the lipid-lipid interactions. Thus, although both symmetric and asymmetric supported bilayers can be prepared from vesicles, the optimal conditions are sensitive to the lipid composition of the system.     

Coupling of bending and stretching deformations in vesicle membranes

Biomimetic membranes are fluid and can undergo two different elastic deformations, bending and stretching. The bending of a membrane is primarily governed by two elastic parameters: its spontaneous (or preferred) curvature m and its bending rigidity κ. These two parameters define an intrinsic tension scale, the spontaneous tension 2 κm². Membrane stretching and compression, on the other hand, are determined by the mechanical tension acting within the membrane. For vesicle membranes, the two elastic deformations are coupled via the enclosed vesicle volume even in the absence of mechanical forces as shown here by minimizing the combined bending and stretching energy with respect to membrane area for fixed vesicle volume. As a consequence, the mechanical tension within a vesicle membrane depends on the spontaneous curvature and on the bending rigidity. This interdependence, which is difficult to grasp intuitively, is then illustrated for a variety of simple vesicle shapes. Depending on the vesicle morphology, the magnitude of the mechanical tension can be comparable to or can be much smaller than the spontaneous tension.     

Fluorescent and highly stable unimodal DMPC based unilamellar vesicles formed by spontaneous curvature

The formation of uniform and highly stable unilamellar vesicles (ULVs) and the theory behind it are ongoing tasks within the vesicle community. Herein, we report the formation of highly stable, fluorescent, and unimodal 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) based ULVs with an average size of ~100 nm, as determined by cryogenic transmission electron microscopy (cryo-TEM) and dynamic light scattering (DLS). The ULVs are formed by mixing a two-component powder mixture or mixed lipid film of DMPC and 5 mol % of a novel amphiphilic carbenium salt, sodium 2-didecylamino-6,10-bis(N-methyltaruino)-4,8,12-trioxatriangulenium (Na-DSA) in aqueous solution when subjected to shaking. We propose that the high stability and the unimodal size distribution of the 5% DSA ULVs confirmed by DLS studies are a product of spontaneous curvature. UV-vis absorption/emission studies reveal that the structure of DSA promotes a strong interaction between the DMPC and the DSA to take place due to the complementary charge distribution of the DSA and DMPC head groups. The strong interaction may introduce an asymmetric amphiphile composition in the inner and outer leaflet of the bilayer which drives the spontaneous curvature.     

Dynamics of spontaneous vesicle formation in fluorocarbon and hydrocarbon surfactant mixtures

The spontaneous self-assembly of unilamellar vesicles was investigated by means of time-resolved synchrotron small-angle X-ray scattering. The self-assembly process was initiated by rapid mixing of anionic surfactant micelles with either zwitterionic or cationic surfactant micelles in equimolar ratio using a stopped-flow device. For the zwitteranionic systems, transient disklike mixed micelles are observed as structural intermediates prior to the onset of vesiculation. These disklike micelles display an exponential growth law, and above a critical size they close to form unilamellar vesicles. In the catanionic system, the earliest observable structures within the mixing time of 4 ms are unilamellar vesicles. Nevertheless, in both systems a narrow distribution of the vesicle size was observed at the initial stages of their formation. The subsequent evolution of the vesicle size distribution depends on the subtle differences in the bilayer composition and properties.     

Influence of nanotopography on phospholipid bilayer formation on silicon dioxide

We have investigated the effect of well-defined nanoscale topography on the 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid vesicle adsorption and supported phospholipid bilayer (SPB) formation on SiO2 surfaces using a quartz crystal microbalance with dissipation monitoring (QCM-D) and atomic force microscopy (AFM). Unilamellar lipid vesicles with two different sizes, 30 and 100 nm, were adsorbed on pitted surfaces with two different pit diameters, 110 and 190 nm, as produced by colloidal lithography, and the behavior was compared to results obtained on flat surfaces. In all cases, complete bilayer formation was observed after a critical coverage of adsorbed vesicles had been reached. However, the kinetics of the vesicle-to-bilayer transformation, including the critical coverage, was significantly altered by surface topography for both vesicle sizes. Surface topography hampered the overall bilayer formation kinetics for the smaller vesicles, but promoted SPB formation for the larger vesicles. Depending on vesicle size, we propose two modifications of the precursor-mediated vesicle-to-bilayer transformation mechanism used to describe supported lipid bilayer formation on the corresponding flat surface. Our results may have important implications for various lipid-membrane-based applications using rough or topographically structured surfaces.     

Shape fluctuations of large unilamellar lipid vesicles observed by laser light scattering: influence of the small-scale structure

In the present paper, we apply the dynamic laser light scattering technique to investigate the dependence of the characteristic times of thermally induced shape fluctuation of large unilamellar vesicles (LUVs) on bilayer composition. After addressing single-component LUVs made of two common phospholipids, dimyristoylphosphatidylcholine (DMPC) and dipalmitoylphosphatidylcholine (DPPC), we investigate the changes in vesicle shape fluctuation times due to the presence of cholesterol and gangliosides (GM1), added in small amounts. The experimental results show that the addition of a second component, even in small amount, to DMPC vesicles induces a change in membrane fluctuation times. Moreover, in the case of ganglioside addition, also the disposition of GM1 within the bilayer is of importance. Quite unexpectedly, the symmetric or asymmetric disposition of GM1 has opposite effects on bilayer dynamics, the first resulting in a "hardening" and the second in a "softening" of the membrane. Those results support that the small-scale structure of the bilayer is important in determining the overall dynamics of the vesicle. They also suggest that the physiological disposition of GM1 in the outer leaflet of real cells has a significative result in mechanical terms, positively affecting the dynamics of the membrane.     

Zero spontaneous curvature and its effects on lamellar phase morphology and vesicle size distributions

Equimolar mixtures of dodecyltrimethylammonium chloride (DTAC) and sodium octyl sulfonate (SOSo) show a vesicle phase at >99 wt % water and a single, fluid lamellar phase for water fractions below 80 wt %. This combination is consistent with the bilayer bending elasticity kappa approximately k(B)T and zero bilayer spontaneous curvature. Caillé line shape analysis of the small-angle X-ray scattering from the lamellar phase shows that the effective kappa depends on the lamellar d spacing consistent with a logarithmic renormalization of kappa, with kappa(o) = (0.8 +/- 0.1)k(B)T. The vesicle size distribution determined by cryogenic transmission electron microscopy is well fit by models with zero spontaneous curvature to give (kappa + (kappa/2)) = (1.7 +/- 0.1)k(B)T, resulting in kappa = (1.8 +/- 0.2)k(B)T. The positive value of kappa and the lack of spontaneous curvature act to eliminate the spherulite defects found in the lamellar gel phases found in other catanionic mixtures. Current theories of spontaneous bilayer curvature require an excess of one or more components on opposite sides of the bilayer; the absence of such an excess at equimolar surfactant ratios explains the zero spontaneous curvature.     

Shape transformation of lipid vesicles induced by diffusing macromolecules

The attachment of macromolecules to the surface of a lipid vesicle may cause its deformations such as budding or creation of cylindrical protrusions. Diffusion of the macromolecules in the membranes may cause its shape transformations. The process of shrinking the protrusions due to diffusion of the macromolecules is investigated. It is assumed that macromolecules modify locally the spontaneous curvature and bending rigidity of the lipid membrane. Both spontaneous curvature and bending rigidities depend on the concentration of membrane components. It has been shown that cylindrical protrusions are created when the macromolecules which induce large spontaneous curvature are accumulated at a piece of the vesicle surface. It has been observed that here the elastic constants influence very little the evolution of the vesicle shape caused by diffusing macromolecules and the most important is the value the spontaneous curvature imposed by the macromolecules.     

Internally catalyzed separation of adhered lipid membranes

When a giant vesicle composed of POPC (rendered anionic with 5 mol % POPG) touches a giant POPC vesicle (rendered cationic with 5 mol % of DDAB), the two vesicles adhere strongly. When, however, low levels (0.1-2 mol %) of a perylene-substituted lipid are incorporated in to the bilayer, the vesicles separate at a rate that depends on the additive concentration. The vesicles that drift apart lose charge, indicating that the anionic and cationic components of the vesicles have interchanged upon contact. Presumably, the large perylene disrupts bilayer packing to allow the intervesicular exchange, and subsequent charge neutralization, to occur with up to 104 rate increases. It is possible that adhered living cells release one another by, similarly, producing low levels of a membrane-bound lipid or protein that induces so-called "kiss-and-run" vesicle events by promoting the release of adhesive elements.     

Molecular dynamics simulation of the spontaneous formation of a small DPPC vesicle in water in atomistic detail

Molecular dynamics simulations have been used to study the spontaneous aggregation of a concentrated solution of dipalmitoylphosphatidylcholine (DPPC) molecules in water into a small vesicle. The molecules were represented in atomistic detail. Starting from a DPPC solution in water, an oblong vesicle with a long axis of 15 nm and short axes of 10 nm was formed spontaneously. After 90 ns of simulation, the vesicle contained a number of water pores. Water pores were shown to facilitate exchange of lipids between inner and outer leaflets. Lipid tails were shown to be less ordered in the inner leaflet of the vesicle, as compared to those in the outer leaflet of the vesicle and an equilibrated lamellar bilayer.     

A Comparison of the Molecular Dynamics of Fluorescent Probes in Curved Lipid Vesicles and Planar Multibilayers

The dynamic behaviour of the probe molecules DPH and TMA-DPH embedded in small unilamellar vesicles and planar multibilayers of POPC has been studied by time-resolved fluorescence depolarization techniques. The molecular dynamics of the probe molecules was analysed in terms of the rotational diffusion model. It is found that analysis of the time-dependent fluorescence anisotropy from the vesicle system yields two distinct, though statistically equivalent solutions. On the other hand the measurements on planar multibilayers can be interpreted unequivocally. It is shown that the order parameters of the probe molecules are higher in the multibilayers than in the vesicles. A reconstruction of the orientational distribution function reveals that the TMA-DPH molecules have a more pronounced tendency to lie with their long axes parallel to the bilayer surface in the curved vesicles than in the planar multibilayers. An intriguing finding is that the reorientational motion of the probes is considerably slower in the multibilayer samples than in the vesicles. These differences are attributed to the curvature and higher hydration of the bilayers in the vesicle systems.     

KINETICS OF CHLOROPHYLL PHOTOSENSITIZED ELECTRON TRANSFER REACTIONS IN PHOspHOLIPID BILAYER VESICLES

Abstract The effects of electrostatic surface charge and valinomycin addition in the presence of K* on the kinetics and the inside-outside asymmetry properties of light-induced electron transfer reactions between chlorophyll triplet state and benzoquinone, ferricyanide and methyl viologen in large unilamellar vesicles have been investigated using laser flash photolysis. Modifying the surface charge of the bilayers by incorporating charged surfactants or decreasing the ionic strength of the suspending medium caused large changes in the dynamics of the electron transfer reactions, which could be interpreted in terms of electrostatic interactions between reactants, products and membrane components, and the existence of a spontaneous transmembrane electrical potential corresponding to an excess of negative charge at the outer surface of the vesicle bilayer. The presence of valinomycin had more specific effects on these reactions, which were consistent with an electrostatic influence of the presence of the positively-charged K⁺-valinomycin complex within the bilayer on the dynamics of only those triplet quenching and radical formation and decay processes which occur in this region of the vesicle structure.