Sample records for bilayer fluid membrane

  1. Efficient tunable generic model for self-assembling fluid bilayer membranes (United States)

    Deserno, Markus


    We present a new model for the simulation of generic lipid bilayers in the mesoscopic regime (between a few nanometers and many tens of nanometers), which is very robust, versatile, and extremely efficient, since it avoids the need for an embedding solvent. Based entirely on simple pair potentials, it features a wide region of unassisted self assembly into fluid bilayers without the need for careful parameter tuning. The resulting membranes display the correct continuum elastic behavior with bending constants in the experimentally relevant range. It can be readily used to study events like bilayer fusion, bilayer melting, lipid mixtures, rafts, and protein-bilayer interactions.

  2. Fluid lipid bilayers: Intermonolayer coupling and its thermodynamic manifestations

    DEFF Research Database (Denmark)

    Hansen, Per Lyngs; Miao, Ling; Ipsen, John Hjorth


    A fluid membrane of lipid bilayer consists of two individual molecular monolayers physically opposed to each other. This unique molecular architecture naturally necessitates the need to treat a lipid-bilayer membrane as one entity of two coupled two-dimensional systems (monolayers), each of which...

  3. Planar bilayer membranes from photoactivable phospholipids. (United States)

    Borle, F; Sänger, M; Sigrist, H


    Planar bilayer membranes formed from photoactivable phospholipids have been characterized by low frequency voltametry. Cyclic voltametric measurements were applied for simultaneous registration of planar membrane conductivity and capacitance. The procedure has been utilized to characterize the formation and stability of planar bilayer membranes. Bilayer membranes were formed from N'-(1,2-dimyristoyl-sn-glycero-3-phosphoethyl)-N-((m-3- trifluoromethyldiazirine)phenyl)thiourea (C14-PED), a head-group photosensitive phospholipid. In situ photoactivation of C14-PED at wavelengths greater than or equal to 320 nm altered neither the mean conductivity nor the capacitance of the bilayer. Ionophore (valinomycin) and ion channel (gramicidin) activities were not impaired upon photoactivation. In contrast, bilayer membranes formed from 1,2-bis(hexadeca-2,4-dienoyl)-sn- glycero-3-phosphocholine (C16-DENPC) revealed short life times. In situ photopolymerization of the diene fatty acids significantly increased the membrane conductivity or led to membrane rupture.

  4. PI3 kinase enzymology on fluid lipid bilayers. (United States)

    Dutta, Debjit; Pulsipher, Abigail; Luo, Wei; Yousaf, Muhammad N


    We report the use of fluid lipid bilayer membrane as a model platform to study the influence of the bilayer microenvironment and composition on the enzymology in membrane. As a model system we determined the enzyme kinetics on membranes for the transformation of bilayers containing phosphoinositol(4,5)-bisphosphate (PI(4,5)P2) to phosphoinositol(3,4,5)-trisphosphate (PI(3,4,5)P3) by the enzyme phosphoinositol-3-kinase (PI3K) using radiolabeled ATP. The activity of the enzyme was monitored as a function of the radioactivity incorporated within the bilayer. The transformation of PI(4,5)P2 to PI(3,4,5)P3 was determined using a mass strip assay. The fluidity of the bilayer was confirmed by Fluorescence Recovery After Photobleaching (FRAP) experiments. Kinetic simulations were performed based on Langmuir adsorption and Michaelis-Menton kinetics equations to generate the rate constants for the enzymatic reaction. The effect of cholesterol on the enzyme kinetics was studied by doping the bilayer with 1% cholesterol. This leads to significant reduction in reaction rate due to change in membrane microenvironment. This strategy provides a method to study the enzymology of various kinases and phosphatases occurring at the membrane and also how these reactions are affected by the membrane composition and surface microenvironment.

  5. Equilibrium Configurations of Lipid Bilayer Membranes and Carbon Nanostructures

    Institute of Scientific and Technical Information of China (English)

    Iva(i)lo M.Mladenov; Peter A.Djondjorov; Mariana Ts.Hadzhilazova; Vassil M.Vassilev


    The present article concerns the continuum modelling of the mechanical behaviour and equilibrium shapes of two types of nano-scale objects:fluid lipid bilayer membranes and carbon nanostructures.A unified continuum model is used to handle four different case studies.Two of them consist in representing in analytic form cylindrical and axisymmetric equilibrium configurations of single-wall carbon nanotubes and fluid lipid bilayer membranes subjected to uniform hydrostatic pressure.The third one is concerned with determination of possible shapes of junctions between a single-wall carbon nanotube and a fiat graphene sheet or another single-wall carbon nanotube.The last one deals with the mechanical behaviour of closed fluid lipid bilayer membranes (vesicles) adhering onto a fiat homogeneous rigid substrate subjected to micro-injection and uniform hydrostatic pressure.

  6. Photon correlation spectroscopy of bilayer lipid membranes. (United States)

    Crilly, J F; Earnshaw, J C


    Light scattering by thermal fluctuations on simple monoglyceride bilayer membranes has been used to investigate the viscoelastic properties of these structures. Spectroscopic analysis of these fluctuations (capillary waves) permits the nonperturbative measurement of the interfacial tension and a shear interfacial viscosity acting normal to the membrane plane. The methods were established by studies of solvent and nonsolvent bilayers of glycerol monooleate (GMO). Changes in the tension of GMO/n-decane membranes induced by altering the composition of the parent solution were detected and quantified. In a test of the reliability of the technique controlled variations of the viscosity of the aqueous bathing solution were accurately monitored. The technique was applied to solvent-free bilayers formed from dispersions of GMO in squalane. The lower tensions observed attested to the comparative absence of solvent in such bilayers. In contrast to the solvent case, the solvent-free membranes exhibited a significant transverse shear viscosity, indicative of the enhanced intermolecular interactions within the bilayer.

  7. Biotechnology Applications of Tethered Lipid Bilayer Membranes

    Directory of Open Access Journals (Sweden)

    Joshua A. Jackman


    Full Text Available The importance of cell membranes in biological systems has prompted the development of model membrane platforms that recapitulate fundamental aspects of membrane biology, especially the lipid bilayer environment. Tethered lipid bilayers represent one of the most promising classes of model membranes and are based on the immobilization of a planar lipid bilayer on a solid support that enables characterization by a wide range of surface-sensitive analytical techniques. Moreover, as the result of molecular engineering inspired by biology, tethered bilayers are increasingly able to mimic fundamental properties of natural cell membranes, including fluidity, electrical sealing and hosting transmembrane proteins. At the same time, new methods have been employed to improve the durability of tethered bilayers, with shelf-lives now reaching the order of weeks and months. Taken together, the capabilities of tethered lipid bilayers have opened the door to biotechnology applications in healthcare, environmental monitoring and energy storage. In this review, several examples of such applications are presented. Beyond the particulars of each example, the focus of this review is on the emerging design and characterization strategies that made these applications possible. By drawing connections between these strategies and promising research results, future opportunities for tethered lipid bilayers within the biotechnology field are discussed.

  8. Bilayer-thickness-mediated interactions between integral membrane proteins

    CERN Document Server

    Kahraman, Osman; Klug, William S; Haselwandter, Christoph A


    Hydrophobic thickness mismatch between integral membrane proteins and the surrounding lipid bilayer can produce lipid bilayer thickness deformations. Experiment and theory have shown that protein-induced lipid bilayer thickness deformations can yield energetically favorable bilayer-mediated interactions between integral membrane proteins, and large-scale organization of integral membrane proteins into protein clusters in cell membranes. Within the continuum elasticity theory of membranes, the energy cost of protein-induced bilayer thickness deformations can be captured by considering compression and expansion of the bilayer hydrophobic core, membrane tension, and bilayer bending, resulting in biharmonic equilibrium equations describing the shape of lipid bilayers for a given set of bilayer-protein boundary conditions. Here we develop a combined analytic and numerical methodology for the solution of the equilibrium elastic equations associated with protein-induced lipid bilayer deformations. Our methodology al...

  9. Fractional polymerization of a suspended planar bilayer creates a fluid, highly stable membrane for ion channel recordings. (United States)

    Heitz, Benjamin A; Jones, Ian W; Hall, Henry K; Aspinwall, Craig A; Saavedra, S Scott


    Suspended planar lipid membranes (or black lipid membranes (BLMs)) are widely used for studying reconstituted ion channels, although they lack the chemical and mechanical stability needed for incorporation into high-throughput biosensors and biochips. Lipid polymerization enhances BLM stability but is incompatible with ion channel function when membrane fluidity is required. Here, we demonstrate the preparation of a highly stable BLM that retains significant fluidity by using a mixture of polymerizable and nonpolymerizable phospholipids. Alamethicin, a voltage-gated peptide channel for which membrane fluidity is required for activity, was reconstituted into mixed BLMs prepared using bis-dienoyl phosphatidylcholine (bis-DenPC) and diphytanoyl phosphatidylcholine (DPhPC). Polymerization yielded BLMs that retain the fluidity required for alamethicin activity yet are stable for several days as compared to a few hours prior to polymerization. Thus, these polymerized, binary composition BLMs feature both fluidity and long-term stability.

  10. Design of Asymmetric Peptide Bilayer Membranes. (United States)

    Li, Sha; Mehta, Anil K; Sidorov, Anton N; Orlando, Thomas M; Jiang, Zhigang; Anthony, Neil R; Lynn, David G


    Energetic insights emerging from the structural characterization of peptide cross-β assemblies have enabled the design and construction of robust asymmetric bilayer peptide membranes. Two peptides differing only in their N-terminal residue, phosphotyrosine vs lysine, coassemble as stacks of antiparallel β-sheets with precisely patterned charged lattices stabilizing the bilayer leaflet interface. Either homogeneous or mixed leaflet composition is possible, and both create nanotubes with dense negative external and positive internal solvent exposed surfaces. Cross-seeding peptide solutions with a preassembled peptide nanotube seed leads to domains of different leaflet architecture within single nanotubes. Architectural control over these cross-β assemblies, both across the bilayer membrane and along the nanotube length, provides access to highly ordered asymmetric membranes for the further construction of functional mesoscale assemblies.

  11. Engineering Lipid Bilayer Membranes for Protein Studies

    Directory of Open Access Journals (Sweden)

    Muhammad Shuja Khan


    Full Text Available Lipid membranes regulate the flow of nutrients and communication signaling between cells and protect the sub-cellular structures. Recent attempts to fabricate artificial systems using nanostructures that mimic the physiological properties of natural lipid bilayer membranes (LBM fused with transmembrane proteins have helped demonstrate the importance of temperature, pH, ionic strength, adsorption behavior, conformational reorientation and surface density in cellular membranes which all affect the incorporation of proteins on solid surfaces. Much of this work is performed on artificial templates made of polymer sponges or porous materials based on alumina, mica, and porous silicon (PSi surfaces. For example, porous silicon materials have high biocompatibility, biodegradability, and photoluminescence, which allow them to be used both as a support structure for lipid bilayers or a template to measure the electrochemical functionality of living cells grown over the surface as in vivo. The variety of these media, coupled with the complex physiological conditions present in living systems, warrant a summary and prospectus detailing which artificial systems provide the most promise for different biological conditions. This study summarizes the use of electrochemical impedance spectroscopy (EIS data on artificial biological membranes that are closely matched with previously published biological systems using both black lipid membrane and patch clamp techniques.

  12. Bilayer-thickness-mediated interactions between integral membrane proteins. (United States)

    Kahraman, Osman; Koch, Peter D; Klug, William S; Haselwandter, Christoph A


    Hydrophobic thickness mismatch between integral membrane proteins and the surrounding lipid bilayer can produce lipid bilayer thickness deformations. Experiment and theory have shown that protein-induced lipid bilayer thickness deformations can yield energetically favorable bilayer-mediated interactions between integral membrane proteins, and large-scale organization of integral membrane proteins into protein clusters in cell membranes. Within the continuum elasticity theory of membranes, the energy cost of protein-induced bilayer thickness deformations can be captured by considering compression and expansion of the bilayer hydrophobic core, membrane tension, and bilayer bending, resulting in biharmonic equilibrium equations describing the shape of lipid bilayers for a given set of bilayer-protein boundary conditions. Here we develop a combined analytic and numerical methodology for the solution of the equilibrium elastic equations associated with protein-induced lipid bilayer deformations. Our methodology allows accurate prediction of thickness-mediated protein interactions for arbitrary protein symmetries at arbitrary protein separations and relative orientations. We provide exact analytic solutions for cylindrical integral membrane proteins with constant and varying hydrophobic thickness, and develop perturbative analytic solutions for noncylindrical protein shapes. We complement these analytic solutions, and assess their accuracy, by developing both finite element and finite difference numerical solution schemes. We provide error estimates of our numerical solution schemes and systematically assess their convergence properties. Taken together, the work presented here puts into place an analytic and numerical framework which allows calculation of bilayer-mediated elastic interactions between integral membrane proteins for the complicated protein shapes suggested by structural biology and at the small protein separations most relevant for the crowded membrane

  13. Thermotropic and barotropic phase transitions on diacylphosphatidylethanolamine bilayer membranes. (United States)

    Matsuki, Hitoshi; Endo, Shigeru; Sueyoshi, Ryosuke; Goto, Masaki; Tamai, Nobutake; Kaneshina, Shoji


    The bilayer phase transitions of four diacylphosphatidylethanolamines (PEs) with matched saturated acyl chains (Cn=12, 14, 16 and 18) and two PEs with matched unsaturated acyl chains containing a different kind of double bonds were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The temperature-pressure phase diagrams for these PE bilayer membranes were constructed from the obtained phase-transition data. The saturated PE bilayer membranes underwent two different phase transitions related to the liquid crystalline (Lα) phase, the transition from the hydrated crystalline (Lc) phase and the chain melting (gel (Lβ) to Lα) transition, depending on the thermal history. Pressure altered the gel-phase stability of the bilayer membranes of PEs with longer chains at a low pressure. Comparing the thermodynamic quantities of the saturated PE bilayer membranes with those of diacylphosphatidylcholine (PC) bilayer membranes, the PE bilayer membranes showed higher phase-transition temperatures and formed more stable Lc phase, which originates from the strong interaction between polar head groups of PE molecules. On the other hand, the unsaturated PE bilayer membranes underwent the transition from the Lα phase to the inverted hexagonal (HII) phase at a high temperature and this transition showed a small transition enthalpy but high pressure-responsivity. It turned out that the kind of double bonds markedly affects both bilayer-bilayer and bilayer-nonbilayer transitions and the Lα/HII transition is a volume driven transition for the reconstruction of molecular packing. Further, the phase-transition behavior was explained by chemical potential curves of bilayer phases. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Effects of Fatty Acid Inclusion in a DMPC Bilayer Membrane

    DEFF Research Database (Denmark)

    Peters, Günther H.J.; Hansen, Flemming Yssing; Møller, Martin S.


    Free fatty acids in biomembranes have been proposed to be a central component in several cellular control and regulatory mechanisms. To elucidate some fundamental elements underlying this, we have applied molecular dynamics simulations and experimental density measurements to study the molecular...... packing and structure of oleic acid (HOA) and stearic acid (HSA) in fluid bilayers of dimyristoylphosphatidylcholine (DMPC). The experimental data show a small but consistent positive excess volume for fatty acid concentrations below 10 mol %. At higher concentrations the fatty acids mix ideally...... with fluid DMPC. The simulations, which were benchmarked against the densitometric data, revealed interesting differences in the structure and location of the fatty acids depending on their protonation status. Thus, the protonated (uncharged) acid is located rather deeply in the membrane with an average...

  15. Topologically-Mediated Membrane Dynamics in Supported Lipid Bilayers (United States)

    Gilmore, Sean Fitzpatrick


    This thesis is primarily design driven. It describes the development and application of dynamically tunable class of solid-fluid interfaces, which serves as a test-bed configuration for fundamental studies of soft condensed matter in reduced dimension. My specific focus is in developing these interfaces to recapitulate topology-mediated phenomenon in biological lipid membranes. The phenomena that the interfacial topology manifest in diffusional characteristics in model membranes are probed using wide-area epifluorescence microscopy and a semi-quantitative analysis of dynamic recovery following photobleaching. Furthermore, real-time remodeling of the membrane-substrate interface topology is shown to provide fundamental information regarding curvature-dependent molecular sorting and resorting. Specifically our experiments using putative raft composition mixtures confirm the conformation-dependent alignment of liquid-ordered domains and moreover reveal domain-domain interactions for the first time in model bilayers. Ongoing work aimed at delineating these inter-domain interactions in terms of membrane elastic properties is being performed. Future work that includes peptide-driven membrane deformation and sorting, as well large-scale, curvature-driven in vivo sorting of lipids is proposed and discussed.

  16. Mechanism of unassisted ion transport across membrane bilayers (United States)

    Wilson, M. A.; Pohorille, A.


    To establish how charged species move from water to the nonpolar membrane interior and to determine the energetic and structural effects accompanying this process, we performed molecular dynamics simulations of the transport of Na+ and Cl- across a lipid bilayer located between two water lamellae. The total length of molecular dynamics trajectories generated for each ion was 10 ns. Our simulations demonstrate that permeation of ions into the membrane is accompanied by the formation of deep, asymmetric thinning defects in the bilayer, whereby polar lipid head groups and water penetrate the nonpolar membrane interior. Once the ion crosses the midplane of the bilayer the deformation "switches sides"; the initial defect slowly relaxes, and a defect forms in the outgoing side of the bilayer. As a result, the ion remains well solvated during the process; the total number of oxygen atoms from water and lipid head groups in the first solvation shell remains constant. A similar membrane deformation is formed when the ion is instantaneously inserted into the interior of the bilayer. The formation of defects considerably lowers the free energy barrier to transfer of the ion across the bilayer and, consequently, increases the permeabilities of the membrane to ions, compared to the rigid, planar structure, by approximately 14 orders of magnitude. Our results have implications for drug delivery using liposomes and peptide insertion into membranes.

  17. Tethered and Polymer Supported Bilayer Lipid Membranes: Structure and Function

    Directory of Open Access Journals (Sweden)

    Jakob Andersson


    Full Text Available Solid supported bilayer lipid membranes are model systems to mimic natural cell membranes in order to understand structural and functional properties of such systems. The use of a model system allows for the use of a wide variety of analytical tools including atomic force microscopy, impedance spectroscopy, neutron reflectometry, and surface plasmon resonance spectroscopy. Among the large number of different types of model membranes polymer-supported and tethered lipid bilayers have been shown to be versatile and useful systems. Both systems consist of a lipid bilayer, which is de-coupled from an underlying support by a spacer cushion. Both systems will be reviewed, with an emphasis on the effect that the spacer moiety has on the bilayer properties.

  18. Laurdan fluorescence senses mechanical strain in the lipid bilayer membrane. (United States)

    Zhang, Yan-Liang; Frangos, John A; Chachisvilis, Mirianas


    The precise molecular mechanisms by which cells transduce a mechanical stimulus into an intracellular biochemical response have not yet been established. Here, we show for the first time that the fluorescence emission of an environment-sensitive membrane probe Laurdan is modulated by mechanical strain of the lipid bilayer membrane. We have measured fluorescence emission of Laurdan in phospholipid vesicles of 30, 50, and 100 nm diameter to show that osmotically induced membrane tension leads to an increase in polarity (hydration depth) of the phospholipid bilayer interior. Our data indicate that the general polarization of Laurdan emission is linearly dependent on membrane tension. We also show that higher membrane curvature leads to higher hydration levels. We anticipate that the proposed method will facilitate future studies of mechanically induced changes in physical properties of lipid bilayer environment both in vitro and in vivo.

  19. Anthrax toxin-induced rupture of artificial lipid bilayer membranes (United States)

    Nablo, Brian J.; Panchal, Rekha G.; Bavari, Sina; Nguyen, Tam L.; Gussio, Rick; Ribot, Wil; Friedlander, Art; Chabot, Donald; Reiner, Joseph E.; Robertson, Joseph W. F.; Balijepalli, Arvind; Halverson, Kelly M.; Kasianowicz, John J.


    We demonstrate experimentally that anthrax toxin complexes rupture artificial lipid bilayer membranes when isolated from the blood of infected animals. When the solution pH is temporally acidified to mimic that process in endosomes, recombinant anthrax toxin forms an irreversibly bound complex, which also destabilizes membranes. The results suggest an alternative mechanism for the translocation of anthrax toxin into the cytoplasm.

  20. The lipid bilayer membrane and its interactions with additives

    NARCIS (Netherlands)

    Meijer, L.A.


    The aim of this study was to make accurate predictions on the interaction of biologically relevant molecules with lipid bilayer membranes. We emphasised on the partitioning of these molecules between the membrane phase, and the aqueous phase quantified by the partition coefficient. To make

  1. The lipid bilayer membrane and its interactions with additives.

    NARCIS (Netherlands)

    Meijer, L.A.


    The aim of this study was to make accurate predictions on the interaction of biologically relevant molecules with lipid bilayer membranes. We emphasised on the partitioning of these molecules between the membrane phase, and the aqueous phase quantified by the partition coefficient. To make detailed

  2. Motional Coherence in Fluid Phospholipid Membranes

    CERN Document Server

    Rheinstadter, Maikel C; Flenner, Elijah J; Bruening, Beate; Seydel, Tilo; Kosztin, Ioan


    We report a high energy-resolution neutron backscattering study, combined with in-situ diffraction, to investigate slow molecular motions on nanosecond time scales in the fluid phase of phospholipid bilayers of 1,2-dimyristoyl-sn-glycero-3-phoshatidylcholine (DMPC) and DMPC/40% cholesterol (wt/wt). A cooperative structural relaxation process was observed. From the in-plane scattering vector dependence of the relaxation rates in hydrogenated and deuterated samples, combined with results from a 0.1 microsecond long all atom molecular dynamics simulation, it is concluded that correlated dynamics in lipid membranes occurs over several lipid distances, spanning a time interval from pico- to nanoseconds.

  3. Inducing morphological changes in lipid bilayer membranes with microfabricated substrates (United States)

    Liu, Fangjie; Collins, Liam F.; Ashkar, Rana; Heberle, Frederick A.; Srijanto, Bernadeta R.; Collier, C. Patrick


    Lateral organization of lipids and proteins into distinct domains and anchoring to a cytoskeleton are two important strategies employed by biological membranes to carry out many cellular functions. However, these interactions are difficult to emulate with model systems. Here we use the physical architecture of substrates consisting of arrays of micropillars to systematically control the behavior of supported lipid bilayers - an important step in engineering model lipid membrane systems with well-defined functionalities. Competition between attractive interactions of supported lipid bilayers with the underlying substrate versus the energy cost associated with membrane bending at pillar edges can be systematically investigated as functions of pillar height and pitch, chemical functionalization of the microstructured substrate, and the type of unilamellar vesicles used for assembling the supported bilayer. Confocal fluorescent imaging and AFM measurements highlight correlations that exist between topological and mechanical properties of lipid bilayers and lateral lipid mobility in these confined environments. This study provides a baseline for future investigations into lipid domain reorganization on structured solid surfaces and scaffolds for cell growth.

  4. Protein-lipid interactions in bilayer membranes: a lattice model. (United States)

    Pink, D A; Chapman, D


    A lattice model has been developed to study the effects of intrinsic membrane proteins upon the thermodynamic properties of a lipid bilayer membrane. We assume that only nearest-neighbor van der Waals and steric interactions are important and that the polar group interactions can be represented by effective pressure-area terms. Phase diagrams, the temperature T(0), which locates the gel-fluid melting, the transition enthalpy, and correlations were calculated by mean field and cluster approximations. Average lipid chain areas and chain areas when the lipid is in a given protein environment were obtained. Proteins that have a "smooth" homogeneous surface ("cholesterol-like") and those that have inhomogeneous surfaces or that bind lipids specifically were considered. We find that T(0) can vary depending upon the interactions and that another peak can appear upon the shoulder of the main peak which reflects the melting of a eutectic mixture. The transition enthalpy decreases generally, as was found before, but when a second peak appears departures from this behavior reflect aspects of the eutectic mixture. We find that proteins have significant nonzero probabilities for being adjacent to one another so that no unbroken "annulus" of lipid necessarily exists around a protein. If T(0) does not increase much, or decreases, with increasing c, then lipids adjacent to a protein cannot all be all-trans on the time scale (10(-7) sec) of our system. Around a protein the lipid correlation depth is about one lipid layer, and this increases with c. Possible consequences of ignoring changes in polar group interactions due to clustering of proteins are discussed.

  5. Bilayer thickness mismatch controls domain size in biomimetic membranes (United States)

    Heberle, Frederick A.; Petruzielo, Robin S.; Pan, Jianjun; Drazba, Paul; Kučerka, Norbert; Standaert, Robert F.; Feigenson, Gerald W.; Katsara, John


    In order to promote functionality, cells may alter the spatial organization of membrane lipids and proteins, including separation of liquid phases into distinct domains. In model membranes, domain size and morphology depend strongly on composition and temperature, but the physicochemical mechanisms controlling them are poorly understood. Theoretical work suggests a role for interfacial energy at domain boundaries, which may be driven in part by thickness mismatch between a domain and its surrounding bilayer. However, no direct evidence linking thickness mismatch to domain size in free-standing bilayers has been reported. We describe the use of Small Angle Neutron Scattering (SANS) to detect domains in simplified lipid-only models that mimic the composition of plasma membrane. We find that domain size is controlled by the degree of acyl chain unsaturation of low-melting temperature lipids, and that this size transition is correlated to changes in the thickness mismatch between coexisting liquid phases.

  6. Forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces. (United States)

    Zhang, Ying; Wang, Lei; Wang, Xuejing; Qi, Guodong; Han, Xiaojun


    A novel method of forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces is introduced. Polyelectrolyte films were fabricated by the layer-by-layer technique on a silicon oxide surface modified with a 3-aminopropyltriethoxysilane (APTES) monolayer. The surface pK(a) value of the APTES monolayer was determined by cyclic voltammetry to be approximately 5.61, on the basis of which a pH value of 2.0 was chosen for layer-by-layer assembly. Micropatterned polyelectrolyte films were obtained by deep-UV (254 nm) photolysis though a mask. Absorbed fluorescent latex beads were used to visualize the patterned surfaces. Lipid bilayer arrays were fabricated on the micropatterned surfaces by immersing the patterned substrates into a solution containing egg phosphatidylcholine vesicles. Fluorescence recovery after photobleaching studies yielded a lateral diffusion coefficient for probe molecules of 1.31±0.17 μm(2) s(-1) in the bilayer region, and migration of the lipid NBD PE in bilayer lipid membrane arrays was observed in an electric field.

  7. Relaxation dynamics of a compressible bilayer vesicle containing highly viscous fluid. (United States)

    Sachin Krishnan, T V; Okamoto, Ryuichi; Komura, Shigeyuki


    We study the relaxation dynamics of a compressible bilayer vesicle with an asymmetry in the viscosity of the inner and outer fluid medium. First we explore the stability of the vesicle free energy which includes a coupling between the membrane curvature and the local density difference between the two monolayers. Two types of instabilities are identified: a small wavelength instability and a larger wavelength instability. Considering the bulk fluid viscosity and the inter-monolayer friction as the dissipation sources, we next employ Onsager's variational principle to derive the coupled equations both for the membrane and the bulk fluid. The three relaxation modes are coupled to each other due to the bilayer and the spherical structure of the vesicle. Most importantly, a higher fluid viscosity inside the vesicle shifts the crossover mode between the bending and the slipping to a larger value. As the vesicle parameters approach the unstable regions, the relaxation dynamics is dramatically slowed down, and the corresponding mode structure changes significantly. In some limiting cases, our general result reduces to the previously obtained relaxation rates.

  8. Relaxation dynamics of a compressible bilayer vesicle containing highly viscous fluid (United States)

    Sachin Krishnan, T. V.; Okamoto, Ryuichi; Komura, Shigeyuki


    We study the relaxation dynamics of a compressible bilayer vesicle with an asymmetry in the viscosity of the inner and outer fluid medium. First we explore the stability of the vesicle free energy which includes a coupling between the membrane curvature and the local density difference between the two monolayers. Two types of instabilities are identified: a small wavelength instability and a larger wavelength instability. Considering the bulk fluid viscosity and the inter-monolayer friction as the dissipation sources, we next employ Onsager's variational principle to derive the coupled equations both for the membrane and the bulk fluid. The three relaxation modes are coupled to each other due to the bilayer and the spherical structure of the vesicle. Most importantly, a higher fluid viscosity inside the vesicle shifts the crossover mode between the bending and the slipping to a larger value. As the vesicle parameters approach the unstable regions, the relaxation dynamics is dramatically slowed down, and the corresponding mode structure changes significantly. In some limiting cases, our general result reduces to the previously obtained relaxation rates.

  9. Modeling the Elastic Properties of Lipid Bilayer Membranes (United States)

    Barry, Edward; Gibaud, Thomas; Zakhary, Mark; Dogic, Zvonimir


    Model membranes such as lipid bilayers have been indispensable tools for our understanding of the elastic properties of biological membranes. In this talk, I will introduce a colloidal model for membranes and demonstrate that the physical properties of these colloidal membranes are identical to lipid bilayers. The model system is unique in that the constituent molecules are homogenous and non-amphiphilic, yet their self-assembly into membranes and other hierarchical assemblages, such as a lamellar type phases and chiral ribbons, proceeds spontaneously in solution. Owing to the large size of the constituent molecules, individual molecules can be directly visualized and simultaneous observations at the continuum and molecular lengthscales are used to characterize the behavior of model membranes with unprecedented detail. Moreover, once assembled in solution, molecular interactions can be controlled in situ. In particular, the strength of chiral interactions can be varied, leading to fascinating transitions in behavior that resembles the formation of starfish vesicles. These observations point towards the important role of line tension, and have potential implications for phase separated lipid mixtures or lipid rafts.

  10. Protein-membrane interactions: blood clotting on nanoscale bilayers. (United States)

    Morrissey, J H; Pureza, V; Davis-Harrison, R L; Sligar, S G; Rienstra, C M; Kijac, A Z; Ohkubo, Y Z; Tajkhorshid, E


    The clotting cascade requires the assembly of protease-cofactor complexes on membranes with exposed anionic phospholipids. Despite their importance, protein-membrane interactions in clotting remain relatively poorly understood. Calcium ions are known to induce anionic phospholipids to cluster, and we propose that clotting proteins assemble preferentially on such anionic lipid-rich microdomains. Until recently, there was no way to control the partitioning of clotting proteins into or out of specific membrane microdomains, so experimenters only knew the average contributions of phospholipids to blood clotting. The development of nanoscale membrane bilayers (Nanodiscs) has now allowed us to probe, with nanometer resolution, how local variations in phospholipid composition regulate the activity of key protease-cofactor complexes in blood clotting. Furthermore, exciting new progress in solid-state NMR and large-scale molecular dynamics simulations allow structural insights into interactions between proteins and membrane surfaces with atomic resolution.

  11. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs. (United States)

    Richards, Mark J; Hsia, Chih-Yun; Singh, Rohit R; Haider, Huma; Kumpf, Julia; Kawate, Toshimitsu; Daniel, Susan


    Membrane protein interactions with lipids are crucial for their native biological behavior, yet traditional characterization methods are often carried out on purified protein in the absence of lipids. We present a simple method to transfer membrane proteins expressed in mammalian cells to an assay-friendly, cushioned, supported lipid bilayer platform using cell blebs as an intermediate. Cell blebs, expressing either GPI-linked yellow fluorescent proteins or neon-green fused transmembrane P2X2 receptors, were induced to rupture on glass surfaces using PEGylated lipid vesicles, which resulted in planar supported membranes with over 50% mobility for multipass transmembrane proteins and over 90% for GPI-linked proteins. Fluorescent proteins were tracked, and their diffusion in supported bilayers characterized, using single molecule tracking and moment scaling spectrum (MSS) analysis. Diffusion was characterized for individual proteins as either free or confined, revealing details of the local lipid membrane heterogeneity surrounding the protein. A particularly useful result of our bilayer formation process is the protein orientation in the supported planar bilayer. For both the GPI-linked and transmembrane proteins used here, an enzymatic assay revealed that protein orientation in the planar bilayer results in the extracellular domains facing toward the bulk, and that the dominant mode of bleb rupture is via the "parachute" mechanism. Mobility, orientation, and preservation of the native lipid environment of the proteins using cell blebs offers advantages over proteoliposome reconstitution or disrupted cell membrane preparations, which necessarily result in significant scrambling of protein orientation and typically immobilized membrane proteins in SLBs. The bleb-based bilayer platform presented here is an important step toward integrating membrane proteomic studies on chip, especially for future studies aimed at understanding fundamental effects of lipid interactions

  12. Tethered bilayer lipid membranes (tBLMs): interest and applications for biological membrane investigations. (United States)

    Rebaud, Samuel; Maniti, Ofelia; Girard-Egrot, Agnès P


    Biological membranes play a central role in the biology of the cell. They are not only the hydrophobic barrier allowing separation between two water soluble compartments but also a supra-molecular entity that has vital structural functions. Notably, they are involved in many exchange processes between the outside and inside cellular spaces. Accounting for the complexity of cell membranes, reliable models are needed to acquire current knowledge of the molecular processes occurring in membranes. To simplify the investigation of lipid/protein interactions, the use of biomimetic membranes is an approach that allows manipulation of the lipid composition of specific domains and/or the protein composition, and the evaluation of the reciprocal effects. Since the middle of the 80's, lipid bilayer membranes have been constantly developed as models of biological membranes with the ultimate goal to reincorporate membrane proteins for their functional investigation. In this review, after a brief description of the planar lipid bilayers as biomimetic membrane models, we will focus on the construction of the tethered Bilayer Lipid Membranes, the most promising model for efficient membrane protein reconstitution and investigation of molecular processes occurring in cell membranes. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  13. Bilayer Thickness Mismatch Controls Domain Size in Model Membranes

    Energy Technology Data Exchange (ETDEWEB)

    Heberle, Frederick A [ORNL; Petruzielo, Robin S [ORNL; Pan, Jianjun [ORNL; Drazba, Paul [ORNL; Kucerka, Norbert [Canadian Neutron Beam Centre and Comelius University (Slovakia); Feigenson, Gerald [Cornell University; Katsaras, John [ORNL


    The observation of lateral phase separation in lipid bilayers has received considerable attention, especially in connection to lipid raft phenomena in cells. It is widely accepted that rafts play a central role in cellular processes, notably signal transduction. While micrometer-sized domains are observed with some model membrane mixtures, rafts much smaller than 100 nm beyond the reach of optical microscopy are now thought to exist, both in vitro and in vivo. We have used small-angle neutron scattering, a probe free technique, to measure the size of nanoscopic membrane domains in unilamellar vesicles with unprecedented accuracy. These experiments were performed using a four-component model system containing fixed proportions of cholesterol and the saturated phospholipid 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), mixed with varying amounts of the unsaturated phospholipids 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1,2-dioleoylsn- glycero-3-phosphocholine (DOPC). We find that liquid domain size increases with the extent of acyl chain unsaturation (DOPC:POPC ratio). Furthermore, we find a direct correlation between domain size and the mismatch in bilayer thickness of the coexisting liquid-ordered and liquid-disordered phases, suggesting a dominant role for line tension in controlling domain size. While this result is expected from line tension theories, we provide the first experimental verification in free-floating bilayers. Importantly, we also find that changes in bilayer thickness, which accompany changes in the degree of lipid chain unsaturation, are entirely confined to the disordered phase. Together, these results suggest how the size of functional domains in homeothermic cells may be regulated through changes in lipid composition.

  14. The impact of cell-penetrating peptides on membrane bilayer structure during binding and insertion. (United States)

    Hirst, Daniel J; Lee, Tzong-Hsien; Kulkarni, Ketav; Wilce, Jacqueline A; Aguilar, Marie-Isabel


    We have studied the effect of penetratin and a truncated analogue on the bilayer structure using dual polarisation interferometry, to simultaneously measure changes in mass per unit area and birefringence (an optical parameter representing bilayer order) with high sensitivity during the binding and dissociation from the membrane. Specifically, we studied penetratin (RQIKIWFQNRRMKWKK), along with a shortened and biotinylated version known as R8K-biotin (RRMKWKKK(Biotin)-NH2). Overall both peptides bound only weakly to the neutral DMPC and POPC bilayers, while much higher binding was observed for the anionic DMPC/DMPG and POPC/POPG. The binding of penetratin to gel-phase DMPC/DMPG was adequately represented by a two-state model, whereas on the fluid-phase POPC/POPG it exhibited a distinctly different binding pattern, best represented by a three-state kinetic model. However, R8K-biotin did not bind well to DMPC/DMPG and showed a more transitory and superficial binding to POPC/POPG. Comparing the modelling results for both peptides binding to POPC/POPG suggests an important role for a securely bound intermediate prior to penetratin insertion and translocation. Overall these results further elucidate the mechanism of penetratin, and provide another example of the significance of the ability of DPI to measure structural changes and the use of kinetic analysis to investigate the stages of peptide-membrane interactions.

  15. Lateral organization, bilayer asymmetry, and inter-leaflet coupling of biological membranes. (United States)

    Nickels, Jonathan D; Smith, Jeremy C; Cheng, Xiaolin


    Understanding of cell membrane organization has evolved significantly from the classic fluid mosaic model. It is now recognized that biological membranes are highly organized structures, with differences in lipid compositions between inner and outer leaflets and in lateral structures within the bilayer plane, known as lipid rafts. These organizing principles are important for protein localization and function as well as cellular signaling. However, the mechanisms and biophysical basis of lipid raft formation, structure, dynamics and function are not clearly understood. One key question, which we focus on in this review, is how lateral organization and leaflet compositional asymmetry are coupled. Detailed information elucidating this question has been sparse because of the small size and transient nature of rafts and the experimental challenges in constructing asymmetric bilayers. Resolving this mystery will require advances in both experimentation and modeling. We discuss here the preparation of model systems along with experimental and computational approaches that have been applied in efforts to address this key question in membrane biology. We seek to place recent and future advances in experimental and computational techniques in context, providing insight into in-plane and transverse organization of biological membranes.

  16. Carrier-mediated ion transport in lipid bilayer membranes. (United States)

    Laprade, R; Grenier, F; Pagé-Dansereau, M; Dansereau, J


    The electrical properties predicted by a widely accepted model for carrier-mediated ion transport in lipid bilayers are described. The different steps leading to ion transport and their associated rate constants are reaction at the interface between an ion in the aqueous phase and a carrier in the membrane (kRi), followed by translocation of the ion-carrier complex across the membrane interior (kis) and its dissociation at the other interface (kDi) after which the free carrier crosses back the membrane interior (ks). Results on glyceryl monooleate (GMO) membranes for a family of homologue carriers, the macrotetralide actin antibiotics (nonactin, monactin, dinactin, trinactin, and tetranactin) and a variety of ions (Na+, Cs+, Rb+, K+, NH4+, and Tl+) are presented. Internally consistent data obtained from steady-state electrical measurements (zero-current potential and conductance, current-voltage relationship) allow us to obtain the equilibrium permeability ratios for the different ions and show that for a given carrier kRi is relatively invariant from one ion to the other, except for Tl+ (larger), which implies that the ionic selectivity is controlled by the dissociation of the complex. The values of the individual rate constants obtained from current relaxation experiments are also presented and confirm the findings from steady-state measurements, as well as the isostericity concept for complexes of different ions with the same carrier (kis invariant). These also allow us to determine the aqueous phase membrane and torus membrane partition coefficients. Finally, the observed increase in kis from nonactin to tetranactin and, for all homologues, from GMO-decane to solvent-free GMO membranes, together with the concomitant decrease in kDi, can be explained in terms of modifications of electrostatic energy profiles induced by variations in carrier size and membrane thickness.

  17. Detergent interaction with tethered bilayer lipid membranes for protein reconstitution (United States)

    Broccio, Matteo; Zan Goh, Haw; Loesche, Mathias


    Tethered bilayer lipid membranes (tBLMs) are self-assembled biomimetic structures in which the membrane is separated from a solid substrate by a nm-thick hydrated submembrane space. These model systems are being used in binding studies of peripheral proteins and exotoxins. Here we aim at their application for the reconstitution of water-insoluble integral membrane proteins. As an alternative to fusion of preformed proteoliposomes we study the direct reconstitution of such proteins for applications in biosensing and pharmaceutical screening. For reconstitution, highly insulating tBLMs (R˜10^5-10^6 φ) were temporarily incubated with a detergent to screen for conditions that keep the detergent-saturated membranestable and ready to incorporate detergent-solubilized proteins. We assess the electrical characteristics, i.e. specific resistance and capacitance, by means of electrochemical impedance spectroscopy (EIS) under timed incubation with decylmaltoside and dodecylmaltoside detergents in a regime around their critical micelle concentration, 1.8 mM and 0.17 mM respectively and demonstrate the restoration of the tBLM upon detergent removal. Thereby a range of concentration and incubation times was identified, that represents optimal conditions for the subsequent membrane protein reconstitution.

  18. Brownian dynamics simulations of lipid bilayer membrane with hydrodynamic interactions in LAMMPS (United States)

    Fu, Szu-Pei; Young, Yuan-Nan; Peng, Zhangli; Yuan, Hongyan


    Lipid bilayer membranes have been extensively studied by coarse-grained molecular dynamics simulations. Numerical efficiencies have been reported in the cases of aggressive coarse-graining, where several lipids are coarse-grained into a particle of size 4 6 nm so that there is only one particle in the thickness direction. Yuan et al. proposed a pair-potential between these one-particle-thick coarse-grained lipid particles to capture the mechanical properties of a lipid bilayer membrane (such as gel-fluid-gas phase transitions of lipids, diffusion, and bending rigidity). In this work we implement such interaction potential in LAMMPS to simulate large-scale lipid systems such as vesicles and red blood cells (RBCs). We also consider the effect of cytoskeleton on the lipid membrane dynamics as a model for red blood cell (RBC) dynamics, and incorporate coarse-grained water molecules to account for hydrodynamic interactions. The interaction between the coarse-grained water molecules (explicit solvent molecules) is modeled as a Lennard-Jones (L-J) potential. We focus on two sets of LAMMPS simulations: 1. Vesicle shape transitions with varying enclosed volume; 2. RBC shape transitions with different enclosed volume. This work is funded by NSF under Grant DMS-1222550.

  19. Molecular dynamics study of lipid bilayers modeling the plasma membranes of mouse hepatocytes and hepatomas (United States)

    Andoh, Yoshimichi; Aoki, Noriyuki; Okazaki, Susumu


    Molecular dynamics (MD) calculations of lipid bilayers modeling the plasma membranes of normal mouse hepatocytes and hepatomas in water have been performed under physiological isothermal-isobaric conditions (310.15 K and 1 atm). The changes in the membrane properties induced by hepatic canceration were investigated and were compared with previous MD calculations included in our previous study of the changes in membrane properties induced by murine thymic canceration. The calculated model membranes for normal hepatocytes and hepatomas comprised 23 and 24 kinds of lipids, respectively. These included phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. We referred to previously published experimental values for the mole fraction of the lipids adopted in the present calculations. The calculated structural and dynamic properties of the membranes such as lateral structure, order parameters, lateral self-diffusion constants, and rotational correlation times all showed that hepatic canceration causes plasma membranes to become more ordered laterally and less fluid. Interestingly, this finding contrasts with the less ordered structure and increased fluidity of plasma membranes induced by thymic canceration observed in our previous MD study.

  20. Acyl chain composition and coexisting fluid phases in lipid bilayers (United States)

    Gu, Yongwen; Bradley, Miranda; Mitchell, Drake


    At room temperature phospholipid bilayers enriched in sphingolipids and cholesterol may form a solid phase as well as two coexisting fluid phases. These are the standard fluid phase, or the liquid-disordered phase, ld, and the liquid-ordered phase, lo, which is commonly associated with lipid rafts. Ternary mixtures of palmitoyl-oleoyl-phosphocholine (POPC; 16:0,18:1 PC), sphingomyelin (SPM), and cholesterol (Chol) form coexisting lo, ld and solid phases over a wide range of molar ratios. We are examining the ability of two fluorescent probes to detect these 2 phases: NBD linked to di-16:0 PE which partitions strongly into the lo phase and NBD linked to di-18:1 PE which partitions strongly into the ld phase. We are also examining the effect of the highly polyunsaturated phospholipid stearoyl-docosahexanoyl-phosphocholine (SDPC; 18:0, 22:6 PC) on the ternary phase diagram of POPC/SPM/Chol with particular focus on the functionally important lo/ld coexistence region. We report on the fluorescence lifetime and anisotropy decay dynamics of these two fluorescent probes.

  1. Multiscale modeling of droplet interface bilayer membrane networks. (United States)

    Freeman, Eric C; Farimani, Amir B; Aluru, Narayana R; Philen, Michael K


    Droplet interface bilayer (DIB) networks are considered for the development of stimuli-responsive membrane-based materials inspired by cellular mechanics. These DIB networks are often modeled as combinations of electrical circuit analogues, creating complex networks of capacitors and resistors that mimic the biomolecular structures. These empirical models are capable of replicating data from electrophysiology experiments, but these models do not accurately capture the underlying physical phenomena and consequently do not allow for simulations of material functionalities beyond the voltage-clamp or current-clamp conditions. The work presented here provides a more robust description of DIB network behavior through the development of a hierarchical multiscale model, recognizing that the macroscopic network properties are functions of their underlying molecular structure. The result of this research is a modeling methodology based on controlled exchanges across the interfaces of neighboring droplets. This methodology is validated against experimental data, and an extension case is provided to demonstrate possible future applications of droplet interface bilayer networks.

  2. Force dynamique des membranes fluides (United States)

    Evans, Evan; Heinrich, Volkmar


    Rupturing fluid membrane vesicles with a steady ramp of micropipette suction yields a tension distribution that images the kinetic process of membrane failure. When plotted on a log scale of tension loading rate, the distribution peaks (membrane strengths) define a dynamic tension spectrum with distinct regimes that reflect passage of prominent energy barriers along the pathway to rupture. Demonstrated here by tests on giant PC lipid vesicles over loading rates from 0.06-60 mN/m/s, the stochastic process of rupture can be modelled as a causal sequence of two thermally-activated transitions where each transition governs membrane strength on separate scales of loading rate. Under fast ramps of tension, a steep linear regime appears in each spectrum at high strengths which implies that failure requires nucleation of a rare nanoscale defect. The slope and projected intercept yield defect size and spontaneous production rate respectively. However, under slow ramps of loading, the spectrum crosses over to a shallow-curved regime at lower strength, which is consistent with the kinetic impedance to opening an unstable hole in a fluid film. The dependence of rupture tension on rate reveals hole edge energy and frequency scale for thermal fluctuations in size. To cite this article: E. Evans, V. Heinrich, C. R. Physique 4 (2003).

  3. Enhanced sensitivity of a microfabricated resonator using a graphene-polystyrene bilayer membrane

    Energy Technology Data Exchange (ETDEWEB)

    Yun, Minhyuk; Lee, Eunho; Cho, Kilwon; Jeon, Sangmin, E-mail: [Department of Chemical Engineering, Pohang University of Science and Technology (POSTECH), Pohang (Korea, Republic of)


    A graphene layer was synthesized using chemical vapor deposition methods and a polystyrene solution was spin-cast onto the graphene film. The graphene-polystyrene bilayer membrane was attached between the two tines of a microfabricated quartz tuning fork (QTF). The modulus of the graphene-polystyrene bilayer was measured to be twice that of a pristine polystyrene membrane. Exposure of the membrane-coated QTF to ethanol vapor decreased the resonance frequency of the microresonator. The bilayer membrane-coated QTF produced a frequency change that was three times the change obtained using a polystyrene membrane-coated QTF, with a lower degree of degradation in the Q factor. The limit of detection of the bilayer membrane-coated QTF to ethanol vapor was determined to be 20 ppm.

  4. Enhanced sensitivity of a microfabricated resonator using a graphene-polystyrene bilayer membrane (United States)

    Yun, Minhyuk; Lee, Eunho; Cho, Kilwon; Jeon, Sangmin


    A graphene layer was synthesized using chemical vapor deposition methods and a polystyrene solution was spin-cast onto the graphene film. The graphene-polystyrene bilayer membrane was attached between the two tines of a microfabricated quartz tuning fork (QTF). The modulus of the graphene-polystyrene bilayer was measured to be twice that of a pristine polystyrene membrane. Exposure of the membrane-coated QTF to ethanol vapor decreased the resonance frequency of the microresonator. The bilayer membrane-coated QTF produced a frequency change that was three times the change obtained using a polystyrene membrane-coated QTF, with a lower degree of degradation in the Q factor. The limit of detection of the bilayer membrane-coated QTF to ethanol vapor was determined to be 20 ppm.

  5. Mechanical properties of electrospun bilayer fibrous membranes as potential scaffolds for tissue engineering. (United States)

    Pu, Juan; Komvopoulos, Kyriakos


    Bilayer fibrous membranes of poly(l-lactic acid) (PLLA) were fabricated by electrospinning, using a parallel-disk mandrel configuration that resulted in the sequential deposition of a layer with fibers aligned across the two parallel disks and a layer with randomly oriented fibers, both layers deposited in a single process step. Membrane structure and fiber alignment were characterized by scanning electron microscopy and two-dimensional fast Fourier transform. Because of the intricacies of the generated electric field, bilayer membranes exhibited higher porosity than single-layer membranes consisting of randomly oriented fibers fabricated with a solid-drum collector. However, despite their higher porosity, bilayer membranes demonstrated generally higher elastic modulus, yield strength and toughness than single-layer membranes with random fibers. Bilayer membrane deformation at relatively high strain rates comprised multiple abrupt microfracture events characterized by discontinuous fiber breakage. Bilayer membrane elongation yielded excessive necking of the layer with random fibers and remarkable fiber stretching (on the order of 400%) in the layer with fibers aligned in the stress direction. In addition, fibers in both layers exhibited multiple localized necking, attributed to the nonuniform distribution of crystalline phases in the fibrillar structure. The high membrane porosity, good mechanical properties, and good biocompatibility and biodegradability of PLLA (demonstrated in previous studies) make the present bilayer membranes good scaffold candidates for a wide range of tissue engineering applications.

  6. Thermal fluctuations and bending rigidity of bilayer membranes. (United States)

    Tarazona, Pedro; Chacón, Enrique; Bresme, Fernando


    We present a new scheme to estimate the elastic properties of biological membranes in computer simulations. The method analyzes the thermal fluctuations in terms of a coupled undulatory mode, which disentangle the mixing of the mesoscopic undulations and the high-q protrusions. This approach makes possible the accurate estimation of the bending modulus both for membranes under stress and in tensionless conditions; it also extends the applicability of the fluctuation analysis to the small membrane areas normally used in atomistic simulations. Also we clarify the difference between the surface tension imposed in simulations through a pressure coupling barostat, and the surface tension that can be extracted from the analysis of the low wave vector dependence of the coupled undulatory fluctuation spectrum. The physical analysis of the peristaltic mode is also refined, by separating the bulk and protrusions contributions. We illustrate the procedure by analyzing 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine bilayers. The bending moduli obtained from our analysis, shows good agreement with available experiments.

  7. Pressure effects on the equilibrium configurations of bilayer lipid membranes (United States)

    DeVita, Raffaella; Stewart, Iain W.; Leo, Donald J.


    Planar bilayer lipid membranes (BLMs) are currently employed to construct many bio-inspired material systems and structures. In order to characterize the pressure effects on the equilibrium configurations of these biological membranes, a novel continuum model is proposed. The BLM is assumed to be a two-layer smectic A liquid crystal. The mean orientation of the amphiphilic molecules comprising the membrane is postulated to be perpendicular to the layers and each layer is idealized as a two-dimensional liquid. Moreover, the BLM is modeled as a simply supported plate undergoing small deformations. It is subjected to a pressure load that acts perpendicularly to the layers. The equilibrium equations and boundary conditions are derived from the bulk elastic energy for smectic A liquid crystals as described by de Gennes and Prost (1993 The Physics of Liquid Crystals 2nd edn (Oxford Science Publications)) by using variational methods. The resulting fourth-order linear partial differential equation is solved by employing cylindrical functions and the series solution is proved to be convergent. The solution is numerically computed for values of the model parameters that are reported in the literature. This paper is dedicated to the memory of our colleagues, Professors Kevin P Granata and Liviu Librescv, who lost their lives during the sensless tragedy on 16 April, 2007 at Virginia Tech.

  8. Effective interactions between fluid membranes

    CERN Document Server

    Lu, Bing-Sui


    A self-consistent theory is proposed for the general problem of interacting undulating fluid membranes subject to the constraint that they do not interpenetrate. We implement the steric constraint via an exact functional integral representation, and through the use of a saddle-point approximation transform it into a novel effective steric potential. The steric potential is found to consist of two contributions: one generated by zero mode fluctuations of the membranes, and the other by thermal bending fluctuations. For membranes of cross-sectional area $S$, we find that the bending fluctuation part scales with the inter-membrane separation $d$ as $d^{-2}$ for $d \\ll \\sqrt{S}$, but crosses over to $d^{-4}$ scaling for $d \\gg \\sqrt{S}$, whereas the zero mode part of the steric potential always scales as $d^{-2}$. For membranes interacting exclusively via the steric potential, we obtain closed-form expressions for the effective interaction potential and for the rms undulation amplitude $\\sigma$, which becomes sma...

  9. Lipid bilayer regulation of membrane protein function: gramicidin channels as molecular force probes

    DEFF Research Database (Denmark)

    Lundbæk, Jens August; Collingwood, S.A.; Ingolfsson, H.I.


    physical properties. This advance is because of the introduction of new tools for studying lipid bilayer regulation of protein function. The present review provides an introduction to the regulation of membrane protein function by the bilayer physical properties. We further describe the use of gramicidin...... channels as molecular force probes for studying this mechanism, with a unique ability to discriminate between consequences of changes in monolayer curvature and bilayer elastic moduli....

  10. Formation of "solvent-free" black lipid bilayer membranes from glyceryl monooleate dispersed in squalene. (United States)

    White, S H


    A simple technique for forming "black" lipid bilayer membranes containing negligible amounts of alkyl solvent is described. The membranes are formed by the method of Mueller et al (Circulation. 1962. 26:1167.) from glyceryl monooleate (GMO) dispersed in squalene. The squalene forms an annulus to satisfy the boundary conditions of the planar bilayer but does not appear to dissolve noticeably in the bilayer itself. The specific geometric capacitance (Cg) of the membranes at 20 degrees C formed by this technique is 0.7771 +/- 0.0048 muF/cm2. Theoretical estimates of Cg for solvent-free bilayers range from 0.75 to 0.81 muF/cm2. Alkane-free GMO bilayers formed from n-octadecane by the solvent freeze-out method of White (Biochim. Biophys. Acta. 1974. 356:8) have values of Cg = 0.7903 +/- 0.0013 muF/cm2 at 20.5 degrees C. The agreement between the various values of Cg strongly suggests that the bilayers are free of squalene. DC potentials applied to the bilayers have no detectable effect on the value of Cg, as expected for solvent-free films. The ability to form bilayers essentially free of the solvent used in the forming solution makes it possible to determine the area per molecule of the surface active lipid in the bilayer. The area per molecule of GMO at 20 degrees C is estimated to be 37.9 +/- 0.2 A2.

  11. Particle-based simulations of bilayer membranes: self-assembly, structural analysis, and shock-wave damage (United States)

    Steinhauser, Martin O.; Schindler, Tanja


    We report on the results of particle-based, coarse-grained molecular dynamics simulations of amphiphilic lipid molecules in aqueous environment where the membrane structures at equilibrium are subsequently exposed to strong shock waves, and their damage is analyzed. The lipid molecules self-assemble from unbiased random initial configurations to form stable bilayer membranes, including closed vesicles. During self-assembly of lipid molecules, we observe several stages of clustering, starting with many small clusters of lipids, gradually merging together to finally form one single bilayer membrane. We find that the clustering of lipids sensitively depends on the hydrophobic interaction h_c of the lipid tails in our model and on temperature T of the system. The self-assembled bilayer membranes are quantitatively analyzed at equilibrium with respect to their degree of order and their local structure. We also show that—by analyzing the membrane fluctuations and using a linearized theory— we obtain area compression moduli K_A and bending stiffnesses κ_B for our bilayer membranes which are within the experimental range of in vivo and in vitro measurements of biological membranes. We also discuss the density profile and the pair correlation function of our model membranes at equilibrium which has not been done in previous studies of particle-based membrane models. Furthermore, we present a detailed phase diagram of our lipid model that exhibits a sol-gel transition between quasi-solid and fluid domains, and domains where no self-assembly of lipids occurs. In addition, we present in the phase diagram the conditions for temperature T and hydrophobicity h_c of the lipid tails of our model to form closed vesicles. The stable bilayer membranes obtained at equilibrium are then subjected to strong shock waves in a shock tube setup, and we investigate the damage in the membranes due to their interaction with shock waves. Here, we find a transition from self

  12. Particle-based simulations of bilayer membranes: self-assembly, structural analysis, and shock-wave damage (United States)

    Steinhauser, Martin O.; Schindler, Tanja


    We report on the results of particle-based, coarse-grained molecular dynamics simulations of amphiphilic lipid molecules in aqueous environment where the membrane structures at equilibrium are subsequently exposed to strong shock waves, and their damage is analyzed. The lipid molecules self-assemble from unbiased random initial configurations to form stable bilayer membranes, including closed vesicles. During self-assembly of lipid molecules, we observe several stages of clustering, starting with many small clusters of lipids, gradually merging together to finally form one single bilayer membrane. We find that the clustering of lipids sensitively depends on the hydrophobic interaction h_c of the lipid tails in our model and on temperature T of the system. The self-assembled bilayer membranes are quantitatively analyzed at equilibrium with respect to their degree of order and their local structure. We also show that—by analyzing the membrane fluctuations and using a linearized theory— we obtain area compression moduli K_A and bending stiffnesses κ _B for our bilayer membranes which are within the experimental range of in vivo and in vitro measurements of biological membranes. We also discuss the density profile and the pair correlation function of our model membranes at equilibrium which has not been done in previous studies of particle-based membrane models. Furthermore, we present a detailed phase diagram of our lipid model that exhibits a sol-gel transition between quasi-solid and fluid domains, and domains where no self-assembly of lipids occurs. In addition, we present in the phase diagram the conditions for temperature T and hydrophobicity h_c of the lipid tails of our model to form closed vesicles. The stable bilayer membranes obtained at equilibrium are then subjected to strong shock waves in a shock tube setup, and we investigate the damage in the membranes due to their interaction with shock waves. Here, we find a transition from self

  13. Influence of the bilayer composition on the binding and membrane disrupting effect of Polybia-MP1, an antimicrobial mastoparan peptide with leukemic T-lymphocyte cell selectivity. (United States)

    dos Santos Cabrera, Marcia Perez; Arcisio-Miranda, Manoel; Gorjão, Renata; Leite, Natália Bueno; de Souza, Bibiana Monson; Curi, Rui; Procopio, Joaquim; Ruggiero Neto, João; Palma, Mario Sérgio


    This study shows that MP-1, a peptide from the venom of the Polybia paulista wasp, is more toxic to human leukemic T-lymphocytes than to human primary lymphocytes. By using model membranes and electrophysiology measurements to investigate the molecular mechanisms underlying this selective action, the porelike activity of MP-1 was identified with several bilayer compositions. The highest average conductance was found in bilayers formed by phosphatidylcholine or a mixture of phosphatidylcholine and phosphatidylserine (70:30). The presence of cholesterol or cardiolipin substantially decreases the MP-1 pore activity, suggesting that the membrane fluidity influences the mechanism of selective toxicity. The determination of partition coefficients from the anisotropy of Trp indicated higher coefficients for the anionic bilayers. The partition coefficients were found to be 1 order of magnitude smaller when the bilayers contain cholesterol or a mixture of cholesterol and sphingomyelin. The blue shift fluorescence, anisotropy values, and Stern-Volmer constants are indications of a deeper penetration of MP-1 into anionic bilayers than into zwitterionic bilayers. Our results indicate that MP-1 prefers to target leukemic cell membranes, and its toxicity is probably related to the induction of necrosis and not to DNA fragmentation. This mode of action can be interpreted considering a number of bilayer properties like fluidity, lipid charge, and domain formation. Cholesterol-containing bilayers are less fluid and less charged and have a tendency to form domains. In comparison to healthy cells, leukemic T-lymphocyte membranes are deprived of this lipid, resulting in decreased peptide binding and lower conductance. We showed that the higher content of anionic lipids increases the level of binding of the peptide to bilayers. Additionally, the absence of cholesterol resulted in enhanced pore activity. These findings may drive the selective toxicity of MP-1 to Jurkat cells.

  14. Method of fabricating lipid bilayer membranes on solid supports (United States)

    Cho, Nam-Joon (Inventor); Frank, Curtis W. (Inventor); Glenn, Jeffrey S. (Inventor); Cheong, Kwang Ho (Inventor)


    The present invention provides a method of producing a planar lipid bilayer on a solid support. With this method, a solution of lipid vesicles is first deposited on the solid support. Next, the lipid vesicles are destabilized by adding an amphipathic peptide solution to the lipid vesicle solution. This destabilization leads to production of a planar lipid bilayer on the solid support. The present invention also provides a supported planar lipid bilayer, where the planar lipid bilayer is made of naturally occurring lipids and the solid support is made of unmodified gold or titanium oxide. Preferably, the supported planar lipid bilayer is continuous. The planar lipid bilayer may be made of any naturally occurring lipid or mixture of lipids, including, but not limited to phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinsitol, cardiolipin, cholesterol, and sphingomyelin.

  15. Filament networks attached to membranes: cytoskeletal pressure and local bilayer deformation (United States)

    Auth, Thorsten; Safran, S. A.; Gov, Nir S.


    Several cell types, among them red blood cells, have a cortical, two-dimensional (2D) network of filaments sparsely attached to their lipid bilayer. In many mammalian cells, this 2D polymer network is connected to an underlying 3D, more rigid cytoskeleton. In this paper, we consider the pressure exerted by the thermally fluctuating, cortical network of filaments on the bilayer and predict the bilayer deformations that are induced by this pressure. We treat the filaments as flexible polymers and calculate the pressure that a network of such linear chains exerts on the bilayer; we then minimize the bilayer shape in order to predict the resulting local deformations. We compare our predictions with membrane deformations observed in electron micrographs of red blood cells. The polymer pressure along with the resulting membrane deformation can lead to compartmentalization, regulate in-plane diffusion and may influence protein sorting as well as transmit signals to the polymerization of the underlying 3D cytoskeleton.

  16. Filament networks attached to membranes: cytoskeletal pressure and local bilayer deformation

    Energy Technology Data Exchange (ETDEWEB)

    Auth, Thorsten [Department of Materials and Interfaces, Weizmann Institute of Science, PO Box 26, Rehovot 76100 (Israel); Safran, S A [Department of Materials and Interfaces, Weizmann Institute of Science, PO Box 26, Rehovot 76100 (Israel); Gov, Nir S [Department of Chemical Physics, Weizmann Institute of Science, PO Box 26, Rehovot 76100 (Israel)


    Several cell types, among them red blood cells, have a cortical, two-dimensional (2D) network of filaments sparsely attached to their lipid bilayer. In many mammalian cells, this 2D polymer network is connected to an underlying 3D, more rigid cytoskeleton. In this paper, we consider the pressure exerted by the thermally fluctuating, cortical network of filaments on the bilayer and predict the bilayer deformations that are induced by this pressure. We treat the filaments as flexible polymers and calculate the pressure that a network of such linear chains exerts on the bilayer; we then minimize the bilayer shape in order to predict the resulting local deformations. We compare our predictions with membrane deformations observed in electron micrographs of red blood cells. The polymer pressure along with the resulting membrane deformation can lead to compartmentalization, regulate in-plane diffusion and may influence protein sorting as well as transmit signals to the polymerization of the underlying 3D cytoskeleton.

  17. Evaluation and biological characterization of bilayer gelatin/chondroitin-6-sulphate/hyaluronic acid membrane. (United States)

    Wang, Tzu-Wei; Sun, Jui-Sheng; Wu, Hsi-Chin; Huang, Yi-Chau; Lin, Feng-Huei


    A biodegradable polymer scaffold was developed using gelatin, chondroitin-6-sulphate, and hyaluronic acid in the form of bilayer network. The bilayer porous structure of gelatin-chondroitin-6-sulphate-hyaluronic acid (G-C6S-HA) membrane was fabricated using different freezing temperatures followed by lyophilization. 1-Ethyl-3(3-dimethylaminopropyl) carbodiimide was used as crosslinking agent to improve the biological stability of the scaffold. The morphology, physical-chemical properties, and biocompatibility of bilayer G-C6S-HA membrane were evaluated in this study. The functional groups change in crosslinked G-C6S-HA scaffold was characterized by fourier transform infrared spectroscopy. The retention of glycosaminoglycan contents and matrix degradation rate were also examined by p-dimethylamino benzaldehyde and 2,4,6-trinitrobenzene sulphonic acid, respectively. Water absorption capacity was carried out to study G-C6S-HA membrane water containing characteristics. The morphology of the bilayer G-C6S-HA membrane was investigated under scanning electron microscope and light microscopy. In vitro biocompatibility was conducted with MTT test, LDH assay, as well as histological analysis. The results showed that the morphology of bilayer G-C6S-HA membrane was well reserved. The physical-chemical properties were also adequate. With good biocompatibility, this bilayer G-C6S-HA membrane would be suitable as a matrix in the application of tissue engineering.

  18. Effect of glucosylceramide on the biophysical properties of fluid membranes. (United States)

    Varela, Ana R P; Gonçalves da Silva, Amélia M P S; Fedorov, Alexander; Futerman, Anthony H; Prieto, Manuel; Silva, Liana C


    Glucosylceramide (GlcCer), a relevant intermediate in the pathways of glycosphingolipid metabolism, plays key roles in the regulation of cell physiology. The molecular mechanisms by which GlcCer regulates cellular processes are unknown, but might involve changes in membrane biophysical properties and formation of lipid domains. In the present study, fluorescence spectroscopy, confocal microscopy and surface pressure-area (π-A) measurements were used to characterize the effect of GlcCer on the biophysical properties of model membranes. We show that C16:0-GlcCer has a high tendency to segregate into highly ordered gel domains and to increase the order of the fluid phase. Monolayer studies support the aggregation propensity of C16:0-GlcCer. π-A isotherms of single C16:0-GlcCer indicate that bilayer domains, or crystal-like structures, coexist within monolayer domains at the air-water interface. Mixtures with POPC exhibit partial miscibility with expansion of the mean molecular areas relative to the additive behavior of the components. Moreover, C16:0-GlcCer promotes morphological alterations in lipid vesicles leading to formation of flexible tubule-like structures that protrude from the fluid region of the bilayer. These results support the hypothesis that alterations in membrane biophysical properties induced by GlcCer might be involved in its mechanism of action.

  19. Microfabrication of hybrid fluid membrane for microengines (United States)

    Chutani, R.; Formosa, F.; de Labachelerie, M.; Badel, A.; Lanzetta, F.


    This paper describes the microfabrication and dynamic characterization of thick membranes providing a technological solution for microengines. The studied membranes are called hybrid fluid-membrane (HFM) and consist of two thin membranes that encapsulate an incompressible fluid. This work details the microelectromechanical system (MEMS) scalable fabrication and characterization of HFMs. The membranes are composite structures based on Silicon spiral springs embedded in a polymer (RTV silicone). The anodic bonding of multiple stacks of Si/glass structures, the fluid filling and the sealing have been demonstrated. Various HFMs were successfully fabricated and their dynamic characterization demonstrates the agreement between experimental and theoretical results.

  20. Evidence that bilayer bending rigidity affects membrane protein folding. (United States)

    Booth, P J; Riley, M L; Flitsch, S L; Templer, R H; Farooq, A; Curran, A R; Chadborn, N; Wright, P


    The regeneration kinetics of the integral membrane protein bacteriorhodopsin have been investigated in a lipid-based refolding system. Previous studies on bacteriorhodopsin regeneration have involved detergent-based systems, and in particular mixed dimyristoylphosphatidylcholine (DMPC)/CHAPS micelles. Here, we show that the short chain lipid dihexanoylphosphatidylcholine (DHPC) can be substituted for the detergent CHAPS and that bacteriorhodopsin can be regenerated to high yield in mixed DMPC/DHPC micelles. Bacteriorhodopsin refolding kinetics are measured in the mixed DMPC/DHPC micelles. Rapid, stopped flow mixing is employed to initiate refolding of denatured bacterioopsin in SDS micelles with mixed DMPC/DHPC micelles and time-resolved fluorescence spectroscopy to follow changes in protein fluorescence during folding. Essentially identical refolding kinetics are observed for mixed DMPC/CHAPS and mixed DMPC/DHPC micelles. Only one second-order retinal/apoprotein reaction is identified, in which retinal binds to a partially folded apoprotein intermediate, and the free energy of this retinal binding reaction is found to be the same in both types of mixed micelles. Formation of the partially folded apoprotein intermediate is a rate-limiting step in protein folding and appears to be biexponential. Both apparent rate constants are found to be dependent on the relative proportion of DMPC present in the mixed DMPC/DHPC micelles as well as on the pH of the aqueous phase. Increasing the DMPC concentration should increase the bending rigidity of the amphiphilic bilayer, and this is found to slow the rate of formation of the partially folded apoprotein intermediate. Increasing the mole fraction of DMPC from 0.3 to 0.6 slows the two apparent rate constants associated with formation of this intermediate from 0.29 and 0.031 to 0.11 and 0.013 s-1, respectively. Formation of the intermediate also slows with increasing pH, from 0.11 and 0.013 s-1 at pH 6 to 0.033 and 0.0053 s-1 at

  1. Study of supported bilayer lipid membranes for use in chemo-electric energy conversion via active proton transport (United States)

    Sarles, Stephen A.; Sundaresan, Vishnu B.; Leo, Donald J.


    Bilayer lipid membranes (BLMs) have been studied extensively due to functional and structural similarities to cell membranes, fostering research to understand ion-channel protein functions, measure bilayer mechanical properties, and identify self-assembly mechanisms. BLMs have traditionally been formed across single pores in substrates such as PTFE (Teflon). The incorporation of ion-channel proteins into the lipid bilayer enables the selective transfer of ions and fluid through the BLM. Processes of this nature have led to the measurement of ion current flowing across the lipid membrane and have been used to develop sensors that signal the presence of a particular reactant (glucose, urea, penicillin), improve drug recognition in cells, and develop materials capable of creating chemical energy from light. Recent research at Virginia Tech has shown that the incorporation of proton transporters in a supported BLM formed across an array of pores can convert chemical energy available in the adenosine triphosphate (ATP) into electricity. Experimental results from this work show that the system-named Biocell-is capable of developing 2µW/cm2 of membrane area with 15μl of ATPase. Efforts to increase the power output and conversion efficiency of this process while moving toward a packaged device present a unique engineering problem. The bilayer, as host to the active proton transporters, must therefore be formed evenly across a porous substrate, remain stable and yet fluid-like for protein interaction, and exhibit a large seal resistance. This article presents the ongoing work to characterize the Biocell using impedance analysis. Electrical impedance spectroscopy (EIS) is used to study the effect of adding ATPase proteins to POPS:POPE bilayer lipid membranes and correlate structural changes evident in the impedance data to the energy-conversion capability of various partial and whole Biocell assemblies. The specific membrane resistance of a pure BLM drops from 40-120k

  2. The Power of Asymmetry: Architecture and Assembly of the Gram-Negative Outer Membrane Lipid Bilayer. (United States)

    Henderson, Jeremy C; Zimmerman, Shawn M; Crofts, Alexander A; Boll, Joseph M; Kuhns, Lisa G; Herrera, Carmen M; Trent, M Stephen


    Determining the chemical composition of biological materials is paramount to the study of natural phenomena. Here, we describe the composition of model gram-negative outer membranes, focusing on the predominant assembly, an asymmetrical bilayer of lipid molecules. We also give an overview of lipid biosynthetic pathways and molecular mechanisms that organize this material into the outer membrane bilayer. An emphasis is placed on the potential of these pathways as targets for antibiotic development. We discuss deviations in composition, through bacterial cell surface remodeling, and alternative modalities to the asymmetric lipid bilayer. Outer membrane lipid alterations of current microbiological interest, such as lipid structures found in commensal bacteria, are emphasized. Additionally, outer membrane components could potentially be engineered to develop vaccine platforms. Observations related to composition and assembly of gram-negative outer membranes will continue to generate novel discoveries, broaden biotechnologies, and reveal profound mysteries to compel future research.

  3. Drag Coefficient of a Liquid Domain in a Fluid Membrane Surrounded by Confined Three-Dimensional Fluids (United States)

    Fujitani, Youhei


    It is thought that, in a biomembrane, some minor lipid constituents are concentrated in a domain called the lipid raft. Some raftlike domains in a lipid-bilayer membrane can be regarded as two-dimensional droplets. The membrane viscosities inside and outside the domain are generally different. The present author previously studied the drag coefficient of a circular liquid domain in a flat fluid membrane surrounded by three-dimensional fluids, which occupy the semi-infinite spaces on both sides of the membrane. Here we generalize this problem by assuming that the surrounding fluids are confined by container walls parallel to the membrane. Errors in the present author's previous studies are also corrected in this paper.

  4. A generic model for lipid monolayers, bilayers, and membranes

    CERN Document Server

    Schmid, F; Lenz, O; West, B


    We describe a simple coarse-grained model which is suited to study lipid layers and their phase transitions. Lipids are modeled by short semiflexible chains of beads with a solvophilic head and a solvophobic tail component. They are forced to self-assemble into bilayers by a computationally cheap `phantom solvent' environment. The model reproduces the most important phases and phase transitions of monolayers and bilayers. Technical issues such as Monte Carlo parallelization schemes are briefly discussed.

  5. Elastic Properties and Line Tension of Self-Assembled Bilayer Membranes

    CERN Document Server

    Li, Jianfeng; Shi, An-Chang; Schmid, Friederike; Zhou, Jiajia


    The elastic properties of a self-assembled bilayer membrane are studied using the self-consistent field theory, applied to a model system composed of flexible amphiphilic chains dissolved in hydrophilic polymeric solvents. Examining the free energy of bilayer membranes with different geometries allows us to calculate their bending modulus, Gaussian modulus, two fourth-order membrane moduli, and the line tension. The dependence of these parameters on the microscopic characteristics of the amphiphilic chain, characterized by the volume fraction of the hydrophilic component, is systematically studied. The theoretical predictions are compared with the results from a simple monolayer model, which approximates a bilayer membrane by two monolayers. Finally the region of validity of the linear elasticity theory is analyzed by examining the higher-order contributions.

  6. Influence of membrane surface charge on adsorption of complement proteins onto supported lipid bilayers. (United States)

    Yorulmaz, Saziye; Jackman, Joshua A; Hunziker, Walter; Cho, Nam-Joon


    The complement system is an important part of the innate immune response, and there is great interest in understanding how complement proteins interact with lipid membrane interfaces, especially in the context of recognizing foreign particulates (e.g., liposomal nanomedicines). Herein, a supported lipid bilayer platform was employed in order to investigate the effect of membrane surface charge (positive, negative, or neutral) on the adsorption of three complement proteins. Quartz crystal microbalance-dissipation (QCM-D) experiments measured the real-time kinetics and total uptake of protein adsorption onto supported lipid bilayers. The results demonstrate that all three proteins exhibit preferential, mainly irreversible adsorption onto negatively charged lipid bilayers, yet there was also significant variation in total uptake and the relative degree of adsorption onto negatively charged bilayers versus neutral and positively charged bilayers. The total uptake was also observed to strongly depend on the bulk protein concentration. Taken together, our findings contribute to a broader understanding of the factors which influence adsorption of complement proteins onto lipid membranes and offer guidance towards the design of synthetic lipid bilayers with immunocompetent features.

  7. Regulation of membrane protein function by lipid bilayer elasticity—a single molecule technology to measure the bilayer properties experienced by an embedded protein

    DEFF Research Database (Denmark)

    Lundbæk, Jens August


    , in the general regulation of membrane protein function, is unclear. This is to a large extent due to lack of a generally accepted framework in which to understand the many observations. The present review summarizes studies which have demonstrated that the hydrophobic interactions between a membrane protein...... and the host lipid bilayer provide an energetic coupling, whereby protein function can be regulated by the bilayer elasticity. The feasibility of this ‘hydrophobic coupling mechanism’ has been demonstrated using the gramicidin channel, a model membrane protein, in planar lipid bilayers. Using voltage...... properties experienced by an embedded protein has been developed. A theoretical and technological framework, to study the regulation of membrane protein function by lipid bilayer elasticity, has been established....

  8. Magnetically aligned phospholipid bilayers with positive ordering: a new model membrane system. (United States)

    Prosser, R S; Hwang, J S; Vold, R R


    A stable smectic phospholipid bilayer phase aligned with the director parallel to the magnetic field can be generated by the addition of certain trivalent paramagnetic lanthanide ions to a bicellar solution of dimyristoylphosphatidylcholine (DMPC) and dihexanoylphosphatidylcholine (DHPC) in water. Suitable lanthanide ions are those with positive anisotropy of their magnetic susceptibility, namely Eu3+, Er3+, Tm3+, and Yb3+. For samples doped with Tm3+, this phase extends over a wide range of Tm3+ concentrations (6-40 mM) and temperatures (35-90 degrees C) and appears to undergo a transition from a fluid nematic discotic to a fluid, but highly ordered, smectic phase at a temperature that depends on the thulium concentration. As a membrane mimetic, these new, positively ordered phospholipid phases have high potential for structural studies using a variety of techniques such as magnetic resonance (EMR and NMR), small-angle x-ray and neutron diffraction, as well as optical and infrared spectroscopy. PMID:9591667

  9. Application of self-consistent field theory to self-assembled bilayer membranes (United States)

    Zhang, Ping-Wen; Shi, An-Chang


    Bilayer membranes self-assembled from amphiphilic molecules such as lipids, surfactants, and block copolymers are ubiquitous in biological and physiochemical systems. The shape and structure of bilayer membranes depend crucially on their mechanical properties such as surface tension, bending moduli, and line tension. Understanding how the molecular properties of the amphiphiles determine the structure and mechanics of the self-assembled bilayers requires a molecularly detailed theoretical framework. The self-consistent field theory provides such a theoretical framework, which is capable of accurately predicting the mechanical parameters of self-assembled bilayer membranes. In this mini review we summarize the formulation of the self-consistent field theory, as exemplified by a model system composed of flexible amphiphilic chains dissolved in hydrophilic polymeric solvents, and its application to the study of self-assembled bilayer membranes. Project supported by the National Natural Science Foundation of China (Grant Nos. 11421101 and 21274005) and the Natural Sciences and Engineering Research Council (NSERC) of Canada.

  10. A comparative differential scanning calorimetry study of the effects of cholesterol and various oxysterols on the thermotropic phase behavior of dipalmitoylphosphatidylcholine bilayer membranes. (United States)

    Benesch, Matthew G K; McElhaney, Ronald N


    We have carried out a comparative differential scanning calorimetric (DSC) study of the effects of cholesterol (C) and the eight most physiologically relevant oxysterols on the thermotropic phase behavior of dipalmitoylphosphatidylcholine (DPPC) bilayer membranes. The structures of these oxysterols differ from that of C by the presence of additional hydroxyl, keto or epoxy groups on the steroid ring system or by the presence of a hydroxyl group in the alkyl side chain. In general, the progressive incorporation of these oxysterols reduces the temperature, cooperativity and enthalpy of the pretransition of DPPC to a greater extent than C, indicating that their presence thermally destabilizes and disorders the gel states of DPPC bilayers to a greater extent than C. Similarly, the incorporation of these oxysterols either increases the temperature of the broad component of the main phase transition to a smaller extent than C or actually decreases it. Again, this indicates that the presence of these compounds is less effective at thermally stabilizing and ordering the sterol-rich domains of DPPC bilayers than is C itself. Moreover, the incorporation of these oxysterols decrease the cooperativity and enthalpy of the main phase transition of DPPC to a smaller extent than C, indicating that they are somewhat less miscible in fluid DPPC bilayers than is C. Particularly notable in this regard is 25-hydroxycholesterol, which exhibits a markedly reduced miscibility in both gel and fluid DPPC bilayers compared to C itself. In general, the effectiveness of these oxysterols in stabilizing and ordering DPPC bilayers decreases as their rate of interbilayer exchange and the polarity of the oxysterol increases. We close by providing a tentative molecular explanation for the results of our DSC studies and of those of previous biophysical studies of the effects of various oxysterol on lipid bilayer model membranes. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  11. Strong influence of periodic boundary conditions on lateral diffusion in lipid bilayer membranes

    Energy Technology Data Exchange (ETDEWEB)

    Camley, Brian A. [Center for Theoretical Biological Physics and Department of Physics, University of California, San Diego, California 92093 (United States); Department of Physics, University of California, Santa Barbara, California 93106 (United States); Lerner, Michael G. [Department of Physics and Astronomy, Earlham College, Richmond, Indiana 47374 (United States); Laboratory of Computational Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892 (United States); Pastor, Richard W. [Laboratory of Computational Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892 (United States); Brown, Frank L. H. [Department of Physics, University of California, Santa Barbara, California 93106 (United States); Department of Chemistry and Biochemistry, University of California, Santa Barbara, California 93106 (United States)


    The Saffman-Delbrück hydrodynamic model for lipid-bilayer membranes is modified to account for the periodic boundary conditions commonly imposed in molecular simulations. Predicted lateral diffusion coefficients for membrane-embedded solid bodies are sensitive to box shape and converge slowly to the limit of infinite box size, raising serious doubts for the prospects of using detailed simulations to accurately predict membrane-protein diffusivities and related transport properties. Estimates for the relative error associated with periodic boundary artifacts are 50% and higher for fully atomistic models in currently feasible simulation boxes. MARTINI simulations of LacY membrane protein diffusion and LacY dimer diffusion in DPPC membranes and lipid diffusion in pure DPPC bilayers support the underlying hydrodynamic model.

  12. Strong influence of periodic boundary conditions on lateral diffusion in lipid bilayer membranes (United States)

    Camley, Brian A.; Lerner, Michael G.; Pastor, Richard W.; Brown, Frank L. H.


    The Saffman-Delbrück hydrodynamic model for lipid-bilayer membranes is modified to account for the periodic boundary conditions commonly imposed in molecular simulations. Predicted lateral diffusion coefficients for membrane-embedded solid bodies are sensitive to box shape and converge slowly to the limit of infinite box size, raising serious doubts for the prospects of using detailed simulations to accurately predict membrane-protein diffusivities and related transport properties. Estimates for the relative error associated with periodic boundary artifacts are 50% and higher for fully atomistic models in currently feasible simulation boxes. MARTINI simulations of LacY membrane protein diffusion and LacY dimer diffusion in DPPC membranes and lipid diffusion in pure DPPC bilayers support the underlying hydrodynamic model.

  13. Solid-supported polymer bilayers as membrane mimics


    Belegrinou, Serena


    Membranes are one of Nature’s most remarkable designs. Due to their importance in numerous cellular processes, they are prominent subjects of biochemical and biophysical fundamental research. In particular, it is crucial to understand the membrane morphology, the role of individual membrane components, and also to correlate the membrane structure to its various functions. Besides, systems inspired by natural membranes are of high interest for technological applications, such as water purifica...

  14. Ethylene glycol as bore fluid for hollow fiber membrane preparation

    KAUST Repository

    Le, Ngoc Lieu


    We proposed the use of ethylene glycol and its mixture with water as bore fluid for the preparation of poly(ether imide) (PEI) hollow fiber membranes and compared their performance and morphology with membranes obtained with conventional coagulants (water and its mixture with the solvent N-methylpyrrolidone (NMP)). Thermodynamics and kinetics of the systems were investigated. Water and 1:1 water:EG mixtures lead to fast precipitation rates. Slow precipitation is observed for both pure EG and 9:1 NMP:water mixture, but the reasons for that are different. While low osmotic driving force leads to slow NMP and water transport when NMP:water is used, the high EG viscosity is the reason for the slow phase separation when EG is the bore fluid. The NMP:water mixture produces fibers with mixed sponge-like and finger-like structure with large pores in the inner and outer layers; and hence leading to a high water permeance and a high MWCO suitable for separation of large-sized proteins. As compared to NMP:water, using EG as bore fluid provides fibers with a finger-like bilayered structure and sponge-like layers near the surfaces, and hence contributing to the higher water permeance. It also induces small pores for better protein rejection.

  15. Quantitative optical microscopy and micromanipulation studies on the lipid bilayer membranes of giant unilamellar vesicles

    DEFF Research Database (Denmark)

    Bagatolli, Luis; Needham, David


    some of their most important contributions to our understanding of lipid bilayer membranes; and (iii) outline studies that would utilize both techniques simultaneously on the same vesicle thus bringing the ability to characterize structure and strain responses together with the direct application......This manuscript discusses basic methodological aspects of optical microscopy and micromanipulation methods to study membranes and reviews methods to generate giant unilamellar vesicles (GUVs). In particular, we focus on the use of fluorescence microscopy and micropipette manipulation techniques...... to study composition-structure-property materials relationships of free-standing lipid bilayer membranes. Because their size (~5 to 100 m diameter) that is well above the resolution limit of regular light microscopes, GUVs are suitable membrane models for optical microscopy and micromanipulation...

  16. [Effect of microwaves on bilayer lipid membranes: role of a membrane-forming hole in the Teflon film]. (United States)

    Alekseev, S I; Ziskin, M S; Fesenko, E E


    The distributions of specific abcorption rate (SAR) and E-field in a membrane-forming hole of Teflon film and surrounding electrolyte were calculated for 0.9 GHz exposure. It was found that the specific absorption rate in the membrane-forming hole increased greatly with increasing thickness of the Teflon film, and electrolyte concentration and decreasing diameter of the hole. The previously demonstrated significant changes in the conductivity of modified bilayer lipid membranes induced by microwave exposure can be explained by a local increase in specific absorption rate and subsequent elevation of temperature in the membrane-forming hole of the Teflon film.

  17. Effect of the dipole potential of a bilayer lipid membrane on gramicidin channel dissociation kinetics.


    Rokitskaya, T I; Antonenko, Y N; Kotova, E A


    A technique of measuring of the light-induced transients of the gramicidin-mediated electric current across a membrane in the presence of a photosensitizer has been applied for the study of the effect of agents modifying the dipole potential of a bilayer lipid membrane (phloretin, 6-ketocholestanol, and RH421) on the processes of the gramicidin channel dissociation and formation. It is shown that phloretin, known to lower the dipole potential, decelerates the flash-induced decrease in the cur...

  18. Undulation instability in a bilayer lipid membrane due to electric field interaction with lipid dipoles

    CERN Document Server

    Bingham, Richard J; Smye, Stephen W


    Bilayer lipid membranes [BLMs] are an essential component of all biological systems, forming a functional barrier for cells and organelles from the surrounding environment. The lipid molecules that form membranes contain both permanent and induced dipoles, and an electric field can induce the formation of pores when the transverse field is sufficiently strong (electroporation). Here, a phenomenological free energy is constructed to model the response of a BLM to a transverse static electric field. The model contains a continuum description of the membrane dipoles and a coupling between the headgroup dipoles and the membrane tilt. The membrane is found to become unstable through buckling modes, which are weakly coupled to thickness fluctuations in the membrane. The thickness fluctuations, along with the increase in interfacial area produced by membrane buckling, increase the probability of localized membrane breakdown, which may lead to pore formation. The instability is found to depend strongly on the strengt...

  19. Fluorescent molecular probes based on excited state prototropism in lipid bilayer membrane (United States)

    Mohapatra, Monalisa; Mishra, Ashok K.


    Excited state prototropism (ESPT) is observed in molecules having one or more ionizable protons, whose proton transfer efficiency is different in ground and excited states. The interaction of various ESPT molecules like naphthols and intramolecular ESPT (ESIPT) molecules like hydroxyflavones etc. with different microheterogeneous media have been studied in detail and excited state prototropism as a probe concept has been gaining ground. The fluorescence of different prototropic forms of such molecules, on partitioning to an organized medium like lipid bilayer membrane, often show sensitive response to the local environment with respect to the local structure, physical properties and dynamics. Our recent work using 1-naphthol as an ESPT fluorescent molecular probe has shown that the incorporation of monomeric bile salt molecules into lipid bilayer membranes composed from dipalmitoylphosphatidylcholine (DPPC, a lung surfactant) and dimyristoylphosphatidylcholine (DMPC), in solid gel and liquid crystalline phases, induce appreciable wetting of the bilayer up to the hydrocarbon core region, even at very low (fisetin, an ESIPT molecule having antioxidant properties, in lipid bilayer membrane has been sensitively monitored from its intrinsic fluorescence behaviour.

  20. Lipid domains control myelin basic protein adsorption and membrane interactions between model myelin lipid bilayers. (United States)

    Lee, Dong Woog; Banquy, Xavier; Kristiansen, Kai; Kaufman, Yair; Boggs, Joan M; Israelachvili, Jacob N


    The surface forces apparatus and atomic force microscope were used to study the effects of lipid composition and concentrations of myelin basic protein (MBP) on the structure of model lipid bilayers, as well as the interaction forces and adhesion between them. The lipid bilayers had a lipid composition characteristic of the cytoplasmic leaflets of myelin from "normal" (healthy) and "disease-like" [experimental allergic encephalomyelitis (EAE)] animals. They showed significant differences in the adsorption mechanism of MBP. MBP adsorbs on normal bilayers to form a compact film (3-4 nm) with strong intermembrane adhesion (∼0.36 mJ/m(2)), in contrast to its formation of thicker (7-8 nm) swelled films with weaker intermembrane adhesion (∼0.13 mJ/m(2)) on EAE bilayers. MBP preferentially adsorbs to liquid-disordered submicron domains within the lipid membranes, attributed to hydrophobic attractions. These results show a direct connection between the lipid composition of membranes and membrane-protein adsorption mechanisms that affects intermembrane spacing and adhesion and has direct implications for demyelinating diseases.

  1. Folding of β-barrel membrane proteins in lipid bilayers - Unassisted and assisted folding and insertion. (United States)

    Kleinschmidt, Jörg H


    In cells, β-barrel membrane proteins are transported in unfolded form to an outer membrane into which they fold and insert. Model systems have been established to investigate the mechanisms of insertion and folding of these versatile proteins into detergent micelles, lipid bilayers and even synthetic amphipathic polymers. In these experiments, insertion into lipid membranes is initiated from unfolded forms that do not display residual β-sheet secondary structure. These studies therefore have allowed the investigation of membrane protein folding and insertion in great detail. Folding of β-barrel membrane proteins into lipid bilayers has been monitored from unfolded forms by dilution of chaotropic denaturants that keep the protein unfolded as well as from unfolded forms present in complexes with molecular chaperones from cells. This review is aimed to provide an overview of the principles and mechanisms observed for the folding of β-barrel transmembrane proteins into lipid bilayers, the importance of lipid-protein interactions and the function of molecular chaperones and folding assistants. This article is part of a Special Issue entitled: Lipid-protein interactions.

  2. Bilayer lipid membrane (BLM) based ion selective electrodes at the meso-, micro-, and nano-scales. (United States)

    Liu, Bingwen; Rieck, Daniel; Van Wie, Bernard J; Cheng, Gary J; Moffett, David F; Kidwell, David A


    This paper presents a novel method for making micron-sized apertures with tapered sidewalls and nano-sized apertures. Their use in bilayer lipid membrane-based ion selective electrode design is demonstrated and compared to mesoscale bilayers and traditional PVC ion selective electrodes. Micron-sized apertures are fabricated in SU-8 photoresist films and vary in diameter from 10 to 40 microm. The tapered edges in SU-8 films are desired to enhance bilayer lipid membrane (BLM) formation and are fabricated by UV-light overexposure. Nano-apertures are made in boron diffused silicon film. The membranes are used as septa to separate two potassium chloride solutions of different concentrations. Lecithin BLMs are assembled on the apertures by ejecting lipid solution. Potassium ionophore, dibenzo-18-crown-6, is incorporated into BLMs by dissolving it in the lipid solution before membrane assembly. Voltage changes with increasing potassium ion concentrations are recorded with an A/D converter. Various ionophore concentrations in BLMs are investigated. At least a 1% concentration is needed for consistent slopes. Electrode response curves are linear over the 10(-6) to 0.1M range with a sub-Nernstian slope of 20mV per Log concentration change. This system shows high selectivity to potassium ions over potential interfering sodium ions. BLMs on the three different aperture sizes at the meso-, micro-, and nano-scales all show similar linear ranges and limits of detection (LODs) as PVC ion selective membranes.

  3. Eicosapentaenoic acid reduces membrane fluidity, inhibits cholesterol domain formation, and normalizes bilayer width in atherosclerotic-like model membranes. (United States)

    Mason, R Preston; Jacob, Robert F; Shrivastava, Sandeep; Sherratt, Samuel C R; Chattopadhyay, Amitabha


    Cholesterol crystalline domains characterize atherosclerotic membranes, altering vascular signaling and function. Omega-3 fatty acids reduce membrane lipid peroxidation and subsequent cholesterol domain formation. We evaluated non-peroxidation-mediated effects of eicosapentaenoic acid (EPA), other TG-lowering agents, docosahexaenoic acid (DHA), and other long-chain fatty acids on membrane fluidity, bilayer width, and cholesterol domain formation in model membranes. In membranes prepared at 1.5:1 cholesterol-to-phospholipid (C/P) mole ratio (creating pre-existing domains), EPA, glycyrrhizin, arachidonic acid, and alpha linolenic acid promoted the greatest reductions in cholesterol domains (by 65.5%, 54.9%, 46.8%, and 45.2%, respectively) compared to controls; other treatments had modest effects. EPA effects on cholesterol domain formation were dose-dependent. In membranes with 1:1 C/P (predisposing domain formation), DHA, but not EPA, dose-dependently increased membrane fluidity. DHA also induced cholesterol domain formation without affecting temperature-induced changes in-bilayer unit cell periodicity relative to controls (d-space; 57Å-55Å over 15-30°C). Together, these data suggest simultaneous formation of distinct cholesterol-rich ordered domains and cholesterol-poor disordered domains in the presence of DHA. By contrast, EPA had no effect on cholesterol domain formation and produced larger d-space values relative to controls (60Å-57Å; pmembrane bilayer width, membrane fluidity, and cholesterol crystalline domain formation; suggesting omega-3 fatty acids with differing chain length or unsaturation may differentially influence membrane lipid dynamics and structural organization as a result of distinct phospholipid/sterol interactions.

  4. Effects of butanol isomers on dipalmitoylphosphatidylcholine bilayer membranes. (United States)

    Reeves, Megan D; Schawel, Adam K; Wang, Weidong; Dea, Phoebe


    Differential scanning calorimetry and (31)P-NMR were used to study the effects of butanol isomers on the thermotropic phase behavior of dipalmitoylphosphatidylcholine (DPPC) bilayers. The threshold concentration for the onset of interdigitation for each isomer was determined by the disappearance of the pretransition and the onset of a large hysteresis between the heating and cooling scans of the gel-to-liquid main transition. The threshold concentration was found to correlate with increased solubility of the isomers in the aqueous phase, led by tert-butanol. However, as the solution concentration of tert-butanol increased, there was an abrupt shrinking of the hysteresis, initially with well-resolved shoulder peaks indicating mixed phases. The eventual disappearance of the shoulder peaks was correlated with a breakdown of the multilamellar structure identified using (31)P-NMR.

  5. Modeling Yeast Organelle Membranes and How Lipid Diversity Influences Bilayer Properties. (United States)

    Monje-Galvan, Viviana; Klauda, Jeffery B


    Membrane lipids are important for the health and proper function of cell membranes. We have improved computational membrane models for specific organelles in yeast Saccharomyces cerevisiae to study the effect of lipid diversity on membrane structure and dynamics. Previous molecular dynamics simulations were performed by Jo et al. [(2009) Biophys J. 97, 50-58] on yeast membrane models having six lipid types with compositions averaged between the endoplasmic reticulum (ER) and the plasma membrane (PM). We incorporated ergosterol, phosphatidic acid, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol lipids in our models to better describe the unique composition of the PM, ER, and trans-Golgi network (TGN) bilayers of yeast. Our results describe membrane structure based on order parameters (SCD), electron density profiles (EDPs), and lipid packing. The average surface area per lipid decreased from 63.8 ± 0.4 Å(2) in the ER to 47.1 ± 0.3 Å(2) in the PM, while the compressibility modulus (KA) varied in the opposite direction. The high SCD values for the PM lipids indicated a more ordered bilayer core, while the corresponding lipids in the ER and TGN models had lower parameters by a factor of at least 0.7. The hydrophobic core thickness (2DC) as estimated from EDPs is the thickest for PM, which is in agreement with estimates of hydrophobic regions of transmembrane proteins from the Orientation of Proteins in Membranes database. Our results show the importance of lipid diversity and composition on a bilayer's structural and mechanical properties, which in turn influences interactions with the proteins and membrane-bound molecules.

  6. Advanced Flicker Spectroscopy of Fluid Membranes (United States)

    Döbereiner, Hans-Günther; Gompper, Gerhard; Haluska, Christopher; Kroll, Daniel; Petrov, Peter; Riske, Karin


    The bending elasticity of a fluid membrane is characterized by its modulus and spontaneous curvature. We present a new method, advanced flicker spectroscopy of giant nonspherical vesicles, which makes it possible to simultaneously measure both parameters for the first time. Our analysis is based on the generation of a large set of reference data from Monte Carlo simulations of randomly triangulated surfaces. As an example of the potential of the procedure, we monitor thermal trajectories of vesicle shapes and discuss the elastic response of zwitterionic membranes to transmembrane pH gradients. Our technique makes it possible to easily characterize membrane curvature as a function of environmental conditions.

  7. NMR structural studies of the bacterial outer membrane protein OmpX in oriented lipid bilayer membranes. (United States)

    Mahalakshmi, Radhakrishnan; Franzin, Carla M; Choi, Jungyuen; Marassi, Francesca M


    The beta-barrels found in the outer membranes of prokaryotic and eukaryotic organisms constitute an important functional class of proteins. Here we present solid-state NMR spectra of the bacterial outer membrane protein OmpX in oriented lipid bilayer membranes. We show that OmpX is folded in both glass-supported oriented lipid bilayers and in lipid bicelles that can be magnetically oriented with the membrane plane parallel or perpendicular to the direction of the magnetic field. The presence of resolved peaks in these spectra demonstrates that OmpX undergoes rotational diffusion around an axis perpendicular to the membrane surface. A tightly hydrogen-bonded domain of OmpX resists exchange with D2O for days and is assigned to the transmembrane beta-barrel, while peaks at isotropic resonance frequencies that disappear rapidly in D2O are assigned to the extracellular and periplasmic loops. The two-dimensional 1H/15N separated local field spectra of OmpX have several resolved peaks, and agree well with the spectra calculated from the crystal structure of OmpX rotated with the barrel axis nearly parallel (5 degrees tilt) to the direction of the magnetic field. The data indicate that it will be possible to obtain site-specific resonance assignments and to determine the structure, tilt, and rotation of OmpX in membranes using the solid-state NMR methods that are currently being applied to alpha-helical membrane proteins.

  8. Affinity of four polar neurotransmitters for lipid bilayer membranes

    DEFF Research Database (Denmark)

    Wang, Chunhua; Ye, Fengbin; Valardez, Gustavo F.


    Weak interactions of neurotransmitters and the lipid matrix in the synaptic membrane have been hypothesized to play a role in synaptic transmission of nerve signals, particularly with respect to receptor desensitization (Cantor, R. S. Biochemistry 2003, 42, 11891). The strength of such interactions......, however, was not measured, and this is an obvious impediment for further evaluation and understanding of a possible role for desensitization. We have used dialysis equilibrium to directly measure the net affinity of selected neurotransmitters for lipid membranes and analyzed this affinity data...... with respect to calorimetric measurements and molecular dynamics simulations. We studied an anionic (glutamate), a cationic (acetylcholine), and two zwitterionic (-aminobutyric acid and glycine) neurotransmitters, and membranes of pure dimyristoyl phosphatidylcholine (DMPC), DMPC doped with 10% anionic lipid...

  9. Molecular dynamics simulations of the interactions of medicinal plant extracts and drugs with lipid bilayer membranes

    DEFF Research Database (Denmark)

    Kopec, Wojciech; Telenius, Jelena; Khandelia, Himanshu


    Several small drugs and medicinal plant extracts, such as the Indian spice extract curcumin, have a wide range of useful pharmacological properties that cannot be ascribed to binding to a single protein target alone. The lipid bilayer membrane is thought to mediate the effects of many...... studies of the interactions of drugs and plant extracts are therefore of interest. Molecular dynamics simulations, which can access time and length scales that are not simultaneously accessible by other experimental methods, are often used to obtain quantitative molecular and thermodynamic descriptions...... such molecules directly via perturbation of the plasma membrane structure and dynamics, or indirectly by modulating transmembrane protein conformational equilibria. Furthermore, for bioavailability, drugs must interact with and eventually permeate the lipid bilayer barrier on the surface of cells. Biophysical...

  10. JNK3 phosphorylates Bax protein and induces ability to form pore on bilayer lipid membrane

    Directory of Open Access Journals (Sweden)

    Rajeev Gupta


    Full Text Available Bax is a pro-apoptotic cytosolic protein. In this work native (unphosphorylated and JNK3 phosphorylated Bax proteins are studied on artificial bilayer membranes for pore formation. Phosphorylated Bax formed pore on the bilayer lipid membrane whereas native one does not. In cells undergoing apoptosis the pore formed by the phosphorylated Bax could be important in cytochrome c release from the mitochondrial intermembrane space to the cytosol. The low conductance (1.5 nS of the open state of the phosphorylated Bax pore corresponds to pore diameter of 0.9 nm which is small to release cytochrome c (∼3.4 nm. We hypothesized that JNK3 phosphorylated Bax protein can form bigger pores after forming complexes with other mitochondrial proteins like VDAC, t-Bid etc. to release cytochrome c.

  11. The Effect of Lidocaine · HCl on the Fluidity of Native and Model Membrane Lipid Bilayers. (United States)

    Park, Jun-Seop; Jung, Tae-Sang; Noh, Yang-Ho; Kim, Woo-Sung; Park, Won-Ick; Kim, Young-Soo; Chung, In-Kyo; Sohn, Uy Dong; Bae, Soo-Kyung; Bae, Moon-Kyoung; Jang, Hye-Ock; Yun, Il


    The purpose of this study is to investigated the mechanism of pharmacological action of local anesthetic and provide the basic information about the development of new effective local anesthetics. Fluorescent probe techniques were used to evaluate the effect of lidocaine·HCl on the physical properties (transbilayer asymmetric lateral and rotational mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex, and liposomes of total lipids (SPMVTL) and phospholipids (SPMVPL) extracted from the SPMV. An experimental procedure was used based on selective quenching of 1,3-di(1-pyrenyl)propane (Py-3-Py) and 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups, and radiationless energy transfer from the tryptophans of membrane proteins to Py-3-Py. Lidocaine·HCl increased the bulk lateral and rotational mobility of neuronal and model membrane lipid bilayes, and had a greater fluidizing effect on the inner monolayer than the outer monolayer. Lidocaine·HCl increased annular lipid fluidity in SPMV lipid bilayers. It also caused membrane proteins to cluster. The most important finding of this study is that there is far greater increase in annular lipid fluidity than that in lateral and rotational mobilities by lidocaine·HCl. Lidocaine·HCl alters the stereo or dynamics of the proteins in the lipid bilayers by combining with lipids, especially with the annular lipids. In conclusion, the present data suggest that lidocaine, in addition to its direct interaction with proteins, concurrently interacts with membrane lipids, fluidizing the membrane, and thus inducing conformational changes of proteins known to be intimately associated with membrane lipid.

  12. Cyclotides insert into lipid bilayers to form membrane pores and destabilize the membrane through hydrophobic and phosphoethanolamine-specific interactions. (United States)

    Wang, Conan K; Wacklin, Hanna P; Craik, David J


    Cyclotides are a family of plant-derived circular proteins with potential therapeutic applications arising from their remarkable stability, broad sequence diversity, and range of bioactivities. Their membrane-binding activity is believed to be a critical component of their mechanism of action. Using isothermal titration calorimetry, we studied the binding of the prototypical cyclotides kalata B1 and kalata B2 (and various mutants) to dodecylphosphocholine micelles and phosphoethanolamine-containing lipid bilayers. Although binding is predominantly an entropy-driven process, suggesting that hydrophobic forces contribute significantly to cyclotide-lipid complex formation, specific binding to the phosphoethanolamine-lipid headgroup is also required, which is evident from the enthalpic changes in the free energy of binding. In addition, using a combination of dissipative quartz crystal microbalance measurements and neutron reflectometry, we elucidated the process by which cyclotides interact with bilayer membranes. Initially, a small number of cyclotides bind to the membrane surface and then insert first into the outer membrane leaflet followed by penetration through the membrane and pore formation. At higher concentrations of cyclotides, destabilization of membranes occurs. Our results provide significant mechanistic insight into how cyclotides exert their bioactivities.

  13. Photoelectric Effects in Lipid Bilayer Membranes. A Pedagogical Review. (United States)

    Huebner, Jay S.; And Others


    Provides information appropriate for introductory lectures on photoelectric effects in membranes. Describes the apparatus and supplies required for laboratory exercises. Outlines typical laboratory exercises. Identifies the chromophores known to induce photoelectric effects. Concludes that this topic can provide useful subjects for undergraduate…

  14. Membrane order parameters for interdigitated lipid bilayers measured via polarized total-internal-reflection fluorescence microscopy. (United States)

    Ngo, An T; Jakubek, Zygmunt J; Lu, Zhengfang; Joós, Béla; Morris, Catherine E; Johnston, Linda J


    Incorporating ethanol in lipid membranes leads to changes in bilayer structure, including the formation of an interdigitated phase. We have used polarized total-internal-reflection fluorescence microscopy (pTIRFM) to measure the order parameter for Texas Red DHPE incorporated in the ethanol-induced interdigitated phase (LβI) formed from ternary lipid mixtures comprising dioleoylphosphatidylcholine, cholesterol and egg sphingomyelin or dipalmitoylphosphatidylcholine. These lipid mixtures have 3 co-existing phases in the presence of ethanol: liquid-ordered, liquid-disordered and LβI. pTIRFM using Texas Red DHPE shows a reversal in fluorescence contrast between the LβI phase and the surrounding disordered phase with changes in the polarization angle. The contrast reversal is due to changes in the orientation of the dye, and provides a rapid method to identify the LβI phase. The measured order parameters for the LβI phase are consistent with a highly ordered membrane environment, similar to a gel phase. An acyl-chain labeled BODIPY-FL-PC was also tested for pTIRFM studies of ethanol-treated bilayers; however, this probe is less useful since the order parameters of the interdigitated phase are consistent with orientations that are close to random, either due to local membrane disorder or to a mixture of extended and looping conformations in which the fluorophore is localized in the polar headgroup region of the bilayer. In summary, we demonstrate that order parameter measurements via pTIRFM using Texas Red-DHPE can rapidly identify the interdigitated phase in supported bilayers. We anticipate that this technique will aid further research in the effects of alcohols and other additives on membranes.

  15. Bilayer vesicles of amphiphilic cyclodextrins: host membranes that recognize guest molecules. (United States)

    Falvey, Patrick; Lim, Choon Woo; Darcy, Raphael; Revermann, Tobias; Karst, Uwe; Giesbers, Marcel; Marcelis, Antonius T M; Lazar, Adina; Coleman, Anthony W; Reinhoudt, David N; Ravoo, Bart Jan


    A family of amphiphilic cyclodextrins (6, 7) has been prepared through 6-S-alkylation (alkyl=n-dodecyl and n-hexadecyl) of the primary side and 2-O-PEGylation of the secondary side of alpha-, beta-, and gamma-cyclodextrins (PEG=poly(ethylene glycol)). These cyclodextrins form nonionic bilayer vesicles in aqueous solution. The bilayer vesicles were characterized by transmission electron microscopy, dynamic light scattering, dye encapsulation, and capillary electrophoresis. The molecular packing of the amphiphilic cyclodextrins was investigated by using small-angle X-ray diffraction of bilayers deposited on glass and pressure-area isotherms obtained from Langmuir monolayers on the air-water interface. The bilayer thickness is dependent on the chain length, whereas the average molecular surface area scales with the cyclodextrin ring size. The alkyl chains of the cyclodextrins in the bilayer are deeply interdigitated. Molecular recognition of a hydrophobic anion (adamantane carboxylate) by the cyclodextrin vesicles was investigated by using capillary electrophoresis, thereby exploiting the increase in electrophoretic mobility that occurs when the hydrophobic anions bind to the nonionic cyclodextrin vesicles. It was found that in spite of the presence of oligo(ethylene glycol) substituents, the beta-cyclodextrin vesicles retain their characteristic affinity for adamantane carboxylate (association constant K(a)=7.1 x 10(3) M(-1)), whereas gamma-cyclodextrin vesicles have less affinity (K(a)=3.2 x 10(3) M(-1)), and alpha-cyclodextrin or non-cyclodextrin, nonionic vesicles have very little affinity (K(a) approximately 100 M(-1)). Specific binding of the adamantane carboxylate to beta-cyclodextrin vesicles was also evident in competition experiments with beta-cyclodextrin in solution. Hence, the cyclodextrin vesicles can function as host bilayer membranes that recognize small guest molecules by specific noncovalent interaction.

  16. Reparameterization of all-atom dipalmitoylphosphatidylcholine lipid parameters enables simulation of fluid bilayers at zero tension

    DEFF Research Database (Denmark)

    Sonne, Jacob; Jensen, M.Ø.; Hansen, Flemming Yssing;


    represented by the CHARMM energy function in this ensemble, we reparameterized the atomic partial charges in the lipid headgroup and upper parts of the acyl chains. The new charges were determined from the electron structure using both the Mulliken method and the restricted electrostatic potential fitting...... method. We tested the derived charges in molecular dynamics simulations of a fully hydrated DPPC bilayer. Only the simulation with the new restricted electrostatic potential charges shows significant improvements compared with simulations using the original CHARMM27 force field resulting in an area per...... fluid phase of DPPC bilayers can now be simulated in all-atom simulations in the NPT ensemble by employing our modified CHARMM27 force field....

  17. Formation and fluidity measurement of supported lipid bilayer on polyvinyl chloride membrane (United States)

    Kobayashi, Takuji; Kono, Akiteru; Futagawa, Masato; Sawada, Kazuaki; Tero, Ryugo


    We prepared an artificial lipid bilayer on a plasticized poly(vinyl chloride) (PVC) membrane on a Si3N4 layer deposited on a Si wafer. We optimized the experimental condition for the fabrication of the PVC membrane, and obtained a PVC membrane with a flat and uniform surface on the scale of several hundreds of micrometer suitable for a substrate for supported lipid bilayers (SLBs). The SLB of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) was formed on the PVC membrane by the vesicle fusion method. The observation with a conventional epi-fluorescence microscope and a confocal laser scanning microscope gave geometrically uniform images of the SLB on the PVC membrane. The fluidity and the mobile fraction of the SLB was evaluated by the fluorescence recovery after photobleaching method, and compared with that on a thermally oxidized SiO2/Si substrate. The SLB on the PVC membrane contained immobile fraction ˜30%, but the diffusion in the mobile fraction was two times faster than that in the SLB on SiO2/Si, which had little immobile fraction.

  18. Formation and fluidity measurement of supported lipid bilayer on polyvinyl chloride membrane

    Energy Technology Data Exchange (ETDEWEB)

    Kobayashi, Takuji, E-mail:; Kono, Akiteru, E-mail:; Sawada, Kazuaki [Department of Electrical and Electronic Information Engineering, Toyohashi University of Technology, 1-1 Hibarigaoka Tempaku-cho, Toyohashi, 441-8580 (Japan); Futagawa, Masato [Department of Electrical and Electronic Information Engineering and Head Office for the Tailor-Made and Baton-Zone Graduate Course, Toyohashi University of Technology, 1-1 Hibarigaoka Tempaku-cho, Toyohashi, 441-8580 (Japan); Tero, Ryugo, E-mail: [Electronics-Inspired Interdisciplinary Research Institute and Department of Environmental and Life Sciences, Toyohashi University of Technology, 1-1 Hibarigaoka Tempaku-cho, Toyohashi, 441-8580 (Japan)


    We prepared an artificial lipid bilayer on a plasticized poly(vinyl chloride) (PVC) membrane on a Si3N4 layer deposited on a Si wafer. We optimized the experimental condition for the fabrication of the PVC membrane, and obtained a PVC membrane with a flat and uniform surface on the scale of several hundreds of micrometer suitable for a substrate for supported lipid bilayers (SLBs). The SLB of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) was formed on the PVC membrane by the vesicle fusion method. The observation with a conventional epi-fluorescence microscope and a confocal laser scanning microscope gave geometrically uniform images of the SLB on the PVC membrane. The fluidity and the mobile fraction of the SLB was evaluated by the fluorescence recovery after photobleaching method, and compared with that on a thermally oxidized SiO{sub 2}/Si substrate. The SLB on the PVC membrane contained immobile fraction ∼30%, but the diffusion in the mobile fraction was two times faster than that in the SLB on SiO{sub 2}/Si, which had little immobile fraction.

  19. Lanthanide Chelates as Bilayer Alignment Tools in NMR Studies of Membrane-Associated Peptides (United States)

    Prosser, R. S.; Bryant, H.; Bryant, R. G.; Vold, Regitze R.


    Theequimolar complex, consisting of the lipid-like, amphiphilic chelating agent 1,11-bis[distearylamino]-diethylenetriamine pentaacetic acid (DTPA-18) and Tm3+, is shown by deuterium (2H) NMR to be useful in aligning bicelle-like model membranes, consisting of dimyristoylphosphatidylcholine (DMPC) and dihexanoylphosphatidylcholine (DHPC). As shown previously (1996, R. S. Prosser et al., J. Am. Chem. Soc. 118, 269-270), in the absence of chelate, the lanthanide ions bind loosely with the lipid phosphate groups and confer the membrane with a sufficient positive magnetic anisotropy to result in parallel alignment (i.e., average bilayer normal along the field). Apparently, DTPA-18 sequesters the lanthanide ions and inserts into the phospholipid bilayer in such a manner that bilayer morphology is preserved over a wide temperature range (35-70°C). The inherent paramagnetic shifts and line broadening effects are illustrated by 2H NMR spectra of the membrane binding peptide, Leu-enkephalin (Lenk-d2, Tyr-(Gly-d2)-Gly-Phe-Leu-OH), in the presence of varying concentrations of Tm3+, and upon addition of DTPA-18. Two conclusions could be drawn from this study: (1) The addition of Tm3+ to the bicelle system is consistent with a conformational change in the surface associated peptide, and this effect is shown to be reversed by addition of the chelate, and (2) The paramagnetic shifts are shown to be significantly reduced by addition of chelate.

  20. Defining the Free-Energy Landscape of Curvature-Inducing Proteins on Membrane Bilayers

    CERN Document Server

    Tourdot, Richard W; Radhakrishnan, Ravi


    Curvature-sensing and curvature-remodeling proteins are known to reshape cell membranes, and this remodeling event is essential for key biophysical processes such as tubulation, exocytosis, and endocytosis. Curvature-inducing proteins can act as curvature sensors as well as induce curvature in cell membranes to stabilize emergent high curvature, non-spherical, structures such as tubules, discs, and caveolae. A definitive understanding of the interplay between protein recruitment and migration, the evolution of membrane curvature, and membrane morphological transitions is emerging but remains incomplete. Here, within a continuum framework and using the machinery of Monte Carlo simulations, we introduce and compare three free-energy methods to delineate the free-energy landscape of curvature-inducing proteins on bilayer membranes. We demonstrate the utility of the Widom test-particle/field insertion methodology in computing the excess chemical potentials associated with curvature-inducing proteins on the membra...

  1. A New Method for Measuring Edge Tensions and Stability of Lipid Bilayers: Effect of Membrane Composition

    CERN Document Server

    Portet, Thomas; 10.1016/j.bpj.2010.09.032


    We report a new and facile method for measuring edge tensions of lipid membranes. The approach is based on electroporation of giant unilamellar vesicles and analysis of the pore closure dynamics. We applied this method to evaluate the edge tension in membranes with four different compositions: egg phosphatidylcholine (EggPC), dioleoylphosphatidylcholine (DOPC), and mixtures of the latter with cholesterol and dioleoylphosphatidylethanolamine (DOPE). Our data confirm previous results for EggPC and DOPC. The addition of 17 mol % cholesterol to the DOPC membrane causes an increase in the membrane edge tension. On the contrary, when the same fraction of DOPE is added to the membrane, a decrease in the edge tension is observed, which is an unexpected result considering the inverted-cone shape geometry of the molecule. Presumably, interlipid hydrogen bonding lies in the origin of this behavior. Furthermore, cholesterol was found to lower the lysis tension of DOPC bilayers. This behavior differs from that observed on...

  2. Lipid bilayer-bound conformation of an integral membrane beta barrel protein by multidimensional MAS NMR

    Energy Technology Data Exchange (ETDEWEB)

    Eddy, Matthew T. [The Scripps Research Institute, Department of Integrative Structural and Computational Biology (United States); Su, Yongchao; Silvers, Robert; Andreas, Loren; Clark, Lindsay [Massachusetts Institute of Technology, Department of Chemistry (United States); Wagner, Gerhard [Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology (United States); Pintacuda, Guido; Emsley, Lyndon [Université de Lyon, Centre de RMN à Très Hauts Champs, Institut des Sciences Analytiques (CNRS, ENS Lyon, UCB Lyon 1) (France); Griffin, Robert G., E-mail: [Massachusetts Institute of Technology, Department of Chemistry (United States)


    The human voltage dependent anion channel 1 (VDAC) is a 32 kDa β-barrel integral membrane protein that controls the transport of ions across the outer mitochondrial membrane. Despite the determination of VDAC solution and diffraction structures, a structural basis for the mechanism of its function is not yet fully understood. Biophysical studies suggest VDAC requires a lipid bilayer to achieve full function, motivating the need for atomic resolution structural information of VDAC in a membrane environment. Here we report an essential step toward that goal: extensive assignments of backbone and side chain resonances for VDAC in DMPC lipid bilayers via magic angle spinning nuclear magnetic resonance (MAS NMR). VDAC reconstituted into DMPC lipid bilayers spontaneously forms two-dimensional lipid crystals, showing remarkable spectral resolution (0.5–0.3 ppm for {sup 13}C line widths and <0.5 ppm {sup 15}N line widths at 750 MHz). In addition to the benefits of working in a lipid bilayer, several distinct advantages are observed with the lipid crystalline preparation. First, the strong signals and sharp line widths facilitated extensive NMR resonance assignments for an integral membrane β-barrel protein in lipid bilayers by MAS NMR. Second, a large number of residues in loop regions were readily observed and assigned, which can be challenging in detergent-solubilized membrane proteins where loop regions are often not detected due to line broadening from conformational exchange. Third, complete backbone and side chain chemical shift assignments could be obtained for the first 25 residues, which comprise the functionally important N-terminus. The reported assignments allow us to compare predicted torsion angles for VDAC prepared in DMPC 2D lipid crystals, DMPC liposomes, and LDAO-solubilized samples to address the possible effects of the membrane mimetic environment on the conformation of the protein. Concluding, we discuss the strengths and weaknesses of the

  3. Control of a redox reaction on lipid bilayer surfaces by membrane dipole potential. (United States)

    Alakoskela, J I; Kinnunen, P K


    Nitro-2,1,3-benzoxadiazol-4-yl (NBD) group is a widely used, environment-sensitive fluorescent probe. The negatively charged dithionite rapidly reduces the accessible NBD-labeled lipids in liposomes to their corresponding nonfluorescent derivatives. In this study both the phospholipid headgroup and acyl chain NBD-labeled L-alpha-1,2-dipalmitoyl-sn-glycero-3-phospho-[N-(4-nitrobenz-2-oxa-1,3-diazole)-ethanolamine] (DPPN) and 1-acyl-2-[12-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]dodecanoyl]-sn-glycero-3-phosphocholine (NBD-PC), respectively, were employed. The correlation of both the rate coefficient k(1) of the redox reaction and the fluorescence properties of the two probes with the membrane dipole potential Psi in fluid dipalmitoylglycerophosphocholine (DPPC) liposomes is demonstrated. When Psi of the bilayer was varied (decreased by phloretin or increased by 6-ketocholestanol), the value for k1 decreased for both DPPN and NBD-PC with increasing Psi. For both fluorophores a positive correlation to Psi was evident for the relative fluorescence emission intensity (RFI, normalized to the emission of the fluorophore in a DPPC matrix). The relative changes in emission intensity as a function of Psi were approximately equal for both NBD derivatives. Changes similar to those caused by phloretin were seen when dihexadecylglycerophosphocholine (DHPC) was added to DPPC liposomes, in keeping with the lower dipole potential for the former lipid compound compared with DPPC. These effects of Psi on NBD fluorescence should be taken into account when interpreting data acquired using NBD-labeled lipids as fluorescent probes.

  4. Fluctuations of coupled fluid and solid membranes with application to red blood cells (United States)

    Auth, Thorsten; Safran, S. A.; Gov, Nir S.


    The fluctuation spectra and the intermembrane interaction of two membranes at a fixed average distance are investigated. Each membrane can either be a fluid or a solid membrane, and in isolation, its fluctuations are described by a bare or a wave-vector-dependent bending modulus, respectively. The membranes interact via their excluded-volume interaction; the average distance is maintained by an external, homogeneous pressure. For strong coupling, the fluctuations can be described by a single, effective membrane that combines the elastic properties. For weak coupling, the fluctuations of the individual, noninteracting membranes are recovered. The case of a composite membrane consisting of one fluid and one solid membrane can serve as a microscopic model for the plasma membrane and cytoskeleton of the red blood cell. We find that, despite the complex microstructure of bilayers and cytoskeletons in a real cell, the fluctuations with wavelengths λ≳400nm are well described by the fluctuations of a single, polymerized membrane (provided that there are no inhomogeneities of the microstructure). The model is applied to the fluctuation data of discocytes (“normal” red blood cells), a stomatocyte, and an echinocyte. The elastic parameters of the membrane and an effective temperature that quantifies active, metabolically driven fluctuations are extracted from the experiments.

  5. Tunable Surface Hydrophobicity and Fluid Transport through Nanoporous Membranes (United States)

    Ostrowski, Joseph H. J.

    There are more than three billion people across the globe that struggle to obtain clean drinkable water. One of the most promising avenues for generating potable water is through reverse osmosis and nanofiltration. Both solutions require a semipermeable membrane that prohibits passage of unwanted solute particles but allows passage of the solvent. Atomically thin two-dimensional membranes based on porous graphene show great promise as semipermeable materials, but modeling fluid flow on length scales between the microscopic (nanometer and smaller) and macroscopic (micron and larger) regimes presents formidable challenges. This thesis explores both equilibrium and nonequilibrium aspects of this problem and develops new methodology for simulating systems away from thermal equilibrium. First, we hypothesize that there is a wetting penalty for water as it tries to breach a sheet of graphene that should be naturally hydrophobic. By using equilibrium molecular dynamics simulations, we show that the hydrophobicity depends sensitively on the degree of electrical doping, offering an opportunity to tune the hydrophobic effect of graphene using small amounts of doping. The wetting contact angle, a measure of hydrophobicity, changes dramatically with the voltage applied to single layer graphene. We find that the sensitivity of the hydrophobic effect to voltage depends not on hydrogen bonding motifs at the interface between graphene and water, but instead on a phenomenon known as electrowetting. The theory of electrowetting predicts that the difference in surface tensions that defines the contact angle is quartic in the voltage, rather than quadratic, as it would be in bilayer graphene or in a two-dimensional metal. To explore the nonequilibrium aspects of fluid passage through atomically thin membranes, we developed a molecular dynamics methodology for simulating fluid flow at constant flux based on Gauss's principle of least constraint. This method develops microscopic

  6. Fluid bilayer structure determination: Joint refinement in composition space using X-ray and neutron diffraction data

    Energy Technology Data Exchange (ETDEWEB)

    White, S.H. [Univ. of California, Irvine, CA (United States); Wiener, M.C. [Univ. of California, San Francisco, CA (United States)


    Experimentally-determined structural models of fluid lipid bilayers are essential for verifying molecular dynamics simulations of bilayers and for understanding the structural consequences of peptide interactions. The extreme thermal motion of bilayers precludes the possibility of atomic-level structural models. Defining {open_quote}the structure{close_quote} of a bilayer as the time-averaged transbilayer distribution of the water and the principal lipid structural groups such as the carbonyls and double-bonds (quasimolecular fragments), one can represent the bilayer structure as a sum of Gaussian functions referred to collectively as the quasimolecular structure. One method of determining the structure is by neutron diffraction combined with exhaustive specific deuteration. This method is impractical because of the expense of the chemical syntheses and the limited amount of neutron beam time currently available. We have therefore developed the composition space refinement method for combining X-ray and minimal neutron diffraction data to arrive at remarkably detailed and accurate structures of fluid bilayers. The composition space representation of the bilayer describes the probability of occupancy per unit length across the width of the bilayer of each quasimolecular component and permits the joint refinement of X-ray and neutron lamellar diffraction data by means of a single quasimolecular structure that is fitted simultaneously to both data sets. Scaling of each component by the appropriate neutron or X-ray scattering length maps the composition-space profile to the appropriate scattering length space for comparison to experimental data. The difficulty with the method is that fluid bilayer structures are generally only marginally determined by the experimental data. This means that the space of possible solutions must be extensively explored in conjunction with a thorough analysis of errors.

  7. Interaction of alpha-latroinsectotoxin from Latrodectus mactans venom with bilayer lipid membranes. (United States)

    Shatursky OYa; Pashkov, V N; Bulgacov, O V; Grishin, E V


    alpha-Latroinsectotoxin (LIT) from Latrodectus mactans venom increased the conductance of bilayer lipid membranes (BLM) by inducing channel like activity. The channels formed had a maximal single channel conductance of 5 pS in 10 mM CaCl2 solution. This process occurred more rapidly in symmetrical 10 mM CaCl2 solution than in equimolar KCl or NaCl. The LIT induced conductance showed pronounced rectification, that was dependent upon the face of the BLM to which the LIT was applied. This suggests that the LIT molecules incorporate into the bilayer lipid membrane in an oriented manner. The ion channels formed in bilayer phospholipid membrane by LIT are cation selective. The permeability of divalent cations decreased in the order Ba2+ > Ca2+ > Mg2+ > Cd2+ > Zn2+ (Zn2+ and Cd2+ blocked effectively LIT channels with the ratio of Ca2+trans and Cd2+cis or Zn2+cis of 1:1). Selectivity of LIT to monovalent cations was not high and was Ca2+ sensitive. Our data suggest that LIT has at least two Ca(2+)-binding sites, a high affinity site and low one (pK of binding is 2.4). As a result, the binding kinetics of Ca2+ with the toxin shows a high positive cooperativity (Hill coefficient, (h) = 5.95) and that dimerization might be a prerequisite to channel formation. Temperature dependence of conductance of LIT treated lipid bilayers in 100 mM KCl and 10 mM CaCl2 solutions was also determined: 18.9 +/- 2.11 kJ/mol and 28.537 +/- 1.678 kJ/mol, respectively.

  8. Investigations on membrane perturbation by chrysin and its copper complex using self-assembled lipid bilayers. (United States)

    Selvaraj, Stalin; Krishnaswamy, Sridharan; Devashya, Venkappayya; Sethuraman, Swaminathan; Krishnan, Uma Maheswari


    The mechanism of membrane interactions of most of the flavonoids in the presence of transition-metal ions is not well-understood. To understand this phenomenon, the present work aims to synthesize a chrysin-copper complex at room temperature and investigate its influence on the electrical characteristics of planar lipid bilayers. The chrysin-copper complex was characterized by various spectroscopic techniques and was found to have a metal/ligand ratio of 1:2 and of cationic nature. Its ability to inhibit 1,1'-diphenyl-2-picrylhydrazyl (DPPH) radicals was not significant at alkaline pH because of the involvement of the 5-hydroxy group in coordination with the copper ion compared to its parent flavonoid, chrysin (p copper complex to lipid bilayers decreases the resistance, indicating a strong surface interaction and partial insertion into the bilayer near the lipid-water interface. The dose-dependent reduction in resistance as a result of the chrysin-copper complex is more pronounced in comparison to chrysin, implying that the bulkier and charged chrysin-copper complex displays greater ability to distort the lipid bilayer architecture. These conclusions were further confirmed by curcumin-loaded liposome permeabilization studies, where both chrysin and its Cu(II) complex increased the fluidity in a dose-dependent manner. However, the extent of fluidization by the chrysin-copper complex was nearly twice that of chrysin alone (p copper complex on cell membranes were studied using a hypotonic hemolysis assay. Our results demonstrate that, at low concentrations (20 μM), the chrysin-copper complex exhibited twice the protection against hypotonic stress-induced membrane disruption when compared to chrysin. However, this stabilizing effect gradually decreased and became comparable to chrysin at higher concentrations. This biphasic behavior of the chrysin-copper complex could further be explored for therapeutic applications.

  9. Impedance measurements of self-assembled lipid bilayer membranes on the tip of an electrode. (United States)

    Bordi, F; Cametti, Cesare; Gliozzi, A


    Supported lipid membranes were self-assembled on the tip of a freshly cleaved silver wire, in the presence of an appropriate polarization voltage, to facilitate, during the membrane formation, the organization of the lipids into an ordered structure. Radiowave impedance spectroscopy measurements have been carried out to provide information on the relaxation properties of the system. We have measured the conductometric and dielectric properties of bilayers built up of different lipids [dipalmitoylphosphatidic acid (DPPA), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), linoleic acid (LIN)] in a wide frequency range (from 10(3) to 10(6) Hz) and in electrolyte solutions of different ionic strengths, in the presence of uni-univalent (KCl) and di-univalent (CaCl(2), MgCl(2), ZnCl(2)) electrolytes. This made it possible to measure the influence of different cations and different lipid compositions on the membrane properties. In particular, we have found a different capacitive behaviour of the supported lipid bilayer membrane (s-BLM) structure in the presence of different counterions in the electrolyte solution. This peculiarity offers the opportunity for the preparation of a variety of biosensors with diverse applications in membrane biophysics, biochemistry and biotechnology.

  10. Microchemical device based on microscopic bilayer lipid membranes; Bisho 2 bunshimaku wo mochiiita maikuro kagaku debaisu

    Energy Technology Data Exchange (ETDEWEB)

    Yokoyama, H. [Electrotechnical Lab., Ibaraki (Japan)


    If an organism is regarded as a macromolecular system, the element device to construct the same is the molecular structure of nano meter scale formed by the functional protein existing in biomembranes. A lot of essential functions of organism such as the sense reception including vision, gustation, etc., photosynthesis, energy-substance production and so on are performed therein. In this paper, the structure, preparing process and the functions of the microchemical device using micro-bilipid membranes are described. The simulation of the sense receiving functions of organisms is tried by said microchemical device wherein, same as biomembranes, the base is bilayer lipid molecular membrane and the receptive protein for receiving signals from exterior and output molecules such as ion channels connected to said receptive protein and the like are incorporated in the membranes. Recently, it becomes possible to make a partial imaging of the bilayer lipid membranes fixed on porous membrane by the observation with scanning Maxwell-stress microscope. 4 refs., 3 figs.

  11. Interactions of the baicalin and baicalein with bilayer lipid membranes investigated by cyclic voltammetry and UV-Vis spectroscopy. (United States)

    Zhang, Ying; Wang, Xuejing; Wang, Lei; Yu, Miao; Han, Xiaojun


    The baicalin and baicalein are the major flavonoids found in Radix Scutellariae, an essential herb in traditional Chinese medicine for thousands of years. The interactions of the baicalin and baicalein with lipid bilayer membranes were studied using cyclic voltammetry and UV-Vis spectroscopy. The thickness d of supported bilayer lipid membranes was calculated as d=4.59(±0.36) nm using AC impedance spectroscopy. The baicalein interacted with egg PC bilayer membranes in a dose-dependent manner. The responses of K3Fe(CN)6 on lipid bilayer membrane modified Pt electrode linearly increased in a concentration range of baicalein from 6.25μM to 25μM with a detection limit of 0.1μM and current-concentration sensitivity of 0.11(±0.01) μA/μM, and then reached a plateau from 25μM to 50μM. However the baicalin showed much weaker interactions with egg PC bilayer membranes. UV-Vis spectroscopy also confirmed that the baicalein could interact with egg PC membranes noticeably, but the interaction of baicalin with membranes was hard to be detected. The results provide useful information on understanding the mechanism of action of Radix Scutellariae in vivo. © 2013.

  12. Continuity of Monolayer-Bilayer Junctions for Localization of Lipid Raft Microdomains in Model Membranes. (United States)

    Ryu, Yong-Sang; Wittenberg, Nathan J; Suh, Jeng-Hun; Lee, Sang-Wook; Sohn, Youngjoo; Oh, Sang-Hyun; Parikh, Atul N; Lee, Sin-Doo


    We show that the selective localization of cholesterol-rich domains and associated ganglioside receptors prefer to occur in the monolayer across continuous monolayer-bilayer junctions (MBJs) in supported lipid membranes. For the MBJs, glass substrates were patterned with poly(dimethylsiloxane) (PDMS) oligomers by thermally-assisted contact printing, leaving behind 3 nm-thick PDMS patterns. The hydrophobicity of the transferred PDMS patterns was precisely tuned by the stamping temperature. Lipid monolayers were formed on the PDMS patterned surface while lipid bilayers were on the bare glass surface. Due to the continuity of the lipid membranes over the MBJs, essentially free diffusion of lipids was allowed between the monolayer on the PDMS surface and the upper leaflet of the bilayer on the glass substrate. The preferential localization of sphingomyelin, ganglioside GM1 and cholesterol in the monolayer region enabled to develop raft microdomains through coarsening of nanorafts. Our methodology provides a simple and effective scheme of non-disruptive manipulation of the chemical landscape associated with lipid phase separations, which leads to more sophisticated applications in biosensors and as cell culture substrates.

  13. Bilayer membrane permeability of ionic liquid-filled block copolymer vesicles in aqueous solution. (United States)

    Bai, Zhifeng; Zhao, Bin; Lodge, Timothy P


    The bilayer membrane permeability of block copolymer vesicles ("polymersomes") with ionic liquid interiors dispersed in water is quantified using fluorescence quenching. Poly((1,2-butadiene)-b-ethylene oxide) (PB-PEO) block copolymer vesicles in water with their interiors filled with a common hydrophobic ionic liquid, 1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)amide, were prepared containing a hydrophobic dye, Nile Red, by intact migration of dye-encapsulated vesicles from the ionic liquid to water at room temperature. A small quencher molecule, dichloroacetamide, was added to the aqueous solution of the dye-loaded vesicles, and the permeation of the quencher passing through the membrane into the interior was determined from the fluorescence quenching kinetics. Rapid permeation of the quencher across the nanoscale membrane was observed, consistent with the high fluidity of the liquid polybutadiene membrane. Two different PB-PEO copolymers were employed, in order to vary the thickness of the solvophobic membrane. A significant increase in membrane permeability was also observed with decreasing membrane thickness, which is tentatively attributable to differences in quencher solubility in the membranes. Quantitative migration of the vesicles from the aqueous phase back to an ionic liquid phase was achieved upon heating. These microscopically heterogeneous and thermoresponsive vesicles with permeable and robust membranes have potential as recyclable nanoreactors, in which the high viscosity and capital expense of an ionic liquid reaction medium can be mitigated, while retaining the desirable features of ionic liquids as reaction media, and facile catalyst recovery.

  14. Bilayer registry in a multicomponent asymmetric membrane : dependence on lipid composition and chain length

    CERN Document Server

    Polley, Anirban; Rao, Madan


    A question of considerable interest to cell membrane biology is whether phase segregated domains across an asymmetric bilayer are strongly correlated with each other and whether phase segregation in one leaflet can induce segregation in the other. We answer both these questions in the affirmative, using an atomistic molecular dynamics simulation to study the equilibrium statistical properties of a 3-component {\\em asymmetric} lipid bilayer comprising an unsaturated POPC (palmitoyl-oleoyl-phosphatidyl-choline), a saturated SM (sphingomyelin) and cholesterol with different composition ratios. Our simulations are done by fixing the composition of the upper leaflet to be at the coexistence of the liquid ordered ($l_o$) - liquid disordered ($l_d$) phases, while the composition of the lower leaflet is varied from the phase coexistence regime to the mixed $l_d$ phase, across a first-order phase boundary. In the regime of phase coexistence in each leaflet, we find strong transbilayer correlations of the $l_o$ domains...

  15. Diffusion mediated coagulation and fragmentation based study of domain formation in lipid bilayer membrane

    Energy Technology Data Exchange (ETDEWEB)

    Rao, Laxminarsimha V., E-mail: [Mechanics and Applied Mathematics Group, Department of Mechanical Engineering, Indian Institute of Technology Kanpur, Kanpur 208016 (India); Roy, Subhradeep [Department of Biomedical Engineering and Mechanics (MC 0219), Virginia Tech, 495 Old Turner Street, Blacksburg, VA 24061 (United States); Das, Sovan Lal [Mechanics and Applied Mathematics Group, Department of Mechanical Engineering, Indian Institute of Technology Kanpur, Kanpur 208016 (India)


    We estimate the equilibrium size distribution of cholesterol rich micro-domains on a lipid bilayer by solving Smoluchowski equation for coagulation and fragmentation. Towards this aim, we first derive the coagulation kernels based on the diffusion behaviour of domains moving in a two dimensional membrane sheet, as this represents the reality better. We incorporate three different diffusion scenarios of domain diffusion into our coagulation kernel. Subsequently, we investigate the influence of the parameters in our model on the coagulation and fragmentation behaviour. The observed behaviours of the coagulation and fragmentation kernels are also manifested in the equilibrium domain size distribution and its first moment. Finally, considering the liquid domains diffusing in a supported lipid bilayer, we fit the equilibrium domain size distribution to a benchmark solution.

  16. Molecular aspects of electrical excitation in lipid bilayers and cell membranes. (United States)

    Mueller, P


    Several compounds of fungal or bacterial origin (EIM, alamethicin, monazomycin, DJ400B) can be incorporated into planar lipid bilayers where they form molecular channels and generate voltage-dependent ion conductances. When studied by voltage clamp, the kinetic and steady-state characteristics of these conductance changes are in every respect identical to those found in excitable cell membranes, and their major aspects can be quantitatively described by the Hodgkin-Huxley equations. Thus, the steady-state conductance is an expotential function of the membrane potential, the conductance rises with a sigmoid time course and decays exponentially, and the time constants of the conductance changes go through a maximum as a function of the potential. The conductances also show inactivation as seen in the sodium channels of nerve and the potassium channels of muscle. In addition, there appear for particular pulsing sequences certain kinetic transients that cannot be accounted for by the Hodgkin-Huxley equations but are also seen in identical form in nerve. Because the kinetics are identical in all excitable cell membranes and in these bilayers, it is likely that, in spite of the diverse chemical nature of the channel-forming molecules in the bilayers and the widely differing ion selectivities in the cellular systems, the mechanism by which the membrane opens and closes for the flow of ions is essentially the same in all cases. The kinetic data imply that a cooperative process is involved in the gating action. In principle, two different concepts could account for the kinetics--one involving an intramolecular configurational change within a complex permanent channel, the other, the assembly of a channel through the voltage-dependent aggregation of monomeric channel precursors. In the bilayers the high-order dependence of the steady-state conductance and of the gating time constants on the concentration of the channel formers suggests an aggregation mechanism in which the

  17. Voltage-sensitive styryl dyes as singlet oxygen targets on the surface of bilayer lipid membrane. (United States)

    Sokolov, V S; Gavrilchik, A N; Kulagina, A O; Meshkov, I N; Pohl, P; Gorbunova, Yu G


    Photosensitizers are widely used as photodynamic therapeutic agents killing cancer cells by photooxidation of their components. Development of new effective photosensitive molecules requires profound knowledge of possible targets for reactive oxygen species, especially for its singlet form. Here we studied photooxidation of voltage-sensitive styryl dyes (di-4-ANEPPS, di-8-ANEPPS, RH-421 and RH-237) by singlet oxygen on the surface of bilayer lipid membranes commonly used as cell membrane models. Oxidation was induced by irradiation of a photosensitizer (aluminum phthalocyanine tetrasulfonate) and monitored by the change of dipole potential on the surface of the membrane. We studied the drop of the dipole potential both in the case when the dye molecules were adsorbed on the same side of the lipid bilayer as the photosensitizer (cis-configuration) and in the case when they were adsorbed on the opposite side (trans-configuration). Based on a simple model, we determined the rate of oxidation of the dyes from the kinetics of change of the potential during and after irradiation. This rate is proportional to steady-state concentration of singlet oxygen in the membrane under irradiation. Comparison of the oxidation rates of various dyes reveals that compounds of ANEPPS series are more sensitive to singlet oxygen than RH type dyes, indicating that naphthalene group is primarily responsible for their oxidation.

  18. A bilayer-couple model of bacterial outer membrane vesicle biogenesis. (United States)

    Schertzer, Jeffrey W; Whiteley, Marvin


    Gram-negative bacteria naturally produce outer membrane vesicles (OMVs) that arise through bulging and pinching off of the outer membrane. OMVs have several biological functions for bacteria, most notably as trafficking vehicles for toxins, antimicrobials, and signaling molecules. While their biological roles are now appreciated, the mechanism of OMV formation has not been fully elucidated. We recently demonstrated that the signaling molecule 2-heptyl-3-hydroxy-4-quinolone (PQS) is required for OMV biogenesis in P. aeruginosa. We hypothesized that PQS stimulates OMV formation through direct interaction with the outer leaflet of the outer membrane. To test this hypothesis, we employed a red blood cell (RBC) model that has been used extensively to study small-molecule-membrane interactions. Our results revealed that addition of PQS to RBCs induced membrane curvature, resulting in the formation of membrane spicules (spikes), consistent with small molecules that are inserted stably into the outer leaflet of the membrane. Radiotracer experiments demonstrated that sufficient PQS was inserted into the membrane to account for this curvature and that curvature induction was specific to PQS structure. These data suggest that a low rate of interleaflet flip-flop forces PQS to accumulate in and expand the outer leaflet relative to the inner leaflet, thus inducing membrane curvature. In support of PQS-mediated outer leaflet expansion, the PQS effect was antagonized by chlorpromazine, a molecule known to be preferentially inserted into the inner leaflet. Based on these data, we propose a bilayer-couple model to describe P. aeruginosa OMV biogenesis and suggest that this is a general mechanism for bacterial OMV formation. Despite the ubiquity and importance of outer membrane vesicle (OMV) production in Gram-negative bacteria, the molecular details of OMV biogenesis are not fully understood. Early experiments showed that 2-heptyl-3-hydroxy-4-quinolone (PQS) induces OMV formation

  19. Genomic analysis indicates the presence of an asymmetric bilayer outer membrane in Planctomycetes and Verrucomicrobia

    Directory of Open Access Journals (Sweden)

    Daan R Speth


    Full Text Available Bacteria of the phylum Planctomycetes are of special interest for the study of compartmental cellular organization. Members of this phylum share a very unusual prokaryotic cell plan, featuring several membrane-bound compartments. Recently, it was shown that this cellular organization might extend to certain members of the phylum Verrucomicrobia. The Planctomycete cell plan has been defined as featuring a proteinaceous cell wall, a cytoplasmic membrane surrounding the paryphoplasm and an intracytoplasmic membrane defining the riboplasm. So far it was presumed that Planctomycetes did not have an asymmetric bilayer outer membrane as observed in Gram-negative bacteria. However, recent work on outer membrane biogenesis has provided several marker genes in the outer membrane protein (OMP assembly and the lipopolysaccharide (LPS insertion complexes. Additionally, advances in computational prediction of OMPs provided new tools to perform more accurate genomic screening for such proteins.Here we searched all 22 Planctomycetes and Verrucomicrobia genomes available in Genbank, plus the recently published genome of ‘Candidatus Scalindua profunda’, for markers of outer membrane biogenesis and OMPs. We were able to identify the key components of LPS insertion, OMP assembly and at least eight OMPs in all genomes tested. Additionally, we have analyzed the transcriptome and proteome data of the Planctomycetes ‘Candidatus Kuenenia stuttgartiensis’ and ‘Ca. S. profunda’ and could confirm high expression of several predicted OMPs, including the biomarkers of outer membrane biogenesis.

  20. Thermodynamic Free Energy Methods to Investigate Shape Transitions In Bilayer Membranes

    CERN Document Server

    Ramakrishnan, N; Radhakrishnan, Ravi


    The conformational free energy landscape of a system is a fundamental thermodynamic quantity of importance particularly in the study of soft matter and biological systems, in which the entropic contributions play a dominant role. While computational methods to delineate the free energy landscape are routinely used to analyze the relative stability of conformational states, to determine phase boundaries, and to compute ligand-receptor binding energies its use in problems involving the cell membrane is limited. Here, we present an overview of four different free energy methods to study morphological transitions in bilayer membranes, induced either by the action of curvature remodeling proteins or due to the application of external forces. Using a triangulated surface as a model for the cell membrane and using the framework of dynamical triangulation Monte Carlo, we have focused on the methods of Widom insertion, thermodynamic integration, Bennett acceptance scheme, and umbrella sampling and weighted histogram a...

  1. Triglyceride Blisters in Lipid Bilayers: Implications for Lipid Droplet Biogenesis and the Mobile Lipid Signal in Cancer Cell Membranes

    DEFF Research Database (Denmark)

    Khandelia, Himanshu; Duelund, Lars; Pakkanen, Kirsi Inkeri


    Triglycerides have a limited solubility, around 3%, in phosphatidylcholine lipid bilayers. Using millisecond-scale course grained molecular dynamics simulations, we show that the model lipid bilayer can accommodate a higher concentration of triolein (TO) than earlier anticipated, by sequestering...... aggregates of unknown function present in malignant cells, and to the early biogenesis of lipid droplets accommodated between the two leaflets of the endoplasmic reticulum membrane. The TO aggregates give the bilayer a blister-like appearance, and will hinder the formation of multi-lamellar phases in model...

  2. Determining the orientation of protegrin-1 in DLPC bilayers using an implicit solvent-membrane model.

    Directory of Open Access Journals (Sweden)

    Abdallah Sayyed-Ahmad

    Full Text Available Continuum models that describe the effects of solvent and biological membrane molecules on the structure and behavior of antimicrobial peptides, holds a promise to improve our understanding of the mechanisms of antimicrobial action of these peptides. In such methods, a lipid bilayer model membrane is implicitly represented by multiple layers of relatively low dielectric constant embedded in a high dielectric aqueous solvent, while an antimicrobial peptide is accounted for by a dielectric cavity with fixed partial charge at the center of each one of its atoms. In the present work, we investigate the ability of continuum approaches to predict the most probable orientation of the beta-hairpin antimicrobial peptide Protegrin-1 (PG-1 in DLPC lipid bilayers by calculating the difference in the transfer free energy from an aqueous environment to a membrane-water environment for multiple orientations. The transfer free energy is computed as a sum of two terms; polar/electrostatic and non-polar. They both include energetic and entropic contributions to the free energy. We numerically solve the Poisson-Boltzmann equation to calculate the electrostatic contribution to the transfer free energy, while the non-polar contribution to the free energy is approximated using a linear solvent accessible surface area relationships. The most probable orientation of PG-1 is that with the lowest relative transfer free energy. Our simulation results indicate that PG-1 assumes an oblique orientation in DLPC lipid bilayers. The predicted most favorable orientation was with a tilt angle of 19 degrees, which is in qualitative agreement with the experimentally observed orientations derived from solid-state NMR data.

  3. Chemically-activatable alkyne-tagged probe for imaging microdomains in lipid bilayer membranes (United States)

    Yamaguchi, Satoshi; Matsushita, Taku; Izuta, Shin; Katada, Sumika; Ura, Manami; Ikeda, Taro; Hayashi, Gosuke; Suzuki, Yuta; Kobayashi, Koya; Tokunaga, Kyoya; Ozeki, Yasuyuki; Okamoto, Akimitsu


    A chemically-activatable alkynyl steroid analogue probe has been synthesized for visualizing the lipid raft membrane domains by Raman microscopy. The Raman probe, in which ring A of its steroid backbone is replaced with an alkynyl group, was designed to enable activation of the alkyne signal through the Eschenmoser-Tanabe fragmentation reaction of the oxidized cholesterol precursor in lipid bilayer membranes. The alkynyl steroid analogue was observed to form liquid-ordered raft-like domains on a model giant-liposome system in a similar manner as cholesterol, and the large alkyne signal of the accumulated probe at 2120 cm−1 was mapped on the microdomains with a Raman microscope. The alkyne moiety of the probe was confirmed to be converted from the α,β-epoxy ketone group of its precursor by reaction with p-toluensulfonyl hydrazine under a mild condition. Through the reaction, the alkyne signal of the probe was activated on the lipid bilayer membrane of liposomes. Furthermore, the signal activation of the probe was also detected on living cells by stimulated Raman scattering microscopy. The ring-A-opened alkyne steroid analogue, thus, provides a first chemically-activatable Raman probe as a promising tool for potentially unravelling the intracellular formation and trafficking of cholesterol-rich microdomains. PMID:28117375

  4. Fluid and air-stable lipopolymer membranes for biosensor applications. (United States)

    Albertorio, Fernando; Diaz, Arnaldo J; Yang, Tinglu; Chapa, Vanessa A; Kataoka, Sho; Castellana, Edward T; Cremer, Paul S


    The behavior of poly(ethylene glycol) (PEG) conjugated lipids was investigated in planar supported egg phosphatidylcholine bilayers as a function of lipopolymer density, chain length of the PEG moiety, and type of alkyl chains on the PEG lipid. Fluorescence recovery after photobleaching measurements verified that dye-labeled lipids in the membrane as well as the lipopolymer itself maintained a substantial degree of fluidity under most conditions that were investigated. PEG densities exceeding the onset of the mushroom-to-brush phase transition were found to confer air stability to the supported membrane. On the other hand, substantial damage or complete delamination of the lipid bilayer was observed at lower polymer densities. The presence of PEG in the membrane did not substantially hinder the binding of streptavidin to biotinylated lipids present in the bilayer. Furthermore, above the onset of the transition into the brush phase, the protein binding properties of these membranes were found to be very resilient upon removal of the system from water, rigorous drying, and rehydration. These results indicate that supported phospholipid bilayers containing lipopolymers show promise as rugged sensor platforms for ligand-receptor binding.

  5. Structure and dynamics of water and lipid molecules in charged anionic DMPG lipid bilayer membranes

    DEFF Research Database (Denmark)

    Rønnest, A. K.; Peters, Günther H.J.; Hansen, Flemming Yssing


    in the gel phase and 17.5 molecules in the fluid phase, considerably smaller than inferred experimentally for 1,2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC) membranes but comparable to the number inferred for 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine (DLPE) membranes. Some of the properties...

  6. Diffusion of water and selected atoms in DMPC lipid bilayer membranes

    DEFF Research Database (Denmark)

    Hansen, Flemming Yssing; Peters, Günther H.J.; Taub, H.;


    with the diffusion rate of selected atoms in the lipid molecules shows that ∼6 water molecules per lipid molecule move on the same time scale as the lipids and may therefore be considered to be tightly bound to them. The quasielastic neutron scattering functions for water and selected atoms in the lipid molecule......Molecular dynamics simulations have been used to determine the diffusion of water molecules as a function of their position in a fully hydrated freestanding 1,2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC) bilayer membrane at 303 K and 1 atm. The diffusion rate of water in a ∼10 Å thick layer...

  7. Study of water diffusion on single-supported bilayer lipid membranes by quasielastic neutron scattering

    DEFF Research Database (Denmark)

    Bai, M.; Miskowiec, A.; Hansen, F. Y.


    High-energy-resolution quasielastic neutron scattering has been used to elucidate the diffusion of water molecules in proximity to single bilayer lipid membranes supported on a silicon substrate. By varying sample temperature, level of hydration, and deuteration, we identify three different types...... of diffusive water motion: bulk-like, confined, and bound. The motion of bulk-like and confined water molecules is fast compared to those bound to the lipid head groups (7-10 H2O molecules per lipid), which move on the same nanosecond time scale as H atoms within the lipid molecules. Copyright (C) EPLA, 2012...

  8. Fluid physics of a rotating membrane separator (United States)

    Akonur, Alp

    Rotating membrane separation is a powerful dynamic filtration technique used in separation and filtration of suspensions. Rotating membrane separation is superior compared to the conventional filtration techniques owing to the special character of the flow field, namely the supercritical cylindrical Couette flow, observed in the form of nonwavy and wavy toroidal vortices. The underlying physics are investigated by extending the previous particle image velocity (PIV) measurements performed in a radial-axial plane to a radial-azimuthal plane for nonwavy Taylor Couette flow and wavy cylindrical Couette flow. These measurements are matched to previous measurements to obtain the first time- resolved, three-dimensional, three-component velocity field for cylindrical Couette flow. The nonwavy toroidal vortices of Taylor-Couette flow become stronger with increasing Taylor number. The azimuthal velocity varies axially due to the redistribution of the azimuthal momentum by the vortical motion, which results in a substantial increase in the angular momentum at outflow regions and a decrease at inflow regions. For wavy vortex flow, the waviness of the vortices results in a variation of the azimuthal velocity in any given latitudinal place. Streams of axial flow carry fluid along the length of the annulus winding around the vortices radially from the inner cylinder to the outer cylinder, and azimuthally about one-half wavelength. The azimuthal velocity near the centers of the vortices is similar to the velocity of the traveling azimuthal wave. Large shear stresses occur near the inner and outer cylinders especially at the high Taylor numbers. In the middle of the annulus, the shear stress is substantially less. In filtration flow, where radial and axial flows are imposed on cylindrical Couette flow, simultaneous use of particle image velocimetry (PIV) and particle tracking velocimetry (PTV) provides fluid and particle velocities. Results indicate no major differences in the

  9. Fabrication and characterization of an integrated ionic device from suspended polypyrrole and alamethicin-reconstituted lipid bilayer membranes (United States)

    Northcutt, Robert; Sundaresan, Vishnu-Baba


    Conducting polymers are electroactive materials that undergo conformal relaxation of the polymer backbone in the presence of an electrical field through ion exchange with solid or aqueous electrolytes. This conformal relaxation and the associated morphological changes make conducting polymers highly suitable for actuation and sensing applications. Among smart materials, bioderived active materials also use ion transport for sensing and actuation functions via selective ion transport. The transporter proteins extracted from biological cell membranes and reconstituted into a bilayer lipid membrane in bioderived active materials regulate ion transport for engineering functions. The protein transporter reconstituted in the bilayer lipid membrane is referred to as the bioderived membrane and serves as the active component in bioderived active materials. Inspired by the similarities in the physics of transduction in conducting polymers and bioderived active materials, an integrated ionic device is formed from the bioderived membrane and the conducting polymer membrane. This ionic device is fabricated into a laminated thin-film membrane and a common ion that can be processed by the bioderived and the conducting polymer membranes couple the ionic function of these two membranes. An integrated ionic device, fabricated from polypyrrole (PPy) doped with sodium dodecylbenzenesulfonate (NaDBS) and an alamethicin-reconstituted DPhPC bilayer lipid membrane, is presented in this paper. A voltage-gated sodium current regulates the electrochemical response in the PPy(DBS) layer. The integrated device is fabricated on silicon-based substrates through microfabrication, electropolymerization, and vesicle fusion, and ionic activity is characterized through electrochemical measurements.

  10. Single Lipid Molecule Dynamics on Supported Lipid Bilayers with Membrane Curvature

    Directory of Open Access Journals (Sweden)

    Philip P. Cheney


    Full Text Available The plasma membrane is a highly compartmentalized, dynamic material and this organization is essential for a wide variety of cellular processes. Nanoscale domains allow proteins to organize for cell signaling, endo- and exocytosis, and other essential processes. Even in the absence of proteins, lipids have the ability to organize into domains as a result of a variety of chemical and physical interactions. One feature of membranes that affects lipid domain formation is membrane curvature. To directly test the role of curvature in lipid sorting, we measured the accumulation of two similar lipids, 1,2-Dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DHPE and hexadecanoic acid (HDA, using a supported lipid bilayer that was assembled over a nanopatterned surface to obtain regions of membrane curvature. Both lipids studied contain 16 carbon, saturated tails and a head group tag for fluorescence microscopy measurements. The accumulation of lipids at curvatures ranging from 28 nm to 55 nm radii was measured and fluorescein labeled DHPE accumulated more than fluorescein labeled HDA at regions of membrane curvature. We then tested whether single biotinylated DHPE molecules sense curvature using single particle tracking methods. Similar to groups of fluorescein labeled DHPE accumulating at curvature, the dynamics of single molecules of biotinylated DHPE was also affected by membrane curvature and highly confined motion was observed.

  11. Alginate/chitosan based bi-layer composite membrane as potential sustained-release wound dressing containing ciprofloxacin hydrochloride (United States)

    Han, Fei; Dong, Yang; Song, Aihua; Yin, Ran; Li, Sanming


    The aims of this research were to develop and evaluate a novel ciprofloxacin hydrochloride loaded bi-layer composite membrane based on alginate and chitosan. In vitro antimicrobial activity, drug permeation study, morphology, cytotoxicity, primary skin irritation and in vivo pharmacodynamics were investigated. Results showed that the membranes could inhibit the growth of microorganisms for longer than 7 days. And there was no significant decrease in the metabolic activity of the Hacat fibroblasts cells were treated with the membranes. No edema and erythema were observed after administration of membranes on the rabbit skin after 14 days. Moreover, the results of pharmacodynamics showed that the membranes were more effective in improving the wound healing process. In conclusion, a novel bi-layer composite membrane was developed and results suggested that it could be exploited as sustained-release wound dressings.

  12. Chemotherapy Drugs Thiocolchicoside and Taxol Permeabilize Lipid Bilayer Membranes by Forming Ion Pores (United States)

    Ashrafuzzaman, Md; Duszyk, M.; Tuszynski, J. A.


    We report ion channel formation by chemotherapy drugs: thiocolchicoside (TCC) and taxol (TXL) which primarily target tubulin but not only. For example, TCC has been shown to interact with GABAA, nuclear envelope and strychnine-sensitive glycine receptors. TXL interferes with the normal breakdown of microtubules inducing mitotic block and apoptosis. It also interacts with mitochondria and found significant chemotherapeutic applications for breast, ovarian and lung cancer. In order to better understand the mechanisms of TCC and TXL actions, we examined their effects on phospholipid bilayer membranes. Our electrophysiological recordings across membranes constructed in NaCl aqueous phases consisting of TCC or TXL under the influence of an applied transmembrane potential (V) indicate that both molecules induce stable ion flowing pores/channels in membranes. Their discrete current versus time plots exhibit triangular shapes which is consistent with a spontaneous time-dependent change of the pore conductance in contrast to rectangular conductance events usually induced by ion channels. These events exhibit conductance (~0.01-0.1 pA/mV) and lifetimes (~5-30 ms) within the ranges observed in e.g., gramicidin A and alamethicin channels. The channel formation probability increases linearly with TCC/TXL concentration and V and is not affected by pH (5.7 - 8.4). A theoretical explanation on the causes of chemotherapy drug induced ion pore formation and the pore stability has also been found using our recently discovered binding energy between lipid bilayer and the bilayer embedded ion channels using gramicidin A channels as tools. This picture of energetics suggests that as the channel forming agents approach to the lipids on bilayer the localized charge properties in the constituents of both channel forming agents (e.g., chemotherapy drugs in this study) and the lipids determine the electrostatic drug-lipid coupling energy through screened Coulomb interactions between the drug

  13. Self-assembly of azobenzene bilayer membranes in binary ionic liquid-water nanostructured media. (United States)

    Kang, Tejwant Singh; Ishiba, Keita; Morikawa, Masa-aki; Kimizuka, Nobuo


    Anionic azobenzene-containing amphiphile 1 (sodium 4-[4-(N-methyl-N-dodecylamino)phenylazo]benzenesulfonate) forms ordered bilayer membranes in binary ionic liquid (1-ethyl-3-methylimidazolium ethyl sulfate, [C2mim][C2OSO3])-water mixtures. The binary [C2mim][C2OSO3]-water mixture is macroscopically homogeneous at any mixing ratio; however, it possesses fluctuating nanodomains of [C2mim][C2OSO3] molecules as observed by dynamic light scattering (DLS). These nanodomains show reversible heat-induced mixing behavior with water. Although the amphiphile 1 is substantially insoluble in pure water, it is dispersible in the [C2mim][C2OSO3]-water mixtures. The concentration of [C2mim][C2OSO3] and temperature exert significant influences on the self-assembling characteristics of 1 in the binary media, as shown by DLS, transmission electron microscopy (TEM), UV-vis spectroscopy, and zeta-potential measurements. Bilayer membranes with rod- or dotlike nanostructures were formed at a lower content of [C2mim][C2OSO3] (2-30 v/v %), in which azobenzene chromophores adopt parallel molecular orientation regardless of temperature. In contrast, when the content of [C2mim][C2OSO3] is increased above 60 v/v %, azobenzene bilayers showed thermally reversible gel-to-liquid crystalline phase transition. The self-assembly of azobenzene amphiphiles is tunable depending on the volume fraction of [C2mim][C2OSO3] and temperature, which are associated with the solvation by nanoclusters in the binary [C2mim][C2OSO3]-water media. These observations clearly indicate that mixtures of water-soluble ionic liquids and water provide unique and valiant environments for ordered molecular self-assembly.

  14. Membrane Binding of HIV-1 Matrix Protein: Dependence on Bilayer Composition and Protein Lipidation (United States)

    Barros, Marilia; Nanda, Hirsh


    ABSTRACT By assembling in a protein lattice on the host's plasma membrane, the retroviral Gag polyprotein triggers formation of the viral protein/membrane shell. The MA domain of Gag employs multiple signals—electrostatic, hydrophobic, and lipid-specific—to bring the protein to the plasma membrane, thereby complementing protein-protein interactions, located in full-length Gag, in lattice formation. We report the interaction of myristoylated and unmyristoylated HIV-1 Gag MA domains with bilayers composed of purified lipid components to dissect these complex membrane signals and quantify their contributions to the overall interaction. Surface plasmon resonance on well-defined planar membrane models is used to quantify binding affinities and amounts of protein and yields free binding energy contributions, ΔG, of the various signals. Charge-charge interactions in the absence of the phosphatidylinositide PI(4,5)P2 attract the protein to acidic membrane surfaces, and myristoylation increases the affinity by a factor of 10; thus, our data do not provide evidence for a PI(4,5)P2 trigger of myristate exposure. Lipid-specific interactions with PI(4,5)P2, the major signal lipid in the inner plasma membrane, increase membrane attraction at a level similar to that of protein lipidation. While cholesterol does not directly engage in interactions, it augments protein affinity strongly by facilitating efficient myristate insertion and PI(4,5)P2 binding. We thus observe that the isolated MA protein, in the absence of protein-protein interaction conferred by the full-length Gag, binds the membrane with submicromolar affinities. IMPORTANCE Like other retroviral species, the Gag polyprotein of HIV-1 contains three major domains: the N-terminal, myristoylated MA domain that targets the protein to the plasma membrane of the host; a central capsid-forming domain; and the C-terminal, genome-binding nucleocapsid domain. These domains act in concert to condense Gag into a membrane

  15. Comparing ion conductance recordings of synthetic lipid bilayers with cell membranes containing TRP channels

    CERN Document Server

    Laub, Katrine R; Blicher, Andreas; Madsen, Soren B; Luckhoff, Andreas; Heimburg, Thomas


    In this article we compare electrical conductance events from single channel recordings of three TRP channel proteins (TRPA1, TRPM2 and TRPM8) expressed in human embryonic kidney cells with channel events recorded on synthetic lipid membranes close to melting transitions. Ion channels from the TRP family are involved in a variety of sensory processes including thermo- and mechano-reception. Synthetic lipid membranes close to phase transitions display channel-like events that respond to stimuli related to changes in intensive thermodynamic variables such as pressure and temperature. TRP channel activity is characterized by typical patterns of current events dependent on the type of protein expressed. Synthetic lipid bilayers show a wide spectrum of electrical phenomena that are considered typical for the activity of protein ion channels. We find unitary currents, burst behavior, flickering, multistep-conductances, and spikes behavior in both preparations. Moreover, we report conductances and lifetimes for lipi...

  16. Bilayer lipid membranes supported on Teflon filters: a functional environment for ion channels. (United States)

    Phung, Thai; Zhang, Yanli; Dunlop, James; Dalziel, Julie


    Many ion channel proteins have binding sites for toxins and pharmaceutical drugs and therefore have much promise as the sensing entity in high throughput technologies and biosensor devices. Measurement of ionic conductance changes through ion channels requires a robust biological membrane with sufficient longevity for practical applications. The conventional planar BLM is 100-300 μm in diameter and typically contains fewer than a dozen channels whereas pharmaceutical screening methods in cells use current recordings for many ion channels. We present a new, simple method for the fabrication of a disposable porous-supported bilayer lipid membrane (BLM) ion channel biosensor using hydrated Teflon (polytetrafluoroethylene, PTFE) filter material (pore size 5 μm, filter diameter=1 mm). The lipid layer was monitored for its thickness and mechanical stability by electrical impedance spectroscopy. The results showed membrane capacitances of 1.8±0.2 nF and membrane resistances of 25.9±4.1 GΩ, indicating the formation of lipid bilayers. The current level increased upon addition of the pore-forming peptide gramicidin. Following addition of liposomes containing voltage-gated sodium channels, small macroscopic sodium currents (1-80 pA) could be recorded. By preloading the porous Teflon with sodium channel proteoliposomes, prior to BLM formation, currents of 1-10 nA could be recorded in the presence of the activator veratridine that increased with time, and were inhibited by tetrodotoxin. A lack of rectification suggests that the channels incorporated in both orientations. This work demonstrates that PTFE filters can support BLMs that provide an environment in which ion channels can maintain their functional activity relevant for applications in drug discovery, toxin detection, and odour sensing.

  17. Coupling Optical and Electrical Measurements in Artificial Membranes: Lateral Diffusion of Lipids and Channel Forming Peptides in Planar Bilayers

    Directory of Open Access Journals (Sweden)

    Duclohier H


    Full Text Available Planar lipid bilayers (PLB were prepared by the Montal-Mueller technique in a FRAP system designed to simultaneously measure conductivity across, and lateral diffusion of, the bilayer. In the first stage of the project the FRAP system was used to characterise the lateral dynamics of bilayer lipids with regards to phospholipid composition (headgroup, chain unsaturation etc., presence of cholesterol and the effect of divalent cations on negatively-charged bilayers. In the second stage of the project, lateral diffusion of two fluorescently-labelled voltage-dependent pore-forming peptides (alamethicin and S4s from Shaker K+ channel was determined at rest and in the conducting state. This study demonstrates the feasibility of such experiments with PLBs, amenable to physical constraints, and thus offers new opportunities for systematic studies of structure-function relationships in membrane-associating molecules.

  18. Computer Simulations of Lipid Bilayers and Proteins

    DEFF Research Database (Denmark)

    Sonne, Jacob


    , Pressure profile calculations in lipid bilayers: A lipid bilayer is merely $\\sim$5~nm thick, but the lateral pressure (parallel to the bilayer plane) varies several hundred bar on this short distance (normal to the bilayer). These variations in the lateral pressure are commonly referred to as the pressure...... of neglecting pressure contributions from long range electrostatic interactions. The first issue is addressed by comparing two methods for calculating pressure profiles, and judged by the similar results obtained by these two methods the pressure profile appears to be well-defined for fluid phase lipid bilayers......The importance of computer simulations in lipid bilayer research has become more prominent for the last couple of decades and as computers get even faster, simulations will play an increasingly important part of understanding the processes that take place in and across cell membranes. This thesis...

  19. Steady-state oxidation of cholesterol catalyzed by cholesterol oxidase in lipid bilayer membranes on platinum electrodes

    Energy Technology Data Exchange (ETDEWEB)

    Bokoch, Michael P.; Devadoss, Anando; Palencsar, Mariela S.; Burgess, James D


    Cholesterol oxidase is immobilized in electrode-supported lipid bilayer membranes. Platinum electrodes are initially modified with a self-assembled monolayer of thiolipid. A vesicle fusion method is used to deposit an outer leaflet of phospholipids onto the thiolipid monolayer forming a thiolipid/lipid bilayer membrane on the electrode surface. Cholesterol oxidase spontaneously inserts into the electrode-supported lipid bilayer membrane from solution and is consequently immobilized to the electrode surface. Cholesterol partitions into the membrane from buffer solutions containing cyclodextrin. Cholesterol oxidase catalyzes the oxidation of cholesterol by molecular oxygen, forming hydrogen peroxide as a product. Amperometric detection of hydrogen peroxide for continuous solution flow experiments are presented, where flow was alternated between cholesterol solution and buffer containing no cholesterol. Steady-state anodic currents were observed during exposures of cholesterol solutions ranging in concentration from 10 to 1000 {mu}M. These data are consistent with the Michaelis-Menten kinetic model for oxidation of cholesterol as catalyzed by cholesterol oxidase immobilized in the lipid bilayer membrane. The cholesterol detection limit is below 1 {mu}M for cholesterol solution prepared in buffered cyclodextrin. The response of the electrodes to low density lipoprotein solutions is increased upon addition of cyclodextrin. Evidence for adsorption of low density lipoprotein to the electrode surface is presented.

  20. Tetra- and Penta-Cyclic Triterpenes Interaction with Lipid Bilayer Membrane: A Structural Comparative Study. (United States)

    Abboud, Rola; Charcosset, Catherine; Greige-Gerges, Hélène


    The effect of tetracyclic (cortisol, prednisolone, and 9-fluorocortisol acetate) and pentacyclic (uvaol and erythrodiol) triterpenes (TTPs) on the fluidity of dipalmitoyl phosphatidyl choline (DPPC) liposome membrane was investigated by differential scanning calorimetry, Raman spectroscopy, and fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH). Liposomes were prepared in the absence and presence of TTPs at molar ratios DPPC:TTP 100:1, 100:2.5, and 100:10. All the studied TTPs abolished the pre-transition and modified the intensity of the Raman peak at 715 cm(-1) proving the interaction of TTP molecules with the choline head group of phospholipids. An increase in the Raman height intensity ratios of the peaks I 2935/2880, I 2844/2880, and I 1090/1130, giving information about the ratio disorder/order of the alkyl chains, and a decrease of the main transition temperature demonstrated the interaction of TTPs with the alkyl chains. The tetracyclic TTPs produced broadening of the phase transition profile. Besides, a scarcely splitting of the main transition peak was obtained with prednisolone and 9-fluorocortisol acetate. The results of fluorescence depolarization of DPH showed that the studied molecules fluidized the liposomal membrane at 25, 41, and 50 °C. Pentacyclic TTPs, being more hydrophobic than tetracyclic ones, demonstrated higher fluidizing effect than tetracyclic TTPs in the liquid crystalline phase suggesting a deeper incorporation in the lipid bilayer. The presence of a free polar head group at the ring D seems to control the TTP incorporation in the bilayer and consequently its effect on the membrane fluidity.

  1. Study of the ion-channel behavior on glassy carbon electrode supported bilayer lipid membranes stimulated by perchlorate anion

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Zhiquan; Shi, Jun; Huang, Weimin, E-mail:


    In this paper, a kind of didodecyldimethylammonium bromide (DDAB) layer membranes was supported on a glassy carbon electrode (GCE). We studied the ion channel behavior of the supported bilayer lipid membrane by scanning electrochemical microscopy (SCEM) in tris(2,2′-bipyridine) ruthenium(II) solution. Perchlorate anion was used as a presence of stimulus and ruthenium(II) complex cations as the probing ions for the measurement of SECM, the lipid membrane channel was opened and exhibited the behavior of distinct SECM positive feedback curve. The channel was in a closed state in the absence of perchlorate anions while reflected the behavior of SECM negative feedback curve. The rates of electron transfer reaction in the lipid membranes surface were detected and it was dependant on the potential of SECM. - Highlights: • The rates of electron transfer reaction in the lipid membranes surface were detected. • Dynamic investigations of ion-channel behavior of supported bilayer lipid membranes by scanning electrochemical microscopy • A novel way to explore the interaction between molecules and supported bilayer lipid membranes.

  2. On the freezing behavior and diffusion of water in proximity to single-supported zwitterionic and anionic bilayer lipid membranes

    DEFF Research Database (Denmark)

    Miskowiec, A.; Buck, Z. N.; Brown, M. C.


    We compare the freezing/melting behavior of water hydrating single-supported bilayers of a zwitterionic lipid DMPC with that of an anionic lipid DMPG. For both membranes, the temperature dependence of the elastically scattered neutron intensity indicates distinct water types undergoing translatio...

  3. Membrane fluidity and the surface properties of the lipid bilayer: ESR experiment and computer simulation. (United States)

    Man, Dariusz; Olchawa, Ryszard; Kubica, Krystian


    Penetration of the liposome membranes formed in the gel phase from DPPC (DPPC liposomes) and in the liquid-crystalline phase from egg yolk lecithin (EYL liposomes) by the TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl) and 16 DOXYL (2-ethyl-2-(15-methoxy-oxopentadecyl)-4,4-dimethyl-3-oxazolidinyloxy) spin probes has been investigated. The penetration process was followed by 120 hours at 24(0)C, using the electron spin resonance (ESR) method. The investigation of the kinetics of the TEMPO probe building into the membranes of both types of liposomes revealed differences appearing 30 minutes after the start of the experiment. The number of TEMPO particles built into the EYL liposome membranes began to clearly rise, aiming asymptotically to a constant value after about 100 minutes, whereas the number of the TEMPO particles built into the DPPC liposome membranes was almost constant in time. The interpretation of the obtained experimental results was enriched with those of computer simulation, following the behavior of the polar heads (dipoles) of the lipid particles forming a lipid layer due to the change in the value of the model parameter, k, determining the mobility of the dipoles. The possibility of the formation of an irregular ordering of the polar part of lipid membranes was proved, which leads to the appearance of spaces filled with of water for k > 0.4. The appearance of these defects enables the penetration of the bilayer by the TEMPO particles. The limited mobility of lipid polar heads (k < 0.2) prevents the appearance of such areas facilitating the penetration of the lipid membrane by alien particles in the gel phase.

  4. Simulations of a Membrane-Anchored Peptide: Structure, Dynamics, and Influence on Bilayer Properties

    DEFF Research Database (Denmark)

    Jensen, Morten Østergaard; Mouritsen, O.G.; Peters, Günther H.J.


    A three-dimensional structure of a model decapeptide is obtained by performing molecular dynamics simulations of the peptide in explicit water. Interactions between an N-myristoylated form of the folded peptide anchored to dipalmitoylphosphatidylcholine fluid phase lipid membranes are studied at ...

  5. A Parallel Sensing Technique for Automatic Bilayer Lipid Membrane Arrays Monitoring

    Directory of Open Access Journals (Sweden)

    Michele ROSSI


    Full Text Available Ion channels are transmembrane proteins responsible of cell signaling and a large part of pharmaceutical compounds are interacting with them. In-vitro testing of ion channels is a promising technique for high throughput screening (HTS in drug discovery and personalized medicine. Automated tests of single ion channels embedded in artificial bilayer lipid membranes (BLM is gaining attention over patch clamp technique due to its characteristic of performing parallel tests on selected sets of channels or multiple pharmaceutical targets. However, BLM arrays formation is a critical process based on manual and time-consuming techniques. In this paper, an automatic liquid dispensing system for BLM formation monitored in real-time by using low-noise current amplifiers is presented. As proof of this approach, concurrent formation of BLMs is shown. Additionally, single ion channel recordings on an automatically formed BLM is presented and discussed.

  6. Bax and Bif-1 proteins interact on Bilayer Lipid Membrane and form pore. (United States)

    Gupta, Rajeev; Ghosh, Subhendu


    Bax and Bax interacting factor-1(Bif-1) are cytosolic proteins, which translocate towards mitochondria during mitochondria-mediated apoptosis. Bif-1 has been identified to co-immunoprecipitate with Bax in apoptotic cells. We have studied the interaction of Bax and Bif-1 on Bilayer Lipid Membrane (BLM) through electrophysiological experiments. It has been observed that Bax-Bif-1 equimolar mixture can form a pore. The pore conductance is in the range of 4.96-5.41 nS. It also displays a sub-state with a conductance of 2.6 nS. No pore activity is observed on BLM when monomeric Bax and Bif-1 proteins are tested independently. The above-mentioned pore forming activity could be relevant in mitochondria-mediated apoptosis. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Optimization of the hybrid bilayer membrane method for immobilization of avidin on quartz crystal microbalance. (United States)

    Mun, Saem; Choi, Suk-Jung


    Hybrid bilayer membrane (HBM), comprising a lipid monolayer fused to a hydrophobic self-assembled monolayer (SAM), has a potential capability to provide a convenient tool for the preparation of functionalized sensor surfaces. In this work, the HBM approach was optimized for the preparation of avidin-containing quartz crystal microbalance (QCM) sensor chip which would be available for immobilization of biotinylated molecules. Lipid layer of HBM was composed of background lipid such as egg phosphatidyl choline and biotinylated lipid to which avidin was attached. Highest performance was obtained at 1:1 ratio of the biotinylated lipid and the background lipid, and sensitivity and stability of the resulting sensor chip was comparable to a sensor chip prepared by the conventional carbodiimide reaction. By utilizing the HBM method, construction of a stable avidin sensor chip was achieved within 40 min without any chemical steps. Thus the HBM method was proven to be a convenient and efficient way to immobilize avidin on sensor surfaces.

  8. Tubular lipid membranes pulled from vesicles: Dependence of system equilibrium on lipid bilayer curvature (United States)

    Golushko, I. Yu.; Rochal, S. B.


    Conditions of joint equilibrium and stability are derived for a spherical lipid vesicle and a tubular lipid membrane (TLM) pulled from this vesicle. The obtained equations establish relationships between the geometric and physical characteristics of the system and the external parameters, which have been found to be controllable in recent experiments. In particular, the proposed theory shows that, in addition to the pressure difference between internal and external regions of the system, the variable spontaneous average curvature of the lipid bilayer (forming the TLM) also influences the stability of the lipid tube. The conditions for stability of the cylindrical phase of TLMs after switching off the external force that initially formed the TLM from a vesicle are discussed. The loss of system stability under the action of a small axial force compressing the TLM is considered.

  9. Syringotoxin pore formation and inactivation in human red blood cell and model bilayer lipid membranes. (United States)

    Szabó, Zsófia; Gróf, Pál; Schagina, Ludmila V; Gurnev, Philip A; Takemoto, Jon Y; Mátyus, Edit; Blaskó, Katalin


    The effect of syringotoxin (ST), a member of the cyclic lipodepsipeptides family (CLPs) produced by Pseudomonas syringae pv. syringae on the membrane permeability of human red blood cells (RBCs) and model bilayer lipid membranes (BLMs) was studied and compared to that of two recently investigated CLPs, syringomycin E (SRE) and syringopeptin 22A (SP22A) [Biochim. Biophys. Acta 1466 (2000) 79 and Bioelectrochemistry 52 (2000) 161]. The permeability-increasing effect of ST on RBCs was the least among the three CLPs. A time-dependent ST pore inactivation was observed on RBCs at 20 and 37 degrees C but not at 8 degrees C. From the kinetic model worked out parameters as permeability coefficient of RBC membrane for 86Rb(+) and pores mean lifetime were calculated. A shorter pores mean lifetime was calculated at 37 degrees C then at 20 degrees C, which gave us an explanation for the unusual slower rate of tracer efflux measured at 37 degrees C then that at 20 degrees C. The results obtained on BLM showed that the pore inactivation was due to a decrease in the number of pores but not to a change of their dwell time or conductance.

  10. Na+/D-glucose cotransporter based bilayer lipid membrane sensor for D-glucose. (United States)

    Sugao, N; Sugawara, M; Minami, H; Uto, M; Umezawa, Y


    A new type of amperometric blosensor for glucose was fabricated using a Na+/D-glucose cotransporter as the signal-transducing sensory element that exploits the D-glucose-triggered Na+ ion current through bilayer lipid membranes (BLMs). The planar BLM was formed by the folding method across a small aperture of a thin Teflon film. The Na+/D-glucose cotransporter, isolated and purified from small intestinal brush border membrane of guinea pigs, was embedded into BLMs through proteoliposomes. The number of the protein molecules thus incorporated in the present sensing membrane was estimated to be ca. 10(7). The sensor response was measured as an ionic current through the BLM arising from cotransported Na+ ion flux under a constant applied potential and was only induced by D-glucose above 10(-9) M, but not by the other monosaccharides except for D-galactose. The effect of applied potentials, Na+ and K+ ion concentrations, and the addition of a competitive inhibitor, phlorizin, were scrutinized to characterize the sensor output. The results were briefly discussed in terms of the potential use of the Na+/D-glucose cotransporter as a sensory element for D-glucose.

  11. Triglyceride blisters in lipid bilayers: implications for lipid droplet biogenesis and the mobile lipid signal in cancer cell membranes.

    Directory of Open Access Journals (Sweden)

    Himanshu Khandelia

    Full Text Available Triglycerides have a limited solubility, around 3%, in phosphatidylcholine lipid bilayers. Using millisecond-scale course grained molecular dynamics simulations, we show that the model lipid bilayer can accommodate a higher concentration of triolein (TO than earlier anticipated, by sequestering triolein molecules to the bilayer center in the form of a disordered, isotropic, mobile neutral lipid aggregate, at least 17 nm in diameter, which forms spontaneously, and remains stable on at least the microsecond time scale. The results give credence to the hotly debated existence of mobile neutral lipid aggregates of unknown function present in malignant cells, and to the early biogenesis of lipid droplets accommodated between the two leaflets of the endoplasmic reticulum membrane. The TO aggregates give the bilayer a blister-like appearance, and will hinder the formation of multi-lamellar phases in model, and possibly living membranes. The blisters will result in anomalous membrane probe partitioning, which should be accounted for in the interpretation of probe-related measurements.

  12. Fluctuations in lipid bilayers: Are they understood?

    CERN Document Server

    Schmid, Friederike


    We review recent computer simulation studies of undulating lipid bilayers. Theoretical interpretations of such fluctuating membranes are most commonly based on generalized Helfrich-type elastic models, with additional contributions of local "protrusions" and/or density fluctuations. Such models provide an excellent basis for describing the fluctuations of tensionless bilayers in the fluid phase at a quantitative level. However, this description is found to fail for membranes in the gel phase and for membranes subject to high tensions. The fluctuations of tilted gel membranes show a signature of the modulated ripple structure, which is a nearby phase observed in the pretransition regime between the fluid and tilted gel state. This complicates a quantitative analysis on mesoscopic length scales. In the case of fluid membranes under tension, the large-wavelength fluctuation modes are found to be significantly softer than predicted by theory. In the latter context, we also address the general problem of the relat...

  13. Supported Lipid Bilayer Platform To Test Inhibitors of the Membrane Attack Complex: Insights into Biomacromolecular Assembly and Regulation. (United States)

    Yorulmaz, Saziye; Jackman, Joshua A; Hunziker, Walter; Cho, Nam-Joon


    Complement activation plays an important role in innate immune defense by triggering formation of the membrane attack complex (MAC), which is a biomacromolecular assembly that exhibits membrane-lytic activity against foreign invaders including various pathogens and biomaterials. Understanding the details of MAC structure and function has been the subject of extensive work involving bulk liposome and erythrocyte assays. However, it is difficult to characterize the mechanism of action of MAC inhibitor drug candidates using the conventional assays. To address this issue, we employ a biomimetic supported lipid bilayer platform to investigate how two MAC inhibitors, vitronectin and clusterin, interfere with MAC assembly in a sequential addition format, as monitored by the quartz crystal microbalance-dissipation (QCM-D) technique. Two experimental strategies based on modular assembly were selected, precincubation of inhibitor and C5b-7 complex before addition to the lipid bilayer or initial addition of inhibitor followed by the C5b-7 complex. The findings indicate that vitronectin inhibits membrane association of C5b-7 via a direct interaction with C5b-7 and via competitive membrane association onto the supported lipid bilayer. On the other hand, clusterin directly interacts with C5b-7 such that C5b-7 is still able to bind to the lipid bilayer, and clusterin affects the subsequent binding of other complement proteins involved in the MAC assembly. Taken together, the findings in this study outline a biomimetic approach based on supported lipid bilayers to explore the interactions between complement proteins and inhibitors, thereby offering insight into MAC assembly and regulation.

  14. Drug binding and mobility relating to the thermal fluctuation in fluid lipid membranes (United States)

    Okamura, Emiko; Yoshii, Noriyuki


    Drug binding and mobility in fluid lipid bilayer membranes are quantified in situ by using the multinuclear solution NMR combined with the pulsed-field-gradient technique. One-dimensional and pulsed-field-gradient F19 and H1 NMR signals of an anticancer drug, 5-fluorouracil (5FU) are analyzed at 283-313 K in the presence of large unilamellar vesicles (LUVs) of egg phosphatidylcholine (EPC) as model cell membranes. The simultaneous observation of the membrane-bound and free 5FU signals enables to quantify in what amount of 5FU is bound to the membrane and how fast 5FU is moving within the membrane in relation to the thermal fluctuation of the soft, fluid environment. It is shown that the mobility of membrane-bound 5FU is slowed down by almost two orders of magnitude and similar to the lipid movement in the membrane, the movement closely related to the intramembrane fluidity. The mobility of 5FU and EPC is, however, not similar at 313 K; the 5FU movement is enhanced in the membrane as a result of the loose binding of 5FU in the lipid matrices. The membrane-bound fraction of 5FU is ˜0.1 and almost unaltered over the temperature range examined. It is also independent of the 5FU concentration from 2 to 30 mM with respect to the 40-50 mM LUV. The free energy of the 5FU binding is estimated at -4 to -2 kJ/mol, the magnitude always close to the thermal fluctuation, 2.4-2.6 kJ/mol.

  15. Lipid Bilayers in the Gel Phase Become Saturated by Triton X-100 at Lower Surfactant Concentrations Than Those in the Fluid Phase (United States)

    Ahyayauch, Hasna; Collado, M. Isabel; Alonso, Alicia; Goñi, Felix M.


    It has been repeatedly observed that lipid bilayers in the gel phase are solubilized by lower concentrations of Triton X-100, at least within certain temperature ranges, or other nonionic detergents than bilayers in the fluid phase. In a previous study, we showed that detergent partition coefficients into the lipid bilayer were the same for the gel and the fluid phases. In this contribution, turbidity, calorimetry, and 31P-NMR concur in showing that bilayers in the gel state (at least down to 13–20°C below the gel-fluid transition temperature) become saturated with detergent at lower detergent concentrations than those in the fluid state, irrespective of temperature. The different saturation may explain the observed differences in solubilization. PMID:22713566

  16. Design and characterization of a membrane protein unfolding platform in lipid bilayers.

    Directory of Open Access Journals (Sweden)

    Vincent G Nadeau

    Full Text Available Accurate measurement of membrane protein stability--and particularly how it may vary as a result of disease-phenotypic mutations--ideally requires a denaturant that can unfold a membrane-embedded structure while leaving the solubilizing environment unaffected. The steric trap method fulfills this requirement by using monovalent streptavidin (mSA molecules to unfold membrane proteins engineered with two spatially close biotin tags. Here we adapted this method to an 87-residue helix-loop-helix (hairpin construct derived from helices 3 and 4 in the transmembrane domain of the human cystic fibrosis transmembrane conductance regulator (CFTR, wherein helix-helix tertiary interactions are anticipated to confer a portion of construct stability. The wild type CFTR TM3/4 hairpin construct was modified with two accessible biotin tags for mSA-induced unfolding, along with two helix-terminal pyrene labels to monitor loss of inter-helical contacts by pyrene excimer fluorescence. A series of eight constructs with biotin tags at varying distances from the helix-terminal pyrene labels were expressed, purified and labeled appropriately; all constructs exhibited largely helical circular dichroism spectra. We found that addition of mSA to an optimized construct in lipid vesicles led to a complete and reversible loss in pyrene excimer fluorescence and mSA binding, and hence hairpin unfolding--results further supported by SDS-PAGE visualization of mSA bound and unbound species. While some dimeric/oligomeric populations persist that may affect quantitation of the unfolding step, our characterization of the design yields a promising prototype of a future platform for the systematic study of membrane protein folding in a lipid bilayer environment.

  17. Monitoring of cholesterol oxidation in a lipid bilayer membrane using streptolysin O as a sensing and signal transduction element. (United States)

    Shoji, Atsushi; Ikeya, Kana; Aoyagi, Miki; Takatsuji, Ryutaro; Yanagida, Akio; Shibusawa, Yoichi; Sugawara, Masao


    Streptolysin O (SLO), which recognizes sterols and forms nanopores in lipid membranes, is proposed as a sensing element for monitoring cholesterol oxidation in a lipid bilayer. The structural requirements of eight sterols for forming nanopores by SLO confirmed that a free 3-OH group in the β-configuration of sterols is required for recognition by SLO in a lipid bilayer. The extent of nanopore formation by SLO in lipid bilayers increased in the order of cholestanolbilayer. The potential of the SLO nanopore-based method for monitoring cholesterol oxidation in a lipid bilayer by other oxidative enzymes is also discussed.

  18. Modulation of folding and assembly of the membrane protein bacteriorhodopsin by intermolecular forces within the lipid bilayer. (United States)

    Curran, A R; Templer, R H; Booth, P J


    Three different lipid systems have been developed to investigate the effect of physicochemical forces within the lipid bilayer on the folding of the integral membrane protein bacteriorhodopsin. Each system consists of lipid vesicles containing two lipid species, one with phosphatidylcholine and the other with phosphatidylethanolamine headgroups, but the same hydrocarbon chains: either L-alpha-1, 2-dioleoyl, L-alpha-1,2-dipalmitoleoyl, or L-alpha-1,2-dimyristoyl. Increasing the mole fraction of the phosphatidylethanolamine lipid increases the desire of each monolayer leaflet in the bilayer to curve toward water. This increases the torque tension of such monolayers, when they are constrained to remain flat in the vesicle bilayer. Consequently, the lateral pressure in the hydrocarbon chain region increases, and we have used excimer fluorescence from pyrene-labeled phosphatidylcholine lipids to probe these pressure changes. We show that bacteriorhodopsin regenerates to about 95% yield in vesicles of 100% phosphatidylcholine. The regeneration yield decreases as the mole fraction of the corresponding phosphatidylethanolamine component is increased. The decrease in yield correlates with the increase in lateral pressure which the lipid chains exert on the refolding protein. We suggest that the increase in lipid chain pressure either hinders insertion of the denatured state of bacterioopsin into the bilayer or slows a folding step within the bilayer, to the extent that an intermediate involved in bacteriorhodopsin regeneration is effectively trapped.

  19. Diffusion studies on permeable nitroxyl spin probes through bilayer lipid membranes: A low frequency ESR study

    Energy Technology Data Exchange (ETDEWEB)

    Meenakumari, V.; Benial, A. Milton Franklin, E-mail: [Department of Physics, NMSSVN College, Nagamalai, Madurai-625019, Tamilnadu (India); Utsumi, Hideo; Ichikawa, Kazuhiro; Yamada, Ken-ichi [Department of Bio-functional Science, Kyushu University, Fukuoka (Japan); Hyodo, Fuminori [Innovation Center for Medical Redox Navigation, Kyushu University, Fukuoka (Japan); Jawahar, A. [Department of Chemistry, NMSSVN College, Nagamalai, Madurai-625019, Tamilnadu (India)


    Electron spin resonance (ESR) studies were carried out for permeable 2mM {sup 14}N-labeled deutrated 3 Methoxy carbonyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl (MC-PROXYL) in pure water and 1mM, 2mM, 3mM, 4mM concentration of 14N-labeled deutrated MC-PROXYL in 400mM concentration of liposomal solution by using a 300 MHz ESR spectrometer. The ESR parameters such as linewidth, hyperfine coupling constant, g-factor, partition parameter and permeability were reported for these samples. The line broadening was observed for the nitroxyl spin probe in the liposomal solution. The line broadening indicates that the high viscous nature of the liposomal solution. The partition parameter and permeability values indicate the maximum diffusion of nitroxyl spin probes in the bilayer lipid membranes at 2 mM concentration of nitroxyl radical. This study illustrates that ESR can be used to differentiate between the intra and extra- membrane water by loading the liposome vesicles with a lipid-permeable nitroxyl spin probe. From the ESR results, the spin probe concentration was optimized as 2mM in liposomal solution for ESR phantom studies/imaging, invivo and invitro experiments.

  20. Correlating anomalous diffusion with lipid bilayer membrane structure using single molecule tracking and atomic force microscopy (United States)

    Skaug, Michael J.; Faller, Roland; Longo, Marjorie L.


    Anomalous diffusion has been observed abundantly in the plasma membrane of biological cells, but the underlying mechanisms are still unclear. In general, it has not been possible to directly image the obstacles to diffusion in membranes, which are thought to be skeleton bound proteins, protein aggregates, and lipid domains, so the dynamics of diffusing particles is used to deduce the obstacle characteristics. We present a supported lipid bilayer system in which we characterized the anomalous diffusion of lipid molecules using single molecule tracking, while at the same time imaging the obstacles to diffusion with atomic force microscopy. To explain our experimental results, we performed lattice Monte Carlo simulations of tracer diffusion in the presence of the experimentally determined obstacle configurations. We correlate the observed anomalous diffusion with obstacle area fraction, fractal dimension, and correlation length. To accurately measure an anomalous diffusion exponent, we derived an expression to account for the time-averaging inherent to all single molecule tracking experiments. We show that the length of the single molecule trajectories is critical to the determination of the anomalous diffusion exponent. We further discuss our results in the context of confinement models and the generating stochastic process.

  1. Tethered bilayer lipid membranes studied by simultaneous attenuated total reflectance infrared spectroscopy and electrochemical impedance spectroscopy (United States)

    Erbe, Andreas; Bushby, Richard J.; Evans, Stephen D.; Jeuken, Lars J. C.


    The formation of tethered lipid bilayer membranes (tBLMs) from unilamelar vesicles of egg yolk phosphatidylcholine (EggPC) on mixed self–assembled monolayers (SAMs) from varying ratios of 6-mercaptohexanol and EO3Cholesteryl on gold has been monitored by simultaneous attenuated total reflectance fourier transform infrared (ATR–FTIR) spectroscopy and electrochemical impedance spectroscopy (EIS). The influence of the lipid orientation (and hence the anisotropy) of lipids on a gold film on the dichroic ratio was studied by simulations of spectra with a matrix method for anisotropic layers. It is shown that for certain tilt angles of the dielectric tensor of the adsorbed anisotropic layer dispersive and negative absorption bands are possible. The experimental data indicates that the structure of the assemblies obtained varies with varying SAM composition. On SAMs with a high content of EO3Cholesteryl, tBLMs with reduced fluidity are formed. For SAMs with high content of 6-mercaptohexanol, the results are consistent with the adsorption of flattened vesicles, while spherical vesicles have been found in a small range of surface compositions. The kinetics of the adsorption process is consistent with the assumption of spherical vesicles as long–living intermediates for surfaces of high 6-mercaptohexanol content. No long–living spherical vesicles have been detected for surfaces with large fraction of EO3Cholesteryl tethers. The observed differences between the surfaces suggest that for the formation of tBLMs (unlike supported BLMs) no critical surface coverage of vesicles is needed prior to lipid bilayer formation. PMID:17388505

  2. X-Ray Kinematography of Temperature-Jump Relaxation Probes the Elastic Properties of Fluid Bilayers

    CERN Document Server

    Pabst, G; Amenitsch, H; Bernstorff, S; Laggner, P; Pabst, Georg; Rappolt, Michael; Amenitsch, Heinz; Bernstorff, Sigrid; Laggner, Peter


    The response kinetics of liquid crystalline phosphatidylcholine bilayerstacks to rapid, IR-laser induced temperature jumps has been studied bymillisecond time-resolved x-ray diffraction. The system reacts on the fasttemperature change by a discrete bilayer compression normal to its surface anda lateral bilayer expansion. Since water cannot diffuse from the excess phaseinto the interbilayer water region within the 2 ms duration of the laser pulse,the water layer has to follow the bilayer expansion, by an anomalous thinning.Structural analysis of a 20 ms diffraction pattern from the intermediate phaseindicates that the bilayer thickness remains within the limits of isothermalequilibrium values. Both, the intermediate structure and its relaxation intothe original equilibrium L_(alpha)-phase, depend on the visco-elasticproperties of the bilayer/water system. We present an analysis of therelaxation process by an overdamped one-dimensional oscillation model revealingthe concepts of Hooke's law for phospholipid bila...

  3. Coarse-grained molecular dynamics simulations of shear-induced instabilities of lipid bilayer membranes in water (United States)

    Hanasaki, Itsuo; Walther, Jens H.; Kawano, Satoyuki; Koumoutsakos, Petros


    We study shear-induced instabilities of lipid bilayers immersed in water using coarse-grained molecular dynamics simulations. The shear imposed by the flow of the water induces initially microscopic structural changes of the membrane, starting with tilting of the molecules in the direction of the shear. The tilting propagates in the spanwise direction when the shear rate exceeds a critical value and the membrane undergoes a bucklinglike deformation in the direction perpendicular to the shear. The bucklinglike undulation continues until a localized Kelvin-Helmholtz-like instability leads to membrane rupture. We study the different modes of membrane undulation using membranes of different geometries and quantify the relative importance of the bucklinglike bending and the Kelvin-Helmholtz-like instability of the membrane.

  4. Free vibration of membrane/bounded incompressible fluid

    Institute of Scientific and Technical Information of China (English)



    Vibration of a circular membrane in contact with a fluid has extensive applications in industry.The natural vibration frequencies for the asymmetric free vibration of a circular membrane in contact with a bounded incompressible fluid are derived in this paper.Considering small oscillations induced by the membrane vibration in an incompressible and inviscid fluid,the velocity potential function is used to describe the fluid field.Two approaches are used to derive the free vibration frequencies of the system,which include a variational formulation and an approximate solution employing the Rayleigh quotient method.A good correlation is found between free vibration frequencies evaluated by these methods.Finally,the effects of the fluid depth,the mass density,and the radial tension on the free vibration frequencies of the coupled system are investigated.

  5. Lateral diffusion of peripheral membrane proteins on supported lipid bilayers is controlled by the additive frictional drags of (1) bound lipids and (2) protein domains penetrating into the bilayer hydrocarbon core. (United States)

    Ziemba, Brian P; Falke, Joseph J


    Peripheral membrane proteins bound to lipids on bilayer surfaces play central roles in a wide array of cellular processes, including many signaling pathways. These proteins diffuse in the plane of the bilayer and often undergo complex reactions involving the binding of regulatory and substrate lipids and proteins they encounter during their 2D diffusion. Some peripheral proteins, for example pleckstrin homology (PH) domains, dock to the bilayer in a relatively shallow position with little penetration into the bilayer. Other peripheral proteins exhibit more complex bilayer contacts, for example classical protein kinase C isoforms (PKCs) bind as many as six lipids in stepwise fashion, resulting in the penetration of three PKC domains (C1A, C1B, C2) into the bilayer headgroup and hydrocarbon regions. A molecular understanding of the molecular features that control the diffusion speeds of proteins bound to supported bilayers would enable key molecular information to be extracted from experimental diffusion constants, revealing protein-lipid and protein-bilayer interactions difficult to study by other methods. The present study investigates a range of 11 different peripheral protein constructs comprised by 1-3 distinct domains (PH, C1A, C1B, C2, anti-lipid antibody). By combining these constructs with various combinations of target lipids, the study measures 2D diffusion constants on supported bilayers for 17 different protein-lipid complexes. The resulting experimental diffusion constants, together with the known membrane interaction parameters of each complex, are used to analyze the molecular features correlated with diffusional slowing and bilayer friction. The findings show that both (1) individual bound lipids and (2) individual protein domains that penetrate into the hydrocarbon core make additive contributions to the friction against the bilayer, thereby defining the 2D diffusion constant. An empirical formula is developed that accurately estimates the diffusion

  6. Biochemical studies of pigments from a pathogenic fungus Microsporum cookei. V. Evidence for the transmembrane permeability of xanthomegnin across phospholipid bilayer membranes. (United States)

    Kawai, K; Akita, T; Nozawa, Y


    Direct evidence is provided for the transmembrane permeation of xanthomegnin across phospholipid bilayer membranes using ascorbate-loaded liposomes. This process may be associated with an uncoupling effect on the oxidative phosphorylation of mitochondria.

  7. Modeling anisotropic elasticity of fluid membranes

    CERN Document Server

    Ramakrishnan, N; Ipsen, John H; 10.1002/mats.201100002


    The biological membrane, which compartmentalizes the cell and its organelles, exhibit wide variety of macroscopic shapes of varying morphology and topology. A systematic understanding of the relation of membrane shapes to composition, external field, environmental conditions etc. have important biological relevance. Here we review the triangulated surface model, used in the macroscopic simulation of membranes and the associated Monte Carlo (DTMC) methods. New techniques to calculate surface quantifiers, that will facilitate the study of additional in-plane orientational degrees of freedom, has been introduced. The mere presence of a polar and nematic fields in the ordered phase drives the ground state conformations of the membrane to a cylinder and tetrahedron respectively.

  8. Fluid Mosaic Membranes and the Light Reactions of Photosynthesis. (United States)

    Hannay, Jack


    Discusses: (1) the fluid mosaic membrane structure and light reactions of photosynthesis as exemplified by the Hill and Bendall "Z-scheme"; (2) the arrangement of light-harvesting pigments, electron transport components, and ATP synthesis on chloroplast membranes; and (3) how these topics are treated in A-level textbooks. (JN)

  9. Reduction in lateral lipid mobility of lipid bilayer membrane by atmospheric pressure plasma irradiation (United States)

    Suda, Yoshiyuki; Tero, Ryugo; Yamashita, Ryuma; Yusa, Kota; Takikawa, Hirofumi


    Plasma medicine is an emerging research field in which various applications of electrical discharge, especially in the form of nonequilibrium plasma at atmospheric pressure, are examined, for example, the application of plasma to biological targets for various purposes such as selective killing of tumor cells and blood stanching. We have focused on the behavior of an artificial cell membrane system at the solid-liquid interface. To evaluate the lateral lipid mobility, we measured the diffusion coefficient of the supported lipid bilayer (SLB) composed of dioleoylphosphatidylcholine with fluorescence recovery after photobleaching by confocal laser scanning microscopy. It was found that the diffusion coefficient was decreased by plasma irradiation and that the diffusion coefficient decreasing rate proceeded with increasing plasma power. We investigated the effects of stimulation with an equilibrium chemical, H2O2, on the SLB and confirmed that the diffusion coefficient did not change at least up to a H2O2 concentration of 5 mM. These results indicate that transient active species generated by plasma play critical roles in the reduction in SLB fluidity. The effects of the two generated major oxidized lipid species, hydroxyl- or hydroperoxy-phosphatidylcholine (PC) and acyl-chain-truncated PCs terminated with aldehyde or carboxyl group, on lateral lipid mobility are discussed.

  10. Nanosized CaP-silk fibroin-PCL-PEG-PCL/PCL based bilayer membranes for guided bone regeneration. (United States)

    Türkkan, Sibel; Pazarçeviren, A Engin; Keskin, Dilek; Machin, Nesrin E; Duygulu, Özgür; Tezcaner, Ayşen


    Guided bone regeneration (GBR) concept has been developed to prevent the formation of non-functional scar tissue layer on defect site by undertaking barrier role. In this study, a new bilayer membrane which consisted of one layer of electrospun silk fibroin/PCL-PEG-PCL incorporating nanocalcium phosphate (SPCA)(1) and one layer of PCL membrane was developed for GBR. To improve the osteoconductivity of membranes, nanosized calcium phosphate particles synthesized by Flame Spray Pyrolysis method were incorporated into membranes at 10% (wt) (SPCA10) and 20% (wt) (SPCA20) of the polymer content. The structural and chemical analyses revealed the well-integrated two layers of membranes with a total thickness of ca 100μm. In the regenerative layer, the highly porous mesh structure had a thickness of 12.6μm with randomly oriented fibers having diameters around 760nm, and nanoparticles dispersed homogenously. The mechanical test results showed remarkable improvement on the tensile strength of membranes with incorporation of nanoparticles. Higher water affinity of nanoCaP included membranes was proved by lower contact angle values and higher percent water uptake capacity. Biomineralization assay revealed that nucleation and growth of apatites around fibers of SPCA10 and SPCA20 were apparent while on SPCA0 apatite minerals were barely detected after 10days. Human dental pulp stem cells (DPSC) were seeded on electrospun layer of the bilayer membranes for biocompatibility and osteo-compatibility study. Increasing nanoCaP amount resulted in higher cell adhesion, proliferation, ALP activity and calcium deposition on membranes. These overall results confirmed the biocompatibility and potential applicability of proposed membranes for GBR treatments. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Diffusion and spectroscopy of water and lipids in fully hydrated dimyristoylphosphatidylcholine bilayer membranes

    Energy Technology Data Exchange (ETDEWEB)

    Yang, J.; Martí, J., E-mail: [Department of Physics and Nuclear Engineering, Technical University of Catalonia-Barcelona Tech, B4-B5 Northern Campus, Jordi Girona 1-3, 08034 Barcelona, Catalonia (Spain); Calero, C. [Department of Physics and Nuclear Engineering, Technical University of Catalonia-Barcelona Tech, B4-B5 Northern Campus, Jordi Girona 1-3, 08034 Barcelona, Catalonia (Spain); Center for Polymer Studies, Department of Physics, Boston University, 590 Commonwealth Avenue, Boston, Massachusetts 02215 (United States)


    Microscopic structure and dynamics of water and lipids in a fully hydrated dimyristoylphosphatidylcholine phospholipid lipid bilayer membrane in the liquid-crystalline phase have been analyzed with all-atom molecular dynamics simulations based on the recently parameterized CHARMM36 force field. The diffusive dynamics of the membrane lipids and of its hydration water, their reorientational motions as well as their corresponding spectral densities, related to the absorption of radiation, have been considered for the first time using the present force field. In addition, structural properties such as density and pressure profiles, a deuterium-order parameter, surface tension, and the extent of water penetration in the membrane have been analyzed. Molecular self-diffusion, reorientational motions, and spectral densities of atomic species reveal a variety of time scales playing a role in membrane dynamics. The mechanisms of lipid motion strongly depend on the time scale considered, from fast ballistic translation at the scale of picoseconds (effective diffusion coefficients of the order of 10{sup −5} cm{sup 2}/s) to diffusive flow of a few lipids forming nanodomains at the scale of hundreds of nanoseconds (diffusion coefficients of the order of 10{sup −8} cm{sup 2}/s). In the intermediate regime of sub-diffusion, collisions with nearest neighbors prevent the lipids to achieve full diffusion. Lipid reorientations along selected directions agree well with reported nuclear magnetic resonance data and indicate two different time scales, one about 1 ns and a second one in the range of 2–8 ns. We associated the two time scales of reorientational motions with angular distributions of selected vectors. Calculated spectral densities corresponding to lipid and water reveal an overall good qualitative agreement with Fourier transform infrared spectroscopy experiments. Our simulations indicate a blue-shift of the low frequency spectral bands of hydration water as a result of

  12. Study of the Ion Channel Behavior of Didodecyldimethylammonium Bromide Formed Bilayer Lipid Membrane Stimulated by PF-6

    Institute of Scientific and Technical Information of China (English)

    TONG,Yue-Hong; HAN,Xiao-Jun; WANG,Er-Kang


    Bilayer lipid membranes ( BLM ) formed from didodecyldimethylammonium bromide were made on the freshly exposed surface ofa glassy carbon (GC) ani were demonstrated by the ac impedance spectroscopy. The ion channels of membrane properties induced by PF6- were studied by the cyclic voltammetric methods.Experimental results indicated that the ion channel of BLM was open in the presence of the PF6- due to the interaction of PF6- with the BLM, while it was switched offin the absence of PF6-. Because the ion channel behavior was affected by the concentration of PF6-,a sensor for PF6- can be developed.

  13. Nanodiscs for immobilization of lipid bilayers and membrane receptors: kinetic analysis of cholera toxin binding to a glycolipid receptor

    DEFF Research Database (Denmark)

    Borch, Jonas; Torta, Federico; Sligar, Stephen G;


    nanodiscs and their incorporated membrane receptors can be attached to surface plasmon resonance sensorchips and used to measure the kinetics of the interaction between soluble molecules and membrane receptors inserted in the bilayer of nanodiscs. Cholera toxin and its glycolipid receptor G(M1) constitute...... partner cholera toxin B subunit to the receptor with the sensorchip-based surface plasmon resonance (SPR) technology. The measured stoichiometric and kinetic values of the interaction are in agreement with those reported by previous studies, thus providing proof-of-principle that nanodiscs can be employed...

  14. Oxidation of Membrane Curvature-Regulating Phosphatidylethanolamine Lipid Results in Formation of Bilayer and Cubic Structures. (United States)

    Sankhagowit, Shalene; Lee, Ernest Y; Wong, Gerard C L; Malmstadt, Noah


    Oxidation is associated with conditions related to chronic inflammations and aging. Cubic structures have been observed in the smooth endoplasmic reticulum and mitochondrial membranes of cells under oxidative stress (e.g., tumor cells and virus-infected cells). It has been previously suspected that oxidation can result in the rearrangement of lipids from a fluid lamellar phase to a cubic structure in organelles containing membranes enriched with amphiphiles that have nonzero intrinsic curvature, such as phosphatidylethanolamine (PE) and cardiolipin. This study focuses on the oxidation of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), a lipid that natively forms an inverted hexagonal phase at physiological conditions. The oxidized samples contain an approximately 3:2 molar ratio of nonoxidized to oxidized DOPE. Optical microscopy images collected during the hydration of this mixture from a dried film suggest that the system evolves into a coexistence of a stable fluid lamellar phase and transient square lattice structures with unit cell sizes of 500-600 nm. Small-angle X-ray scattering of the same lipid mixture yielded a body-centered Im3m cubic phase with the lattice parameter of 14.04 nm. On average, the effective packing parameter of the oxidized DOPE species was estimated to be 0.657 ± 0.069 (standard deviation). This suggests that the oxidation of PE leads to a group of species with inverted molecular intrinsic curvature. Oxidation can create amphiphilic subpopulations that potently impact the integrity of the membrane, since negative Gaussian curvature intrinsic to cubic phases can enable membrane destabilization processes.

  15. Fundamental Studies of Assembly and Mechanical Properties of Lipid Bilayer Membranes and Unilamellar Vesicles (United States)

    Wang, Xi

    This dissertation work focuses on: (i) obtaining a phospholipid bilayer membrane (LBM)/conducting electrode system with low defect density and optimized rigidity; (ii) investigating vesicle stability and mechanical properties. LBM is a simplified yet representative cell membrane model. LBMs assembled on conductive surfaces can probe protein-LBM interactions activities electrochemically. Sterically stabilized vesicles could be used as cell models or for drug delivery. The main challenges for LBM assembly on gold are vesicles do not spontaneously rupture to form LBMs on gold and the roughness of the gold substrate has considerable influence on molecular film defect density. In this study, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) vesicles were functionalized with 1,2-distearoyl-sn-glycero-3-phosphoethanolamine- N-poly(ethylene glycol)-2000-N-[3-(2-pyridyldithio)propionate] (DSPE-PEG-PDP) to yield stable LBMs on gold without surface modification. A template-stripping method was used to obtain atomically flat and pristine gold surfaces. The critical force to initiate vesicle rupture decreases with increasing DSPE-PEG-PDP concentration, indicating that gold-thiolate bonding between DSPE-PEG-PDP and gold substrates promotes LBM formation. Mechanical properties of LBMs and vesicles were investigated as a function of DSPE-PEG-PDP concentration via Atomic Force Microscopy. The elastic moduli of LBMs were determined with DSPE-PEG-PDP concentration ranging from 0mol% to 24mol% and were found to depend on PEG chain conformation. Incorporating DSPE-PEG-PDP molecules with PEG in mushroom conformation results in a decrease of LBM rigidity, while incorporating PEG in brush conformation leads to LBM stiffening. Contrarily, mechanical properties of functionalized vesicles did not vary significantly by varying DSPE-PEG-PDP concentration. LBM with tunable rigidity by adjusting DSPE-PEG-PDP concentration provides a versatile cell membrane model for studying protein or

  16. Experimental Investigations of Direct and Converse Flexoelectric Effect in Bilayer Lipid Membranes. (United States)

    Todorov, Angelio Todorov

    Flexoelectric coefficients (direct and converse), electric properties (capacitance and resistivity) and mechanical properties (thickness and elastic coefficients) have been determined for bilayer lipid membranes (BLMs) prepared from egg yolk lecithin (EYL), glycerol monoleate (GMO), phosphatidyl choline (PC) and phosphatidyl serine (PS) as a function of frequency, pH and surface charge modifiers. Direct flexoelectric effect manifested itself in the development of microvolt range a.c. potential (U_{f}) upon subjecting one side of a BLM to an oscillating hydrostatic pressure, in the 100-1000 Hz range. Operationally, the flexoelectric coefficient (f) is expressed by the ratio between U_{f} and the change of curvature (c) which accompanied the flexing of the membrane. Membrane curvature was determined by means of either the electric method (capacitance microphone effect) or by the newly developed method of stroboscopic interferometry. Real-time stroboscopic interferometry coupled with simultaneous electric measurements, provided a direct method for the determination of f. Two different frequency regimes of f were recognized. At low frequencies (free mobility of the surfactant, f-values of 24.1 times 10^{-19} and 0.87 times 10^ {-19} Coulombs were obtained for PC and GMO BLMs. At high frequencies (>300 Hz), associated with blocked mobility of the surfactant, f-values of 16.5 times 10^ {-19} and 0.30 times 10^{-19} Coulombs were obtained for PC and GMO BLMs. The theoretically calculated value for the GMO BLM oscillating at high frequency (0.12 times 10^{-19 } Coulombs) agreed well with that determined experimentally (0.3 times 10 ^{-19} Coulombs). For charged bovine brain PS BLM the observed flexocoefficient was f = 4.0 times 10^{ -18} Coulombs. Converse flexoelectric effect manifested itself in voltage-induced BLM curvature. Observations were carried out on uranyl acetate (UA) stabilized PS BLM under a.c. excitation. Frequency dependence of f was revealed by means of

  17. Influence of lysophospholipid hydrolysis by the catalytic domain of neuropathy target esterase on the fluidity of bilayer lipid membranes. (United States)

    Greiner, Aaron J; Richardson, Rudy J; Worden, R Mark; Ofoli, Robert Y


    Neuropathy target esterase (NTE) is an integral membrane protein localized in the endoplasmic reticulum in neurons. Irreversible inhibition of NTE by certain organophosphorus compounds produces a paralysis known as organophosphorus compound-induced delayed neuropathy. In vitro, NTE has phospholipase/lysophospholipase activity that hydrolyses exogenously added single-chain lysophospholipids in preference to dual-chain phospholipids, and NTE mutations have been associated with motor neuron disease. NTE's physiological role is not well understood, although recent studies suggest that it may control the cytotoxic accumulation of lysophospholipids in membranes. We used the NTE catalytic domain (NEST) to hydrolyze palmitoyl-2-hydroxy-sn-glycero-3-phosphocholine (p-lysoPC) to palmitic acid in bilayer membranes comprising 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and the fluorophore 1-oleoyl-2-[12-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]dodecanoyl]-sn-glycero-3-phosphocholine (NBD-PC). Translational diffusion coefficients (D(L)) in supported bilayer membranes were measured by fluorescence recovery after pattern photobleaching (FRAPP). The average D(L) for DOPC/p-lysoPC membranes without NEST was 2.44 microm(2)s(-1)+/-0.09; the D(L) for DOPC/p-lysoPC membranes containing NEST and diisopropylphosphorofluoridate, an inhibitor, was nearly identical at 2.45+/-0.08. By contrast, the D(L) for membranes comprising NEST, DOPC, and p-lysoPC was 2.28+/-0.07, significantly different from the system with inhibited NEST, due to NEST hydrolysis. Likewise, a system without NEST containing the amount of palmitic acid that would have been produced by NEST hydrolysis of p-lysoPC was identical at 2.26+/-0.06. These results indicate that NTE's catalytic activity can alter membrane fluidity. Copyright 2010 Elsevier B.V. All rights reserved.

  18. Localization and interaction of hydroxyflavones with lipid bilayer model membranes: a study using DSC and multinuclear NMR. (United States)

    Sinha, Ragini; Joshi, Akshada; Joshi, Urmila J; Srivastava, Sudha; Govil, Girjesh


    The localization and interaction of six naturally occurring flavones (FLV, 5HF, 6HF, 7HF, CHY and BLN) in DPPC bilayers were studied using DSC and multi-nuclear NMR. DSC results indicate that FLV and 6HF interact with alkyl chains. The (1)H NMR shows interaction of flavones with the sn-glycero region. Ring current induced chemical shifts indicate that 6HF and BLN acquire parallel orientation in bilayers. 2D NOESY spectra indicate partitioning of the B-ring into the alkyl chain region. The DSC, NMR and binding studies indicate that 5HF and 7HF are located near head group region, while 6HF, CHY and BLN are located in the vicinity of sn-glycero region, and FLV is inserted deepest in the membrane.

  19. Spin-label studies on the anchoring and lipid-protein interactions of avidin with N-biotinylphosphatidylethanolamines in lipid bilayer membranes. (United States)

    Swamy, M J; Marsh, D


    The specific binding of hen egg white avidin to phosphatidylcholine lipid membranes containing spin-labeled N-biotinylphosphatidylethanolamines (biotin-PESLs) was investigated by using ESR spectroscopy. Spin-labeled biotin-PEs were prepared with the nitroxide group at position C-5, C-8, C-10, C-12, or C-14 of the sn-2 chain and were incorporated at 1 mol % in lipid bilayer membranes of dimyristoylphosphatidylcholine. Binding of avidin produced a strong and selective restriction of the biotin-PE lipid mobility at all positions of chain labeling, as shown by the ESR spectra recorded in the fluid lipid phase. The spectral components of the fraction of the biotin-PESLs that were not complexed by avidin indicated that the mobility of the bulk membrane lipids was unperturbed by binding avidin, as demonstrated by difference spectroscopy. Comparison of the positional profiles and temperature dependences of the outer hyperfine splittings from the biotin-PESLs suggests that the C-12 and C-14 positions of the avidin-bound biotin-PEs are in register with the C-5 and C-7/C-6 positions, respectively, of the chains of the bulk membrane lipids. This indicates that the biotin-PEs are partially withdrawn from the membrane, with a vertical displacement of ca. 7-8 A, on complexation with avidin. In addition, the specific lipid-protein interaction with avidin results in a selective reduction in the rates of lipid chain motion, as shown by the increased ESR line widths. These data define the way in which avidin is anchored to lipid membranes containing biotin-PEs.

  20. Electrostatics of cell membrane recognition: structure and activity of neutral and cationic rigid push-pull rods in isoelectric, anionic, and polarized lipid bilayer membranes. (United States)

    Sakai, N; Gerard, D; Matile, S


    Design, synthesis, and structural and functional studies of rigid-rod ionophores of different axial electrostatic asymmetry are reported. The employed design strategy emphasized presence of (a) a rigid scaffold to minimize the conformational complexity, (b) a unimolecular ion-conducting pathway to minimize the suprastructural complexity and monitor the function, (c) an extended fluorophore to monitor structure, (d) variable axial rod dipole, and (e) variable terminal charges to create axial asymmetry. Studies in isoelectric, anionic, and polarized bilayer membranes confirmed a general increase in activity of uncharged rigid push-pull rods in polarized bilayers. The similarly increased activity of cationic rigid push-pull rods with an electrostatic asymmetry comparable to that of alpha-helical bee toxin melittin (positive charge near negative axial dipole terminus) is shown by fluorescence-depth quenching experiments to originate from the stabilization of transmembrane rod orientation by the membrane potential. The reduced activity of rigid push-pull rods having an electrostatic asymmetry comparable to that in alpha-helical natural antibiotics (a positive charge near the positive axial dipole terminus) is shown by structural studies to originate from rod "ejection" by membrane potentials comparable to that found in mammalian plasma membranes. This structural evidence for cell membrane recognition by asymmetric rods is unprecedented and of possible practical importance with regard to antibiotic resistance.

  1. Study of pH (low) insertion peptides (pHLIPs) interaction with lipid bilayer of membrane (United States)

    Weerakkody, Dhammika

    The pH-dependent interactions of pHLIPsRTM (pH (Low) Insertion Peptides) with lipid bilayer of membrane provides an opportunity to study and address fundamental questions of protein folding/insertion into membrane and unfolding/exit, as well as develop novel approach to target acidic diseased tissue such as cancer, ischemic myocardium, infection and others. The main goal of the work presented here is to answer the following questions: - What is the molecular mechanism of spontaneous insertion and folding of a peptide in a lipid bilayer of membrane; - What is the molecular mechanism of unfolding and exit of a peptide from a lipid bilayer of membrane; - How polar cargo attached to a peptide's inserting end might affect the process of insertion into a lipid bilayer of membrane; How sequence variation will affect a peptide's interactions with a lipid bilayer of membrane (partitioning into bilayer at neutral and low pH; apparent pK of insertion) with the main goal to identify the best pHLIP variants for imaging and therapy of pathological states such as cancer and others. It has been demonstrated that pHLIP insertion into a membrane is associated with the protonation of Asp/Glu residues, which leads to an increase of hydrophobicity that triggers the folding and insertion of the peptide across a lipid bilayer. The insertion of the pHLIP is unidirectional and it is accompanied by the release of energy. Therefore, the energy of membrane associated-folding can be used to favor the movement of cell-impermeable polar cargo molecules across the hydrophobic membrane bilayer when they are attached to the inserting end of pHLIP. Both pH-targeting behavior and molecular translocation have been demonstrated in cultured cells and in vivo. Thus, there is an opportunity to develop a novel concept in drug delivery, which is based on the use of a monomeric, pH-sensitive peptide molecular transporter, to deliver agents that are significantly more polar than conventional drugs

  2. Measurement of the glucose permeation rate across phospholipid bilayers using small unilamellar vesicles. Effect of membrane composition and temperature. (United States)

    Bresseleers, G J; Goderis, H L; Tobback, P P


    Small unilamellar vesicles were used to measure the permeability of saturated phosphatidylcholine bilayers to glucose. The presented method circumvents most of the common restriction of classical permeability experiments. Increasing the fatty acid chain length of the lipids reduced the permeation rate significantly. Raising the temperature above that of the lipid phase transition drastically increased membrane permeability. Arrhenius plots demonstrated the activation energy to be independent of membrane composition and the phase-state of the lipids. The permeation process is discussed in terms of a constant energy to disrupt all hydrogen bonds between permeant and aqueous solvent prior to penetrating the membrane. The magnitude of the permeability coefficient is partly determined by a unfavourable change in entropy of activation on crossing the water/lipid interface. All results indicate that the penetration of the dehydrated permeant into the hydrophobic barrier is the rate-limiting step in the permeation of glucose.

  3. General model of phospholipid bilayers in fluid phase within the single chain mean field theory

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Yachong; Baulin, Vladimir A. [Departament d’Enginyeria Química, Universitat Rovira i Virgili, Av. dels Paisos Catalans 26, 43007 Tarragona (Spain); Pogodin, Sergey [Institute of Chemical Research of Catalonia, ICIQ, Av. Paisos Catalans 16, 43007 Tarragona (Spain)


    Coarse-grained model for saturated phospholipids: 1,2-didecanoyl-sn-glycero-3-phosphocholine (DCPC), 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and unsaturated phospholipids: 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1,2- dioleoyl-sn-glycero-3-phosphocholine (DOPC) is introduced within the single chain mean field theory. A single set of parameters adjusted for DMPC bilayers gives an adequate description of equilibrium and mechanical properties of a range of saturated lipid molecules that differ only in length of their hydrophobic tails and unsaturated (POPC, DOPC) phospholipids which have double bonds in the tails. A double bond is modeled with a fixed angle of 120°, while the rest of the parameters are kept the same as saturated lipids. The thickness of the bilayer and its hydrophobic core, the compressibility, and the equilibrium area per lipid correspond to experimentally measured values for each lipid, changing linearly with the length of the tail. The model for unsaturated phospholipids also fetches main thermodynamical properties of the bilayers. This model is used for an accurate estimation of the free energies of the compressed or stretched bilayers in stacks or multilayers and gives reasonable estimates for free energies. The proposed model may further be used for studies of mixtures of lipids, small molecule inclusions, interactions of bilayers with embedded proteins.

  4. Membrane docking geometry of GRP1 PH domain bound to a target lipid bilayer: an EPR site-directed spin-labeling and relaxation study.

    Directory of Open Access Journals (Sweden)

    Huai-Chun Chen

    Full Text Available The second messenger lipid PIP(3 (phosphatidylinositol-3,4,5-trisphosphate is generated by the lipid kinase PI3K (phosphoinositide-3-kinase in the inner leaflet of the plasma membrane, where it regulates a broad array of cell processes by recruiting multiple signaling proteins containing PIP(3-specific pleckstrin homology (PH domains to the membrane surface. Despite the broad importance of PIP(3-specific PH domains, the membrane docking geometry of a PH domain bound to its target PIP(3 lipid on a bilayer surface has not yet been experimentally determined. The present study employs EPR site-directed spin labeling and relaxation methods to elucidate the membrane docking geometry of GRP1 PH domain bound to bilayer-embedded PIP(3. The model target bilayer contains the neutral background lipid PC and both essential targeting lipids: (i PIP(3 target lipid that provides specificity and affinity, and (ii PS facilitator lipid that enhances the PIP(3 on-rate via an electrostatic search mechanism. The EPR approach measures membrane depth parameters for 18 function-retaining spin labels coupled to the PH domain, and for calibration spin labels coupled to phospholipids. The resulting depth parameters, together with the known high resolution structure of the co-complex between GRP1 PH domain and the PIP(3 headgroup, provide sufficient constraints to define an optimized, self-consistent membrane docking geometry. In this optimized geometry the PH domain engulfs the PIP(3 headgroup with minimal bilayer penetration, yielding the shallowest membrane position yet described for a lipid binding domain. This binding interaction displaces the PIP(3 headgroup from its lowest energy position and orientation in the bilayer, but the headgroup remains within its energetically accessible depth and angular ranges. Finally, the optimized docking geometry explains previous biophysical findings including mutations observed to disrupt membrane binding, and the rapid lateral

  5. Formation of cholesterol bilayer domains precedes formation of cholesterol crystals in cholesterol/dimyristoylphosphatidylcholine membranes: EPR and DSC studies. (United States)

    Mainali, Laxman; Raguz, Marija; Subczynski, Witold K


    Saturation-recovery EPR along with DSC were used to determine the cholesterol content at which pure cholesterol bilayer domains (CBDs) and cholesterol crystals begin to form in dimyristoylphosphatidylcholine (DMPC) membranes. To preserve compositional homogeneity throughout the membrane suspension, lipid multilamellar dispersions were prepared using a rapid solvent exchange method. The cholesterol content increased from 0 to 75 mol %. With spin-labeled cholesterol analogues, it was shown that the CBDs begin to form at ~50 mol % cholesterol. It was confirmed by DSC that the cholesterol solubility threshold for DMPC membranes is detected at ~66 mol % cholesterol. At levels above this cholesterol content, monohydrate cholesterol crystals start to form. The major finding is that the formation of CBDs precedes formation of cholesterol crystals. The region of the phase diagram for cholesterol contents between 50 and 66 mol % is described as a structured one-phase region in which CBDs have to be supported by the surrounding DMPC bilayer saturated with cholesterol. Thus, the phase boundary located at 66 mol % cholesterol separates the structured one-phase region (liquid-ordered phase of DMPC with CBDs) from the two-phase region where the structured liquid-ordered phase of DMPC coexists with cholesterol crystals. It is likely that CBDs are precursors of monohydrate cholesterol crystals.

  6. An investigation of the effect of membrane curvature on transmembrane-domain dependent protein sorting in lipid bilayers (United States)

    Fossati, Matteo; Goud, Bruno; Borgese, Nica; Manneville, Jean-Baptiste


    Sorting of membrane proteins within the secretory pathway of eukaryotic cells is a complex process involving discrete sorting signals as well as physico-chemical properties of the transmembrane domain (TMD). Previous work demonstrated that tail-anchored (TA) protein sorting at the interface between the Endoplasmic Reticulum (ER) and the Golgi complex is exquisitely dependent on the length and hydrophobicity of the transmembrane domain, and suggested that an imbalance between TMD length and bilayer thickness (hydrophobic mismatch) could drive long TMD-containing proteins into curved membrane domains, including ER exit sites, with consequent export of the mismatched protein out of the ER. Here, we tested a possible role of curvature in TMD-dependent sorting in a model system consisting of Giant Unilamellar Vesicles (GUVs) from which narrow membrane tubes were pulled by micromanipulation. Fluorescent TA proteins differing in TMD length were incorporated into GUVs of uniform lipid composition or made of total ER lipids, and TMD-dependent sorting and diffusion, as well as the bending rigidity of bilayers made of microsomal lipids, were investigated. Long and short TMD-containing constructs were inserted with similar orientation, diffused equally rapidly in GUVs and in tubes pulled from GUVs, and no difference in their final distribution between planar and curved regions was detected. These results indicate that curvature alone is not sufficient to drive TMD-dependent sorting at the ER-Golgi interface, and set the basis for the investigation of the additional factors that must be required. PMID:25210649

  7. Guided periodontal regeneration using bilayered collagen membranes and bovine bone mineral in fenestration defects in the canine. (United States)

    Tal, Haim; Artzi, Zvi; Moses, Ofer; Nemcovsky, Carlos; Kozlovsky, Avital


    This study was performed to evaluate the effect of deproteinized bovine porous bone mineral (BBM) and BBM-collagen (BBMC) used alone or in combination with a bilayer collagen membrane in guided periodontal regeneration. In 12 dogs, contralateral surgical circular fenestration defects 5 mm in diameter were produced at the midbuccal aspect of the alveolar bone in 24 maxillary canines. Bone, periodontal ligament, and cementum were completely removed. Experimental sites were filled with BBM or BBMC. Bilayered collagen membranes covered half the experimental sites (BBM+M and BBMC+M), and the other half were left uncovered. Control sites remained empty; half were covered with collagen membranes (cont+M) and the underlying space spontaneously filled with blood, and half were left uncovered (cont). Three months postsurgery, undecalcified sections were prepared. Measurements were made using a caliper on a projection microscope, and the surface area of new bone and BBM particles within the healed surgical defect was evaluated using the point-counting method. In the experimental defects, new cementum covered 31% to 67% of the exposed dentin, with a significant difference between defects covered with membranes and defects that were not covered (P tissue in the covered defects than in the uncovered defects (P tissue/bone marrow, and bovine bone particles. New bone area fraction was 23.4% to 25.2% in defects filled with BBMC and BBM, respectively (P = NS). Bone fraction area in membrane-covered defects ranged from 34.4% to 36.8% in experimental defects (P = NS). All membrane-treated defects showed higher values for bone area fraction in comparison to the uncovered control defects. Particle area fraction ranged between 17.4% and 26.2%, with only BBMC and BBM+M defects showing a statistically significant difference (P regeneration than experimental defects filled with BBM or BBMC. Treatment of defects with BBM or BBMC showed similar influences on bone and cementum regeneration in

  8. A DSC and FTIR spectroscopic study of the effects of the epimeric coprostan-3-ols and coprostan-3-one on the thermotropic phase behaviour and organization of dipalmitoylphosphatidylcholine bilayer membranes: Comparison with their 5-cholesten analogues. (United States)

    Benesch, Matthew G K; Lewis, Ruthven N A H; Mannock, David A; McElhaney, Ronald N


    We present the results of a comparative differential calorimetric and Fourier transform infrared spectroscopic study of the effect of cholesterol and five analogues on the thermotropic phase behaviour and organization of dipalmitoylphosphatidylcholine bilayer membranes. These sterols/steroids differ in both the nature and stereochemistry of the polar head group at C3 (β-OH, α-OH or CO) and in the presence or absence of a double bond in ring B and in the orientation of rings A and B. The Δ(5) sterols/steroid have a trans rather than a cis ring A/B junction, and the concentration of these compounds required to abolish the DPPC pretransition, inversely related to their relative ability to disorder gel state DPPC bilayers, decreases in the order β-OH > α-OH > CO. However, in the saturated ring junction-inverted (cis) series, these concentrations are much more similar, regardless of polar head group chemical structure. Similarly, the residual enthalpy of the DPPC main phase transition at 50 mol% sterol/steroid, which is inversely related to the miscibility of these compounds in fluid DPPC bilayers, also increases in the order β-OH > α-OH > CO, but this effect is attenuated in the saturated series with an inverted ring A/B orientation. Moreover, replacement of the double bond at C5-C6 with a saturated linkage and inversion of the ring A/B junction reduces both sterol/steroid solubility and the ability to order the hydrocarbon chains of fluid DPPC molecules all cases. Thus, the characteristic effects of sterols/steroids on fluid lipid bilayers are generally optimal when an OH group rather than CO group is present at C3, and when this OH group is in the equatorial (β) orientation, and when the orientation of the ring A/B fusion is trans rather than cis. Overall, these results demonstrate that variations in the saturation and stereochemistry of the steroid ring system influence the effect of variations in the nature and stereochemistry of the polar headgroup at C3

  9. Mesoscale computational studies of membrane bilayer remodeling by curvature-inducing proteins (United States)

    Ramakrishnan, N.; Sunil Kumar, P. B.; Radhakrishnan, Ravi


    Biological membranes constitute boundaries of cells and cell organelles. These membranes are soft fluid interfaces whose thermodynamic states are dictated by bending moduli, induced curvature fields, and thermal fluctuations. Recently, there has been a flood of experimental evidence highlighting active roles for these structures in many cellular processes ranging from trafficking of cargo to cell motility. It is believed that the local membrane curvature, which is continuously altered due to its interactions with myriad proteins and other macromolecules attached to its surface, holds the key to the emergent functionality in these cellular processes. Mechanisms at the atomic scale are dictated by protein-lipid interaction strength, lipid composition, lipid distribution in the vicinity of the protein, shape and amino acid composition of the protein, and its amino acid contents. The specificity of molecular interactions together with the cooperativity of multiple proteins induce and stabilize complex membrane shapes at the mesoscale. These shapes span a wide spectrum ranging from the spherical plasma membrane to the complex cisternae of the Golgi apparatus. Mapping the relation between the protein-induced deformations at the molecular scale and the resulting mesoscale morphologies is key to bridging cellular experiments across various length scales. In this review, we focus on the theoretical and computational methods used to understand the phenomenology underlying protein-driven membrane remodeling. Interactions at the molecular scale can be computationally probed by all atom and coarse grained molecular dynamics (MD, CGMD), as well as dissipative particle dynamics (DPD) simulations, which we only describe in passing. We choose to focus on several continuum approaches extending the Canham-Helfrich elastic energy model for membranes to include the effect of curvature-inducing proteins and explore the conformational phase space of such systems. In this description, the

  10. Mesoscale computational studies of membrane bilayer remodeling by curvature-inducing proteins

    Energy Technology Data Exchange (ETDEWEB)

    Ramakrishnan, N., E-mail: [Department of Chemical and Biomolecular Engineering, University of Pennsylvania, Philadelphia, PA-19104 (United States); Department of Bioengineering, University of Pennsylvania, Philadelphia, PA-19104 (United States); Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA-19104 (United States); Sunil Kumar, P.B., E-mail: [Department of Physics, Indian Institute of Technology Madras, Chennai, 600036 (India); Radhakrishnan, Ravi, E-mail: [Department of Chemical and Biomolecular Engineering, University of Pennsylvania, Philadelphia, PA-19104 (United States); Department of Bioengineering, University of Pennsylvania, Philadelphia, PA-19104 (United States); Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA-19104 (United States)


    Biological membranes constitute boundaries of cells and cell organelles. These membranes are soft fluid interfaces whose thermodynamic states are dictated by bending moduli, induced curvature fields, and thermal fluctuations. Recently, there has been a flood of experimental evidence highlighting active roles for these structures in many cellular processes ranging from trafficking of cargo to cell motility. It is believed that the local membrane curvature, which is continuously altered due to its interactions with myriad proteins and other macromolecules attached to its surface, holds the key to the emergent functionality in these cellular processes. Mechanisms at the atomic scale are dictated by protein–lipid interaction strength, lipid composition, lipid distribution in the vicinity of the protein, shape and amino acid composition of the protein, and its amino acid contents. The specificity of molecular interactions together with the cooperativity of multiple proteins induce and stabilize complex membrane shapes at the mesoscale. These shapes span a wide spectrum ranging from the spherical plasma membrane to the complex cisternae of the Golgi apparatus. Mapping the relation between the protein-induced deformations at the molecular scale and the resulting mesoscale morphologies is key to bridging cellular experiments across various length scales. In this review, we focus on the theoretical and computational methods used to understand the phenomenology underlying protein-driven membrane remodeling. Interactions at the molecular scale can be computationally probed by all atom and coarse grained molecular dynamics (MD, CGMD), as well as dissipative particle dynamics (DPD) simulations, which we only describe in passing. We choose to focus on several continuum approaches extending the Canham–Helfrich elastic energy model for membranes to include the effect of curvature-inducing proteins and explore the conformational phase space of such systems. In this description

  11. Dispersion of atmospheric fine particulate matters in simulated lung fluid and their effects on model cell membranes. (United States)

    Zhou, Qiuhua; Wang, Lixin; Cao, Zhaoyu; Zhou, Xuehua; Yang, Fan; Fu, Pingqing; Wang, Zhenhua; Hu, Jingtian; Ding, Lei; Jiang, Wei


    Atmospheric fine particulate matter (PM2.5) was collected to investigate its dispersion in simulated lung fluid (SLF) and its interaction with model cell membranes. Organic acids, NH4(+), SO4(2-) and NO3(-) were detected in PM2.5 soluble fraction, and heavy metals were detected from the total mass. The insoluble fraction contained kaolinite, CaCO3, aliphatic carbons, aromatic rings, carboxyl and hydroxyl groups reflected by the infrared spectra. Proteins dispersed PM2.5 in SLF, resulted in smaller hydrodynamic diameter (dH) and slower sedimentation rate. Conversely, phospholipids increased dH value and accelerated sedimentation rate. Giant unilamellar vesicles (GUVs) and supported lipid bilayers (SLBs) were used as model cell membranes. PM2.5 adhered on and disrupted the membrane containing positively-charged lipids but not the membrane containing neutrally- and negatively-charged lipids, which was monitored by microscopy and a quartz crystal microbalance with dissipation (QCM-D). The cationic sites on membrane were necessary for PM2.5 adhesion, but membrane should be disrupted by the combined action of electrostatic forces and hydrogen bonds between PM2.5 oxygen containing groups and the lipid phosphate groups. Our results specified the roles of proteins and phospholipids in PM2.5 dispersion and transport, highly suggested that the health hazard of PM2.5 was related to the biomolecules in the lung fluid and the particle surface groups.

  12. Specific volume and compressibility of bilayer lipid membranes with incorporated Na,K-ATPase. (United States)

    Hianik, Tibor; Rybár, Peter; Krivánek, Roland; Petríková, Mária; Roudna, Milena; Apell, Hans Jürgen


    Ultrasound velocimetry and densitometry methods were used to study the interactions of the Na,K-ATPase with the lipid bilayer in large unilamellar liposomes composed of dioleoyl phosphatidylcholine (DOPC). The ultrasound velocity increased and the specific volume of the phospholipids decreased with increasing concentrations of protein. These experiments allowed us to determine the reduced specific apparent compressibility of the lipid bilayer, which decreased by approx. 11% with increasing concentrations of the Na,K-ATPase up to an ATPase/DOPC molar ratio = 2 × 10⁻⁴. Assuming that ATPase induces rigidization of the surrounding lipid molecules one can obtain from the compressibility data that 3.7 to 100 times more lipid molecules are affected by the protein in comparison with annular lipids. However, this is in contradiction with the current theories of the phase transitions in lipid bilayers. It is suggested that another physical mechanisms should be involved for explanation of observed effect.

  13. A comparison of coarse-grained and continuum models for membrane bending in lipid bilayer fusion pores. (United States)

    Yoo, Jejoong; Jackson, Meyer B; Cui, Qiang


    To establish the validity of continuum mechanics models quantitatively for the analysis of membrane remodeling processes, we compare the shape and energies of the membrane fusion pore predicted by coarse-grained (MARTINI) and continuum mechanics models. The results at these distinct levels of resolution give surprisingly consistent descriptions for the shape of the fusion pore, and the deviation between the continuum and coarse-grained models becomes notable only when the radius of curvature approaches the thickness of a monolayer. Although slow relaxation beyond microseconds is observed in different perturbative simulations, the key structural features (e.g., dimension and shape of the fusion pore near the pore center) are consistent among independent simulations. These observations provide solid support for the use of coarse-grained and continuum models in the analysis of membrane remodeling. The combined coarse-grained and continuum analysis confirms the recent prediction of continuum models that the fusion pore is a metastable structure and that its optimal shape is neither toroidal nor catenoidal. Moreover, our results help reveal a new, to our knowledge, bowing feature in which the bilayers close to the pore axis separate more from one another than those at greater distances from the pore axis; bowing helps reduce the curvature and therefore stabilizes the fusion pore structure. The spread of the bilayer deformations over distances of hundreds of nanometers and the substantial reduction in energy of fusion pore formation provided by this spread indicate that membrane fusion can be enhanced by allowing a larger area of membrane to participate and be deformed. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  14. Effect of chirality and length on the penetrability of single-walled carbon nanotubes into lipid bilayer cell membranes. (United States)

    Skandani, A Alipour; Zeineldin, R; Al-Haik, M


    The ability of carbon nanotubes to enter the cell membrane acting as drug-delivery vehicles has yielded a plethora of experimental investigations, mostly with inconclusive results because of the wide spectra of carbon nanotube structures. Because of the virtual impossibility of synthesizing CNTs with distinct chirality, we report a parametric study on the use of molecular dynamics to provide better insight into the effect of the carbon nanotube chirality and the aspect ratio on the interaction with a lipid bilayer membrane. The simulation results indicated that a single-walled carbon nanotube utilizes different time-evolving mechanisms to facilitate their internalization within the membrane. These mechanisms comprise both penetration and endocytosis. It was observed that carbon nanotubes with higher aspect ratios penetrate the membrane faster whereas shorter nanotubes undergo significant rotation during the final stages of endocytosis. Furthermore, nanotubes with lower chiral indices developed significant adhesion with the membrane. This adhesion is hypothesized to consume some of the carbon nanotube energy, thus resulting in longer times for the nanotube to translocate through the membrane.

  15. Cell membrane fluid-mosaic structure and cancer metastasis. (United States)

    Nicolson, Garth L


    Cancer cells are surrounded by a fluid-mosaic membrane that provides a highly dynamic structural barrier with the microenvironment, communication filter and transport, receptor and enzyme platform. This structure forms because of the physical properties of its constituents, which can move laterally and selectively within the membrane plane and associate with similar or different constituents, forming specific, functional domains. Over the years, data have accumulated on the amounts, structures, and mobilities of membrane constituents after transformation and during progression and metastasis. More recent information has shown the importance of specialized membrane domains, such as lipid rafts, protein-lipid complexes, receptor complexes, invadopodia, and other cellular structures in the malignant process. In describing the macrostructure and dynamics of plasma membranes, membrane-associated cytoskeletal structures and extracellular matrix are also important, constraining the motion of membrane components and acting as traction points for cell motility. These associations may be altered in malignant cells, and probably also in surrounding normal cells, promoting invasion and metastatic colonization. In addition, components can be released from cells as secretory molecules, enzymes, receptors, large macromolecular complexes, membrane vesicles, and exosomes that can modify the microenvironment, provide specific cross-talk, and facilitate invasion, survival, and growth of malignant cells.

  16. A Fluid Membrane-Based Soluble Ligand Display System for Live CellAssays

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jwa-Min; Nair, Pradeep N.; Neve, Richard M.; Gray, Joe W.; Groves, Jay T.


    Cell communication modulates numerous biological processes including proliferation, apoptosis, motility, invasion and differentiation. Correspondingly, there has been significant interest in the development of surface display strategies for the presentation of signaling molecules to living cells. This effort has primarily focused on naturally surface-bound ligands, such as extracellular matrix components and cell membranes. Soluble ligands (e.g. growth factors and cytokines) play an important role in intercellular communications, and their display in a surface-bound format would be of great utility in the design of array-based live cell assays. Recently, several cell microarray systems that display cDNA, RNAi, or small molecules in a surface array format were proven to be useful in accelerating high-throughput functional genetic studies and screening therapeutic agents. These surface display methods provide a flexible platform for the systematic, combinatorial investigation of genes and small molecules affecting cellular processes and phenotypes of interest. In an analogous sense, it would be an important advance if one could display soluble signaling ligands in a surface assay format that allows for systematic, patterned presentation of soluble ligands to live cells. Such a technique would make it possible to examine cellular phenotypes of interest in a parallel format with soluble signaling ligands as one of the display parameters. Herein we report a ligand-modified fluid supported lipid bilayer (SLB) assay system that can be used to functionally display soluble ligands to cells in situ (Figure 1A). By displaying soluble ligands on a SLB surface, both solution behavior (the ability to become locally enriched by reaction-diffusion processes) and solid behavior (the ability to control the spatial location of the ligands in an open system) could be combined. The method reported herein benefits from the naturally fluid state of the supported membrane, which allows

  17. Low concentration of DMSO stabilizes the bilayer gel phase rather than the interdigitated gel phase in dihexadecylphosphatidylcholine membrane. (United States)

    Yamashita, Y; Kinoshita, K; Yamazaki, M


    We have investigated effects of dimethylsulfoxide (DMSO) on the phase stability of multilamellar vesicles of the ether-linked 1,2-dihexadecyl-sn-glycero-3-phosphatidylcholine (DHPC-MLV), which is known to be in the interdigitated gel (LbetaI) phase in excess water at 20 degrees C. The results of X-ray diffraction experiments indicate that the DHPC membrane was in the Lbeta, phase at X> or =0.12 (X=mole fraction of DMSO in DMSO/water mixture). The result of differential scanning calorimetry indicate that the gel to liquid-crystalline phase transition temperature increased, but the LbetaI to Pbeta, phase transition temperature decreased with an increase in DMSO concentration. These results show that DMSO stabilizes the bilayer gel phase rather than the LbetaI phase at its low concentration. The solubility of phosphorylcholine, which is the same structure as the headgroup of DHPC, decreased with an increase in DMSO concentration, indicating that the interaction free energy of the hydrophilic segments of the membrane with solvents increases with an increase in DMSO concentration. On the basis of the thermodynamic analysis, the mechanism of the stabilization of the bilayer gel phase of DHPC-MLV by DMSO is discussed. The decrease in the repulsive interaction between the headgroups of the phospholipid induced by the low concentrations of DMSO in water plays an important role in this stabilization.

  18. Solid-state NMR of the Yersinia pestis outer membrane protein Ail in lipid bilayer nanodiscs sedimented by ultracentrifugation

    Energy Technology Data Exchange (ETDEWEB)

    Ding, Yi; Fujimoto, L. Miya; Yao, Yong; Marassi, Francesca M., E-mail: [Sanford-Burnham Medical Research Institute (United States)


    Solid-state NMR studies of sedimented soluble proteins has been developed recently as an attractive approach for overcoming the size limitations of solution NMR spectroscopy while bypassing the need for sample crystallization or precipitation (Bertini et al. Proc Natl Acad Sci USA 108(26):10396–10399, 2011). Inspired by the potential benefits of this method, we have investigated the ability to sediment lipid bilayer nanodiscs reconstituted with a membrane protein. In this study, we show that nanodiscs containing the outer membrane protein Ail from Yersinia pestis can be sedimented for solid-state NMR structural studies, without the need for precipitation or lyophilization. Optimized preparations of Ail in phospholipid nanodiscs support both the structure and the fibronectin binding activity of the protein. The same sample can be used for solution NMR, solid-state NMR and activity assays, facilitating structure–activity correlation experiments across a wide range of timescales.

  19. [Isolation and purification of human blood plasma proteins able to form potassium channels in artificial bilayer lipid membrane]. (United States)

    Venediktova, N I; Kuznetsov, K V; Gritsenko, E N; Gulidova, G P; Mironova, G D


    Protein fraction able to induce K(+)-selective transport across bilayer lipid membrane was isolated from human blood plasma with the use of the detergent and proteolytic enzyme-free method developed at our laboratory. After addition of the studied sample to the artificial membrane in the presence of 100 mM KCl, a discrete current change was observed. No channel activity was recorded in the presence of calcium and sodium ions. Channel forming activity of fraction was observed only in the presence of K+. Using a threefold gradient of KCl in the presence of studied proteins the potassium-selective potential balanced by voltage of -29 mV was registered. This value is very close to the theoretical Nernst potential in this case. This means that the examined ion channel is cation-selective. According to data obtained with MS-MALDI-TOF/TOF and database NCBI three protein components were identified in isolated researched sample.

  20. Bilayer Vesicles of Amphiphilic Cyclodextrins: Host Membranes That Recognize Guest Molecules

    NARCIS (Netherlands)

    Falvey, Patrick; Lim, Choon Woo; Darcy, Raphael; Revermann, Tobias; Karst, Uwe; Giesbers, Marcel; Marcelis, Antonius T.M.; Lazar, Adina; Coleman, Anthony W.; Reinhoudt, David N.; Ravoo, Bart Jan


    A family of amphiphilic cyclodextrins (6, 7) has been prepared through 6-S-alkylation (alkyl=n-dodecyl and n-hexadecyl) of the primary side and 2-O-PEGylation of the secondary side of a-, B-, and Y-cyclodextrins (PEG=poly(ethylene glycol)). These cyclodextrins form nonionic bilayer vesicles in aqueo

  1. Bilayer vesicles of amphiphilic cyclodextrines: host membranes that recognize guest molecules

    NARCIS (Netherlands)

    Falvey, P.; Lim, C.W.; Darcy, R.; Revermann, T.; Karst, U.; Marcelis, A.T.M.; Coleman, A.W.; Reinhoudt, D.N.; Ravoo, B.J.


    A family of amphiphilic cyclodextrins (6, 7) has been prepared through 6-S-alkylation (alkyl=n-dodecyl and n-hexadecyl) of the primary side and 2-O-PEGylation of the secondary side of alpha-, beta-, and gamma-cyclodextrins (PEG=poly(ethylene glycol)). These cyclodextrins form nonionic bilayer vesicl

  2. Bilayer vesicles of amphiphilic cyclodextrines: host membranes that recognize guest molecules

    NARCIS (Netherlands)

    Falvey, P.; Lim, C.W.; Darcy, R.; Revermann, T.; Karst, U.; Marcelis, A.T.M.; Coleman, A.W.; Reinhoudt, D.N.; Ravoo, B.J.


    A family of amphiphilic cyclodextrins (6, 7) has been prepared through 6-S-alkylation (alkyl=n-dodecyl and n-hexadecyl) of the primary side and 2-O-PEGylation of the secondary side of alpha-, beta-, and gamma-cyclodextrins (PEG=poly(ethylene glycol)). These cyclodextrins form nonionic bilayer

  3. Bilayer Vesicles of Amphiphilic Cyclodextrins: Host Membranes That Recognize Guest Molecules

    NARCIS (Netherlands)

    Falvey, Patrick; Lim, C.W.; Darcy, Raphael; Revermann, T.; Karst, U.; Giesbers, Marcel; Marcelis, Antonius T.M.; Lazar, Adina; Coleman, Anthony W.; Reinhoudt, David; Ravoo, B.J.


    A family of amphiphilic cyclodextrins (6, 7) has been prepared through 6-S-alkylation (alkyl=n-dodecyl and n-hexadecyl) of the primary side and 2-O-PEGylation of the secondary side of a-, B-, and Y-cyclodextrins (PEG=poly(ethylene glycol)). These cyclodextrins form nonionic bilayer vesicles in

  4. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells. (United States)

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi


    Molecular dynamics (MD) calculations for the plasma membranes of normal murine thymocytes and thymus-derived leukemic GRSL cells in water have been performed under physiological isothermal-isobaric conditions (310.15K and 1 atm) to investigate changes in membrane properties induced by canceration. The model membranes used in our calculations for normal and leukemic thymocytes comprised 23 and 25 kinds of lipids, respectively, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. The mole fractions of the lipids adopted here were based on previously published experimental values. Our calculations clearly showed that the membrane area was increased in leukemic cells, and that the isothermal area compressibility of the leukemic plasma membranes was double that of normal cells. The calculated membranes of leukemic cells were thus considerably bulkier and softer in the lateral direction compared with those of normal cells. The tilt angle of the cholesterol and the conformation of the phospholipid fatty acid tails both showed a lower level of order in leukemic cell membranes compared with normal cell membranes. The lateral radial distribution function of the lipids also showed a more disordered structure in leukemic cell membranes than in normal cell membranes. These observations all show that, for the present thymocytes, the lateral structure of the membrane is considerably disordered by canceration. Furthermore, the calculated lateral self-diffusion coefficient of the lipid molecules in leukemic cell membranes was almost double that in normal cell membranes. The calculated rotational and wobbling autocorrelation functions also indicated that the molecular motion of the lipids was enhanced in leukemic cell membranes. Thus, here we have demonstrated that the membranes of thymocyte leukemic cells are more disordered and more fluid than normal cell membranes.

  5. Spatiotemporal Pattern Formation in BioFluids I: Cell Shape Perturbants As Evidence of Spatially-Organised Membrane Flows

    CERN Document Server

    Lofthouse, J T


    I show the assumed Bilayer structure of cell membranes is Topologically falsified by known aminophospholipid dynamics in metabolically-active, Far from Equilibrium cells. The sensitivity of lipid and cytoplasmic flows to temperature, surfactants, viscosity and the gravity vector are used to suggest that rather than being random viscous fluids as currently assumed, both are actually spatially-organised by convective and shear driven mechanisms in vivo. I show how protein-lipid feedback provokes a Gestalt Shift in Cell Mechanics by demonstrating that the primary forces involved in shape changes are generated by bifurcations in fluid flow Topology, which induce affine deformations of the cytoskeletal lattice. The feedback model allows the transduction of Gravitational information into biological form, is universally applicable, and provides a rationale for Homeoviscous Adaptation, and the extensive lipid polymorphism observed in Nature.

  6. Capillary electrophoresis with noncovalently bilayer-coated capillaries for stability study of allergenic proteins in simulated gastrointestinal fluids. (United States)

    Zheng, Chang; Liu, Youping; Zhou, Qiuhong; Di, Xin


    A novel noncovalently bilayer-coated capillary using cationic polymer polybrene (PB) and anionic polymer (sodium 4-styrenesulfonate) (PSS) as coatings was prepared. This PB-PSS coating showed good migration-time reproducibility for proteins and high stability in the range of pH 2-10 and in the presence of 1M NaOH, acetonitrile and methanol. Capillary electrophoresis with PB-PSS coated capillaries was successfully applied to quantitatively investigate the stability of bovine serum albumin, ovomucoid, β-lactoglobulin and lysozyme in simulated gastrointestinal fluids. β-lactoglobulin A and β-lactoglobulin B were both stable in simulated gastric fluid with degradation percentages of 34.3% and 17.2% after 60min of incubation, respectively. Bovine serum albumin, ovomucoid and lysozyme were stable in simulated intestinal fluid with degradation percentages of 17.7%, 23.4% and 22.8% after 60min of incubation, respectively. The superiority of the proposed method over sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and capillary electrophoresis with untreated fused silica capillaries was demonstrated and emphasized.

  7. Raised Vaginal Fluid Fibronectin Level Indicates Premature Rupture of Membrane

    Directory of Open Access Journals (Sweden)

    Amrita Bhowmik


    Full Text Available Background: Premature rupture of membrane (PROM is one of the common complications of pregnancy that has major impact on fetal and neonatal outcome. It is the commonest clinical event where a normal pregnancy becomes suddenly a high-risk one for mother and fetus or neonate. Objective: The study was undertaken to investigate whether raised fibronectin level in vaginal fluid may indicate premature rupture of membrane. Materials and Methods: This cross sectional study was conducted in the department of Obstetrics and Gynecology in Sir Salimullah Medical College & Mitford Hospital, Dhaka during the period of January 2006 to December 2007. A total of 114 pregnant women with gestational age 28th week up to 40th week were included. Sixty were PROM (Group I and 54 were non-PROM (Group II subjects. Fibronectin in vaginal fluid was measured by an immunochemical reaction by nephelometer. Statistical analysis was done by SPSS version 10.0. Results: The PROM patients had significantly higher concentration of fibronectin (225.77 ± 115.18 ng/mL compared to that in non-PROM subjects (8.04 ± 16.17 ng/mL (p < 0.001. Conclusion: It can be concluded that in cases of unequivocal rupture or intactness of the membranes, the result of the fibronectin test corresponds well with the clinical situation. So fibronectin is a sensitive test for detection of amniotic fluid in the vagina.

  8. A two-helix motif positions the lysophosphatidic acid acyltransferase active site for catalysis within the membrane bilayer. (United States)

    Robertson, Rosanna M; Yao, Jiangwei; Gajewski, Stefan; Kumar, Gyanendra; Martin, Erik W; Rock, Charles O; White, Stephen W


    Phosphatidic acid (PA), the central intermediate in membrane phospholipid synthesis, is generated by two acyltransferases in a pathway conserved in all life forms. The second step in this pathway is catalyzed by 1-acyl-sn-glycerol-3-phosphate acyltransferase, called PlsC in bacteria. Here we present the crystal structure of PlsC from Thermotoga maritima, revealing an unusual hydrophobic/aromatic N-terminal two-helix motif linked to an acyltransferase αβ-domain that contains the catalytic HX4D motif. PlsC dictates the acyl chain composition of the 2-position of phospholipids, and the acyl chain selectivity 'ruler' is an appropriately placed and closed hydrophobic tunnel. We confirmed this by site-directed mutagenesis and membrane composition analysis of Escherichia coli cells that expressed mutant PlsC. Molecular dynamics (MD) simulations showed that the two-helix motif represents a novel substructure that firmly anchors the protein to one leaflet of the membrane. This binding mode allows the PlsC active site to acylate lysophospholipids within the membrane bilayer by using soluble acyl donors.

  9. Fluid lipid membranes: from differential geometry to curvature stresses. (United States)

    Deserno, Markus


    A fluid lipid membrane transmits stresses and torques that are fully determined by its geometry. They can be described by a stress- and torque-tensor, respectively, which yield the force or torque per length through any curve drawn on the membrane's surface. In the absence of external forces or torques the surface divergence of these tensors vanishes, revealing them as conserved quantities of the underlying Euler-Lagrange equation for the membrane's shape. This review provides a comprehensive introduction into these concepts without assuming the reader's familiarity with differential geometry, which instead will be developed as needed, relying on little more than vector calculus. The Helfrich Hamiltonian is then introduced and discussed in some depth. By expressing the quest for the energy-minimizing shape as a functional variation problem subject to geometric constraints, as proposed by Guven (2004), stress- and torque-tensors naturally emerge, and their connection to the shape equation becomes evident. How to reason with both tensors is then illustrated with a number of simple examples, after which this review concludes with four more sophisticated applications: boundary conditions for adhering membranes, corrections to the classical micropipette aspiration equation, membrane buckling, and membrane mediated interactions.

  10. Solid supported membranes doped with PIP2: influence of ionic strength and pH on bilayer formation and membrane organization. (United States)

    Braunger, Julia A; Kramer, Corinna; Morick, Daniela; Steinem, Claudia


    Phosphoinositides and in particular L-α-phosphatidylinositol-4,5-bisphosphate (PIP2) are key lipids controlling many cellular events and serve as receptors for a large number of intracellular proteins. To quantitatively analyze protein-PIP2 interactions in vitro in a time-resolved manner, planar membranes on solid substrates are highly desirable. Here, we describe an optimized protocol to form PIP2 containing planar solid supported membranes on silicon surfaces by vesicle spreading. Supported lipid bilayers (SLBs) were obtained by spreading POPC/PIP2 (92:8) small unilamellar vesicles onto hydrophilic silicon substrates at a low pH of 4.8. These membranes were capable of binding ezrin, resulting in large protein coverage as concluded from reflectometric interference spectroscopy and fluorescence microscopy. As deduced from fluorescence microscopy, only under low pH conditions, a homogeneously appearing distribution of fluorescently labeled PIP2 molecules in the membrane was achieved. Fluorescence recovery after photobleaching experiments revealed that PIP2 is not mobile in the bottom layer of the SLBs, while PIP2 is fully mobile in the top layer with diffusion coefficients of about 3 μm(2)/s. This diffusion coefficient was considerably reduced by a factor of about 3 if ezrin has been bound to PIP2 in the membrane.

  11. The influence of oscillating electromagnetic fields on membrane structure and function: Synthetic liposome and natural membrane bilayer systems with direct application to the controlled delivery of chemical agents

    Energy Technology Data Exchange (ETDEWEB)

    Liburdy, R.P.; de Manincor, D.; Fingado, B.


    Investigations have been conducted to determine if an imposed electromagnetic field can influence membrane transport, and ion and drug permeability in both synthetic and natural cell membrane systems. Microwave fields enhance accumulation of sodium in the lymphocyte and induce protein shedding at Tc. Microwaves also trigger membrane permeability of liposome systems under specific field exposure conditions. Sensitivity varies in a defined way in bilayers displaying a membrane structural phase transition temperature, Tc; maximal release was observed at or near Tc. Significantly, liposome systems without a membrane phase transition were also found to experience permeability increases but, in contrast, this response was temperature independent. The above results indicate that field-enhanced drug release occurs in liposome vesicles that possess a Tc as well as non-Tc liposomes. Additional studies extend non-Tc liposome responses to the in vivo case in which microwaves trigger Gentamicin release from a liposome depot'' placed subcutaneously in the rat hind leg. In addition, evidence is provided that cell surface sequestered liposomes can be triggered by microwave fields to release drugs directly into target cells. 24 refs., 6 figs.

  12. Steroid hormones partition to distinct sites in a model membrane bilayer: direct demonstration by small-angle X-ray diffraction. (United States)

    Golden, G A; Rubin, R T; Mason, R P


    The classical, genomic mechanisms of steroid hormone action cannot account for their rapid cellular effects. Membrane-bound steroid receptors have been partially characterized, but many rapid steroid effects occur in the absence of steroid-protein binding. Although it has been proposed that these effects could be due to steroid-induced biophysical alterations of the cell membrane, only indirect supporting evidence for this hypothesis has been forthcoming. In the present study, the ability of cortisol and estradiol (E2), natural steroids of different lipophilicity, to induce alterations in a model membrane (lecithin) bilayer was examined directly by small-angle X-ray diffraction under physiologic-like conditions. Within minutes, both steroids partitioned to distinct sites in the membrane. With increasing membrane cholesterol content, cortisol was displaced toward the polar headgroup region of the phospholipid bilayer, whereas E2 was displaced in the opposite direction, toward the nonpolar hydrocarbon core. Membrane-based partition coefficients (Kp[mem]) for both steroids (>100:1) were highest at those cholesterol concentrations that displaced the steroids toward the headgroup region (high cholesterol for cortisol; low for E2). Both steroids, when located in the headgroup region, increased overall bilayer width by 3-4 A, a change that could modulate the structure and function of integral membrane proteins independent from steroid effects on the genome.

  13. Structure and fluctuations of a single floating lipid bilayer. (United States)

    Daillant, J; Bellet-Amalric, E; Braslau, A; Charitat, T; Fragneto, G; Graner, F; Mora, S; Rieutord, F; Stidder, B


    A single lipid molecular bilayer of 17 or 18 carbon chain phosphocholines, floating in water near a flat wall, is prepared in the bilayer gel phase and then heated to the fluid phase. Its structure (electron density profile) and height fluctuations are determined by using x-ray reflectivity and non-specular scattering. By fitting the off-specular signal to that calculated for a two-dimensional membrane using a Helfrich Hamiltonian, we determine the three main physical quantities that govern the bilayer height fluctuations: The wall attraction potential is unexpectedly low; the surface tension, roughly independent on chain length and temperature, is moderate (approximately 5 x 10(-4) J.m(-2)) but large enough to dominate the intermediate range of the fluctuation spectrum; and the bending modulus abruptly decreases by an order-of-magnitude from 10(-18) J to 10(-19) J at the bilayer gel-to-fluid transition.

  14. Nuclear envelope remnants: fluid membranes enriched in sterols and polyphosphoinositides.

    Directory of Open Access Journals (Sweden)

    Marie Garnier-Lhomme

    Full Text Available BACKGROUND: The cytoplasm of eukaryotic cells is a highly dynamic compartment where membranes readily undergo fission and fusion to reorganize the cytoplasmic architecture, and to import, export and transport various cargos within the cell. The double membrane of the nuclear envelope that surrounds the nucleus, segregates the chromosomes from cytoplasm and regulates nucleocytoplasmic transport through pores. Many details of its formation are still unclear. At fertilization the sperm devoid of nuclear envelope pores enters the egg. Although most of the sperm nuclear envelope disassembles, remnants of the envelope at the acrosomal and centriolar fossae do not and are subsequently incorporated into the newly forming male pronuclear envelope. Remnants are conserved from annelid to mammalian sperm. METHODOLOGY/PRINCIPAL FINDINGS: Using lipid mass spectrometry and a new application of deuterium solid-state NMR spectroscopy we have characterized the lipid composition and membrane dynamics of the sperm nuclear envelope remnants in isolated sperm nuclei. CONCLUSIONS/SIGNIFICANCE: We report nuclear envelope remnants are relatively fluid membranes rich in sterols, devoid of sphingomyelin, and highly enriched in polyphosphoinositides and polyunsaturated phospholipids. The localization of the polybasic effector domain of MARCKS illustrates the non-nuclear aspect of the polyphosphoinositides. Based on their atypical biophysical characteristics and phospholipid composition, we suggest a possible role for nuclear envelope remnants in membrane fusion leading to nuclear envelope assembly.

  15. Biological Evaluation (In Vitro and In Vivo) of Bilayered Collagenous Coated (Nano Electrospun and Solid Wall) Chitosan Membrane for Periodontal Guided Bone Regeneration. (United States)

    Lotfi, Ghogha; Shokrgozar, Mohammad Ali; Mofid, Rasoul; Abbas, Fatemeh Mashhadi; Ghanavati, Farzin; Baghban, Alireza Akbarzadeh; Yavari, Seyedeh Kimia; Pajoumshariati, Seyedramin


    The application of barrier membranes in guided bone regeneration (GBR) has become a commonly used surgical technique in periodontal research. The objectives of this study were to evaluate the in vitro biocompatibility and osteogenic differentiation of mesenchymal stem cells (MSCs) on two different collagenous coatings (nano electrospun fibrous vs. solid wall) of bilayered collagen/chitosan membrane and their histological evaluation on bone regeneration in rabbit calvarial defects. It was found that chitosan-nano electrospun collagen (CNC) membranes had higher proliferation/metabolic activity compared to the chitosan-collagen (CC) and pristine chitosan membranes. The qRT-PCR analysis demonstrated the CNC membranes induced significant expression of osteogenic genes (Osteocalcin, RUNX2 and Col-α1) in MSCs. Moreover, higher calcium content and alkaline phosphatase activity of MSCs were observed compared to the other groups. Histologic and histomorphometric evaluations were performed on the uncovered (negative control) as well as covered calvarial defects of ten adult white rabbits with different membranes (CNC, CC, BioGide (BG, positive control)) at 1 and 2 months after surgery. More bone formation was detected in the defects covered with CNC and BG membranes than those covered by CC and the negative control. No inflammation and residual biomaterial particles were observed on the membrane surface or in the surrounding tissues in the surgical areas. These results suggest that bilayer CNC membrane can have the potential for use as a GBR membrane material facilitating bone formation.

  16. Antimicrobial peptides at work: interaction of myxinidin and its mutant WMR with lipid bilayers mimicking the P. aeruginosa and E. coli membranes (United States)

    Lombardi, Lucia; Stellato, Marco Ignazio; Oliva, Rosario; Falanga, Annarita; Galdiero, Massimiliano; Petraccone, Luigi; D'Errico, Geradino; de Santis, Augusta; Galdiero, Stefania; Del Vecchio, Pompea


    Antimicrobial peptides are promising candidates as future therapeutics in order to face the problem of antibiotic resistance caused by pathogenic bacteria. Myxinidin is a peptide derived from the hagfish mucus displaying activity against a broad range of bacteria. We have focused our studies on the physico-chemical characterization of the interaction of myxinidin and its mutant WMR, which contains a tryptophan residue at the N-terminus and four additional positive charges, with two model biological membranes (DOPE/DOPG 80/20 and DOPE/DOPG/CL 65/23/12), mimicking respectively Escherichia coli and Pseudomonas aeruginosa membrane bilayers. All our results have coherently shown that, although both myxinidin and WMR interact with the two membranes, their effect on membrane microstructure and stability are different. We further have shown that the presence of cardiolipin plays a key role in the WMR-membrane interaction. Particularly, WMR drastically perturbs the DOPE/DOPG/CL membrane stability inducing a segregation of anionic lipids. On the contrary, myxinidin is not able to significantly perturb the DOPE/DOPG/CL bilayer whereas interacts better with the DOPE/DOPG bilayer causing a significant perturbing effect of the lipid acyl chains. These findings are fully consistent with the reported greater antimicrobial activity of WMR against P. aeruginosa compared with myxinidin.

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

    Suwalsky, Mario; Belmar, Jessica; Villena, Fernando; Gallardo, María José; Jemiola-Rzeminska, Malgorzata; Strzalka, Kazimierz


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

  18. Sorption of Cationic Surfactants to Artificial Cell Membranes: Comparing Phospholipid Bilayers with Monolayer Coatings and Molecular Simulations. (United States)

    Timmer, Niels; Droge, Steven T J


    This study reports the distribution coefficient between phospholipid bilayer membranes and phosphate buffered saline (PBS) medium (DMW,PBS) for 19 cationic surfactants. The method used a sorbent dilution series with solid supported lipid membranes (SSLMs). The existing SSLM protocol, applying a 96 well plate setup, was adapted to use 1.5 mL glass autosampler vials instead, which facilitated sampling and circumvented several confounding loss processes for some of the cationic surfactants. About 1% of the phospholipids were found to be detached from the SSLM beads, resulting in nonlinear sorption isotherms for compounds with log DMW values above 4. Renewal of the medium resulted in linear sorption isotherms. DMW values determined at pH 5.4 demonstrated that cationic surfactant species account for the observed DMW,PBS. Log DMW,PBS values above 5.5 are only experimentally feasible with lower LC-MS/MS detection limits and/or concentrated extracts of the aqueous samples. Based on the number of carbon atoms, dialkylamines showed a considerably lower sorption affinity than linear alkylamine analogues. These SSLM results closely overlapped with measurements on a chromatographic tool based on immobilized artificial membranes (IAM-HPLC) and with quantum-chemistry based calculations with COSMOmic. The SSLM data suggest that IAM-HPLC underestimates the DMW of ionized primary and secondary alkylamines by 0.8 and 0.5 log units, respectively.

  19. Monitoring the Transmembrane Proton Gradient Generated by Cytochrome bo3 in Tethered Bilayer Lipid Membranes Using SEIRA Spectroscopy. (United States)

    Wiebalck, Swantje; Kozuch, Jacek; Forbrig, Enrico; Tzschucke, C Christoph; Jeuken, Lars J C; Hildebrandt, Peter


    Membrane proteins act as biocatalysts or ion/proton pumps to convert and store energy from ubiquitous environmental sources. Interfacing these proteins to electrodes allows utilizing the energy for enzymatic biofuel cells or other auspicious biotechnological applications. To optimize the efficiency of these devices, appropriate membrane models are required that ensure structural and functional integrity of the embedded enzymes and provide structural insight. We present a spectroelectrochemical surface-enhanced infrared absorption (SEIRA) and electrical impedance spectroscopy (EIS) study of the bacterial respiratory ubiquinol/cytochrome bo3 (cyt bo3) couple incorporated into a tethered bilayer lipid membrane (tBLM). Here, we employed a new lipid tether (WK3SH, dihydrocholesteryl (2-(2-(2-ethoxy)ethoxy)ethanethiol), which was synthesized using a three-step procedure with very good yield and allowed measuring IR spectra without significant spectral interference of the tBLM. The functional integrity of the incorporated cyt bo3 was demonstrated by monitoring the enzymatic O2 reduction current and the formation of the transmembrane proton gradient. Based on a SEIRA-spectroscopic redox titration, a shift of the pH-dependent redox potential of the ubiquinones under turnover conditions was correlated with an alkalinization of the submembrane reservoir by +0.8 pH units. This study demonstrates the high potential of tBLMs and the SEIRA spectroscopic approach to study bioenergetic processes.

  20. Marine sponge cyclic peptide theonellamide A disrupts lipid bilayer integrity without forming distinct membrane pores. (United States)

    Espiritu, Rafael Atillo; Cornelio, Kimberly; Kinoshita, Masanao; Matsumori, Nobuaki; Murata, Michio; Nishimura, Shinichi; Kakeya, Hideaki; Yoshida, Minoru; Matsunaga, Shigeki


    Theonellamides (TNMs) are antifungal and cytotoxic bicyclic dodecapeptides derived from the marine sponge Theonella sp. These peptides specifically bind to 3β-hydroxysterols, resulting in 1,3-β-D-glucan overproduction and membrane damage in yeasts. The inclusion of cholesterol or ergosterol in phosphatidylcholine membranes significantly enhanced the membrane affinity of theonellamide A (TNM-A) because of its direct interaction with 3β-hydroxyl groups of sterols. To better understand TNM-induced membrane alterations, we investigated the effects of TNM-A on liposome morphology. (31)P nuclear magnetic resonance (NMR) and dynamic light scattering (DLS) measurements revealed that the premixing of TNM-A with lipids induced smaller vesicle formation. When giant unilamellar vesicles were incubated with exogenously added TNM-A, confocal micrographs showed dynamic changes in membrane morphology, which were more frequently observed in cholesterol-containing than sterol-free liposomes. In conjunction with our previous data, these results suggest that the membrane action of TNM-A proceeds in two steps: 1) TNM-A binds to the membrane surface through direct interaction with sterols and 2) accumulated TNM-A modifies the local membrane curvature in a concentration-dependent manner, resulting in dramatic membrane morphological changes and membrane disruption.

  1. Multiscale Modeling of Supported Lipid Bilayers (United States)

    Hoopes, Matthew I.; Xing, Chenyue; Faller, Roland

    Cell membranes consist of a multitude of lipid molecules that serve as a framework for the even greater variety of membrane associated proteins [1-4]. As this highly complex (nonequilibrium) system cannot easily be understood and studied in a controlled way, a wide variety of model systems have been devised to understand the dynamics, structure, and thermodynamics in biological membranes. One such model system is a supported lipid bilayer (SLB), a two-dimensional membrane suspended on a surface. SLBs have been realized to be manageable experimentally while reproducing many of the key features of real biological membranes [5,6]. One of the main advantages of supported bilayers is the physical stability due to the solid support that enables a wide range of surface characterization techniques not available to free or unsupported membranes. As SLBs maintain some of the crucial structural and dynamic properties of biological membranes, they provide an important bridge to natural systems. In order to mimic cell membranes reliably, certain structural and dynamic features have to be reliably reproduced in the artificially constructed lipid bilayers. SLBs should display lateral mobility as in living cells, because many membrane activities involve transport, recruitment, or assembly of specific components. It is also critical for membranes to exhibit the correct thermodynamic phase, namely, a fluid lipid bilayer, to respond to environmental stress such as temperature and pressure changes [7]. There are several ways to fabricate supported lipid bilayers (SLBs) on planar substrates. One can use vesicle fusion on solid substrates [5,8-10] as well as Langmuir-Blodgett deposition [11,12]. Proteoliposome adsorption and subsequent membrane formation on a mica surface was first demonstrated by Brian and McConnell [13]. Because of its simplicity and reproducibility, this is one of the most common approaches to prepare supported membranes. A diverse range of different solid substrates

  2. Viscoelastic deformation of lipid bilayer vesicles† (United States)

    Wu, Shao-Hua; Sankhagowit, Shalene; Biswas, Roshni; Wu, Shuyang; Povinelli, Michelle L.


    Lipid bilayers form the boundaries of the cell and its organelles. Many physiological processes, such as cell movement and division, involve bending and folding of the bilayer at high curvatures. Currently, bending of the bilayer is treated as an elastic deformation, such that its stress-strain response is independent of the rate at which bending strain is applied. We present here the first direct measurement of viscoelastic response in a lipid bilayer vesicle. We used a dual-beam optical trap (DBOT) to stretch 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) giant unilamellar vesicles (GUVs). Upon application of a step optical force, the vesicle membrane deforms in two regimes: a fast, instantaneous area increase, followed by a much slower stretching to an eventual plateau deformation. From measurements of dozens of GUVs, the average time constant of the slower stretching response was 0.225 ± 0.033 s (standard deviation, SD). Increasing the fluid viscosity did not affect the observed time constant. We performed a set of experiments to rule out heating by laser absorption as a cause of the transient behavior. Thus, we demonstrate here that the bending deformation of lipid bilayer membranes should be treated as viscoelastic. PMID:26268612

  3. Viscoelastic deformation of lipid bilayer vesicles. (United States)

    Wu, Shao-Hua; Sankhagowit, Shalene; Biswas, Roshni; Wu, Shuyang; Povinelli, Michelle L; Malmstadt, Noah


    Lipid bilayers form the boundaries of the cell and its organelles. Many physiological processes, such as cell movement and division, involve bending and folding of the bilayer at high curvatures. Currently, bending of the bilayer is treated as an elastic deformation, such that its stress-strain response is independent of the rate at which bending strain is applied. We present here the first direct measurement of viscoelastic response in a lipid bilayer vesicle. We used a dual-beam optical trap (DBOT) to stretch 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) giant unilamellar vesicles (GUVs). Upon application of a step optical force, the vesicle membrane deforms in two regimes: a fast, instantaneous area increase, followed by a much slower stretching to an eventual plateau deformation. From measurements of dozens of GUVs, the average time constant of the slower stretching response was 0.225 ± 0.033 s (standard deviation, SD). Increasing the fluid viscosity did not affect the observed time constant. We performed a set of experiments to rule out heating by laser absorption as a cause of the transient behavior. Thus, we demonstrate here that the bending deformation of lipid bilayer membranes should be treated as viscoelastic.

  4. Molecular Dynamics of a Water-Lipid Bilayer Interface (United States)

    Wilson, Michael A.; Pohorille, Andrew


    We present results of molecular dynamics simulations of a glycerol 1-monooleate bilayer in water. The total length of analyzed trajectories is 5ns. The calculated width of the bilayer agrees well with the experimentally measured value. The interior of the membrane is in a highly disordered fluid state. Atomic density profile, orientational and conformational distribution functions, and order parameters indicate that disorder increases toward the center of the bilayer. Analysis of out-of-plane thermal fluctuations of the bilayer surfaces occurring at the time scale of the present calculations reveals that the distribution of modes agrees with predictions of the capillary wave model. Fluctuations of both bilayer surfaces are uncorrelated, yielding Gaussian distribution of instantaneous widths of the membrane. Fluctuations of the width produce transient thinning defects in the bilayer which occasionally span almost half of the membrane. The leading mechanism of these fluctuations is the orientational and conformational motion of head groups rather than vertical motion of the whole molecules. Water considerably penetrates the head group region of the bilayer but not its hydrocarbon core. The total net excess dipole moment of the interfacial water points toward the aqueous phase, but the water polarization profile is non-monotonic. Both water and head groups significantly contribute to the surface potential across the interface. The calculated sign of the surface potential is in agreement with that from experimental measurements, but the value is markedly overestimated. The structural and electrical properties of the water-bilayer system are discussed in relation to membrane functions, in particular transport of ions and nonelectrolytes across membranes.

  5. Molecular Dynamics of a Water-Lipid Bilayer Interface (United States)

    Wilson, Michael A.; Pohorille, Andrew


    We present results of molecular dynamics simulations of a glycerol 1-monooleate bilayer in water. The total length of analyzed trajectories is 5ns. The calculated width of the bilayer agrees well with the experimentally measured value. The interior of the membrane is in a highly disordered fluid state. Atomic density profile, orientational and conformational distribution functions, and order parameters indicate that disorder increases toward the center of the bilayer. Analysis of out-of-plane thermal fluctuations of the bilayer surfaces occurring at the time scale of the present calculations reveals that the distribution of modes agrees with predictions of the capillary wave model. Fluctuations of both bilayer surfaces are uncorrelated, yielding Gaussian distribution of instantaneous widths of the membrane. Fluctuations of the width produce transient thinning defects in the bilayer which occasionally span almost half of the membrane. The leading mechanism of these fluctuations is the orientational and conformational motion of head groups rather than vertical motion of the whole molecules. Water considerably penetrates the head group region of the bilayer but not its hydrocarbon core. The total net excess dipole moment of the interfacial water points toward the aqueous phase, but the water polarization profile is non-monotonic. Both water and head groups significantly contribute to the surface potential across the interface. The calculated sign of the surface potential is in agreement with that from experimental measurements, but the value is markedly overestimated. The structural and electrical properties of the water-bilayer system are discussed in relation to membrane functions, in particular transport of ions and nonelectrolytes across membranes.

  6. Exploring the Local Elastic Properties of Bilayer Membranes Using Molecular Dynamics Simulations

    DEFF Research Database (Denmark)

    Pieffet, Gilles; Botero, Alonso; Peters, Günther H.J.;


    of mean force (PMF) allowed us to dissect the elastic contribution. With this information, we calculated an effective linear spring constant of 44 +/- 4 kJ.nm-2.mol-1 for the DOPC membrane, in agreement with experimental estimates. The membrane deformation profile was determined independently during...... the stretching process in molecular detail, allowing us to fit this profile to a previously proposed continuum elastic model. Through this approach, we calculated an effective membrane spring constant of 42 kJ-2.mol-1, which is in good agreement with the PMF calculation. Furthermore, the solvation energy we...... derived from the data is shown to match the solvation energy estimated from critical micelle formation constants. This methodology can be used to determine how changes in lipid composition or the presence of membrane modifiers can affect the elastic properties of a membrane at a local level....

  7. Structure and dynamics of water and lipid molecules in charged anionic DMPG lipid bilayer membranes

    DEFF Research Database (Denmark)

    Rønnest, A. K.; Peters, Günther H.J.; Hansen, Flemming Yssing;


    the sensitivity to confirm the diffusion of a small component of water bound to the lipids as found in the simulations. In addition, the orientation of the dipole moment of the water molecules has been determined as a function of their depth in the membrane. Previous indirect estimates of the electrostatic...... compared to experimental results and used to determine an average diffusion constant for all water molecules in the system. On extrapolating the diffusion constants inferred experimentally to a temperature of 310 K, reasonable agreement with the simulations is obtained. However, the experiments do not have...... potential within phospholipid membranes imply an enormous electric field of 108-109 V m-1, which is likely to have great significance in controlling the conformation of translocating membrane proteins and in the transfer of ions and molecules across the membrane. We have calculated the membrane potential...

  8. Effects of pentanol isomers on the phase behavior of phospholipid bilayer membranes. (United States)

    Griffin, Kathryn L; Cheng, Chih-Yin; Smith, Eric A; Dea, Phoebe K


    Differential scanning calorimetry (DSC) was used to analyze the thermotropic phase behavior of dipalmitoylphosphatidylcholine (DPPC) bilayers in the presence of pentanol isomers. The concentration of each pentanol isomer needed to induce the interdigitated phase was determined by the appearance of a biphasic effect in the main transition temperatures, the onset of a hysteresis associated with the main transition from the gel-to-liquid crystalline phase, and the disappearance of the pretransition. Lower threshold concentrations were found to correlate with isomers of greater alkyl chain length while branching of the alkyl chain was found to increase biphasic behavior. The addition of a methyl group to butanol systems drastically decreased threshold concentrations. However, as demonstrated in the DPPC/neopentanol system, branching of the alkyl chain away from the -OH group lowers the threshold concentration while maintaining a biphasic effect.

  9. Low pH induces an interdigitated gel to bilayer gel phase transition in dihexadecylphosphatidylcholine membrane. (United States)

    Furuike, S; Levadny, V G; Li, S J; Yamazaki, M


    We have investigated the influence of pH on the structures and phase behaviors of multilamellar vesicles of the ether-linked dihexadecylphosphatidylcholine (DHPC-MLV). This phospholipid is known to be in the interdigitated gel (L(beta)I) phase in excess water at 20 degrees C at neutral pH. The results of X-ray diffraction experiments indicate that a phase transition from L(beta)I phase to the bilayer gel phase occurred in DHPC-MLV in 0.5 M KCl around pH 3.9 with a decrease in pH, and that at low pH values, less than pH 2.2, DHPC-MLVs were in L(beta') phase. The results of fluorescence and light scattering method indicate that the gel to liquid-crystalline phase transition temperature (T(m)) of DHPC-MLV increased with a decrease in pH. On the basis of a thermodynamic analysis, we conclude that the main mechanism of the low-pH induced L(beta)I to bilayer gel phase transition in DHPC-MLV and the increase in its T(m) is connected with the decrease in the repulsive interaction between the headgroups of these phospholipids. As pH decreases, the phosphate groups of the headgroups begin to be protonated, and as a result, the apparent positive surface charges appear. However, surface dipoles decrease and the interaction free energy of the hydrophilic segments with water increases. The latter effect dominates the pure electrostatic repulsion between the charged headgroups, and thereby, the total repulsive interaction in the interface decreases.

  10. Mesoscale computational studies of membrane bilayer remodeling by curvature-inducing proteins

    CERN Document Server

    Ramakrishnan, N; Radhakrishnan, Ravi


    Biological membranes constitute boundaries of cells and cell organelles. Physico-chemical mechanisms at the atomic scale are dictated by protein-lipid interaction strength, lipid composition, lipid distribution in the vicinity of the protein, shape and amino acid composition of the protein, and its amino acid contents. The specificity of molecular interactions together with the cooperativity of multiple proteins induce and stabilize complex membrane shapes at the mesoscale. These shapes span a wide spectrum ranging from the spherical plasma membrane to the complex cisternae of the Golgi apparatus. Mapping the relation between the protein-induced deformations at the molecular scale and the resulting mesoscale morphologies is key to bridging cellular experiments across the various length scales. In this review, we focus on the theoretical and computational methods used to understand the phenomenology underlying protein-driven membrane remodeling. The suite of methods discussed here can be tailored to applicatio...

  11. Catalytic photoinduced electron transport across a lipid bilayer mediated by a membrane-soluble electron relay. (United States)

    Limburg, B; Bouwman, E; Bonnet, S


    Unidirectional photocatalytic electron transfer from a hydrophilic electron donor encapsulated in the interior of a liposome, to a hydrophilic electron acceptor on the other side of the membrane, has been achieved using the simple membrane-soluble electron relay 1-methoxy-N-methylphenazinium (MMP(+)). The total amount of photoproduct (>140 nmol) exceeds the number of moles of MMP(+) present (125 nmol), thus showing that the transport of electrons is catalytic.

  12. Texture of lipid bilayer domains

    DEFF Research Database (Denmark)

    Jensen, Uffe Bernchou; Brewer, Jonathan R.; Midtiby, Henrik Skov


    chains. By imaging the intensity variations as a function of the polarization angle, we map the lateral variations of the lipid tilt within domains. Results reveal that gel domains are composed of subdomains with different lipid tilt directions. We have applied a Fourier decomposition method......We investigate the texture of gel (g) domains in binary lipid membranes composed of the phospholipids DPPC and DOPC. Lateral organization of lipid bilayer membranes is a topic of fundamental and biological importance. Whereas questions related to size and composition of fluid membrane domain...... are well studied, the possibility of texture in gel domains has so far not been examined. When using polarized light for two-photon excitation of the fluorescent lipid probe Laurdan, the emission intensity is highly sensitive to the angle between the polarization and the tilt orientation of lipid acyl...

  13. Is the fluid mosaic (and the accompanying raft hypothesis a suitable model to describe fundamental features of biological membranes? What may be missing?

    Directory of Open Access Journals (Sweden)

    Luis Alberto Bagatolli


    Full Text Available The structure, dynamics, and stability of lipid bilayers are controlled by thermodynamic forces, leading to overall tensionless membranes with a distinct lateral organization and a conspicuous lateral pressure profile. Bilayers are also subject to built-in curvature-stress instabilities that may be released locally or globally in terms of morphological changes leading to the formation of non-lamellar and curved structures. A key controller of the bilayer’s propensity to form curved structures is the average molecular shape of the different lipid molecules. Via the curvature stress, molecular shape mediates a coupling to membrane-protein function and provides a set of physical mechanisms for formation of lipid domains and laterally differentiated regions in the plane of the membrane. Unfortunately, these relevant physical features of membranes are often ignored in the most popular models for biological membranes. Results from a number of experimental and theoretical studies emphasize the significance of these fundamental physical properties and call for a refinement of the fluid mosaic model (and the accompanying raft hypothesis.

  14. Size-selective permeation of water-soluble polymers through the bilayer membrane of cyclodextrin vesicles investigated by PFG-NMR. (United States)

    Himmelein, Sabine; Sporenberg, Nora; Schönhoff, Monika; Ravoo, Bart Jan


    Cyclodextrin vesicles (CDVs) consist of a bilayer of amphiphilic cyclodextrins (CDs). CDVs exhibit CD cavities at their surface that are able to recognize and bind hydrophobic guest molecules via size-selective inclusion. In this study, the permeability of α- and β-CDVs is investigated by pulsed field gradient-stimulated echo (PFG-STE) nuclear magnetic resonance. Diffusion experiments with water and two types of water-soluble polymers, polyethylene glycol (PEG) and polypropylene glycol (PPG), revealed three main factors that influence the exchange rate and permeability of CDVs. First, the length of the hydrophobic chain of the CD amphiphile plays a crucial role. Reasonably, vesicles consisting of amphiphiles with a longer aliphatic chain are less permeable since both membrane thickness and melting temperature T(m) increase. Second, the exchange rate through the bilayer membrane depends on the molecular weight of the polymer and decreases with increasing weight of the polymer. Most interestingly, a size-selective distinction of permeation due to the embedded CDs in the bilayer membrane was found. The mechanism of permeation is shown to occur through the CD cavity, such that depending on the size of the cavity, permeation of polymers with different cross-sectional diameters takes place. Whereas PPG permeates through the membrane of β-CD vesicles, it does not permeate α-CD vesicles.

  15. Cationic Au Nanoparticle Binding with Plasma Membrane-like Lipid Bilayers: Potential Mechanism for Spontaneous Permeation to Cells Revealed by Atomistic Simulations

    DEFF Research Database (Denmark)

    Heikkila, E.; Martinez-Seara, H.; Gurtovenko, A. A.


    Despite being chemically inert as a bulk material, nanoscale gold can pose harmful side effects to living organisms. In particular, cationic Au nanoparticles (AuNP+) of 2 nm diameter or less permeate readily through plasma membranes and induce cell death. We report atomistic simulations of cationic...... Au nanoparticles interacting with realistic membranes and explicit solvent using a model system that comprises two cellular compartments, extracellular and cytosolic, divided by two asymmetric lipid bilayers. The membrane-AuNP+ binding and membrane reorganization processes are discovered...... to be governed by cooperative effects where AuNP+, counterions, water, and the two membrane leaflets all contribute. On the extracellular side, we find that the nanoparticle has to cross a free energy barrier of about 5 k(B)T prior forming a stable contact with the membrane. This results in a rearrangement...

  16. Structure of a fluid dioleoylphosphatidylcholine bilayer determined by joint refinement of x-ray and neutron diffraction data. II. Distribution and packing of terminal methyl groups.


    Wiener, M C; White, S H


    We continue in this paper the presentation of theoretical and experimental methods for the joint refinement of neutron and x-ray lamellar diffraction data for the analysis of fluid (L alpha phase) bilayer structure (Wiener, M. C., and S. H. White. 1991 a, b, c. Biophys. J. 59:162-173 and 174-185; Biochemistry. 30:6997-7008; Wiener, M. C., G. I. King, and S. H. White. Biophys. J. 60: 568-576). We show how to obtain the distribution and packing of the terminal methyls in the interior of a fluid...

  17. Stress analysis and fail-safe design of bilayered tubular supported ceramic membranes

    DEFF Research Database (Denmark)

    Kwok, Kawai; Frandsen, Henrik Lund; Søgaard, Martin


    . Stress distributions in two membrane systems have been analyzed and routes to minimize stress are proposed. For a Ba0.5Sr0.5Co0.8Fe0.2O3−δBa0.5Sr0.5Co0.8Fe0.2O3−δ membrane supported on a porous substrate of the same material under pressure-vacuum operation, the optimal configuration in terms...... gradient. Tailoring the thermal expansion coefficient of the support is an effective method to alleviate the total stress. Failure criteria for membrane fracture under compression are thereafter presented. It is found that the tolerable flaw size for fracture in compression is in the millimeter range...

  18. Effect of primycin on monovalent cation transport of erythrocyte membrane and lipid bilayer. (United States)

    Blaskó, K; Györgyi, S; Horváth, I


    The effects of primycin were investigated on the alkali-cation transport of human erythrocytes and on the electric conduction of bimolecular lipid membranes. In the concentration range of 3.10(-6) approximately 10(-5) M primycin increased the permeability of erythrocytes to alkali-cations according to the sequences Cs+ greater than Rb+ approximately K+ greater than Na+, while the conductance of the negatively charged phosphatidylserine bimolecular lipid membrane increased by 2 approximately 3 orders of magnitude. The resistance-lowering effect of primycin strongly depended on the cationic species applied and a selectivity order Na+ greater than K+ greater than Rb+ greater than Cs+ was found. A possible mechanism of the primycin-membrane interaction is suggested on the basis of experimental data.

  19. The distribution of lipid attached spin probes in bilayers: application to membrane protein topology. (United States)

    Vogel, Alexander; Scheidt, Holger A; Huster, Daniel


    The distribution of the lipid-attached doxyl electron paramagnetic resonance (EPR) spin label in 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine membranes has been studied by (1)H and (13)C magic angle spinning nuclear magnetic resonance relaxation measurements. The doxyl spin label was covalently attached to the 5th, 10th, and 16th carbons of the sn-2 stearic acid chain of a 1-palmitoyl-2-stearoyl-(5/10/16-doxyl)-sn-glycero-3-phosphocholine analog. Due to the unpaired electron of the spin label, (1)H and (13)C lipid relaxation rates are enhanced by paramagnetic relaxation. For all lipid segments the influence of paramagnetic relaxation is observed even at low probe concentrations. Paramagnetic relaxation rates provide a measure for the interaction strength between lipid segments and the doxyl group. Plotted along the membrane director a transverse distribution profile of the EPR probe is obtained. The chain-attached spin labels are broadly distributed in the membrane with a maximum at the approximate chain position of the probe. Both (1)H and (13)C relaxation measurements show these broad distributions of the doxyl group in the membrane indicating that (1)H spin diffusion does not influence the relaxation measurements. The broad distributions of the EPR label result from the high degree of mobility and structural heterogeneity in liquid-crystalline membranes. Knowing the distribution profiles of the EPR probes, their influence on relaxation behavior of membrane inserted peptide and protein segments can be studied by (13)C magic angle spinning nuclear magnetic resonance. As an example, the location of Ala residues positioned at three sites of the transmembrane WALP-16 peptide was investigated. All three doxyl-labeled phospholipid analogs induce paramagnetic relaxation of the respective Ala site. However, for well ordered secondary structures the strongest relaxation enhancement is observed for that doxyl group in the closest proximity to the respective Ala. Thus

  20. Molecular dynamics of leucine and dopamine transporter proteins in a model cell membrane lipid bilayer. (United States)

    Gedeon, Patrick C; Indarte, Martín; Surratt, Christopher K; Madura, Jeffry D


    The dopamine transporter (DAT) operates via facilitated diffusion, harnessing an inward Na(+) gradient to drive dopamine from the extracellular synaptic cleft to the neuron interior. The DAT is relevant to central nervous system disorders such as Parkinson disease and attention-deficit hyperactivity disorder and is the primary site of action for the abused psychostimulants cocaine and amphetamines. Crystallization of a DAT homolog, the bacterial leucine transporter LeuT, provided the first reliable 3-D DAT template. Here, the LeuT crystal structure and the DAT molecular model have been combined with their respective substrates, leucine and dopamine, in lipid bilayer molecular dynamics simulations toward tracking substrate movement along the protein's substrate/ion permeation pathway. Specifically, movement of residue pairs that comprise the "external gate" was followed as a function of substrate presence. The transmembrane (TM) 1 arginine-TM 10 aspartate strut formed less readily in DAT compared with LeuT, with or without substrate present. For LeuT but not DAT, the addition of substrate enhanced the chances of forming the TM 1-10 bridge. Also, movement of the fourth extracellular loop EL-4 in the presence of substrate was more pronounced for DAT, the EL-4 unwinding to a degree. The overall similarity between the LeuT and DAT molecular dynamics simulations indicated that LeuT was a legitimate model to guide DAT structure-function predictions. There were, nevertheless, differences significant enough to allow for DAT-unique insights, which may include how cocaine, methylphenidate (Ritalin, NIDA Drug Supply, Rockville, MD), and other DAT blockers are not recognized as substrates even though they can access the primary substrate binding pocket. Proteins 2010. (c) 2009 Wiley-Liss, Inc.

  1. The structural role of cholesterol in cell membranes: from condensed bilayers to lipid rafts. (United States)

    Krause, Martin R; Regen, Steven L


    CONSPECTUS: Defining the two-dimensional structure of cell membranes represents one of the most daunting challenges currently facing chemists, biochemists, and biophysicists. In particular, the time-averaged lateral organization of the lipids and proteins that make up these natural enclosures has yet to be established. As the classic Singer-Nicolson model of cell membranes has evolved over the past 40 years, special attention has focused on the structural role played by cholesterol, a key component that represents ca. 30% of the total lipids that are present. Despite extensive studies with model membranes, two fundamental issues have remained a mystery: (i) the mechanism by which cholesterol condenses low-melting lipids by uncoiling their acyl chains and (ii) the thermodynamics of the interaction between cholesterol and high- and low-melting lipids. The latter bears directly on one of the most popular notions in modern cell biology, that is, the lipid raft hypothesis, whereby cholesterol is thought to combine with high-melting lipids to form "lipid rafts" that float in a "sea" of low-melting lipids. In this Account, we first describe a chemical approach that we have developed in our laboratories that has allowed us to quantify the interactions between exchangeable mimics of cholesterol and low- and high-melting lipids in model membranes. In essence, this "nearest-neighbor recognition" (NNR) method involves the synthesis of dimeric forms of these lipids that contain a disulfide moiety as a linker. By means of thiolate-disulfide interchange reactions, equilibrium mixtures of dimers are then formed. These exchange reactions are initiated either by adding dithiothreitol to a liposomal dispersion to generate a small amount of thiol monomer or by including a small amount of thiol monomer in the liposomes at pH 5.0 and then raising the pH to 7.4. We then show how such NNR measurements have allowed us to distinguish between two very different mechanisms that have been

  2. Controlled Modulation of Lipid Bilayer State by a Photosensitive Membrane Effector

    DEFF Research Database (Denmark)

    Shen, Chen; Jørgensen, Lars; Zargarani, Dordaneh;


    The lipid membrane matrix represents a 2-D liquid-crystal, the properties of which, at fixed other conditions, are locally modulated by the presence of effectors as e.g. cholesterol (passive) or proteins (passive and active). Not only does the incorporation of effectors into the host matrix locally...... by a combination of spectroscopic (UV-vis, NMR, mass spectroscopy), thermodynamic (Langmuir compression, calorimetry) and structural studies (X-ray/neutron reflectometry, grazing incidence X-ray diffraction). The conformational change of the guest upon illumination is coupled into the host system, inducing...... as a response to the conformational switching of the guest effector via external light illumination. In a more general context, similar behavior may be found upon the conformational changes of membrane proteins during work....

  3. The Effect of Lidocaine · HCl on the Fluidity of Native and Model Membrane Lipid Bilayers


    Park, Jun-Seop; Jung, Tae-Sang; Noh, Yang-Ho; Kim, Woo-Sung; Park, Won-Ick; Kim, Young-Soo; Chung, In-Kyo; Sohn, Uy Dong; Bae, Soo-Kyung; Bae, Moon-Kyoung; Jang, Hye-Ock; Yun, Il


    The purpose of this study is to investigated the mechanism of pharmacological action of local anesthetic and provide the basic information about the development of new effective local anesthetics. Fluorescent probe techniques were used to evaluate the effect of lidocaine·HCl on the physical properties (transbilayer asymmetric lateral and rotational mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex, a...

  4. Fatty acid composition of membrane bilayers: importance of diet polyunsaturated fat balance. (United States)

    Abbott, Sarah K; Else, Paul L; Atkins, Taleitha A; Hulbert, A J


    In one of the most extensive analyses to date we show that the balance of diet n-3 and n-6 polyunsaturated fatty acids (PUFA) is the most important determinant of membrane composition in the rat under 'normal' conditions. Young adult male Sprague-Dawley rats were fed one of twelve moderate-fat diets (25% of total energy) for 8weeks. Diets differed only in fatty acid (FA) profiles, with saturate (SFA) content ranging 8-88% of total FAs, monounsaturate (MUFA) 6-65%, total PUFA 4-81%, n-6 PUFA 3-70% and n-3 PUFA 1-70%. Diet PUFA included only essential FAs 18:2n-6 and 18:3n-3. Balance between n-3 and n-6 PUFA is defined as the PUFA balance (n-3 PUFA as % of total PUFA) and ranged 1-86% in the diets. FA composition was measured for brain, heart, liver, skeletal muscle, erythrocytes and plasma phospholipids, as well as adipose tissue and plasma triglycerides. The conformer-regulator model was used (slope=1 indicates membrane composition completely conforming to diet). Extensive changes in diet SFA, MUFA and PUFA had minimal effect on membranes (average slopes 0.01, 0.07, 0.07 respectively), but considerable influence on adipose tissue and plasma triglycerides (average slopes 0.27, 0.53, 0.47 respectively). Diet balance between n-3 and n-6 PUFA had a biphasic influence on membrane composition. When n-3 PUFAdiet (average slope 0.95), while diet PUFA balance>10% had little influence (average slope 0.19). The modern human diet has an average PUFA balance ~10% and this will likely have significant health implications.

  5. Hydrophobic thickness of fluid planar monooleylglycerol membran maximally thinned by inversed micellisation

    DEFF Research Database (Denmark)

    Knudsen, P. J.; Mouritsen, Ole G.


    A procedure of making membranes of amphiphilic materials at the bottom of a U-shaped flexible plastic tube within an aqueous medium is described. The membranes were made sufficiently large in order for the annulus area to be neglected. Consequently the hydrophobic thickness of the membrane could...... be measured by a capacitance technique assuming the relative permittivity of the hydrophobic part of the bilayer. Introduction of an AC microvolt technique allowed manufacture of stable thick membranes by quenching the electroconstriction observed when DC electrical potentials in the millivolt range are used....... By continuously monitoring the hydrophobic thickness and by use of the AC microvolt technique the membrane-thinning process by chemical means could be studied in isolation because the electroconstriction was quenched. The maximally thinned hydrophobic thickness of a monooleylglycerol membrane measured at 38...

  6. Biomembrane disruption by silica-core nanoparticles: effect of surface functional group measured using a tethered bilayer lipid membrane. (United States)

    Liu, Ying; Zhang, Zhen; Zhang, Quanxuan; Baker, Gregory L; Worden, R Mark


    Engineered nanomaterials (ENM) have desirable properties that make them well suited for many commercial applications. However, a limited understanding of how ENM's properties influence their molecular interactions with biomembranes hampers efforts to design ENM that are both safe and effective. This paper describes the use of a tethered bilayer lipid membrane (tBLM) to characterize biomembrane disruption by functionalized silica-core nanoparticles. Electrochemical impedance spectroscopy was used to measure the time trajectory of tBLM resistance following nanoparticle exposure. Statistical analysis of parameters from an exponential resistance decay model was then used to quantify and analyze differences between the impedance profiles of nanoparticles that were unfunctionalized, amine-functionalized, or carboxyl-functionalized. All of the nanoparticles triggered a decrease in membrane resistance, indicating nanoparticle-induced disruption of the tBLM. Hierarchical clustering allowed the potency of nanoparticles for reducing tBLM resistance to be ranked in the order amine>carboxyl~bare silica. Dynamic light scattering analysis revealed that tBLM exposure triggered minor coalescence for bare and amine-functionalized silica nanoparticles but not for carboxyl-functionalized silica nanoparticles. These results indicate that the tBLM method can reproducibly characterize ENM-induced biomembrane disruption and can distinguish the BLM-disruption patterns of nanoparticles that are identical except for their surface functional groups. The method provides insight into mechanisms of molecular interaction involving biomembranes and is suitable for miniaturization and automation for high-throughput applications to help assess the health risk of nanomaterial exposure or identify ENM having a desired mode of interaction with biomembranes. © 2013. Published by Elsevier B.V. All rights reserved.

  7. Hamilton's equations for a fluid membrane

    Energy Technology Data Exchange (ETDEWEB)

    Capovilla, R [Departamento de Fisica, Centro de Investigacion y de Estudios Avanzados, Apdo. Postal 14-740, 07000 Mexico, DF (Mexico); Guven, J [Instituto de Ciencias Nucleares, Universidad Nacional Autonoma de Mexico, Apdo. Postal 70-543, 04510 Mexico, DF (Mexico); Rojas, E [Facultad de Fisica e Inteligencia Artificial, Universidad Veracruzana, 91000 Xalapa, Veracruz (Mexico)


    Consider a homogeneous fluid membrane described by the Helfrich-Canham energy, quadratic in the mean curvature of the membrane surface. The shape equation that determines equilibrium configurations is fourth order in derivatives and cubic in the mean curvature. We introduce a Hamiltonian formulation of this equation which dismantles it into a set of coupled first-order equations. This involves interpreting the Helfrich-Canham energy as an action; equilibrium surfaces are generated by the evolution of space curves. Two features complicate the implementation of a Hamiltonian framework. (i) The action involves second derivatives. This requires treating the velocity as a phase-space variable and the introduction of its conjugate momentum. The canonical Hamiltonian is constructed on this phase space. (ii) The action possesses a local symmetry-reparametrization invariance. The two labels we use to parametrize points on the surface are themselves physically irrelevant. This symmetry implies primary constraints, one for each label, that need to be implemented within the Hamiltonian. The two Lagrange multipliers associated with these constraints are identified as the components of the acceleration tangential to the surface. The conservation of the primary constraints implies two secondary constraints, fixing the tangential components of the momentum conjugate to the position. Hamilton's equations are derived and the appropriate initial conditions on the phase-space variables are identified. Finally, it is shown how the shape equation can be reconstructed from these equations.

  8. Size Control and Fractionation of Ionic Liquid Filled Polymersomes with Glassy and Rubbery Bilayer Membranes. (United States)

    So, Soonyong; Lodge, Timothy P


    We demonstrate control over the size of ionic liquid (IL) filled polymeric vesicles (polymersomes) by three distinct methods: mechanical extrusion, cosolvent-based processing in an IL, and fractionation of polymersomes in a biphasic system of IL and water. For the representative ionic liquid (1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl) imide ([EMIM][TFSI])), the size and dispersity of polymersomes formed from 1,2-polybutadiene-b-poly(ethylene oxide) (PB-PEO) and polystyrene-b-poly(ethylene oxide) (PS-PEO) diblock copolymers were shown to be sensitive to assembly conditions. During mechanical extrusion through a polycarbonate membrane, the relatively larger polymersomes were broken up and reorganized into vesicles with mean size comparable to the membrane pore (100 nm radius); the distribution width also decreased significantly after only a few passes. Other routes were studied using the solvent-switch or cosolvent (CS) method, whereby the initial content of the cosolvent and the PEO block length of PS-PEO were systemically changed. The nonvolatility of the ionic liquid directly led to the desired concentration of polymersomes in the ionic liquid using a single step, without the dialysis conventionally used in aqueous systems, and the mean vesicle size depended on the amount of cosolvent employed. Finally, selective phase transfer of PS-PEO polymersomes based on size was used to extract larger polymersomes from the IL to the aqueous phase via interfacial tension controlled phase transfer. The interfacial tension between the PS membrane and the aqueous phase was varied with the concentration of sodium chloride (NaCl) in the aqueous phase; then the larger polymersomes were selectively separated to the aqueous phase due to differences in shielding of the hydrophobic core (PS) coverage by the hydrophilic corona brush (PEO). This novel fractionation is a simple separation process without any special apparatus and can help to prepare monodisperse polymersomes

  9. Antioxidant effect of 4-nerolidylcatechol and α-tocopherol in erythrocyte ghost membranes and phospholipid bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Fernandes, K.S.; Silva, A.H.M.; Mendanha, S.A. [Instituto de Física, Universidade Federal de Goiás, Goiânia, GO (Brazil); Rezende, K.R. [Laboratório de Biofarmácia e Farmacocinética de Substâncias Bioativas, Faculdade de Farmácia, Universidade Federal de Goiás, Goiânia, GO (Brazil); Alonso, A. [Instituto de Física, Universidade Federal de Goiás, Goiânia, GO (Brazil)


    4-Nerolidylcatechol (4-NC) is found in Pothomorphe umbellata root extracts and is reported to have a topical protective effect against UVB radiation-induced skin damage, toxicity in melanoma cell lines, and antimalarial activity. We report a comparative study of the antioxidant activity of 4-NC and α-tocopherol against lipid peroxidation initiated by two free radical-generating systems: 2,2′-azobis(2-aminopropane) hydrochloride (AAPH) and FeSO{sub 4}/H{sub 2}O{sub 2}, in red blood cell ghost membranes and in egg phosphatidylcholine (PC) vesicles. Lipid peroxidation was monitored by membrane fluidity changes assessed by electron paramagnetic resonance spectroscopy of a spin-labeled lipid and by the formation of thiobarbituric acid-reactive substances. When lipoperoxidation was initiated by the hydroxyl radical in erythrocyte ghost membranes, both 4-NC and α-tocopherol acted in a very efficient manner. However, lower activities were observed when lipoperoxidation was initiated by the peroxyl radical; and, in this case, the protective effect of α-tocopherol was lower than that of 4-NC. In egg PC vesicles, malondialdehyde formation indicated that 4-NC was effective against lipoperoxidation initiated by both AAPH and FeSO{sub 4}/H{sub 2}O{sub 2}, whereas α-tocopherol was less efficient in protecting against lipoperoxidation by AAPH, and behaved as a pro-oxidant for FeSO{sub 4}/H{sub 2}O{sub 2}. The DPPH (2,2-diphenyl-1-picrylhydrazyl) free-radical assay indicated that two free radicals were scavenged per 4-NC molecule, and one free radical was scavenged per α-tocopherol molecule. These data provide new insights into the antioxidant capacity of 4-NC, which may have therapeutic applications for formulations designed to protect the skin from sunlight irradiation.

  10. Controlled Modulation of Lipid Bilayer State by a Photosensitive Membrane Effector

    DEFF Research Database (Denmark)

    Shen, Chen; Jørgensen, Lars; Zargarani, Dordaneh;

    The lipid membrane matrix represents a 2-D liquid-crystal, the properties of which, at fixed other conditions, are locally modulated by the presence of effectors as e.g. cholesterol (passive) or proteins (passive and active). Not only does the incorporation of effectors into the host matrix locally...... by a combination of spectroscopic (UV-vis, NMR, mass spectroscopy), thermodynamic (Langmuir compression, calorimetry) and structural studies (X-ray/neutron reflectometry, grazing incidence X-ray diffraction). The conformational change of the guest upon illumination is coupled into the host system, inducing...

  11. Interaction of antimicrobial peptide Plantaricin149a and four analogs with lipid bilayers and bacterial membranes

    Directory of Open Access Journals (Sweden)

    José Luiz de Souza Lopes


    Full Text Available The amidated analog of Plantaricin149, an antimicrobial peptide from Lactobacillus plantarum NRIC 149, directly interacts with negatively charged liposomes and bacterial membranes, leading to their lysis. In this study, four Pln149-analogs were synthesized with different hydrophobic groups at their N-terminus with the goal of evaluating the effect of the modifications at this region in the peptide's antimicrobial properties. The interaction of these peptides with membrane models, surface activity, their hemolytic effect on red blood cells, and antibacterial activity against microorganisms were evaluated. The analogs presented similar action of Plantaricin149a; three of them with no hemolytic effect (< 5% until 0.5 mM, in addition to the induction of a helical element when binding to negative liposomes. The N-terminus difference between the analogs and Plantaricin149a retained the antibacterial effect on S. aureus and P. aeruginosa for all peptides (MIC50 of 19 µM and 155 µM to Plantaricin149a, respectively but resulted in a different mechanism of action against the microorganisms, that was bactericidal for Plantaricin149a and bacteriostatic for the analogs. This difference was confirmed by a reduction in leakage action for the analogs. The lytic activity of Plantaricin149a is suggested to be a result of the peptide-lipid interactions from the amphipathic helix and the hydrophobic residues at the N-terminus of the antimicrobial peptide.

  12. Atomic force microscopy and spectroscopy to probe single membrane proteins in lipid bilayers. (United States)

    Sapra, K Tanuj


    The atomic force microscope (AFM) has opened vast avenues hitherto inaccessible to the biological scientist. The high temporal (millisecond) and spatial (nanometer) resolutions of the AFM are suited for studying many biological processes in their native conditions. The AFM cantilever stylus is aptly termed as a "lab on a tip" owing to its versatility as an imaging tool as well as a handle to manipulate single bonds and proteins. Recent examples assert that the AFM can be used to study the mechanical properties and monitor processes of single proteins and single cells, thus affording insight into important mechanistic details. This chapter specifically focuses on practical and analytical protocols of single-molecule AFM methodologies related to high-resolution imaging and single-molecule force spectroscopy of membrane proteins. Both these techniques are operator oriented, and require specialized working knowledge of the instrument, theoretical, and practical skills.

  13. Diffusion studies on permeable nitroxyl spin probe through lipid bilayer membrane

    Energy Technology Data Exchange (ETDEWEB)

    Benial, A. Milton Franklin; Meenakumari, V. [Department of Physics, NMSSVN College, Nagamalai, Madurai-625019 (India); Ichikawa, Kazuhiro; Yamada, Ken-ichi; Utsumi, Hideo, E-mail: [Department of Bio-functional Science, Kyushu University, Fukuoka (Japan); Hyodo, Fuminori [Innovation Center for Medical Redox Navigation, Kyushu University, Fukuoka (Japan); Jawahar, A. [Department of Chemistry, NMSSVN College, Nagamalai, Madurai-625 019 (India)


    Electron spin resonance (ESR) studies were carried out for 2mM {sup 14}N labeled deutrated permeable 3- methoxycarbonyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl (MC-PROXYL) in pure water, 1 mM, 2 mM, 3 mM and 4 mM concentration of MC-PROXYL in 300 mM concentration of liposomal solution by using a L-band ESR spectrometer. The ESR parameters such as linewidth, hyperfine coupling constant, g-factor, partition parameter and permeability were reported. The partition parameter and permeability values indicate the maximum spin distribution in the lipid phase at 2 mM concentration. This study illustrates that ESR can be used to differentiate between the intra and extra-membrane water by loading the liposome vesicles with a lipid-permeable nitroxyl spin probe. From the ESR results, the radical concentration was optimized as 2 mM in liposomal solution for ESR phantom studies and experiments.

  14. Amniotic Fluid Infection in Preterm Pregnancies with Intact Membranes (United States)

    Rahkonen, Leena; Nupponen, Irmeli; Pätäri-Sampo, Anu; Tikkanen, Minna; Sorsa, Timo; Juhila, Juuso; Andersson, Sture; Paavonen, Jorma; Stefanovic, Vedran


    Introduction. Intra-amniotic infection (IAI) is a major cause of preterm labor and adverse neonatal outcome. We evaluated amniotic fluid (AF) proteolytic cascade forming biomarkers in relation to microbial invasion of the amniotic cavity (MIAC) and IAI in preterm pregnancies with intact membranes. Material and Methods. Amniocentesis was made to 73 women with singleton pregnancies; 27 with suspected IAI; and 46 controls. AF biomarkers were divided into three cascades: Cascade 1: matrix metalloproteinase-8 (MMP-8), MMP-9, myeloperoxidase (MPO), and interleukin-6; Cascade 2: neutrophil elastase (HNE), elafin, and MMP-9; Cascade 3: MMP-2, tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), MMP-8/TIMP-1 molar ratio, and C-reactive protein (CRP). MMP-8 was measured by an immunoenzymometric assay and the others were measured by ELISA. Standard biochemical methods, molecular microbiology, and culture techniques were used. Results. MMP-8, MMP-9, MPO, elafin, and TIMP-1 concentrations were higher in IAI suspected cases compared to controls and also in IAI suspected cases with MIAC compared to those without MIAC when adjusted by gestational age at amniocentesis. All biomarkers except elafin and MMP-2 had the sensitivity of 100% with thresholds based on ROC-curve. Odd ratios of biomarkers for MIAC were 1.2-38 and 95% confidential intervals 1.0-353.6. Conclusions. Neutrophil based AF biomarkers were associated with IAI and MIAC. PMID:28167848

  15. Structural studies of mixed lipid bilayers on solid substrates using x-ray reflectivity (United States)

    Chen, Gang; Mukhopadhyay, Mrinmay; Ma, Yicong; Sinha, Sunil; Jiang, Zhang; Decaro, Curt; Berry, Justin; Lurio, Laurence; Brozell, Adrian; Parikh, Atul


    The lipid bilayers of natural membranes generally exist in a fluid state which occurs above the gel to liquid crystalline phase transition temperature. Knowledge of the structure of such bilayers is important for understanding fundamental biological processes mediated by or occurring within membranes. We have performed systematic measurements on bilayers of 1,2-Dipalmitoyl-sn-Glycero-3-Phosphoethanolamine (DPPE) and its mixture with 1,2-Dioleoyl-sn-Glycero-3-Phosphocholine (DOPC) and cholesterol (CH) on silicon substrates with x-ray reflectivity both below and above their phase transition temperatures. Structural variations as a function of temperature are demonstrated by fitting the reflectivity data with both a model dependent and a model independent routine. Studies of Au nanoparticle labeled DOPC and DOPC + DPPE + CH mixture are also performed and the location of Au nanoparticles in these bilayers is established by analyzing the x-ray reflectivity data.

  16. From biological membranes to biomimetic model membranes

    Directory of Open Access Journals (Sweden)

    Eeman, M.


    Full Text Available Biological membranes play an essential role in the cellular protection as well as in the control and the transport of nutrients. Many mechanisms such as molecular recognition, enzymatic catalysis, cellular adhesion and membrane fusion take place into the biological membranes. In 1972, Singer et al. provided a membrane model, called fluid mosaic model, in which each leaflet of the bilayer is formed by a homogeneous environment of lipids in a fluid state including globular assembling of proteins and glycoproteins. Since its conception in 1972, many developments were brought to this model in terms of composition and molecular organization. The main development of the fluid mosaic model was made by Simons et al. (1997 and Brown et al. (1997 who suggested that membrane lipids are organized into lateral microdomains (or lipid rafts with a specific composition and a molecular dynamic that are different to the composition and the dynamic of the surrounding liquid crystalline phase. The discovery of a phase separation in the plane of the membrane has induced an explosion in the research efforts related to the biology of cell membranes but also in the development of new technologies for the study of these biological systems. Due to the high complexity of biological membranes and in order to investigate the biological processes that occur on the membrane surface or within the membrane lipid bilayer, a large number of studies are performed using biomimicking model membranes. This paper aims at revisiting the fundamental properties of biological membranes in terms of membrane composition, membrane dynamic and molecular organization, as well as at describing the most common biomimicking models that are frequently used for investigating biological processes such as membrane fusion, membrane trafficking, pore formation as well as membrane interactions at a molecular level.

  17. General aspects of peptide selectivity towards lipid bilayers and cell membranes studied by variation of the structural parameters of amphipathic helical model peptides. (United States)

    Dathe, Margitta; Meyer, Jana; Beyermann, Michael; Maul, Björn; Hoischen, Christian; Bienert, Michael


    Model compounds of modified hydrophobicity (Eta), hydrophobic moment (mu) and angle subtended by charged residues (Phi) were synthesized to define the general roles of structural motifs of cationic helical peptides for membrane activity and selectivity. The peptide sets were based on a highly hydrophobic, non-selective KLA model peptide with high antimicrobial and hemolytic activity. Variation of the investigated parameters was found to be a suitable method for modifying peptide selectivity towards either neutral or highly negatively charged lipid bilayers. Eta and mu influenced selectivity preferentially via modification of activity on 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) bilayers, while the size of the polar/hydrophobic angle affected the activity against 1-palmitoyl-2-oleoylphosphatidyl-DL-glycerol (POPG). The influence of the parameters on the activity determining step was modest in both lipid systems and the activity profiles were the result of the parameters' influence on the second less pronounced permeabilization step. Thus, the activity towards POPC vesicles was determined by the high permeabilizing efficiency, however, changes in the structural parameters preferentially influenced the relatively moderate affinity. In contrast, intensive peptide accumulation via electrostatic interactions was sufficient for the destabilization of highly negatively charged POPG lipid membranes, but changes in the activity profile, as revealed by the modification of Phi, seem to be preferentially caused by variation of the low permeabilizing efficiency. The parameters proved very effective also in modifying antimicrobial and hemolytic activity. However, their influence on cell selectivity was limited. A threshold value of hydrophobicity seems to exist which restricted the activity modifying potential of mu and Phi on both lipid bilayers and cell membranes.

  18. Solid-state NMR spectroscopy of a membrane protein in biphenyl phospholipid bicelles with the bilayer normal parallel to the magnetic field (United States)

    Park, Sang Ho; Loudet, Cécile; Marassi, Francesca M.; Dufourc, Erick J.; Opella, Stanley J.


    Bicelles composed of the long-chain biphenyl phospholipid TBBPC (1-tetradecanoyl-2-(4-(4-biphenyl)butanoyl)-sn-glycero-3-PC) and the short-chain phospholipid DHPC align with their bilayer normals parallel to the direction of the magnetic field. In contrast, in typical bicelles the long-chain phospholipid is DMPC or DPPC, and the bilayers align with their normals perpendicular to the field. Samples of the membrane-bound form of the major coat protein of Pf1 bacteriophage in TBBPC bicelles are stable for several months, align magnetically over a wide range of temperatures, and yield well-resolved solid-state NMR spectra similar to those obtained from samples aligned mechanically on glass plates or in DMPC bicelle samples "flipped" with lanthanide ions so that their bilayer normals are parallel to the field. The order parameter of the TBBPC bicelle sample decreases from approximately 0.9 to 0.8 upon increasing the temperature from 20 °C to 60 °C. Since the frequency spans of the chemical shift and dipolar coupling interactions are twice as large as those obtained from proteins in DMPC bicelles without lanthanide ions, TBBPC bicelles provide an opportunity for structural studies with higher spectral resolution of the metal-binding membrane proteins without the risk of chemical or spectroscopic interference from the added lanthanide ions. In addition, the large temperature range of these samples is advantageous for the studies of membrane proteins that are unstable at elevated temperatures and for experiments requiring measurements as a function of temperature.

  19. Nanoparticle-lipid bilayer interactions studied with lipid bilayer arrays (United States)

    Lu, Bin; Smith, Tyler; Schmidt, Jacob J.


    The widespread environmental presence and commercial use of nanoparticles have raised significant health concerns as a result of many in vitro and in vivo assays indicating toxicity of a wide range of nanoparticle species. Many of these assays have identified the ability of nanoparticles to damage cell membranes. These interactions can be studied in detail using artificial lipid bilayers, which can provide insight into the nature of the particle-membrane interaction through variation of membrane and solution properties not possible with cell-based assays. However, the scope of these studies can be limited because of the low throughput characteristic of lipid bilayer platforms. We have recently described an easy to use, parallel lipid bilayer platform which we have used to electrically investigate the activity of 60 nm diameter amine and carboxyl modified polystyrene nanoparticles (NH2-NP and COOH-NP) with over 1000 lipid bilayers while varying lipid composition, bilayer charge, ionic strength, pH, voltage, serum, particle concentration, and particle charge. Our results confirm recent studies finding activity of NH2-NP but not COOH-NP. Detailed analysis shows that NH2-NP formed pores 0.3-2.3 nm in radius, dependent on bilayer and solution composition. These interactions appear to be electrostatic, as they are regulated by NH2-NP surface charge, solution ionic strength, and bilayer charge. The ability to rapidly measure a large number of nanoparticle and membrane parameters indicates strong potential of this bilayer array platform for additional nanoparticle bilayer studies.The widespread environmental presence and commercial use of nanoparticles have raised significant health concerns as a result of many in vitro and in vivo assays indicating toxicity of a wide range of nanoparticle species. Many of these assays have identified the ability of nanoparticles to damage cell membranes. These interactions can be studied in detail using artificial lipid bilayers, which

  20. Stokesian jellyfish: Viscous locomotion of bilayer vesicles

    CERN Document Server

    Evans, Arthur A; Lauga, Eric


    Motivated by recent advances in vesicle engineering, we consider theoretically the locomotion of shape-changing bilayer vesicles at low Reynolds number. By modulating their volume and membrane composition, the vesicles can be made to change shape quasi-statically in thermal equilibrium. When the control parameters are tuned appropriately to yield periodic shape changes which are not time-reversible, the result is a net swimming motion over one cycle of shape deformation. For two classical vesicle models (spontaneous curvature and bilayer coupling), we determine numerically the sequence of vesicle shapes through an enthalpy minimization, as well as the fluid-body interactions by solving a boundary integral formulation of the Stokes equations. For both models, net locomotion can be obtained either by continuously modulating fore-aft asymmetric vesicle shapes, or by crossing a continuous shape-transition region and alternating between fore-aft asymmetric and fore-aft symmetric shapes. The obtained hydrodynamic e...

  1. Probing Structural Dynamics and Topology of the KCNE1 Membrane Protein in Lipid Bilayers via Site-Directed Spin Labeling and Electron Paramagnetic Resonance Spectroscopy. (United States)

    Sahu, Indra D; Craig, Andrew F; Dunagan, Megan M; Troxel, Kaylee R; Zhang, Rongfu; Meiberg, Andrew G; Harmon, Corrinne N; McCarrick, Robert M; Kroncke, Brett M; Sanders, Charles R; Lorigan, Gary A


    KCNE1 is a single transmembrane protein that modulates the function of voltage-gated potassium channels, including KCNQ1. Hereditary mutations in the genes encoding either protein can result in diseases such as congenital deafness, long QT syndrome, ventricular tachyarrhythmia, syncope, and sudden cardiac death. Despite the biological significance of KCNE1, the structure and dynamic properties of its physiologically relevant native membrane-bound state are not fully understood. In this study, the structural dynamics and topology of KCNE1 in bilayered lipid vesicles was investigated using site-directed spin labeling (SDSL) and electron paramagnetic resonance (EPR) spectroscopy. A 53-residue nitroxide EPR scan of the KCNE1 protein sequence including all 27 residues of the transmembrane domain (45-71) and 26 residues of the N- and C-termini of KCNE1 in lipid bilayered vesicles was analyzed in terms of nitroxide side-chain motion. Continuous wave-EPR spectral line shape analysis indicated the nitroxide spin label side-chains located in the KCNE1 TMD are less mobile when compared to the extracellular region of KCNE1. The EPR data also revealed that the C-terminus of KCNE1 is more mobile when compared to the N-terminus. EPR power saturation experiments were performed on 41 sites including 18 residues previously proposed to reside in the transmembrane domain (TMD) and 23 residues of the N- and C-termini to determine the topology of KCNE1 with respect to the 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (POPG) lipid bilayers. The results indicated that the transmembrane domain is indeed buried within the membrane, spanning the width of the lipid bilayer. Power saturation data also revealed that the extracellular region of KCNE1 is solvent-exposed with some of the portions partially or weakly interacting with the membrane surface. These results are consistent with the previously published solution NMR

  2. Exploration of interactions between membrane proteins embedded in supported lipid bilayers and their antibodies by reflectometric interference spectroscopy-based sensing. (United States)

    Kurihara, Yoshikazu; Sawazumi, Tsuneo; Takeuchi, Toshifumi


    A microfluidic reflectometric interference spectroscopy (RIfS)-based sensor was fabricated to investigate the activity of multidrug resistance-associated protein 1 (MRP1), applied as a model membrane protein. Vesicles containing MRP1 were immobilized simply by injecting a vesicle solution (50 μg mL(-1)) onto a zirconium oxide (ZrO2) chip under constant flow conditions. Monitoring the shift of the minimum reflectance wavelength (Δλ) of the RIfS demonstrated that the vesicles were adsorbed onto the ZrO2 chip in a Langmuir-like fashion and suggested that the lipid bilayer structure was preserved on the ZrO2 chip. The theoretical maximum physical thickness of the layer was 4.97 nm, which was close to the values previously reported for supported lipid bilayers (4.2 to 5.2 nm). When a model protein, the anti-MRP1 antibody (1-50 μg mL(-1)), was injected onto the MRP1-immobilizing ZrO2 chip a concentration-dependent increase in Δλ was observed. In contrast, a ZrO2 chip on which the supported lipid bilayers did not contain MRP1 exhibited no response. Moreover, an anti-human IgG antibody generated no change in Δλ, confirming that anti-MRP1 antibodies were selectively bound to the MRP1 immobilized on the chip. These results show that the RIfS sensor can follow specific binding events of biologically active membrane proteins and represents a simple, label-free system capable of facilitating biomedical investigations.

  3. Oxygen as a paramagnetic probe for nuclear magnetic resonance: structure and paramagnetic profile of a lipid bilayer/membrane model system

    Energy Technology Data Exchange (ETDEWEB)

    Al-Abdul Wahid, M.S


    Paramagnetic contact shifts and relaxation rate enhancements from molecular oxygen dissolved in a model membrane, were studied by nuclear magnetic resonance spectroscopy. The model membrane system was an isotropic bicelle formed using 1-myristelaidoyl-2-myristoyl-d27-sn- glycero-3-phosphocholine (MLMPC), a custom phospholipid, and 1-2-dihexanoyl-d22-sn-glycero-3-phosphocholine (DHPC). The {sup 13}C and {sup 1}H spectra of MLMPC were assigned. Molecular oxygen was delivered at external pressures of 20 and 50 atm. Paramagnetic contact shifts were found to scale with the oxygen solubility gradient in the lipid bilayer, were found to be invariant to temperature changes in the region studied (288K to 331K), and scaled linearly with changes in oxygen pressure. Relaxation rate enhancements from oxygen were low in the headgroup region and increased to a roughly constant rate in the acyl chain region. Rates were comparable to values predicted by simple thermodynamic theories which take into account the observed gradients in diffusion rates and solubility of oxygen in bilayers. (author)

  4. The action of polyene antibiotics on lipid bilayer membranes in the presence of several cations and anions

    NARCIS (Netherlands)

    Zutphen, H. van; Demel, R.A.; Norman, A.W.; Deenen, L.L.M. van


    1. 1. Filipin complex, filipin II, filipin III, nystatin, etruscomycin, and pimaricin at concentrations of 10−5 M were able to disrupt bimolecular lipid films containing lecithin and cholesterol in a 1:1 molar ratio. 2. 2. The above antibiotics were not able to disrupt lecithin bilayer

  5. Membrane fusion induced by a short fusogenic peptide is assessed by its insertion and orientation into target bilayers

    NARCIS (Netherlands)

    Martin, [No Value; Pecheur, EI; Ruysschaert, JM; Hoekstra, D


    To clarify the molecular mechanism by which an amphipathic negatively charged peptide consisting of 11 residues (WAE) induces fusion, and the relevance of these features for fusion, its mode of insertion and orientation into target bilayers were investigated. Using attenuated total reflection

  6. Pt and PtRu catalyst bilayers increase efficiencies for ethanol oxidation in proton exchange membrane electrolysis and fuel cells (United States)

    Altarawneh, Rakan M.; Pickup, Peter G.


    Polarization curves, product distributions, and reaction stoichiometries have been measured for the oxidation of ethanol at anodes consisting of Pt and PtRu bilayers and a homogeneous mixture of the two catalysts. These anode structures all show synergies between the two catalysts that can be attributed to the oxidation of acetaldehyde produced at the PtRu catalyst by the Pt catalyst. The use of a PtRu layer over a Pt layer produces the strongest effect, with higher currents than a Pt on PtRu bilayer, mixed layer, or either catalyst alone, except for Pt at high potentials. Reaction stoichiometries (average number of electrons transferred per ethanol molecule) were closer to the values for Pt alone for both of the bilayer configurations but much lower for PtRu and mixed anodes. Although Pt alone would provide the highest overall fuel cell efficiency at low power densities, the PtRu on Pt bilayer would provide higher power densities without a significant loss of efficiency. The origin of the synergy between the Pt and PtRu catalysts was elucidated by separation of the total current into the individual components for generation of carbon dioxide and the acetaldehyde and acetic acid byproducts.

  7. A general theory of non-equilibrium dynamics of lipid-protein fluid membranes

    DEFF Research Database (Denmark)

    Lomholt, Michael Andersen; Hansen, Per Lyngs; Miao, L.


    We present a general and systematic theory of non-equilibrium dynamics of multi-component fluid membranes, in general, and membranes containing transmembrane proteins, in particular. Developed based on a minimal number of principles of statistical physics and designed to be a meso...

  8. Aspirin inhibits formation of cholesterol rafts in fluid lipid membranes. (United States)

    Alsop, Richard J; Toppozini, Laura; Marquardt, Drew; Kučerka, Norbert; Harroun, Thad A; Rheinstädter, Maikel C


    Aspirin and other non-steroidal anti-inflammatory drugs have a high affinity for phospholipid membranes, altering their structure and biophysical properties. Aspirin has been shown to partition into the lipid head groups, thereby increasing membrane fluidity. Cholesterol is another well known mediator of membrane fluidity, in turn increasing membrane stiffness. As well, cholesterol is believed to distribute unevenly within lipid membranes leading to the formation of lipid rafts or plaques. In many studies, aspirin has increased positive outcomes for patients with high cholesterol. We are interested if these effects may be, at least partially, the result of a non-specific interaction between aspirin and cholesterol in lipid membranes. We have studied the effect of aspirin on the organization of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) membranes containing cholesterol. Through Langmuir-Blodgett experiments we show that aspirin increases the area per lipid and decreases compressibility at 32.5 mol% cholesterol, leading to a significant increase of fluidity of the membranes. Differential scanning calorimetry provides evidence for the formation of meta-stable structures in the presence of aspirin. The molecular organization of lipids, cholesterol and aspirin was studied using neutron diffraction. While the formation of rafts has been reported in binary DPPC/cholesterol membranes, aspirin was found to locally disrupt membrane organization and lead to the frustration of raft formation. Our results suggest that aspirin is able to directly oppose the formation of cholesterol structures through non-specific interactions with lipid membranes. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Kinetics of the lamellar gel-fluid transition in phosphatidylcholine membranes in the presence of sugars

    Energy Technology Data Exchange (ETDEWEB)

    Lenné, Thomas; Garvey, Christopher J.; Koster, Karen L.; Bryant, Gary [ANSTO; (USD); (RMIT)


    Phase diagrams are presented for dipalmitoylphosphatidylcholine (DPPC) in the presence of sugars (sucrose) over a wide range of relative humidities (RHs). The phase information presented here, determined by small angle X-ray scattering (SAXS), is shown to be consistent with previous results achieved by differential scanning calorimetry (DSC). Both techniques show a significant effect of sucrose concentration on the phase behaviour of this phospholipid bilayer. An experimental investigation into the effect of sugars on the kinetic behaviour of the gel to fluid transition is also presented showing that increasing the sugar content appears to slightly increase the rate at which the transition occurs.

  10. Corrugation of Phase-Separated Lipid Bilayers Supported by Nanoporous Silica Xerogel Surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Goksu, E I; Nellis, B A; Lin, W; Satcher Jr., J H; Groves, J T; Risbud, S H; Longo, M L


    Lipid bilayers supported by substrates with nanometer-scale surface corrugations holds interest in understanding both nanoparticle-membrane interactions and the challenges of constructing models of cell membranes on surfaces with desirable properties, e.g. porosity. Here, we successfully form a two-phase (gel-fluid) lipid bilayer supported by nanoporous silica xerogel. Surface topology, diffusion, and lipid density in comparison to mica-supported lipid bilayers were characterized by AFM, FRAP, FCS, and quantitative fluorescence microscopy, respectively. We found that the two-phase lipid bilayer follows the xerogel surface contours. The corrugation imparted on the lipid bilayer results in a lipid density that is twice that on a flat mica surface. In direct agreement with the doubling of actual bilayer area in a projected area, we find that the lateral diffusion coefficient (D) of lipids on xerogel ({approx}1.7 {micro}m{sup 2}/s) is predictably lower than on mica ({approx}4.1 {micro}m{sup 2}/s) by both FRAP and FCS techniques. Furthermore, the gel-phase domains on xerogel compared to mica were larger and less numerous. Overall, our results suggest the presence of a relatively defect-free continuous two-phase bilayer that penetrates approximately midway into the first layer of {approx}50 nm xerogel beads.

  11. Amniotic membrane and amniotic fluid-derived cells: potential tools for regenerative medicine? (United States)

    Parolini, Ornella; Soncini, Maddalena; Evangelista, Marco; Schmidt, Dörthe


    Human amniotic membranes and amniotic fluid have attracted increasing attention in recent years as a possible reserve of stem cells that may be useful for clinical application in regenerative medicine. Many studies have been conducted to date in terms of the differentiation potential of these cells, with several reports demonstrating that cells from both the amniotic fluid and membrane display high plasticity. In addition, cells from the amniotic membrane have also been shown to display immunomodulatory characteristics both in vivo and in vitro, which could make them useful in an allotransplantation setting. Here, we provide an overview comparing the latest findings regarding the stem characteristics of cells from both the amniotic membrane and amniotic fluid, as well as on the potential utility of these cells for future clinical application in regenerative medicine.

  12. Horizontal Bilayer for Electrical and Optical Recordings

    Directory of Open Access Journals (Sweden)

    Alf Honigmann


    Full Text Available Artificial bilayer containing reconstituted ion channels, transporters and pumps serve as a well-defined model system for electrophysiological investigations of membrane protein structure–function relationship. Appropriately constructed microchips containing horizontally oriented bilayers with easy solution access to both sides provide, in addition, the possibility to investigate these model bilayer membranes and the membrane proteins therein with high resolution fluorescence techniques up to the single-molecule level. Here, we describe a bilayer microchip system in which long-term stable horizontal free-standing and hydrogel-supported bilayers can be formed and demonstrate its prospects particularly for single-molecule fluorescence spectroscopy and high resolution fluorescence microscopy in probing the physicochemical properties like phase behavior of the bilayer-forming lipids, as well as in functional studies of membrane proteins.

  13. Studies of Phase Transitions and Critical Phenomena: I. Origin of Broken Particle-Hole Symmetry in Critical Fluids. I. Phase Transitions of Interacting Membranes. (United States)

    Goldstein, Raymond Ethan

    The longstanding problem of the precise correspondence between critical phenomena in fluids and ferromagnets is resolved in Part I through a synthesis of mean field theory, exact results for lattice models, field-theoretic techniques, and by extensive quantitative comparison with experiment. Emphasis is placed on the origin of broken particle-hole symmetry in fluids as reflected in the form of the critical point scaling fields and in systematic variations in certain nonuniversal critical amplitudes with molecular polarizability. Those trends and the degree to which the scaling axes are linearly mixed versions of the bare "thermal" and "magnetic" fields in particle-hole symmetric systems are shown both for lattice models and real fluids to be intimately related to the presence of many-body interactions of the Axilrod-Teller type. A quantitatively accurate microscopic expression for the field-mixing operator of fluids is derived on the basis of an exact Hubbard-Stratonovich transformation relating the fluid Hamiltonian to that of a Landau-Ginzburg-Wilson model. A phenomenological theory of the phase behavior of multilamellar liquid crystals of hydrated phospholipid bilayers is developed in Part II, and its predictions tested by extensive comparison with experiment. A Ginzburg-Landau free energy functional based on the elastic properties of two coupled monolayers is proposed to describe intrabilayer ordering, and the phenomenon of structural phase transitions driven by membrane interactions is described by incorporating in addition the attractive dispersion interactions and repulsive "hydration" forces acting between membranes. The theory indicates and experiments support a connection between the pseudocriticality of the bilayer transitions and the large susceptibility of the in-plane order to membrane interactions. The pseudocriticality in turn is suggested to arise from the analog of a capillary critical point accessible by finite-size effects. Theoretical phase

  14. A new and robust method of tethering IgG surrogate antigens on lipid bilayer membranes to facilitate the TIRFM based live cell and single molecule imaging experiments.

    Directory of Open Access Journals (Sweden)

    Shaosen Zhang

    Full Text Available Our understanding of cell-cell interactions has been significantly improved in the past years with the help of Total Internal Reflection Fluorescence Microscope (TIRFM in combination with an antigen presenting system supported by planar lipid bilayer (PLB membranes, which are used to mimic the extensive receptor and ligand interactions within cell-cell contact interface. In TIRFM experiments, it is a challenge to uniformly present ligand molecules in monomeric format on the surface of PLB membranes. Here, we introduce a new and robust method of tethering IgG surrogate antigen ligands on the surface of Ni(2+-containing PLB membranes. In this method, we use a modified D domain from staphylococcal protein A molecule that is fused with an N-terminus polyhistidine tag (H12-D-domain to tether IgG surrogate antigens on Ni(2+-containing PLB membranes. We systematically assessed the specificity and capability of H12-D-domain construct to capture IgG molecules from different species through live cell and single molecule TIRFM imaging. We find that these IgG surrogate antigens tethered by H12-D-domain show better lateral mobility and are more uniformly distributed on PLB membranes than the ones tethered by streptavidin. Neither IgM molecules, nor Fab or F(ab'2 fragments of IgG molecules can be tethered on PLB membranes by H12-D-domain construct. These tethered IgG surrogate antigens strongly induce the formation and accumulation of signaling active antigen receptor microclusters within the immunological synapse in B or T lymphocyte cells. Thus our method provides a new and robust method to tether IgG surrogate antigens or other molecules fused with IgG Fc portion on PLB membranes for TIRFM based molecule imaging experiments.

  15. Computational fluid dynamics simulations of flow and concentration polarization in forward osmosis membrane systems

    DEFF Research Database (Denmark)

    Gruber, M.F.; Johnson, C.J.; Tang, C.Y.;


    Forward osmosis is an osmotically driven membrane separation process that relies on the utilization of a large osmotic pressure differential generated across a semi-permeable membrane. In recent years forward osmosis has shown great promise in the areas of wastewater treatment, seawater...... the understanding of membrane systems, models that can accurately encapsulate all significant physical processes occurring in the systems are required. The present study demonstrates a computational fluid dynamics (CFD) model capable of simulating forward osmosis systems with asymmetric membranes. The model...

  16. Tension induced phase transitions in biomimetic fluid membranes (United States)

    Shapiro, Marc; Vlahovska, Petia


    Membranes in eukaryotic cells are mixtures of hundreds of lipid species. The lipid diversity enables membranes to phase separate and form domains, called rafts, which play a critical role in cell functions such as signaling and trafficking. The phase transitions underlying raft formation have been extensively studied as a function of temperature and composition. However, the third dimension of the phase diagram, i.e., the tension (2D pressure), is still unexplored because membrane tension is difficult to control and quantify. To overcome this challenge, we develop two approaches, capillary micromechanics and electrodeformation, in which the tension is regulated by the area dilation accompanying deformation of a vesicle (a closed membrane). The first technique consists of forcing an initially quasi-spherical vesicle through a tapered glass microcapillary, while the second method utilizes uniform electric fields to deform the vesicle into an ellipsoid. Domains are visualized using a fluorescent dye, which preferentially partitions in one of the phases. The experimental results suggest that the miscibility temperature (at which domains form in an initially homogeneous membrane) increases with applied tension. Domain motions and coarsening are also investigated.

  17. DNA nanotechnology: Bringing lipid bilayers into shape (United States)

    Howorka, Stefan


    Lipid bilayers form the thin and floppy membranes that define the boundary of compartments such as cells. Now, a method to control the shape and size of bilayers using DNA nanoscaffolds has been developed. Such designer materials advance synthetic biology and could find use in membrane research.

  18. Membrane finite element method for simulating fluid flow in porous medium

    Institute of Scientific and Technical Information of China (English)

    Mei-li ZHAN; Wen-jie ZHANG; Jin-chang SHENG; Jian-hui LI; Shu-yuan HE


    A new membrane finite element method for modeling fluid flow in a porous medium is presented in order to quickly and accurately simulate the geo-membrane fabric used in civil engineering. It is based on discontinuous finite element theory, and can be easily coupled with the normal Galerkin finite element method. Based on the saturated seepage equation, the element coefficient matrix of the membrane element method is derived, and a geometric transform relation for the membrane element between a global coordinate system and a local coordinate system is obtained. A method for the determination of the fluid flux conductivity of the membrane element is presented. This method provides a basis for determining discontinuous parameters in discontinuous finite element theory. An anti-seepage problem regarding the foundation of a building is analyzed by coupling the membrane finite element method with the normal Galerkin finite element method. The analysis results demonstrate the utility and superiority of the membrane finite element method in fluid flow analysis of a porous medium.

  19. Modelling Meso-Scale Diffusion Processes in Stochastic Fluid Bio-Membranes

    CERN Document Server

    Rafii-Tabar, H


    The space-time dynamics of rigid inhomogeneities (inclusions) free to move in a randomly fluctuating fluid bio-membrane is derived and numerically simulated as a function of the membrane shape changes. Both vertically placed (embedded) inclusions and horizontally placed (surface) inclusions are considered. The energetics of the membrane, as a two-dimensional (2D) meso-scale continuum sheet, is described by the Canham-Helfrich Hamiltonian, with the membrane height function treated as a stochastic process. The diffusion parameter of this process acts as the link coupling the membrane shape fluctuations to the kinematics of the inclusions. The latter is described via Ito stochastic differential equation. In addition to stochastic forces, the inclusions also experience membrane-induced deterministic forces. Our aim is to simulate the diffusion-driven aggregation of inclusions and show how the external inclusions arrive at the sites of the embedded inclusions. The model has potential use in such emerging fields as...

  20. Estimation of the bending rigidity and spontaneous curvature of fluid membranes in simulations (United States)

    Shiba, Hayato; Noguchi, Hiroshi


    Several numerical methods for measuring the bending rigidity and the spontaneous curvature of fluid membranes are studied using two types of meshless membrane models. The bending rigidity is estimated from the thermal undulations of planar and tubular membranes and the axial force of tubular membranes. We found a large dependence of its estimate value from the thermal undulation analysis on the upper-cutoff frequency qcut of the least-squares fit. The inverse power-spectrum fit with an extrapolation to qcut→0 yields the smallest estimation error among the investigated methods. The spontaneous curvature is estimated from the axial force of tubular membranes and the average curvature of bent membrane strips. The results of these methods show good agreement with each other.

  1. A novel CHS/ALG bi-layer composite membrane with sustained antimicrobial efficacy used as wound dressing

    Institute of Scientific and Technical Information of China (English)

    Yang Dong; Hong Zhuo Liu; Lu Xu; Gang Li; Zi Ning Ma; Fei Han; Hui Min Yao; Yan Hui Sun; San Ming Li


    A membrane composed of an alginate (ALG) layer and a chitosan (CHS) layer with sustained antimicrobial efficacy was prepared. Ciprofloxacin HCI (CIP) was incorporated into the ALG layer. Morphological feature of the composite membrane was analyzed by scanning electron microscopy (SEM). Water uptake capacity, in vitro drug release, and in vitro antimicrobial activity were evaluated. The composite membrane exhibited perfect binding characteristic between the two layers. The water uptake capacity of all the membranes was above 800%. The CIP could release from the composite membranes for 48 h. The membrane could control the bacterial growth persistently. The results suggested that this CHS/ALG composite membrane incorporated with CIP had the potential for wound dressing application.

  2. Dissection of LolB function--lipoprotein binding, membrane targeting and incorporation of lipoproteins into lipid bilayers. (United States)

    Tsukahara, Jun; Mukaiyama, Keita; Okuda, Suguru; Narita, Shin-ichiro; Tokuda, Hajime


    Escherichia coli cells express at least 90 species of lipoprotein. LolB is one of the essential outer membrane lipoproteins, being involved in the last step of lipoprotein sorting. It accepts lipoproteins from a periplasmic molecular chaperone, LolA, and mediates the outer membrane anchoring of lipoproteins through a largely unknown mechanism. It has been shown previously that a LolB derivative, mLolB, lacking an N-terminal acyl chain, can bind lipoproteins. We examined how the lack of an N-terminal anchor affects the outer membrane anchoring of lipoproteins. Surprisingly, mLolB compensates for LolB function and supports E. coli growth, indicating that the N-terminal anchor is not essential for its function. Indeed, mLolB correctly localizes lipoproteins to either the inner or outer membrane depending on the sorting signal at the steady state. Furthermore, periplasmic mLolB enables the dissection of LolB function, namely lipoprotein binding, membrane targeting and lipoprotein anchoring. It mediates the transfer of lipoproteins from LolA to the outer membrane, but also the inner membrane and liposomes, indicating that mLolB exhibits no membrane preference and targets to phospholipids. Consequently, an outer membrane-specific lipoprotein is transiently mislocalized to the inner membrane when cells express only mLolB. LolB anchored to the outer membrane does not cause such mislocalization and is more active than mLolB. Phosphatidylethanolamine has been found to stimulate the mLolB-dependent membrane anchoring of lipoproteins. Taken together, these results indicate that lipoprotein binding, membrane targeting and membrane incorporation of lipoproteins are intrinsic functions of LolB.

  3. Bilayer Poly(Lactic-co-glycolic acid/Nano-Hydroxyapatite Membrane with Barrier Function and Osteogenesis Promotion for Guided Bone Regeneration

    Directory of Open Access Journals (Sweden)

    Li Fu


    Full Text Available Guided bone regeneration (GBR is one such treatment that reconstructs neo-bone tissue by using a barrier membrane to prevent the invasion of soft tissue and to create a space for guiding new bone growth into the bone defect. Herein, we report a novel functionally graded bilayer membrane (FGBM for GBR application. To fabricate the novel membrane, the composites of poly(lactic-co-glycolic acid and nano-hydroxyapatite were prepared by phase inversion for the dense layer and by electrospinning for another porous layer, and their corresponding properties were evaluated including surface morphology, mechanics, degradability, cell barrier function, and in vitro osteogenic bioactivity. The results showed that PLGA with 5% nHA in dense layer could meet the requirement of mechanical strength and have excellent barrier function even on condition of post-degradation. Furthermore, PLGA with 30% nHA in porous layer could achieve the good physical and chemical properties. In addition, 30% nHA incorporation would enhance the in vitro mineralization, and have superior capabilities of cell adhesion, proliferation and differentiation compared to other groups. Therefore, the designed FGBM could potentially serve as a barrier for preferential tissue ingrowth and achieve a desirable therapeutic result for bone tissue regeneration.

  4. Determination of structural topology of a membrane protein in lipid bilayers using polarization optimized experiments (POE) for static and MAS solid state NMR spectroscopy. (United States)

    Mote, Kaustubh R; Gopinath, T; Veglia, Gianluigi


    The low sensitivity inherent to both the static and magic angle spinning techniques of solid-state NMR (ssNMR) spectroscopy has thus far limited the routine application of multidimensional experiments to determine the structure of membrane proteins in lipid bilayers. Here, we demonstrate the advantage of using a recently developed class of experiments, polarization optimized experiments, for both static and MAS spectroscopy to achieve higher sensitivity and substantial time-savings for 2D and 3D experiments. We used sarcolipin, a single pass membrane protein, reconstituted in oriented bicelles (for oriented ssNMR) and multilamellar vesicles (for MAS ssNMR) as a benchmark. The restraints derived by these experiments are then combined into a hybrid energy function to allow simultaneous determination of structure and topology. The resulting structural ensemble converged to a helical conformation with a backbone RMSD ~0.44 Å, a tilt angle of 24° ± 1°, and an azimuthal angle of 55° ± 6°. This work represents a crucial first step toward obtaining high-resolution structures of large membrane proteins using combined multidimensional oriented solid-state NMR and magic angle spinning solid-state NMR.

  5. Determination of structural topology of a membrane protein in lipid bilayers using polarization optimized experiments (POE) for static and MAS solid state NMR spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Mote, Kaustubh R. [University of Minnesota, Department of Chemistry (United States); Gopinath, T. [University of Minnesota, Department of Biochemistry, Molecular Biology and Biophysics (United States); Veglia, Gianluigi, E-mail: [University of Minnesota, Department of Chemistry (United States)


    The low sensitivity inherent to both the static and magic angle spinning techniques of solid-state NMR (ssNMR) spectroscopy has thus far limited the routine application of multidimensional experiments to determine the structure of membrane proteins in lipid bilayers. Here, we demonstrate the advantage of using a recently developed class of experiments, polarization optimized experiments, for both static and MAS spectroscopy to achieve higher sensitivity and substantial time-savings for 2D and 3D experiments. We used sarcolipin, a single pass membrane protein, reconstituted in oriented bicelles (for oriented ssNMR) and multilamellar vesicles (for MAS ssNMR) as a benchmark. The restraints derived by these experiments are then combined into a hybrid energy function to allow simultaneous determination of structure and topology. The resulting structural ensemble converged to a helical conformation with a backbone RMSD {approx}0.44 A, a tilt angle of 24 Degree-Sign {+-} 1 Degree-Sign , and an azimuthal angle of 55 Degree-Sign {+-} 6 Degree-Sign . This work represents a crucial first step toward obtaining high-resolution structures of large membrane proteins using combined multidimensional oriented solid-state NMR and magic angle spinning solid-state NMR.

  6. Increased adhesion between neutral lipid bilayers: interbilayer bridges formed by tannic acid. (United States)

    Simon, S A; Disalvo, E A; Gawrisch, K; Borovyagin, V; Toone, E; Schiffman, S S; Needham, D; McIntosh, T J


    Tannic acid (TA) is a naturally occurring polyphenolic compound that aggregates membranes and neutral phosolipid vesicles and precipitates many proteins. This study analyzes TA binding to lipid membranes and the ensuing aggregation. The optical density of dispersions of phosphatidylcholine (PC) vesicles increased upon the addition of TA and electron micrographs showed that TA caused the vesicles to aggregate and form stacks of tightly packed disks. Solution calorimetry showed that TA bound to PC bilayers with a molar binding enthalpy of -8.3 kcal/mol and zeta potential measurements revealed that TA imparted a small negative charge to PC vesicles. Monolayer studies showed that TA bound to PC with a dissociation constant of 1.5 microM and reduced the dipole potential by up to 250 mV. Both the increase in optical density and decrease in dipole potential produced by TA could be reversed by the addition of polyvinylpyrrolidone, a compound that chelates TA by providing H-bond acceptor groups. NMR, micropipette aspiration, and x-ray diffraction experiments showed that TA incorporated into liquid crystalline PC membranes, increasing the area per lipid molecule and decreasing the bilayer thickness by 2 to 4%. 2H-NMR quadrupole splitting measurements also showed that TA associated with a PC molecule for times much less than 10(-4) s. In gel phase bilayers, TA caused the hydrocarbon chains from apposing monolayers to fully interdigitate. X-ray diffraction measurements of both gel and liquid crystalline dispersions showed that TA, at a critical concentration of about 1 mM, reduced the fluid spacing between adjacent bilayers by 8-10 A. These data place severe constraints on how TA can pack between adjacent bilayers and cause vesicles to adhere. We conclude that TA promotes vesicle aggregation by reducing the fluid spacing between bilayers by the formation of transient interbilayer bridges by inserting its digallic acid residues into the interfacial regions of adjacent bilayers

  7. Hydrodynamic Forces on Macromolecules Protruding from Lipid Bilayers Due to External Liquid Flows. (United States)

    Jönsson, Peter; Jönsson, Bengt


    It has previously been observed that an externally applied hydrodynamic shear flow above a fluid lipid bilayer can change the local concentration of macromolecules that are associated with the lipid bilayer. The external liquid flow results in a hydrodynamic force on molecules protruding from the lipid bilayer, causing them to move in the direction of the flow. However, there has been no quantitative study about the magnitude of these forces. We here use finite element simulations to investigate how the magnitude of the external hydrodynamic forces varies with the size and shape of the studied macromolecule. The simulations show that the hydrodynamic force is proportional to the effective hydrodynamic area of the studied molecule, Ahydro, multiplied by the mean hydrodynamic shear stress acting on the membrane surface, σhydro. The parameter Ahydro depends on the size and shape of the studied macromolecule above the lipid bilayer and scales with the cross-sectional area of the molecule. We also investigate how hydrodynamic shielding from other surrounding macromolecules decreases Ahydro when the surface coverage of the shielding macromolecules increases. Experiments where the protein streptavidin is anchored to a supported lipid bilayer on the floor of a microfluidic channel were finally performed at three different surface concentrations, Φ = 1%, 6%, and 10%, where the protein is being moved relative to the lipid bilayer by a liquid flow through the channel. From photobleaching measurements of fluorescently labeled streptavidin we found the experimental drift data to be within good accuracy of the simulated results, less than 12% difference, indicating the validity of the results obtained from the simulations. In addition to giving a deeper insight into how a liquid flow can affect membrane-associated molecules in a lipid bilayer, we also see an interesting potential of using hydrodynamic flow experiments together with the obtained results to study the size and

  8. Mechanical membrane for the separation of a paramagnetic constituent from a fluid

    Energy Technology Data Exchange (ETDEWEB)

    Maurice, David


    The disclosure provides an apparatus and method for the separation of a paramagnetic component from a mixture using a mechanical membrane apparatus. The mechanical membrane comprises a supporting material having a plurality of pores where each pore is surrounded by a plurality of magnetic regions. The magnetic regions augment a magnetic field on one side of the supporting material while mitigating the field to near zero on the opposite side. In operation, a flow of fluid such as air comprising a paramagnetic component such as O.sub.2 is directed toward the mechanical membrane, and the paramagnetic component is typically attracted toward a magnetic field surrounding a pore while dimagnetic components such as N.sub.2 are generally repelled. As some portion of the fluid passes through the plurality of magnetic apertures to the opposite side of the mechanical membrane, the mechanical membrane generates a fluid enriched in the paramagnetic component. Alternately, the magnetic field may act to repel the paramagnetic component while diamagnetic components such as N.sub.2 are generally unaffected and pass to the opposite side of the mechanical membrane.

  9. Dissipative particle dynamics simulation study of the bilayer-vesicle transition

    Institute of Scientific and Technical Information of China (English)


    A bilayer structure is an important immediate for the vesicle formation. However,the mechanism for the bilayer-vesicle transition remains unclear. In this work,a dissipative particle dynamics(DPD) simulation method was employed to study the mechanism of the bilayer-vesicle transition. A coarse-grained model was built based on a lipid molecule termed dimyristoylphosphatidylcholine(DMPC). Simulations were performed from two different initial configurations:a random dispersed solution and a tensionless bilayer. It was found that the bilayer-vesicle transition was driven by the minimization of the water-tail hydrophobic interaction energy,and was accompanied with the increase of the position entropy due to the redistribution of water molecules. The bulk pressure was reduced during the bilayer-vesicle transition,suggesting the evolved vesicle morphology was at the relatively low free energy state. The membrane in the product vesicle was a two-dimensional fluid. It can be concluded that the membrane of a vesicle is not interdigitated and most of the bonds in lipid chains are inclined to orient along the radical axis of the vesicle.

  10. Formation of individual protein channels in lipid bilayers suspended in nanopores. (United States)

    Studer, André; Han, Xiaojun; Winkler, Fritz K; Tiefenauer, Louis X


    Free-standing lipid bilayers are formed in regularly arranged nanopores of 200, 400 and 800 nm in a 300 nm thin hydrophobic silicon nitride membrane separating two fluid compartments. The extraordinary stability of the lipid bilayers allows us to monitor channel formation of the model peptide melittin and alpha-hemolysin from Staphylococcus aureus using electrochemical impedance spectroscopy and chronoamperometry. We observed that melittin channel formation is voltage-dependent and transient, whereas transmembrane heptameric alpha-hemolysin channels in nano-BLMs persist for hours. The onset of alpha-hemolysin-mediated conduction depends on the applied protein concentration and strongly on the diameter of the nanopores. Heptameric channel formation from adsorbed alpha-hemolysin monomers needs more time in bilayers suspended in 200 nm pores compared to bilayers in pores of 400 and 800 nm diameters. Diffusion of sodium ions across alpha-hemolysin channels present in a sufficiently high number in the bilayers was quantitatively and specifically determined using ion selective electrodes. The results demonstrate that relatively small variations of nano-dimensions have a tremendous effect on observable dynamic biomolecular processes. Such nanopore chips are potentially useful as supports for stable lipid bilayers to establish functional assays of membrane proteins needed in basic research and drug discovery.

  11. Dynamics of Lipid Bilayer Vesicles in Viscous Flows (United States)

    Schwalbe, Jonathan; Vlahovska, Petia; Miksis, Michael J.


    An analytical theory is developed to describe the dynamics of a closed lipid bilayer membrane (vesicle) in a general linear viscous flow. The dynamics of the membrane is governed by the Stokes equations in the fluid plus the normal and tangential stress condition along the bilayer interface. The effects of the membrane fluidity, incompressibility and resistance to bending are taken into account. The model is a generalization of the work on planar membranes by Seifert and Langer (Europhys. Lett. vol. 23, 71, 1993), which accounted for the variations in lipid density along both leaflets of the bilayer. Considering a nearly spherical vesicle, a perturbation solution is derived. The leading order analysis results in a nonlinear coupled system of equations for the dynamics of the shape and the mean lipid density difference between the inner and outer monolayer. Multiple solution states are found as a function of viscosity ratio and the monolayer slip coefficient. The dynamics and stability of these solutions is discussed. Comparisons are made to previous works based on the minimal curvature model which did not consider variable lipid density.



    Sengupta, Prabuddha; Baird, Barbara; Holowka, David


    Novel biophysical approaches combined with modeling and new biochemical data have helped to recharge the lipid raft field and have contributed to the generation of a refined model of plasma membrane organization. In this review, we summarize new information in the context of previous literature to provide new insights into the spatial organization and dynamics of lipids and proteins in the plasma membrane of live cells. Recent findings of large-scale separation of liquid-ordered and liquid-di...

  13. Biomineralization studies on cellulose membrane exposed to biological fluids of Anodonta cygnea. (United States)

    Lopes, Anabela; Lopes-Lima, Manuel; Ferreira, Jorge; Araújo, Sandra; Hinzmann, Mariana; Oliveira, José; Rocha, António; Domingues, Bernardo; Bobos, Iulius; Machado, Jorge


    The present work proposes to analyse the results obtained under in vitro conditions where cellulose artificial membranes were incubated with biological fluids from the freshwater bivalve Anodonta cygnea. The membranes were mounted between two half 'Ussing chambers' with different composition solutions in order to simulate epithelial surfaces separating organic fluid compartments. The membrane surfaces were submitted to two synthetic calcium and phosphate solutions on opposite sides, at pH 6.0, 7.0 or 9.0 during a period of 6 hours. Additional assays were accomplished mixing these solutions with haemolymph or extrapallial fluid from A. cygnea, only on the calcium side. A selective ion movement, mainly dependent on the membrane pore size and/or cationic affinity, occurred with higher permeability for calcium ions to the opposite phosphate chamber supported by calcium diffusion forces across the cellulose membrane. In general, this promoted a more intense mineral precipitation on the phosphate membrane surface. A strong deposition of calcium phosphate mineral was observed at pH 9.0 as a primary layer with a homogeneous microstructure, being totally absent at pH 6.0. The membrane showed an additional crystal phase at pH 7.0 exhibiting a very particular hexagonal or cuttlebone shape, mainly on the phosphate surface. When organic fluids of A. cygnea were included, these crystal forms presented a high tendency to aggregate under rosaceous shapes, also predominantly in the phosphate side. The cellulose membrane was permeable to small organic molecules that diffused from the calcium towards the phosphate side. In the calcium side, very few similar crystals were observed. The presence of organic matrix from A. cygnea fluids induced a preliminary apatite-brushite crystal polymorphism. So, the present results suggest that cellulose membranes can be used as surrogates of biological epithelia with preferential ionic diffusion from the calcium to the phosphate side where the main

  14. Interstitial fluid flow:simulation of mechanical environment of cells in the interosseous membrane

    Institute of Scientific and Technical Information of China (English)

    Wei Yao; Guang-Hong Ding


    In vitro experiments have shown that subtle fluid flow environment plays a significant role in living biological tissues,while there is no in vivo practical dynamical measurement of the interstitial fluid flow velocity. On the basis of a new finding that capillaries and collagen fibrils in the interosseous membrane form a parallel array,we set up a porous media model simulating the flow field with FLUENT software,studied the shear stress on interstitial cells' surface due to the interstitial fluid flow,and analyzed the effect of flow on protein space distribution around the cells. The numerical simulation results show that the parallel nature of capillaries could lead to directional interstitial fluid flow in the direction of capillaries. Interstitial fluid flow would induce shear stress on the membrane of interstitial cells,up to 30 Pa or so,which reaches or exceeds the threshold values of cells' biological response observed in vitro. Interstitial fluid flow would induce nonuniform spacial distribution of secretion protein of mast cells. Shear tress on cells could be affected by capillary parameters such as the distance between the adjacent capillaries,blood pressure and the permeability coefficient of capillary's wall. The interstitial pressure and the interstitial porosity could also affect the shear stress on cells. In conclusion,numerical simulation provides an effective way for in vivo dynamic interstitial velocity research,helps to set up the vivid subtle interstitial flow environment of cells,and is beneficial to understanding the physiological functions of interstitial fluid flow.

  15. Electrochemically deposited and etched membranes with precisely sized micropores for biological fluids microfiltration (United States)

    Hamzah, A. A.; Zainal Abidin, H. E.; Yeop Majlis, B.; Mohd Nor, M.; Ismardi, A.; Sugandi, G.; Tiong, T. Y.; Dee, C. F.; Yunas, J.


    This paper presents simple and economical, yet reliable techniques to fabricate a micro-fluidic filter for MEMS lab-on-chip (LoC) applications. The microporous filter is a crucial component in a MEMS LoC system. Microsized components and contaminants in biological fluids are selectively filtered using copper and silicon membranes with precisely controlled microsized pores. Two techniques were explored in microporous membrane fabrication, namely copper electroplating and electrochemical etching (ECE) of silicon. In the first technique, a copper membrane with evenly distributed micropores was fabricated by electroplating the copper layer on the silicon nitride membrane, which was later removed to leave the freestanding microporous membrane structure. The second approach involves the thinning of bulk silicon down to a few micrometers thick using KOH and etching the resulting silicon membrane in 5% HF by ECE to create micropores. Upon testing with nanoparticles of various sizes, it was observed that electroplated copper membrane passes nanoparticles up to 200 nm wide, while porous silicon membrane passes nanoparticles up to 380 nm in size. Due to process compatibility, simplicity, and low-cost fabrication, electroplated copper and porous silicon membranes enable synchronized microfilter fabrication and integration into the MEMS LoC system.

  16. How lidocaine influences the bilayer thickness and bending elasticity of biomembranes

    Energy Technology Data Exchange (ETDEWEB)

    Zheng Yi; Nagao, Michihiro; Bossev, Dobrin P, E-mail:


    We have studied how local anesthetics influence the structural and dynamical properties of model bio-membranes. The measurements of small-angle neutron scattering have been performed on 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) unilamellar vesicles with different concentrations of lidocaine in D{sub 2}O to determine the bilayer thickness as a function of the lidocaine concentration. The neutron-spin echo spectroscopy (NSE) has been used to study the influence of lidocaine on the bending elasticity of DMPC bilayers in fluid crystal phase (L{sub {alpha}}) and the ripple gel (P{sub {beta}}') phase.

  17. Process to remove turbidity-causing components from a fluid by micro-filtration - passes the fluid across an asymmetric membrane with inlet pores larger than those of nominal size, and cleans the membrane by backwashing

    DEFF Research Database (Denmark)


    Process is based on microfiltration using an asymmetric membrane. The membrane has inlet pores which are larger than the nominal pore size; pores of nominal size are located towards the membrane outlet side. The fluid is fed across the membrane. and backwashing is used for cleaning. USE-To remove...... turbidity-causing components from beer, wine, fruit juice, milk and blood, and from bacterial and enzyme suspensions. ADVANTAGE-The process greatly reduces the lost production time associated with earlier filtration methods, and beneficial components can pass through the membrane, thereby improving...... the quality of permeated fluids such as beer and wine. (Amended abstract week 9605)...

  18. Vascular basement membranes as pathways for the passage of fluid into and out of the brain. (United States)

    Morris, Alan W J; Sharp, Matthew MacGregor; Albargothy, Nazira J; Fernandes, Rute; Hawkes, Cheryl A; Verma, Ajay; Weller, Roy O; Carare, Roxana O


    In the absence of conventional lymphatics, drainage of interstitial fluid and solutes from the brain parenchyma to cervical lymph nodes is along basement membranes in the walls of cerebral capillaries and tunica media of arteries. Perivascular pathways are also involved in the entry of CSF into the brain by the convective influx/glymphatic system. The objective of this study is to differentiate the cerebral vascular basement membrane pathways by which fluid passes out of the brain from the pathway by which CSF enters the brain. Experiment 1: 0.5 µl of soluble biotinylated or fluorescent Aβ, or 1 µl 15 nm gold nanoparticles was injected into the mouse hippocampus and their distributions determined at 5 min by transmission electron microscopy. Aβ was distributed within the extracellular spaces of the hippocampus and within basement membranes of capillaries and tunica media of arteries. Nanoparticles did not enter capillary basement membranes from the extracellular spaces. Experiment 2: 2 µl of 15 nm nanoparticles were injected into mouse CSF. Within 5 min, groups of nanoparticles were present in the pial-glial basement membrane on the outer aspect of cortical arteries between the investing layer of pia mater and the glia limitans. The results of this study and previous research suggest that cerebral vascular basement membranes form the pathways by which fluid passes into and out of the brain but that different basement membrane layers are involved. The significance of these findings for neuroimmunology, Alzheimer's disease, drug delivery to the brain and the concept of the Virchow-Robin space are discussed.

  19. Aspirin Increases the Solubility of Cholesterol in Lipid Membranes (United States)

    Alsop, Richard; Barrett, Matthew; Zheng, Sonbo; Dies, Hannah; Rheinstadter, Maikel


    Aspirin (ASA) is often prescribed for patients with high levels of cholesterol for the secondary prevention of myocardial events, a regimen known as the Low-Dose Aspirin Therapy. We have recently shown that Aspirin partitions in lipid bilayers. However, a direct interplay between ASA and cholesterol has not been investigated. Cholesterol is known to insert itself into the membrane in a dispersed state at moderate concentrations (under ~37.5%) and decrease fluidity of membranes. We prepared model lipid membranes containing varying amounts of both ASA and cholesterol molecules. The structure of the bilayers as a function of ASA and cholesterol concentration was determined using high-resolution X-ray diffraction. At cholesterol levels of more than 40mol%, immiscible cholesterol plaques formed. Adding ASA to the membranes was found to dissolve the cholesterol plaques, leading to a fluid lipid bilayer structure. We present first direct evidence for an interaction between ASA and cholesterol on the level of the cell membrane.

  20. Development of modifications to the material point method for the simulation of thin membranes, compressible fluids, and their interactions

    Energy Technology Data Exchange (ETDEWEB)

    York, A.R. II [Sandia National Labs., Albuquerque, NM (United States). Engineering and Process Dept.


    The material point method (MPM) is an evolution of the particle in cell method where Lagrangian particles or material points are used to discretize the volume of a material. The particles carry properties such as mass, velocity, stress, and strain and move through a Eulerian or spatial mesh. The momentum equation is solved on the Eulerian mesh. Modifications to the material point method are developed that allow the simulation of thin membranes, compressible fluids, and their dynamic interactions. A single layer of material points through the thickness is used to represent a membrane. The constitutive equation for the membrane is applied in the local coordinate system of each material point. Validation problems are presented and numerical convergence is demonstrated. Fluid simulation is achieved by implementing a constitutive equation for a compressible, viscous, Newtonian fluid and by solution of the energy equation. The fluid formulation is validated by simulating a traveling shock wave in a compressible fluid. Interactions of the fluid and membrane are handled naturally with the method. The fluid and membrane communicate through the Eulerian grid on which forces are calculated due to the fluid and membrane stress states. Validation problems include simulating a projectile impacting an inflated airbag. In some impact simulations with the MPM, bodies may tend to stick together when separating. Several algorithms are proposed and tested that allow bodies to separate from each other after impact. In addition, several methods are investigated to determine the local coordinate system of a membrane material point without relying upon connectivity data.

  1. Effect of integral proteins in the phase stability of a lipid bilayer: Application to raft formation in cell membranes (United States)

    Gómez, Jordi; Sagués, Francesc; Reigada, Ramon


    The existence of lipid rafts is a controversial issue. The affinity of cholesterol for saturated lipids is manifested in macroscopic phase separation in model membranes, and is believed to be the thermodynamic driving force for raft formation. However, there is no clear reason to explain the small (nanometric) size of raft domains in cell membranes. In a recent paper Yethiraj and Weisshaar [Biophys. J. 93, 3113 (2007)] proposed that the effect of neutral integral membrane proteins may prevent from the formation of large lipid domains. In this paper we extend this approach by studying the effect of the protein size, as well as the lipid-protein interaction. Depending on these factors, two different mechanisms for nanodomain stabilization are shown to be possible for static proteins. The application of these results to a biological context is discussed.

  2. Lipid domain formation and ligand-receptor distribution in lipid bilayer membranes investigated by atomic force microscopy

    DEFF Research Database (Denmark)

    Kaasgaard, Thomas; Mouritsen, O.G.; Jørgensen, K.


    A novel experimental technique, based on atomic force microscopy (AFM), is proposed to visualize the lateral organization of membrane systems in the nanometer range. The technique involves the use of a ligand-receptor pair, biotin-avidin, which introduces a height variation on a solid-supported l......A novel experimental technique, based on atomic force microscopy (AFM), is proposed to visualize the lateral organization of membrane systems in the nanometer range. The technique involves the use of a ligand-receptor pair, biotin-avidin, which introduces a height variation on a solid...

  3. Amniotic fluid cathepsin-G in pregnancies complicated by the preterm prelabor rupture of membranes. (United States)

    Musilova, Ivana; Andrys, Ctirad; Drahosova, Marcela; Soucek, Ondrej; Pliskova, Lenka; Stepan, Martin; Bestvina, Tomas; Maly, Jan; Jacobsson, Bo; Kacerovsky, Marian


    The aim of this study was to evaluate the amniotic fluid cathepsin-G concentrations in women with preterm prelabor rupture of membranes (PPROM) based on the presence of the microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI). A total of 154 women with singleton pregnancies complicated by PPROM were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis. Amniotic fluid cathepsin-G concentrations were assessed by ELISA. MIAC was determined using a non-cultivation approach. IAI was defined as an amniotic fluid bedside interleukin-6 concentration ≥ 745 pg/mL. Women with MIAC had higher amniotic fluid cathepsin-G concentrations than women without MIAC (with MIAC: median 82.7 ng/mL, versus without MIAC: median 64.7 ng/mL; p = 0.0003). Women with IAI had higher amniotic fluid cathepsin-G concentrations than women without this complication (with IAI: median 103.0 ng/mL, versus without IAI: median 66.2 ng/mL; p G concentrations than women with colonization (MIAC without IAI) and women without both MIAC and IAI (p G concentrations in pregnancies complicated by PPROM. Amniotic fluid cathepsin-G appears to be a potential marker of IAI.

  4. Fragmented state of lipid bilayers in water

    DEFF Research Database (Denmark)

    Helfrich, W.; Thimmel, J.; Klösgen, Beate Maria


    The bilayers of some typical biological membrane lipids such as PC and DGDG disintegrate in a large excess of water to form an optically invisible dispersive bilayer phase. `Dark bodies' can be reversibly precipitated from it by raising the temperature. The dispersive phase probably consists...

  5. Pore formation in lipid membrane I: Continuous reversible trajectory from intact bilayer through hydrophobic defect to transversal pore. (United States)

    Akimov, Sergey A; Volynsky, Pavel E; Galimzyanov, Timur R; Kuzmin, Peter I; Pavlov, Konstantin V; Batishchev, Oleg V


    Lipid membranes serve as effective barriers allowing cells to maintain internal composition differing from that of extracellular medium. Membrane permeation, both natural and artificial, can take place via appearance of transversal pores. The rearrangements of lipids leading to pore formation in the intact membrane are not yet understood in details. We applied continuum elasticity theory to obtain continuous trajectory of pore formation and closure, and analyzed molecular dynamics trajectories of pre-formed pore reseal. We hypothesized that a transversal pore is preceded by a hydrophobic defect: intermediate structure spanning through the membrane, the side walls of which are partially aligned by lipid tails. This prediction was confirmed by our molecular dynamics simulations. Conversion of the hydrophobic defect into the hydrophilic pore required surmounting some energy barrier. A metastable state was found for the hydrophilic pore at the radius of a few nanometers. The dependence of the energy on radius was approximately quadratic for hydrophobic defect and small hydrophilic pore, while for large radii it depended on the radius linearly. The pore energy related to its perimeter, line tension, thus depends of the pore radius. Calculated values of the line tension for large pores were in quantitative agreement with available experimental data.

  6. High fidelity neuronal networks formed by plasma masking with a bilayer membrane: analysis of neurodegenerative and neuroprotective processes

    NARCIS (Netherlands)

    Hardelauf, Heike; Sisnaiske, Julia; Taghipour-Anvari, Amir Ali; Jacob, Peter; Drabiniok, Evelyn; Marggraf, Ulrich; Frimat, Jean-Philippe; Hengstler, Jan G.; Neyer, Andreas; Thriel, van Christoph; West, Jonathan


    Spatially defined neuronal networks have great potential to be used in a wide spectrum of neurobiology assays. We present an original technique for the precise and reproducible formation of neuronal networks. A PDMS membrane comprising through-holes aligned with interconnecting microchannels was use

  7. Pattern formation in biological fluids II: cell deformation in shear fields evidences convective membrane organisation

    CERN Document Server

    Lofthouse, J


    The mechanical behaviour and symmetry-breaking shape deformation of red blood cells subjected to shear flows is used to demonstrate that far from being random fluids, both the membrane and cytoplasm of every biological cell undergo spatially organised convective and shear driven flows when the cell maintains a Near Equilibrium state through continuousmetabolic activity. The model demonstrates that fluid bifurcation events drive cell shape changes, rather than a Meccano like cytoskeletal structure, and represents a significant Gestalt shift in models of cell mechanics.

  8. The Balance of Fluid and Osmotic Pressures across Active Biological Membranes with Application to the Corneal Endothelium.

    Directory of Open Access Journals (Sweden)

    Xi Cheng

    Full Text Available The movement of fluid and solutes across biological membranes facilitates the transport of nutrients for living organisms and maintains the fluid and osmotic pressures in biological systems. Understanding the pressure balances across membranes is crucial for studying fluid and electrolyte homeostasis in living systems, and is an area of active research. In this study, a set of enhanced Kedem-Katchalsky (KK equations is proposed to describe fluxes of water and solutes across biological membranes, and is applied to analyze the relationship between fluid and osmotic pressures, accounting for active transport mechanisms that propel substances against their concentration gradients and for fixed charges that alter ionic distributions in separated environments. The equilibrium analysis demonstrates that the proposed theory recovers the Donnan osmotic pressure and can predict the correct fluid pressure difference across membranes, a result which cannot be achieved by existing KK theories due to the neglect of fixed charges. The steady-state analysis on active membranes suggests a new pressure mechanism which balances the fluid pressure together with the osmotic pressure. The source of this pressure arises from active ionic fluxes and from interactions between solvent and solutes in membrane transport. We apply the proposed theory to study the transendothelial fluid pressure in the in vivo cornea, which is a crucial factor maintaining the hydration and transparency of the tissue. The results show the importance of the proposed pressure mechanism in mediating stromal fluid pressure and provide a new interpretation of the pressure modulation mechanism in the in vivo cornea.

  9. Dynamics of multicomponent lipid membranes (United States)

    Camley, Brian Andrew

    We present theoretical and computational descriptions of the dynamics of multicomponent lipid bilayer membranes. These systems are both model systems for "lipid rafts" in cell membranes and interesting physical examples of quasi-two-dimensional fluids. Our chief tool is a continuum simulation that uses a phase field to track the composition of the membrane, and solves the hydrodynamic equations exactly using the appropriate Green's function (Oseen tensor) for the membrane. We apply this simulation to describe the diffusion of domains in phase-separated membranes, the dynamics of domain flickering, and the process of phase separation in lipid membranes. We then derive an analytical theory to describe domain flickering that is consistent with our simulation results, and use this to analyze experimental measurements of membrane domains. Through this method, we measure the membrane viscosity solely from fluorescence microscopy measurements. We study phase separation in quasi-two-dimensional membranes in depth with both simulations and scaling theory, and classify the different scaling regimes and morphologies, which differ from pure two-dimensional fluids. Our results may explain previous inconsistent measurements of the dynamical scaling exponent for phase separation in membranes. We also extend our theory beyond the simplest model, including the possibility that the membrane will be viscoelastic, as well as considering the inertia of the membrane and the fluid surrounding the membrane.

  10. Geometry and Topology of Cell Membranes (United States)

    Bouligand, Y.

    Cells are limited by a membrane which is a fluid bilayer of phospholipids to which are associated numerous components, such as cholesterol, polysaccharides, proteins and, among them, many enzymes. organelles within cells are made for a large part of similar bilayers including phospholipids and various molecules. The cell membrane forms architectures closely related to those observed in liquid crystalline phases given by water-lipid systems (purified amphiphilic molecules in presence of water and oily components). The cell is divided into a series of compartments with definite topological relations, which are rehandled more or less profoundly in diverse circumstances as endocytosis, exocytosis, mitosis etc. There are several geometric arrangements of membrane sets : parallel membranes, hexagonal packing of tubes, cubic systems made of tubes joining either three by three, or four by four, or six by six. There are other arrangements less directly related to liquid crysyalline structures (annulate lamellae, tubes and lamellae with nematic symmetries, randomly joining tubes). Comparisons of structures in cellular membranes and in water-lipid systems reveal important differences. If geometries are often similar, water percentage and scales are distinct and bilayers observed in vitro present a symmetry which is broken in cell membrane bilayers. The curvature effects observed in water-lipid systems mainly come from a density difference between polar heads and corresponding paraffinic chains within a monolayer, whereas, in biological membranes, the asymmetry lies between the two monolayers and their associated molecules. Both systems produce saddle-shaped bilayers arranging into cubic lattices separating two aqueous compartments. In water-lipid systems, the coupling at an interface of two different areas seems to predominate, whereas in biological membranes, mechanisms are different and probably originate from geometric properties of proteins included within bilayers.

  11. Syrinx fluid transport: modeling pressure-wave-induced flux across the spinal pial membrane. (United States)

    Elliott, N S J


    Syrinxes are fluid-filled cavities of the spinal cord that characterize syringomyelia, a disease involving neurological damage. Their formation and expansion is poorly understood, which has hindered successful treatment. Syrinx cavities are hydraulically connected with the spinal subarachnoid space (SSS) enveloping the spinal cord via the cord interstitium and the network of perivascular spaces (PVSs), which surround blood vessels penetrating the pial membrane that is adherent to the cord surface. Since the spinal canal supports pressure wave propagation, it has been hypothesized that wave-induced fluid exchange across the pial membrane may play a role in syrinx filling. To investigate this conjecture a pair of one-dimensional (1-d) analytical models were developed from classical elastic tube theory coupled with Darcy's law for either perivascular or interstitial flow. The results show that transpial flux serves as a mechanism for damping pressure waves by alleviating hoop stress in the pial membrane. The timescale ratio over which viscous and inertial forces compete was explicitly determined, which predicts that dilated PVS, SSS flow obstructions, and a stiffer and thicker pial membrane-all associated with syringomyelia-will increase transpial flux and retard wave travel. It was also revealed that the propagation of a pressure wave is aided by a less-permeable pial membrane and, in contrast, by a more-permeable spinal cord. This is the first modeling of the spinal canal to include both pressure-wave propagation along the spinal axis and a pathway for fluid to enter and leave the cord, which provides an analytical foundation from which to approach the full poroelastic problem.

  12. Validation of an immersed thick boundary method for simulating fluid-structure interactions of deformable membranes (United States)

    Sigüenza, J.; Mendez, S.; Ambard, D.; Dubois, F.; Jourdan, F.; Mozul, R.; Nicoud, F.


    This paper constitutes an extension of the work of Mendez et al. (2014) [36], for three-dimensional simulations of deformable membranes under flow. An immersed thick boundary method is used, combining the immersed boundary method with a three-dimensional modeling of the structural part. The immersed boundary method is adapted to unstructured grids for the fluid resolution, using the reproducing kernel particle method. An unstructured finite-volume flow solver for the incompressible Navier-Stokes equations is coupled with a finite-element solver for the structure. The validation process relying on a number of test cases proves the efficiency of the method, and its robustness is illustrated when computing the dynamics of a tri-leaflet aortic valve. The proposed immersed thick boundary method is able to tackle applications involving both thin and thick membranes/closed and open membranes, in significantly high Reynolds number flows and highly complex geometries.

  13. Unsteady fluid-structure interactions with a heaving compliant membrane wing (United States)

    Alon Tzezana, Gali; Breuer, Kenneth


    Membrane wings have been shown to provide some benefits over rigid wings at the low Reynolds number regime (Re 103 to 105), specifically improved thrust in flapping flight. Here we present results from a theoretical framework used to characterize the unsteady aeroelastic behavior of compliant membrane wings executing a heaving motion. An analytical model is developed using 2D unsteady thin airfoil theory, coupled with an unsteady membrane equation. Chebyshev collocation methods are used to solve the coupled system efficiently. The model is used to explore the effects of wing compliance, inertia (including added mass effect) and flapping kinematics on the aerodynamic performance, identifying optimal conditions for maximum thrust and propulsive efficiency. A resonant frequency of the coupled system is identified and characterized for different fluid-structure interaction regimes. Extensions to pitching kinematics are also discussed.

  14. Membranes having aligned 1-D nanoparticles in a matrix layer for improved fluid separation

    Energy Technology Data Exchange (ETDEWEB)

    Revanur, Ravindra; Lulevich, Valentin; Roh, Il Juhn; Klare, Jennifer E.; Kim, Sangil; Noy, Aleksandr; Bakajin, Olgica


    Membranes for fluid separation are disclosed. These membranes have a matrix layer sandwiched between an active layer and a porous support layer. The matrix layer includes 1-D nanoparticles that are vertically aligned in a porous polymer matrix, and which substantially extend through the matrix layer. The active layer provides species-specific transport, while the support layer provides mechanical support. A matrix layer of this type has favorable surface morphology for forming the active layer. Furthermore, the pores that form in the matrix layer tend to be smaller and more evenly distributed as a result of the presence of aligned 1-D nanoparticles. Improved performance of separation membranes of this type is attributed to these effects.

  15. The influence of non polar and polar molecules in mouse motile cells membranes and pure lipid bilayers.

    Directory of Open Access Journals (Sweden)

    Francisco J Sierra-Valdez

    Full Text Available We report an experimental study of mouse sperm motility that shows chief aspects characteristic of neurons: the anesthetic (produced by tetracaine and excitatory (produced by either caffeine or calcium effects and their antagonic action. While tetracaine inhibits sperm motility and caffeine has an excitatory action, the combination of these two substances balance the effects, producing a motility quite similar to that of control cells. We also study the effects of these agents (anesthetic and excitatory on the melting points of pure lipid liposomes constituted by 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC and dipalmitoyl phosphatidic acid (DPPA. Tetracaine induces a large fluidization of the membrane, shifting the liposomes melting transition temperature to much lower values. The effect of caffeine is null, but its addition to tetracaine-doped liposomes greatly screen the fluidization effect. A high calcium concentration stiffens pure lipid membranes and strongly reduces the effect of tetracaine. Molecular Dynamics Simulations are performed to further understand our experimental findings at the molecular level. We find a strong correlation between the effect of antagonic molecules that could explain how the mechanical properties suitable for normal cell functioning are affected and recovered.

  16. Constant-pH MD Simulations of DMPA/DMPC Lipid Bilayers. (United States)

    Santos, Hugo A F; Vila-Viçosa, Diogo; Teixeira, Vitor H; Baptista, António M; Machuqueiro, Miguel


    Current constant-pH molecular dynamics (CpHMD) simulations provide a proper treatment of pH effects on the structure and dynamics of soluble biomolecules like peptides and proteins. However, addressing such effects on lipid membrane assemblies has remained problematic until now, despite the important role played by lipid ionization at physiological pH in a plethora of biological processes. Modeling (de)protonation events in these systems requires a proper consideration of the physicochemical features of the membrane environment, including a sound treatment of solution ions. Here, we apply our recent CpHMD-L method to the study of pH effects on a 25% DMPA/DMPC bilayer membrane model, closely reproducing the correct lipid phases of this system, namely, gel-fluid coexistence at pH 4 and a fluid phase at pH 7. A significant transition is observed for the membrane ionization and mechanical properties at physiological pH, providing a molecular basis for the well-established role of phosphatidic acid (PA) as a key player in the regulation of many cellular events. Also, as reported experimentally, we observed pH-induced PA-PA lipid aggregation at acidic pH. By including the titration of anionic phospholipids, the current methodology makes possible to simulate lipid bilayers with increased realism. To the best of our knowledge, this is the first simulation study dealing with a continuous phospholipid bilayer with pH titration of all constituent lipids.

  17. High fidelity neuronal networks formed by plasma masking with a bilayer membrane: analysis of neurodegenerative and neuroprotective processes. (United States)

    Hardelauf, Heike; Sisnaiske, Julia; Taghipour-Anvari, Amir Ali; Jacob, Peter; Drabiniok, Evelyn; Marggraf, Ulrich; Frimat, Jean-Philippe; Hengstler, Jan G; Neyer, Andreas; van Thriel, Christoph; West, Jonathan


    Spatially defined neuronal networks have great potential to be used in a wide spectrum of neurobiology assays. We present an original technique for the precise and reproducible formation of neuronal networks. A PDMS membrane comprising through-holes aligned with interconnecting microchannels was used during oxygen plasma etching to dry mask a protein rejecting poly(ethylene glycol) (PEG) adlayer. Patterns were faithfully replicated to produce an oxidized interconnected array pattern which supported protein adsorption. Differentiated human SH-SY5Y neuron-like cells adhered to the array nodes with the micron-scale interconnecting tracks guiding neurite outgrowth to produce neuronal connections and establish a network. A 2.0 μm track width was optimal for high-level network formation and node compliance. These spatially standardized neuronal networks were used to analyse the dynamics of acrylamide-induced neurite degeneration and the protective effects of co-treatment with calpeptin or brain derived neurotrophic factor (BDNF).

  18. Amniotic fluid volume and composition after fetal membrane resection in late-gestation sheep. (United States)

    Brace, Robert A; Cheung, Cecilia Y


    The chronically catheterized fetal sheep is a widely used model for fetal physiologic and pathophysiologic investigations. Catheterization involves opening the amniochorion to gain access to the fetus. In the current study, we explored the role of the amnion and amniochorion in maintaining normal amniotic fluid volume (AFV) and composition and fetal blood-gas status after surgery. Fetal sheep were catheterized at 119.6 ± 0.3 (mean ± SE, n = 25) d gestation (term, approximately 147 d). An opening equal to approximately 5% of total membrane surface area was created by resecting a portion of the amnion or amniochorion during surgery. The uterine wall was closed in all animals. Compared with control sheep (AFV = 992 ± 153 mL, n = 11), resection of the amnion had no significant effect on AFV (745 ± 156 mL, n = 7) measured 5 d after surgery, whereas resection of the amniochorion resulted in extensive loss of amniotic fluid (AFV = 131 ± 38 mL, n = 7). This loss resulted from extensive entry of amniotic fluid into the space between the chorion and uterine wall. Amniotic fluid, fetal plasma, and urinary solute concentrations; arterial pH; oxygen tension; and carbon dioxide tension were unchanged. A small opening in the amnion has minimal effects on ovine AFV, whereas a small opening in the amniochorion results in oligohydramnios. In addition, the amnion appears to be the primary site that limits the rate of amniotic fluid absorption by the chorionic vasculature.

  19. Bipolar tetraether lipids: chain flexibility and membrane polarity gradients from spin-label electron spin resonance. (United States)

    Bartucci, R; Gambacorta, A; Gliozzi, A; Marsh, D; Sportelli, L


    Membranes of thermophilic Archaea are composed of unique tetraether lipids in which C40, saturated, methyl-branched biphytanyl chains are linked at both ends to polar groups. In this paper, membranes composed of bipolar lipids P2 extracted from the acidothermophile archaeon Sulfolobus solfataricus are studied. The biophysical basis for the membrane formation and thermal stability is investigated by using electron spin resonance (ESR) of spin-labeled lipids. Spectral anisotropy and isotropic hyperfine couplings are used to determine the chain flexibility and polarity gradients, respectively. For comparison, similar measurements have been carried out on aqueous dispersions of diacyl reference lipid dipalmitoyl phosphatidylcholine and also of diphytanoyl phosphatidylcholine, which has methyl-branched chains. At a given temperature, the bolaform lipid chains are more ordered and less flexible than in normal bilayer membranes. Only at elevated temperatures (80 degrees C) does the flexibility of the chain environment in tetraether lipid assemblies approach that of fluid bilayer membranes. The height of the hydrophobic barrier formed by a monolayer of archaebacterial lipids is similar to that in conventional fluid bilayer membranes, and the permeability barrier width is comparable to that formed by a bilayer of C16 lipid chains. At a mole ratio of 1:2, the tetraether P2 lipids mix well with dipalmitoyl phosphatidylcholine lipids and stabilize conventional bilayer membranes. The biological as well as the biotechnological relevance of the results is discussed.

  20. Cholesterol rules: direct observation of the coexistence of two fluid phases in native pulmonary surfactant membranes at physiological temperatures

    DEFF Research Database (Denmark)

    Bernardino de la Serna, Jorge; Perez-Gil, Jesus; Simonsen, Adam C


    Pulmonary surfactant, the lipid-protein material that stabilizes the respiratory surface of the lungs, contains approximately equimolar amounts of saturated and unsaturated phospholipid species and significant proportions of cholesterol. Such lipid composition suggests that the membranes taking...... part in the surfactant structures could be organized heterogeneously in the form of inplane domains, originating from particular distributions of specific proteins and lipids. Here we report novel results concerning the lateral organization of bilayer membranes made of native pulmonary surfactant where...... of this material is naturally designed to be at the "edge" of a lateral structure transition under physiological conditions, likely providing particular structural and dynamic properties for its mechanical function. The observed lateral structure in native pulmonary surfactant membranes is dramatically affected...

  1. Rationalization of reduced penetration of drugs through ceramide gel phase membrane. (United States)

    Paloncýová, Markéta; DeVane, Russell H; Murch, Bruce P; Berka, Karel; Otyepka, Michal


    Since computing resources have advanced enough to allow routine molecular simulation studies of drug molecules interacting with biologically relevant membranes, a considerable amount of work has been carried out with fluid phospholipid systems. However, there is very little work in the literature on drug interactions with gel phase lipids. This poses a significant limitation for understanding permeation through the stratum corneum where the primary pathway is expected to be through a highly ordered lipid matrix. To address this point, we analyzed the interactions of p-aminobenzoic acid (PABA) and its ethyl (benzocaine) and butyl (butamben) esters with two membrane bilayers, which differ in their fluidity at ambient conditions. We considered a dioleoylphosphatidylcholine (DOPC) bilayer in a fluid state and a ceramide 2 (CER2, ceramide NS) bilayer in a gel phase. We carried out unbiased (100 ns long) and biased z-constraint molecular dynamics simulations and calculated the free energy profiles of all molecules along the bilayer normal. The free energy profiles converged significantly slower for the gel phase. While the compounds have comparable affinities for both membranes, they exhibit penetration barriers almost 3 times higher in the gel phase CER2 bilayer. This elevated barrier and slower diffusion in the CER2 bilayer, which are caused by the high ordering of CER2 lipid chains, explain the low permeability of the gel phase membranes. We also compared the free energy profiles from MD simulations with those obtained from COSMOmic. This method provided the same trends in behavior for the guest molecules in both bilayers; however, the penetration barriers calculated by COSMOmic did not differ between membranes. In conclusion, we show how membrane fluid properties affect the interaction of drug-like molecules with membranes.

  2. Co-existence of Gel and Fluid Lipid Domains in Single-component Phospholipid Membranes

    Energy Technology Data Exchange (ETDEWEB)

    Armstrong, Clare L [McMaster University; Barrett, M [McMaster University; Toppozini, L [McMaster University; Yamani, Zahra [Canadian Neutron Beam Centre, National Research Council, Chalk River Laboratorie; Kucerka, Norbert [Canadian Neutron Beam Centre and Comelius University (Slovakia); Katsaras, John [ORNL; Fragneto, Giovanna [Institut Laue-Langevin (ILL); Rheinstadter, Maikel C [McMaster University


    Lateral nanostructures in membranes, so-called rafts, are believed to strongly influence membrane properties and functions. The experimental observation of rafts has proven difficult as they are thought to be dynamic structures that likely fluctuate on nano- to microsecond time scales. Using neutron diffraction we present direct experimental evidence for the co-existence of gel and fluid lipid domains in a single-component phospholipid membrane made of DPPC as it undergoes its main phase transition. The coherence length of the neutron beam sets a lower limit for the size of structures that can be observed. Neutron coherence lengths between 30 and 242A used in this study were obtained by varying the incident neutron energy and the resolution of the neutron spectrometer. We observe Bragg peaks corresponding to co-existing nanometer sized structures, both in out-of-plane and in-plane scans, by tuning the neutron coherence length. During the main phase transition, instead of a continuous transition that shows a pseudo-critical behavior, we observe the co-existence of gel and fluid domains.

  3. Evaluation of agonist selectivity for the NMDA receptor ion channel in bilayer lipid membranes based on integrated single-channel currents. (United States)

    Hirano, A; Sugawara, M; Umezawa, Y; Uchino, S; Nakajima-Iijima, S


    A new method for evaluating chemical selectivity of agonists to activate the N-methyl-D-aspartate (NMDA) receptor was presented by using typical agonists NMDA, L-glutamate and (2S, 3R, 4S)-2-(carboxycyclopropyl)glycine (L-CCG-IV) and the mouse epsilon1/zeta1 NMDA receptor incorporated in bilayer lipid membranes (BLMs) as an illustrative example. The method was based on the magnitude of an agonist-induced integrated single-channel current corresponding to the number of total ions passed through the open channel. The very magnitudes of the integrated single-channel currents were compared with the different BLMs as a new measure of agonist selectivity. The epsilon1/zeta1 NMDA receptor was partially purified from Chinese hamster ovary (CHO) cells expressing the epsilon1/zeta1 NMDA receptor and incorporated in BLMs formed by the tip-dip method. The agonist-induced integrated single-channel currents were obtained at 50 microM agonist concentration, where the integrated current for NMDA was shown to reach its saturated value. The obtained integrated currents were found to be (4.5 +/- 0.55) x 10(-13) C/s for NMDA, (5.8 +/- 0.72) x 10(-13) C/s for L-glutamate and (6.6 +/- 0.61) x 10(-13) C/s for L-CCG-IV, respectively. These results suggest that the agonist selectivity in terms of the total ion flux through the single epsilon1/zeta1 NMDA receptor is in the order of L-CCG-IV approximately = L-glutamate > NMDA.

  4. Pair interaction of bilayer-coated nanoscopic particles

    Institute of Scientific and Technical Information of China (English)

    Zhang Qi-Yi


    The pair interaction between bilayer membrane-coated nanosized particles has been explored by using the self-consistent field (SCF) theory. The bilayer membranes are composed of amphiphilic polymers. For different system parameters, the pair-interaction free energies are obtained. Particular emphasis is placcd on the analysis of a sequence of structural transformations of bilayers on spherical particles, which occur during their approaching processes. For different head fractions of amphiphilcs, the asymmetrical morphologies between bilayers on two particles and the inverted micellar intermediates have been found in the membrane fusion pathway. These results can benefit the fabrication of vesicles as encapsulation vectors for drug and gene delivery.

  5. Dynamics, Surface Electrostatics and Phase Properties of Nanoscale Curved Lipid Bilayers (United States)

    Koolivand, Amir

    Surface electrostatic potential of a lipid bilayer governs many vital functions of living cells. Several classes of proteins are known of exhibiting strong binding preferences to curved lipid bilayer surfaces. In this project we employed electron paramagnetic resonance (EPR) of a recently introduced phospholipid (IMTSL-PTE) bearing a pH-sensitive nitroxide covalently attached to the lipid head group to measure the surface electrostatics of the lipid membrane and nanopore-confined lipid bilayers as a function of the bilayer curvature. The pKa of the ionizable group of this lipid-based spin probe is reporting on the bilayer surface electrostatics potential by changes in the EPR spectra. Specifically, both rotational dynamics and magnetic parameters of the nitroxide are affected by the probe protonation. Effect of curvature on the surface electrostatic potential and dynamics of lipid bilayer was studied for POPG and DMPG unilamellar vesicles (ULVs). It was found that the magnitude of the negative surface electrostatic potential increased upon decrease in the vesicle diameter for the bilayers in the fluid phase; however, no significant changes were observed for DMPG ULVs in a gel phase. We speculate that biologically relevant fluid bilayer phase allows for a larger variability in the lipid packing density in the lipid polar head group region than a more ordered gel phase and it is likely that the lipid flip-flop is responsible for pH equilibration of IMTSL-PTE. The kinetic EPR study of nitroxide reduction showed that the rate of flip-flop is in the order of 10-5 s-1. The flip-flop rate constant increases when vesicle size deceases. Oxygen permeability measured by X-ban EPR decreases in higher curved vesicles---an observation that is consistent with a tighter packing in smaller vesicles. Partitioning of a small nitroxide molecule TEMPO into ULVs was measured by X-band (9 GHz) and W-band (95 GHz) EPR spectroscopy. The partitioning coefficient of this probe in the lipid

  6. The Fluid-Mosaic Model of Membrane Structure: still relevant to understanding the structure, function and dynamics of biological membranes after more than 40 years. (United States)

    Nicolson, Garth L


    In 1972 the Fluid-Mosaic Membrane Model of membrane structure was proposed based on thermodynamic principals of organization of membrane lipids and proteins and available evidence of asymmetry and lateral mobility within the membrane matrix [S. J. Singer and G. L. Nicolson, Science 175 (1972) 720-731]. After over 40years, this basic model of the cell membrane remains relevant for describing the basic nano-structures of a variety of intracellular and cellular membranes of plant and animal cells and lower forms of life. In the intervening years, however, new information has documented the importance and roles of specialized membrane domains, such as lipid rafts and protein/glycoprotein complexes, in describing the macrostructure, dynamics and functions of cellular membranes as well as the roles of membrane-associated cytoskeletal fences and extracellular matrix structures in limiting the lateral diffusion and range of motion of membrane components. These newer data build on the foundation of the original model and add new layers of complexity and hierarchy, but the concepts described in the original model are still applicable today. In updated versions of the model more emphasis has been placed on the mosaic nature of the macrostructure of cellular membranes where many protein and lipid components are limited in their rotational and lateral motilities in the membrane plane, especially in their natural states where lipid-lipid, protein-protein and lipid-protein interactions as well as cell-matrix, cell-cell and intracellular membrane-associated protein and cytoskeletal interactions are important in restraining the lateral motility and range of motion of particular membrane components. The formation of specialized membrane domains and the presence of tightly packed integral membrane protein complexes due to membrane-associated fences, fenceposts and other structures are considered very important in describing membrane dynamics and architecture. These structures along

  7. Spatiotemporal Organization of Spin-Coated Supported Model Membranes (United States)

    Simonsen, Adam Cohen

    All cells of living organisms are separated from their surroundings and organized internally by means of flexible lipid membranes. In fact, there is consensus that the minimal requirements for self-replicating life processes include the following three features: (1) information carriers (DNA, RNA), (2) a metabolic system, and (3) encapsulation in a container structure [1]. Therefore, encapsulation can be regarded as an essential part of life itself. In nature, membranes are highly diverse interfacial structures that compartmentalize cells [2]. While prokaryotic cells only have an outer plasma membrane and a less-well-developed internal membrane structure, eukaryotic cells have a number of internal membranes associated with the organelles and the nucleus. Many of these membrane structures, including the plasma membrane, are complex layered systems, but with the basic structure of a lipid bilayer. Biomembranes contain hundreds of different lipid species in addition to embedded or peripherally associated membrane proteins and connections to scaffolds such as the cytoskeleton. In vitro, lipid bilayers are spontaneously self-organized structures formed by a large group of amphiphilic lipid molecules in aqueous suspensions. Bilayer formation is driven by the entropic properties of the hydrogen bond network in water in combination with the amphiphilic nature of the lipids. The molecular shapes of the lipid constituents play a crucial role in bilayer formation, and only lipids with approximately cylindrical shapes are able to form extended bilayers. The bilayer structure of biomembranes was discovered by Gorter and Grendel in 1925 [3] using monolayer studies of lipid extracts from red blood cells. Later, a number of conceptual models were developed to rationalize the organization of lipids and proteins in biological membranes. One of the most celebrated is the fluid-mosaic model by Singer and Nicolson (1972) [4]. According to this model, the lipid bilayer component of

  8. Hamilton's equations for a fluid membrane: axial symmetry

    Energy Technology Data Exchange (ETDEWEB)

    Capovilla, R [Departamento de Fisica, Centro de Investigacion y de Estudios Avanzados del IPN, Apdo Postal 14-740, 07000 Mexico, DF (Mexico); Guven, J [Instituto de Ciencias Nucleares, Universidad Nacional Autonoma de Mexico, Apdo Postal 70-543, 04510 Mexico, DF (Mexico); Rojas, E [Facultad de Fisica e Inteligencia Artificial, Universidad Veracruzana, 91000 Xalapa, Veracruz (Mexico)


    Consider a homogeneous fluid membrane, or vesicle, described by the Helfrich-Canham energy, quadratic in the mean curvature. When the membrane is axially symmetric, this energy can be viewed as an 'action' describing the motion of a particle; the contours of equilibrium geometries are identified with particle trajectories. A novel Hamiltonian formulation of the problem is presented which exhibits the following two features: (i) the second derivatives appearing in the action through the mean curvature are accommodated in a natural phase space and (ii) the intrinsic freedom associated with the choice of evolution parameter along the contour is preserved. As a result, the phase space involves momenta conjugate not only to the particle position but also to its velocity, and there are constraints on the phase space variables. This formulation provides the groundwork for a field theoretical generalization to arbitrary configurations, with the particle replaced by a loop in space.

  9. Validation of computational non-Newtonian fluid model for membrane bioreactor

    DEFF Research Database (Denmark)

    Sørensen, Lasse; Bentzen, Thomas Ruby; Skov, Kristian


    for optimizing MBR-systems is computational fluid dynamics (CFD) modelling, giving the ability to describe the flow in the systems. A parameter which is often neglected in such models is the non-Newtonian properties of active sludge, which is of great importance for MBR systems since they operate at sludge...... concentrations up to a factor 10 compared to conventional activated sludge (CAS) systems, resulting in strongly shear thinning liquids. A CFD-model is validated against measurements conducted in a system with rotating cross flow membranes submerged in non-Newtonian liquids, where tangential velocities...

  10. Profile structures of the voltage-sensor domain and the voltage-gated K+-channel vectorially oriented in a single phospholipid bilayer membrane at the solid-vapor and solid-liquid interfaces determined by x-ray interferometry (United States)

    Gupta, S.; Liu, J.; Strzalka, J.; Blasie, J. K.


    One subunit of the prokaryotic voltage-gated potassium ion channel from Aeropyrum pernix (KvAP) is comprised of six transmembrane α helices, of which S1-S4 form the voltage-sensor domain (VSD) and S5 and S6 contribute to the pore domain (PD) of the functional homotetramer. However, the mechanism of electromechanical coupling interconverting the closed-to-open (i.e., nonconducting-to-K+-conducting) states remains undetermined. Here, we have vectorially oriented the detergent (OG)-solubilized VSD in single monolayers by two independent approaches, namely “directed-assembly” and “self-assembly,” to achieve a high in-plane density. Both utilize Ni coordination chemistry to tether the protein to an alkylated inorganic surface via its C-terminal His6 tag. Subsequently, the detergent is replaced by phospholipid (POPC) via exchange, intended to reconstitute a phospholipid bilayer environment for the protein. X-ray interferometry, in which interference with a multilayer reference structure is used to both enhance and phase the specular x-ray reflectivity from the tethered single membrane, was used to determine directly the electron density profile structures of the VSD protein solvated by detergent versus phospholipid, and with either a moist He (moderate hydration) or bulk aqueous buffer (high hydration) environment to preserve a native structure conformation. Difference electron density profiles, with respect to the multilayer substrate itself, for the VSD-OG monolayer and VSD-POPC membranes at both the solid-vapor and solid-liquid interfaces, reveal the profile structures of the VSD protein dominating these profiles and further indicate a successful reconstitution of a lipid bilayer environment. The self-assembly approach was similarly extended to the intact full-length KvAP channel for comparison. The spatial extent and asymmetry in the profile structures of both proteins confirm their unidirectional vectorial orientation within the reconstituted membrane and

  11. Possible mechanism of adhesion in a mica supported phospholipid bilayer

    Energy Technology Data Exchange (ETDEWEB)

    Pertsin, Alexander, E-mail: [Angewandte Physikalische Chemie, Universität Heidelberg, Im Neuenheimer Feld 253, 69120 Heidelberg (Germany); Institute of Organo-Element Compounds, Russian Academy of Sciences, Vavilov Str. 28, 117991 Moscow (Russian Federation); Grunze, Michael [Angewandte Physikalische Chemie, Universität Heidelberg, Im Neuenheimer Feld 253, 69120 Heidelberg (Germany); Institute for Functional Interfaces, Karlsruhe Institute of Technology, Hermann-von- Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany)


    Phospholipid bilayers supported on hydrophilic solids like silica and mica play a substantial role in fundamental studies and technological applications of phospholipid membranes. In both cases the molecular mechanism of adhesion between the bilayer and the support is of primary interest. Since the possibilities of experimental methods in this specific area are rather limited, the methods of computer simulation acquire great importance. In this paper we use the grand canonical Monte Carlo technique and an atomistic force field to simulate the behavior of a mica supported phospholipid bilayer in pure water as a function of the distance between the bilayer and the support. The simulation reveals a possible adhesion mechanism, where the adhesion is due to individual lipid molecules that protrude from the bilayer and form widely spaced links with the support. Simultaneously, the bilayer remains separated from the bilayer by a thin water interlayer which maintains the bilayer fluidity.

  12. Bilayer Effects of Antimalarial Compounds.

    Directory of Open Access Journals (Sweden)

    Nicole B Ramsey

    Full Text Available Because of the perpetual development of resistance to current therapies for malaria, the Medicines for Malaria Venture developed the Malaria Box to facilitate the drug development process. We tested the 80 most potent compounds from the box for bilayer-mediated effects on membrane protein conformational changes (a measure of likely toxicity in a gramicidin-based stopped flow fluorescence assay. Among the Malaria Box compounds tested, four compounds altered membrane properties (p< 0.05; MMV007384 stood out as a potent bilayer-perturbing compound that is toxic in many cell-based assays, suggesting that testing for membrane perturbation could help identify toxic compounds. In any case, MMV007384 should be approached with caution, if at all.

  13. Surface electrostatics of lipid bilayers by EPR of a pH-sensitive spin-labeled lipid. (United States)

    Voinov, Maxim A; Rivera-Rivera, Izarys; Smirnov, Alex I


    Many biophysical processes such as insertion of proteins into membranes and membrane fusion are governed by bilayer electrostatic potential. At the time of this writing, the arsenal of biophysical methods for such measurements is limited to a few techniques. Here we describe a, to our knowledge, new spin-probe electron paramagnetic resonance (EPR) approach for assessing the electrostatic surface potential of lipid bilayers that is based on a recently synthesized EPR probe (IMTSL-PTE) containing a reversibly ionizable nitroxide tag attached to the lipids' polar headgroup. EPR spectra of the probe directly report on its ionization state and, therefore, on electrostatic potential through changes in nitroxide magnetic parameters and the degree of rotational averaging. Further, the lipid nature of the probe provides its full integration into lipid bilayers. Tethering the nitroxide moiety directly to the lipid polar headgroup defines the location of the measured potential with respect to the lipid bilayer interface. Electrostatic surface potentials measured by EPR of IMTSL-PTE show a remarkable (within ±2%) agreement with the Gouy-Chapman theory for anionic DMPG bilayers in fluid (48°C) phase at low electrolyte concentration (50 mM) and in gel (17°C) phase at 150-mM electrolyte concentration. This agreement begins to diminish for DMPG vesicles in gel phase (17°C) upon varying electrolyte concentration and fluid phase bilayers formed from DMPG/DMPC and POPG/POPC mixtures. Possible reasons for such deviations, as well as the proper choice of an electrostatically neutral reference interface, have been discussed. Described EPR method is expected to be fully applicable to more-complex models of cellular membranes.

  14. Amphiphile regulation of ion channel function by changes in the bilayer spring constant

    DEFF Research Database (Denmark)

    Lundbæk, Jens August; Koeppe, R.E.; Andersen, Oluf Sten


    be predicted from measurements of isolated changes in such properties. Thus, the bilayer contribution to the promiscuous regulation of membrane proteins by drugs and other amphiphiles remains unknown. To overcome this problem, we use gramicidin A (gA) channels as molecular force probes to measure the net......Many drugs are amphiphiles that, in addition to binding to a particular target protein, adsorb to cell membrane lipid bilayers and alter intrinsic bilayer physical properties (e. g., bilayer thickness, monolayer curvature, and elastic moduli). Such changes can modulate membrane protein function...... by altering the energetic cost (Delta G(bilayer)) of bilayer deformations associated with protein conformational changes that involve the protein-bilayer interface. But amphiphiles have complex effects on the physical properties of lipid bilayers, meaning that the net change in Delta G(bilayer) cannot...

  15. Near field fluid coupling between internal motion of the organ of Corti and the basilar membrane

    Energy Technology Data Exchange (ETDEWEB)

    Elliott, Stephen J.; Ni, Guangjian [Institute of Sound and Vibration Research, University of Southampton, Southampton (United Kingdom)


    The pressure distribution in each of the fluid chambers of the cochlea can be decomposed into a 1D, or plane wave, component and a near field component, which decays rapidly away from the excitation point. The transverse motion of the basilar membrane, BM, for example, generates both a 1D pressure field, which couples into the slow wave, and a local near field pressure, proportional to the BM acceleration, that generates an added mass on the BM due to the fluid motion. When the organ of Corti, OC, undergoes internal motion, due for example to outer hair cell activity, this motion will not itself generate any 1D pressure if the OC is incompressible and the BM is constrained not to move volumetrically, and so will not directly couple into the slow wave. This motion will, however, generate a near field pressure, proportional to the OC acceleration, which will act on the OC and thus increases its effective mass. The near field pressure due to this OC motion will also act on the BM, generating a force on the BM proportional to the acceleration of the OC, and thus create a “coupling mass” effect. By reciprocity, this coupling mass is the same as that acting on the OC due to the motion of the BM. This near field fluid coupling is initially observed in a finite element model of a slice of the cochlea. These simulations suggest a simple analytical formulation for the fluid coupling, using higher order beam modes across the width of the cochlear partition. It is well known that the added mass due to the near field pressure dominates the overall mass of the BM, and thus significantly affects the micromechanical dynamics. This work not only quantifies the added mass of the OC due its own motion in the fluid, and shows that this is important, but also demonstrates that the coupling mass effect between the BM and OC significantly affects the dynamics of simple micromechanical models.

  16. Applications of Amniotic Membrane and Fluid in Stem Cell Biology and Regenerative Medicine

    Directory of Open Access Journals (Sweden)

    Kerry Rennie


    Full Text Available The amniotic membrane (AM and amniotic fluid (AF have a long history of use in surgical and prenatal diagnostic applications, respectively. In addition, the discovery of cell populations in AM and AF which are widely accessible, nontumorigenic and capable of differentiating into a variety of cell types has stimulated a flurry of research aimed at characterizing the cells and evaluating their potential utility in regenerative medicine. While a major focus of research has been the use of amniotic membrane and fluid in tissue engineering and cell replacement, AM- and AF-derived cells may also have capabilities in protecting and stimulating the repair of injured tissues via paracrine actions, and acting as vectors for biodelivery of exogenous factors to treat injury and diseases. Much progress has been made since the discovery of AM and AF cells with stem cell characteristics nearly a decade ago, but there remain a number of problematic issues stemming from the inherent heterogeneity of these cells as well as inconsistencies in isolation and culturing methods which must be addressed to advance the field towards the development of cell-based therapies. Here, we provide an overview of the recent progress and future perspectives in the use of AM- and AF-derived cells for therapeutic applications.

  17. Numerical computations of the dynamics of fluidic membranes and vesicles

    CERN Document Server

    Barrett, John W; Nürnberg, Robert


    Vesicles and many biological membranes are made of two monolayers of lipid molecules and form closed lipid bilayers. The dynamical behaviour of vesicles is very complex and a variety of forms and shapes appear. Lipid bilayers can be considered as a surface fluid and hence the governing equations for the evolution include the surface (Navier--)Stokes equations, which in particular take the membrane viscosity into account. The evolution is driven by forces stemming from the curvature elasticity of the membrane. In addition, the surface fluid equations are coupled to bulk (Navier--)Stokes equations. We introduce a parametric finite element method to solve this complex free boundary problem, and present the first three dimensional numerical computations based on the full (Navier--)Stokes system for several different scenarios. For example, the effects of the membrane viscosity, spontaneous curvature and area difference elasticity (ADE) are studied. In particular, it turns out, that even in the case of no viscosit...

  18. Effects of Concentrations on the Transdermal Permeation Enhancing Mechanisms of Borneol: A Coarse-Grained Molecular Dynamics Simulation on Mixed-Bilayer Membranes

    Directory of Open Access Journals (Sweden)

    Xingxing Dai


    Full Text Available Borneol is a natural permeation enhancer that is effective in drugs used in traditional clinical practices as well as in modern scientific research. However, its molecular mechanism is not fully understood. In this study, a mixed coarse-grained model of stratum corneum (SC lipid bilayer comprised of Ceramide-N-sphingosine (CER NS 24:0, cholesterol (CHOL and free fatty acids (FFA 24:0 (2:2:1 was used to examine the permeation enhancing mechanism of borneol on the model drug osthole. We found two different mechanisms that were dependent on concentrations levels of borneol. At low concentrations, the lipid system maintained a bilayer structure. The addition of borneol made the lipid bilayer loosen and improved drug permeation. The “pull” effect of borneol also improved drug permeation. However, for a strongly hydrophobic drug like osthole, the permeation enhancement of borneol was limited. When most borneol molecules permeated into bilayers and were located at the hydrophobic tail region, the spatial competition effect inhibited drug molecules from permeating deeper into the bilayer. At high concentrations, borneol led to the formation of water pores and long-lived reversed micelles. This improved the permeation of osthole and possibly other hydrophobic or hydrophilic drugs through the SC. Our simulation results were supported by Franz diffusion tests and transmission electron microscope (TEM experiments.

  19. Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells. (United States)

    Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C


    As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.


    Pilot-scale demonstration of pervaporation-based removal of volatile organic compounds from a surfactant enhanced aquifer remediation (SEAR) fluid has been conducted at USEPA's Test & Evaluation Facility using hollow fiber membrane modules. The membranes consisted of microporous...

  1. Alcohol's Effects on Lipid Bilayer Properties (United States)

    Ingólfsson, Helgi I.; Andersen, Olaf S.


    Alcohols are known modulators of lipid bilayer properties. Their biological effects have long been attributed to their bilayer-modifying effects, but alcohols can also alter protein function through direct protein interactions. This raises the question: Do alcohol's biological actions result predominantly from direct protein-alcohol interactions or from general changes in the membrane properties? The efficacy of alcohols of various chain lengths tends to exhibit a so-called cutoff effect (i.e., increasing potency with increased chain length, which that eventually levels off). The cutoff varies depending on the assay, and numerous mechanisms have been proposed such as: limited size of the alcohol-protein interaction site, limited alcohol solubility, and a chain-length-dependent lipid bilayer-alcohol interaction. To address these issues, we determined the bilayer-modifying potency of 27 aliphatic alcohols using a gramicidin-based fluorescence assay. All of the alcohols tested (with chain lengths of 1–16 carbons) alter the bilayer properties, as sensed by a bilayer-spanning channel. The bilayer-modifying potency of the short-chain alcohols scales linearly with their bilayer partitioning; the potency tapers off at higher chain lengths, and eventually changes sign for the longest-chain alcohols, demonstrating an alcohol cutoff effect in a system that has no alcohol-binding pocket. PMID:21843475

  2. Fragmented state of lipid bilayers in water

    DEFF Research Database (Denmark)

    Helfrich, W.; Thimmel, J.; Klösgen, Beate Maria


    The bilayers of some typical biological membrane lipids such as PC and DGDG disintegrate in a large excess of water to form an optically invisible dispersive bilayer phase. `Dark bodies' can be reversibly precipitated from it by raising the temperature. The dispersive phase probably consists...... of `knotted sticks', i.e. very thin nodular tubes of bilayer. After reviewing pertinent experimental and theoretical work we report on the discovery of a lower consolute point near room temperature in DGDG/water systems. Its existence shows that the dispersive phase and the dark bodies belong to the same...

  3. Semi-permeable membrane retention of synovial fluid lubricants hyaluronan and proteoglycan 4 for a biomimetic bioreactor. (United States)

    Blewis, Megan E; Lao, Brian J; Jadin, Kyle D; McCarty, William J; Bugbee, William D; Firestein, Gary S; Sah, Robert L


    Synovial fluid (SF) contains lubricant macromolecules, hyaluronan (HA), and proteoglycan 4 (PRG4). The synovium not only contributes lubricants to SF through secretion by synoviocyte lining cells, but also concentrates lubricants in SF due to its semi-permeable nature. A membrane that recapitulates these synovium functions may be useful in a bioreactor system for generating a bioengineered fluid (BF) similar to native SF. The objectives were to analyze expanded polytetrafluoroethylene membranes with pore sizes of 50 nm, 90 nm, 170 nm, and 3 microm in terms of (1) HA and PRG4 secretion rates by adherent synoviocytes, and (2) the extent of HA and PRG4 retention with or without synoviocytes adherent on the membrane. Experiment 1: Synoviocytes were cultured on tissue culture (TC) plastic or membranes +/- IL-1beta + TGF-beta1 + TNF-alpha, a cytokine combination that stimulates lubricant synthesis. HA and PRG4 secretion rates were assessed by analysis of medium. Experiment 2: Bioreactors were fabricated to provide a BF compartment enclosed by membranes +/- adherent synoviocytes, and an external compartment of nutrient fluid (NF). A solution with HA (1 mg/mL, MW ranging from 30 to 4,000 kDa) or PRG4 (50 microg/mL) was added to the BF compartment, and HA and PRG4 loss into the NF compartment after 2, 8, and 24 h was determined. Lubricant loss kinetics were analyzed to estimate membrane permeability. Experiment 1: Cytokine-regulated HA and PRG4 secretion rates on membranes were comparable to those on TC plastic. Experiment 2: Transport of HA and PRG4 across membranes was lowest with 50 nm membranes and highest with 3 microm membranes, and transport of high MW HA was decreased by adherent synoviocytes (for 50 and 90 nm membranes). The permeability to HA mixtures for 50 nm membranes was approximately 20 x 10(-8) cm/s (- cells) and approximately 5 x 10(-8) cm/s (+ cells), for 90 nm membranes was approximately 35 x 10(-8) cm/s (- cells) and approximately 19 x 10(-8) cm

  4. Lipid-protein interactions in plasma membranes of fiber cells isolated from the human eye lens. (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K


    The protein content in human lens membranes is extremely high, increases with age, and is higher in the nucleus as compared with the cortex, which should strongly affect the organization and properties of the lipid bilayer portion of intact membranes. To assess these effects, the intact cortical and nuclear fiber cell plasma membranes isolated from human lenses from 41- to 60-year-old donors were studied using electron paramagnetic resonance spin-labeling methods. Results were compared with those obtained for lens lipid membranes prepared from total lipid extracts from human eyes of the same age group [Mainali, L., Raguz, M., O'Brien, W. J., and Subczynski, W. K. (2013) Biochim. Biophys. Acta]. Differences were considered to be mainly due to the effect of membrane proteins. The lipid-bilayer portions of intact membranes were significantly less fluid than lipid bilayers of lens lipid membranes, prepared without proteins. The intact membranes were found to contain three distinct lipid environments termed the bulk lipid domain, boundary lipid domain, and trapped lipid domain. However, the cholesterol bilayer domain, which was detected in cortical and nuclear lens lipid membranes, was not detected in intact membranes. The relative amounts of bulk and trapped lipids were evaluated. The amount of lipids in domains uniquely formed due to the presence of membrane proteins was greater in nuclear membranes than in cortical membranes. Thus, it is evident that the rigidity of nuclear membranes is greater than that of cortical membranes. Also the permeability coefficients for oxygen measured in domains of nuclear membranes were significantly lower than appropriate coefficients measured in cortical membranes. Relationships between the organization of lipids into lipid domains in fiber cells plasma membranes and the organization of membrane proteins are discussed.

  5. Quantum effects of stringy and membranic nature for the swimming of micro-organisms in a fluid

    CERN Document Server

    Elizalde, E; Nojiri, S; Kawamura, M; Sugamoto, A


    The static potential is investigated in string and membrane theories coupled to U(1) gauge fields (specifically, external magnetic fields) and with antisymmetric tensor fields. The explicit dependence of the potential on the shape of the extended objects is obtained, including a careful calculation of the quantum effects. Noting the features which are common to the dynamics of strings and membranes moving in background fields and to the swimming of micro-organisms in a fluid, the latter problem is studied. The Casimir energy of a micro-organism is estimated, taking into account the quantum effects and the backreaction from the outside fluid.

  6. Bursting Bubbles and Bilayers

    Directory of Open Access Journals (Sweden)

    Steven P. Wrenn, Stephen M. Dicker, Eleanor F. Small, Nily R. Dan, Michał Mleczko, Georg Schmitz, Peter A. Lewin


    Full Text Available This paper discusses various interactions between ultrasound, phospholipid monolayer-coated gas bubbles, phospholipid bilayer vesicles, and cells. The paper begins with a review of microbubble physics models, developed to describe microbubble dynamic behavior in the presence of ultrasound, and follows this with a discussion of how such models can be used to predict inertial cavitation profiles. Predicted sensitivities of inertial cavitation to changes in the values of membrane properties, including surface tension, surface dilatational viscosity, and area expansion modulus, indicate that area expansion modulus exerts the greatest relative influence on inertial cavitation. Accordingly, the theoretical dependence of area expansion modulus on chemical composition - in particular, poly (ethylene glyclol (PEG - is reviewed, and predictions of inertial cavitation for different PEG molecular weights and compositions are compared with experiment. Noteworthy is the predicted dependence, or lack thereof, of inertial cavitation on PEG molecular weight and mole fraction. Specifically, inertial cavitation is predicted to be independent of PEG molecular weight and mole fraction in the so-called mushroom regime. In the “brush” regime, however, inertial cavitation is predicted to increase with PEG mole fraction but to decrease (to the inverse 3/5 power with PEG molecular weight. While excellent agreement between experiment and theory can be achieved, it is shown that the calculated inertial cavitation profiles depend strongly on the criterion used to predict inertial cavitation. This is followed by a discussion of nesting microbubbles inside the aqueous core of microcapsules and how this significantly increases the inertial cavitation threshold. Nesting thus offers a means for avoiding unwanted inertial cavitation and cell death during imaging and other applications such as sonoporation. A review of putative sonoporation mechanisms is then presented




    A numerical study was conducted to analyze fluid flow within hollow fiber membranes of the artificial lungs. The hollow fiber bundle was approximated using a porous media model. In addition, the transport equations were solved using the finite-element formulation based on the Galerkin method of weighted residuals. Comparisons with previously published work on the basis of special cases were performed and found to be in excellent agreement. A Newtonian viscous fluid model for the blood was use...

  8. Membrane-supported sample preparation for chemical analysis in fluid flow systems; Membrangestuetzte Probenvorbereitung zur chemischen Analyse in Fliesssystemen

    Energy Technology Data Exchange (ETDEWEB)

    Kiesow, T.; Frenzel, W. [TU-Berlin, Fachgebiet Luftreinhaltung im Inst. fuer Technischen Umweltschutz, Berlin (Germany)


    Membrane-supported separation processes are described in detail, i.e. gas diffusion, dialysis and liquid membrane extraction in fluid flow systems. The focus is on gas diffusion and on the analysis of ammonium and short-chain amines. (orig.) [German] Im folgendem werden die membrangestuetzten Trennverfahren Gasdiffusion, Dialyse und Fluessigmembranextraktion im Fliesssystem naeher beschrieben. Dabei soll im besonderen auf die Gasdiffusion mit einem Bestimmungsverfahren fuer Ammonium und kurzkettige Amine eingegangen werden. (orig.)

  9. Inverted catenoid as a fluid membrane with two points pulled together. (United States)

    Castro-Villarreal, Pavel; Guven, Jemal


    Under inversion in any (interior) point, a catenoid transforms into a deflated compact geometry which touches at two points (its poles). The catenoid is a minimal surface and, as such, is an equilibrium shape of a symmetric fluid membrane. The conformal symmetry of the Hamiltonian implies that inverted minimal surfaces are also equilibrium shapes. However, they will exhibit curvature singularities at their poles. Such singularities are the geometrical signature of the external forces required to pull the poles together. These forces will set up stresses in the inverted shapes. Tuning the force corresponds geometrically to the translation of the point of inversion. For any fixed surface area, there will be a maximum force. The associated shape is a symmetric discocyte. Lowering the external force will induce a transition from the discocyte to a cup-shaped stomatocyte.

  10. Issues associated with modelling of proton exchange membrane fuel cell by computational fluid dynamics (United States)

    Bednarek, Tomasz; Tsotridis, Georgios


    The objective of the current study is to highlight possible limitations and difficulties associated with Computational Fluid Dynamics in PEM single fuel cell modelling. It is shown that an appropriate convergence methodology should be applied for steady-state solutions, due to inherent numerical instabilities. A single channel fuel cell model has been taken as numerical example. Results are evaluated for quantitative as well qualitative points of view. The contribution to the polarization curve of the different fuel cell components such as bi-polar plates, gas diffusion layers, catalyst layers and membrane was investigated via their effects on the overpotentials. Furthermore, the potential losses corresponding to reaction kinetics, due to ohmic and mas transport limitations and the effect of the exchange current density and open circuit voltage, were also investigated. It is highlighted that the lack of reliable and robust input data is one of the issues for obtaining accurate results.

  11. Transient computation fluid dynamics modeling of a single proton exchange membrane fuel cell with serpentine channel (United States)

    Hu, Guilin; Fan, Jianren

    The proton exchange membrane fuel cell (PEMFC) has become a promising candidate for the power source of electrical vehicles because of its low pollution, low noise and especially fast startup and transient responses at low temperatures. A transient, three-dimensional, non-isothermal and single-phase mathematical model based on computation fluid dynamics has been developed to describe the transient process and the dynamic characteristics of a PEMFC with a serpentine fluid channel. The effects of water phase change and heat transfer, as well as electrochemical kinetics and multicomponent transport on the cell performance are taken into account simultaneously in this comprehensive model. The developed model was employed to simulate a single laboratory-scale PEMFC with an electrode area about 20 cm 2. The dynamic behavior of the characteristic parameters such as reactant concentration, pressure loss, temperature on the membrane surface of cathode side and current density during start-up process were computed and are discussed in detail. Furthermore, transient responses of the fuel cell characteristics during step changes and sinusoidal changes in the stoichiometric flow ratio of the cathode inlet stream, cathode inlet stream humidity and cell voltage are also studied and analyzed and interesting undershoot/overshoot behavior of some variables was found. It was also found that the startup and transient response time of a PEM fuel cell is of the order of a second, which is similar to the simulation results predicted by most models. The result is an important guide for the optimization of PEMFC designs and dynamic operation.

  12. New generation of membrane efficient water-based drilling fluids: pragmatic and cost-effective solutions to borehole stability problems

    Energy Technology Data Exchange (ETDEWEB)

    Tare, U.A. [Haliburton, Calgary, AB (Canada); Mody, F.K. [Shell International E and P Inc., Calgary, AB (Canada); Tan, C.P. [CSIRO Petroleum, Kensington, WA (Australia)


    Drilling and completion operations in shales often suffer as a result of wellbore instability. Mechanical failure of the rock around a wellbore is the primary cause of shale instability. This process can be exacerbated by physico-chemical interactions between drilling fluids and shales. Water-based drilling fluids are used more and more due to environmental awareness that becomes more prevalent. Wellbore instability problems can however result from an improper application of water-based drilling fluids in those cases where drilling occurs in sensitive clay-rich formations. To meet the requirements of the petroleum industry, considerable collaborative efforts were expanded in the development of innovative environmentally acceptable water-based drilling fluids. In this paper, the authors describe the process that leads to the development of these drilling fluids. It is possible to achieve shale stability through an osmotic outflow of pore fluid and prevention/minimization of mud pressure penetration, as laboratory experiments on shale samples under realistic downhole conditions exposed to these drilling fluids prove. High membrane efficiencies, in excess of 80 per cent, were generated by this new generation of membrane efficient water-based drilling fluids. Drilling objectives resulting from an improved application of water-based drilling fluids are made possible by a fundamental understanding of the main drilling fluid-shale interaction mechanisms for shale stability and the application of experimental data to field conditions. The authors indicate that the achievement of trouble-free drilling of shales and notable reductions in non-productive time is accomplished by following the practical guidelines included in this paper for maintaining shale stability with the new generation of water-based drilling fluids. 8 refs., 2 tabs., 4 figs.

  13. Direct in situ measurement of specific capacitance, monolayer tension, and bilayer tension in a droplet interface bilayer. (United States)

    Taylor, Graham J; Venkatesan, Guru A; Collier, C Patrick; Sarles, Stephen A


    Thickness and tension are important physical parameters of model cell membranes. However, traditional methods to measure these quantities require multiple experiments using separate equipment. This work introduces a new multi-step procedure for directly accessing in situ multiple physical properties of droplet interface bilayers (DIB), including specific capacitance (related to thickness), lipid monolayer tension in the Plateau-Gibbs border, and bilayer tension. The procedure employs a combination of mechanical manipulation of bilayer area followed by electrowetting of the capacitive interface to examine the sensitivities of bilayer capacitance to area and contact angle to voltage, respectively. These data allow for determining the specific capacitance of the membrane and surface tension of the lipid monolayer, which are then used to compute bilayer thickness and tension, respectively. The use of DIBs affords accurate optical imaging of the connected droplets in addition to electrical measurements of bilayer capacitance, and it allows for reversibly varying bilayer area. After validating the accuracy of the technique with diphytanoyl phosphatidylcholine (DPhPC) DIBs in hexadecane, the method is applied herein to quantify separately the effects on membrane thickness and tension caused by varying the solvent in which the DIB is formed and introducing cholesterol into the bilayer. Because the technique relies only on capacitance measurements and optical images to determine both thickness and tension, this approach is specifically well-suited for studying the effects of peptides, biomolecules, natural and synthetic nanoparticles, and other species that accumulate within membranes without altering bilayer conductance.

  14. Standard Test Methods for Microscopical Sizing and Counting Particles from Aerospace Fluids on Membrane Filters

    CERN Document Server

    American Society for Testing and Materials. Philadelphia


    1.1 These test methods cover the determination of the size distribution and quantity of particulate matter contamination from aerospace fluids isolated on a membrane filter. The microscopical techniques described may also be applied to other properly prepared samples of small particles. Two test methods are described for sizing particles as follows: 1.1.1 Test Method A—Particle sizes are measured as the diameter of a circle whose area is equal to the projected area of the particle. 1.1.2 Test Method B—Particle sizes are measured by their longest dimension. 1.2 The test methods are intended for application to particle contamination determination of aerospace fluids, gases, surfaces, and environments. 1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard. 1.4 These test methods do not provide for sizing particles smaller than 5 μm. Note 1—Results of these methods are subject to variables inherent in any statistical method. The...

  15. The anti-apoptotic effect of fluid mechanics preconditioning by cells membrane and mitochondria in rats brain microvascular endothelial cells. (United States)

    Tian, Shan; Zhu, Fengping; Hu, Ruiping; Tian, Song; Chen, Xingxing; Lou, Dan; Cao, Bing; Chen, Qiulei; Li, Bai; Li, Fang; Bai, Yulong; Wu, Yi; Zhu, Yulian


    Exercise preconditioning is a simple and effective way to prevent ischemia. This paper further provided the mechanism in hemodynamic aspects at the cellular level. To study the anti-apoptotic effects of fluid mechanics preconditioning, Cultured rats brain microvascular endothelial cells were given fluid intervention in a parallel plate flow chamber before oxygen glucose deprivation. It showed that fluid mechanics preconditioning could inhibit the apoptosis of endothelial cells, and this process might be mediated by the shear stress activation of Tie-2 on cells membrane surface and Bcl-2 on the mitochondria surface. Copyright © 2017. Published by Elsevier B.V.

  16. Validation of computational non-Newtonian fluid model for membrane bioreactor. (United States)

    Sørensen, Lasse; Bentzen, Thomas Ruby; Skov, Kristian


    Membrane bioreactor (MBR) systems are often considered as the wastewater treatment method of the future due to their high effluent quality. One of the main problems with such systems is a relative large energy consumption, compared to conventional activated sludge (CAS) systems, which has led to further research in this specific area. A powerful tool for optimizing MBR-systems is computational fluid dynamics (CFD) modelling, which gives researchers the ability to describe the flow in the systems. A parameter which is often neglected in such models is the non-Newtonian properties of active sludge, which is of great importance for MBR systems since they operate at sludge concentrations up to a factor of 10 compared to CAS systems, resulting in strongly shear thinning liquids. A CFD-model is validated against measurements conducted in a system with rotating cross-flow membranes submerged in non-Newtonian liquids, where tangential velocities are measured with a Laser Doppler Anemometer (LDA). The CFD model is found to be capable of modelling the correct velocities in a range of setups, making CFD models a powerful tool for optimization of MBR systems.

  17. Microfluidic fabrication of addressable tethered lipid bilayer arrays and optimization using SPR with silane-derivatized nanoglassy substrates. (United States)

    Taylor, Joseph D; Phillips, K Scott; Cheng, Quan


    We report the microfluidic fabrication of robust and fluid tethered bilayer arrays within a poly(dimethylsiloxane) (PDMS) chip, and demonstrate its addressability and biosensing by incorporating the GM1 receptor into the bilayer framework for detection of cholera toxin. Rapid optimization of the experimental conditions is achieved by using nanoglassified surfaces in combination with surface plasmon resonance. The ultrathin glassy film on gold mimics glass surfaces employed in microfluidics, allowing real-time monitoring of multiple assembly steps and therefore permitting rapid prototyping of microfluidic arrays. The tethered bilayer array utilizes a covalently immobilized biotinylated protein for generation of well-defined capture zones where a streptavidin link is employed for the immobilization of biotinylated vesicles. Fusion of captured vesicles is accomplished using a concentrated PEG solution, and the lateral diffusion of the tethered bilayer membrane is characterized by fluorescence recovery after photobleaching methods. The tethered membrane arrays demonstrate marked stability and high mobility, which provide an ideal host environment for membrane-associated proteins and open new avenues for high-throughput analysis of these proteins.

  18. Equilibrium insertion of nanoscale objects into phospholipid bilayers

    CERN Document Server

    Pogodin, Sergey


    Certain membrane proteins, peptides, nanoparticles and nanotubes have rigid structure and fixed shape. They are often viewed as spheres and cylinders with certain surface properties. Single Chain Mean Field theory is used to model the equilibrium insertion of nanoscale spheres and rods into the phospholipid bilayer. The equilibrium structures and the resulting free energies of the nano-objects in the bilayer allow to distinguish different orientations in the bilayer and estimate the energy barrier of insertion.

  19. Concentration of Sonication Fluid through Centrifugation Is Superior to Membrane Filtration for Microbial Diagnosis of Orthopedic Implant-Associated Infection. (United States)

    Zitron, Renato; Wajsfeld, Tali; Klautau, Giselle Burlamaqui; da Silva, Cely Barreto; Nigro, Stanley; Mercadante, Marcelo Tomanik; Polesello, Giancarlo Cavalli; Cury, Ricardo de Paula Leite; Salles, Mauro Jose Costa


    Microbial identification of orthopedic implant-associated infections using sonication fluid (SF) submitted to a concentration step by membrane filtration (SMF) was compared with the standard centrifugation (SC) method. Among 33 retrieved infected implants, sonication identified microorganisms in 26 (78.8%). The sensitivity of SC was higher than that of SMF (78.8% versus 30.3%; P < 0.001).

  20. The effect of tail-length mismatch in binary DMPC/DSPC lipid bilayers (United States)

    Ashkar, Rana; Nagao, Michihiro; Butler, Paul


    Bilayer heterogeneity has been long hypothesized to drive raft formation and promote complex functionality in lipid membranes. The highly dynamic nature of the membrane however is thought to play a critical role in this delicate balance between structure and performance. To probe the effect of lateral heterogeneity on membrane dynamics, we investigate the thermal response of unilamellar-vesicle systems of mixed dimyristoylphosphatidylcholine (DMPC) and distearoylphosphatidylcholine (DSPC) with DMPC/DSPC ratios of 50/50 and 70/30. Both lipids experience a transition from an ordered gel phase, with stiff stretched tails, to a melted fluid phase, with more coiled flexible tails, as they are heated through their melting temperature, Tm(DMPC) ~ 21 °C and Tm(DSPC) ~ 51 °C. The distinct Tm's of the two lipids provide a broad gel-fluid phase with a significant mismatch (~ 20 Å) between the tail-lengths of the DMPC and DSPC molecules. The structural properties of the vesicles were determined by small-angle neutron and x-ray scattering and the collective lipid dynamics in the bilayer were investigated by neutron spin-echo (NSE) spectroscopy on selectively deuterated samples. The NSE results indicate a slowdown of thickness fluctuations in the gel-fluid coexistence phase and an intriguingly strong enhancement in the thickness fluctuation amplitude for T >Tm(DSPC) compared to our previous work on single component vesicles.

  1. Lipid bilayer composition influences small multidrug transporters

    Directory of Open Access Journals (Sweden)

    Curnow Paul


    Full Text Available Abstract Background Membrane proteins are influenced by their surrounding lipids. We investigate the effect of bilayer composition on the membrane transport activity of two members of the small multidrug resistance family; the Escherichia coli transporter, EmrE and the Mycobacterium tuberculosis, TBsmr. In particular we address the influence of phosphatidylethanolamine and anionic lipids on the activity of these multidrug transporters. Phosphatidylethanolamine lipids are native to the membranes of both transporters and also alter the lateral pressure profile of a lipid bilayer. Lipid bilayer lateral pressures affect membrane protein insertion, folding and activity and have been shown to influence reconstitution, topology and activity of membrane transport proteins. Results Both EmrE and TBsmr are found to exhibit a similar dependence on lipid composition, with phosphatidylethanolamine increasing methyl viologen transport. Anionic lipids also increase transport for both EmrE and TBsmr, with the proteins showing a preference for their most prevalent native anionic lipid headgroup; phosphatidylglycerol for EmrE and phosphatidylinositol for TBsmr. Conclusion These findings show that the physical state of the membrane modifies drug transport and that substrate translocation is dependent on in vitro lipid composition. Multidrug transport activity seems to respond to alterations in the lateral forces exerted upon the transport proteins by the bilayer.

  2. Thermotropic and Barotropic Phase Behavior of Phosphatidylcholine Bilayers

    Directory of Open Access Journals (Sweden)

    Nobutake Tamai


    Full Text Available Bilayers formed by phospholipids are frequently used as model biological membranes in various life science studies. A characteristic feature of phospholipid bilayers is to undergo a structural change called a phase transition in response to environmental changes of their surroundings. In this review, we focus our attention on phase transitions of some major phospholipids contained in biological membranes, phosphatidylcholines (PCs, depending on temperature and pressure. Bilayers of dipalmitoylphosphatidylcholine (DPPC, which is the most representative lipid in model membrane studies, will first be explained. Then, the bilayer phase behavior of various kinds of PCs with different molecular structures is revealed from the temperature–pressure phase diagrams, and the difference in phase stability among these PC bilayers is discussed in connection with the molecular structure of the PC molecules. Furthermore, the solvent effect on the phase behavior is also described briefly.

  3. Lipid bilayers and interfaces

    NARCIS (Netherlands)

    Kik, R.A.


    In biological systems lipid bilayers are subject to many different interactions with other entities. These can range from proteins that are attached to the hydrophilic region of the bilayer or transmembrane proteins that interact with the hydrophobic region of the lipid bilayer. Interaction between

  4. Blood clotting reactions on nanoscale phospholipid bilayers. (United States)

    Morrissey, James H; Pureza, Vincent; Davis-Harrison, Rebecca L; Sligar, Stephen G; Ohkubo, Y Zenmei; Tajkhorshid, Emad


    Blood clotting reactions, such as those catalyzed by the tissue factor:factor VIIa complex (TF:FVIIa), assemble on membrane surfaces containing anionic phospholipids such as phosphatidylserine (PS). In fact, membrane binding is critical for the function of most of the steps in the blood clotting cascade. In spite of this, our understanding of how the membrane contributes to catalysis, or even how these proteins interact with phospholipids, is incomplete. Making matters more complicated, membranes containing mixtures of PS and neutral phospholipids are known to spontaneously form PS-rich membrane microdomains in the presence of plasma concentrations of calcium ions, and it is likely that blood-clotting proteases such as TF:FVIIa partition into these PS-rich microdomains. Unfortunately, little is known about how membrane microdomain composition influences the activity of blood-clotting proteases, which is typically not under experimental control even in "simple" model membranes. Our laboratories have developed and applied new technologies for studying membrane proteins to gain insights into how blood-clotting protease-cofactor pairs assemble and function on membrane surfaces. This includes using a novel, nanoscale bilayer system (Nanodiscs) that permits assembling blood-clotting protease-cofactor pairs on stable bilayers containing from 65 to 250 phospholipid molecules per leaflet. We have used this system to investigate how local (nanometer-scale) changes in phospholipid bilayer composition modulate TF:FVIIa activity. We have also used detailed molecular-dynamics simulations of nanoscale bilayers to provide atomic-scale predictions of how the membrane-binding domain of factor VIIa interacts with PS in membranes.

  5. Cholesterol orientation and tilt modulus in DMPC bilayers


    Khelashvili, George; Pabst, Georg; Harries, Daniel


    We performed molecular dynamics (MD) simulations of hydrated bilayers containing mixtures of dimyristoylphosphatidylcholine (DMPC) and Cholesterol at various ratios, to study the effect of cholesterol concentration on its orientation, and to characterize the link between cholesterol tilt and overall phospholipid membrane organization. The simulations show a substantial probability for cholesterol molecules to transiently orient perpendicular to the bilayer normal, and suggest that cholesterol...

  6. A prospective study of 20 foot and ankle wounds treated with cryopreserved amniotic membrane and fluid allograft. (United States)

    Werber, Bruce; Martin, Erin


    We reviewed the background information and previous clinical studies that considered the use of allogeneic amniotic tissue and fluid (granulized amniotic membrane and amniotic fluid) in the treatment of chronic diabetic foot wounds. This innovation represents a relatively new approach to wound management by delivering a unique allograft of live human cells in a nonimmunogenic structural tissue matrix. Developed to fill soft tissue defects and bone voids and to convey antimicrobial and anti-inflammatory capabilities, granulized amniotic membrane and amniotic fluid does not require fetal death, because its procurement is performed with maternal consent during birth. In the present investigation, 20 chronic wounds (20 patients) that had been treated with standard wound therapy for a mean of 36.6 ± 31.58 weeks and with a mean baseline area of 10.15 ± 19.54 cm(2) were followed up during a 12-week observation period or until they healed. A total of 18 of the wounds (90%) healed during the 12-week observation period, and none of the wounds progressed to amputation. From our experience with the patients in the present case series, we believe that granulized amniotic membrane and amniotic fluid represents a useful option for the treatment of chronic diabetic foot wounds.

  7. Polydopamine-Supported Lipid Bilayers

    Directory of Open Access Journals (Sweden)

    Souryvanh Nirasay


    Full Text Available We report the formation of lipid membranes supported by a soft polymeric cushion of polydopamine. First, 20 nm thick polydopamine films were formed on mica substrates. Atomic force microscopy imaging indicated that these films were also soft with a surface roughness of 2 nm under hydrated conditions. A zwitterionic phospholipid bilayer was then deposited on the polydopamine cushion by fusion of dimyristoylphosphatidylcholine (DMPC and dioleoylphosphatidylcholine (DOPC vesicles. Polydopamine films preserved the lateral mobility of the phospholipids as shown by fluorescence microscopy recovery after photobleaching (FRAP experiments. Diffusion coefficients of ~5.9 and 7.2 µm2 s−1 were respectively determined for DMPC and DOPC at room temperature, values which are characteristic of lipids in a free standing bilayer system.

  8. Polydopamine-Supported Lipid Bilayers (United States)

    Nirasay, Souryvanh; Badia, Antonella; Leclair, Grégoire; Claverie, Jerome P.; Marcotte, Isabelle


    We report the formation of lipid membranes supported by a soft polymeric cushion of polydopamine. First, 20 nm thick polydopamine films were formed on mica substrates. Atomic force microscopy imaging indicated that these films were also soft with a surface roughness of 2 nm under hydrated conditions. A zwitterionic phospholipid bilayer was then deposited on the polydopamine cushion by fusion of dimyristoylphosphatidylcholine (DMPC) and dioleoylphosphatidylcholine (DOPC) vesicles. Polydopamine films preserved the lateral mobility of the phospholipids as shown by fluorescence microscopy recovery after photobleaching (FRAP) experiments. Diffusion coefficients of ~5.9 and 7.2 µm2 s−1 were respectively determined for DMPC and DOPC at room temperature, values which are characteristic of lipids in a free standing bilayer system.

  9. Multiscale Modeling of supported bilayers (United States)

    Faller, Roland; Xing, Chenyue; Hoopes, Matthew I.


    Supported Lipid Bilayers are an abundant research platform for understanding the behavior of real cell membranes as they allow for additional mechanical stability. We studied systematically the changes that a support induces on a phospholipid bilayer using coarse-grained molecular modeling on different levels. We characterize the density and pressure profiles as well as the density imbalance inflicted on the membrane by the support. We also determine the diffusion coefficients and characterize the influence of different corrugations of the support. We then determine the free energy of transfer of phospholipids between the proximal and distal leaflet of a supported membrane using the coarse-grained Martini model. It turns out that there is at equilibrium about a 2-3% higher density in the proximal leaflet. These results are in favorable agreement with recent data obtained by very large scale modeling using a water free model where flip-flop can be observed directly. We compare results of the free energy of transfer obtained by pulling the lipid across the membrane in different ways. There are small quantitative differences but the overall picture is consistent. We are additionally characterizing the intermediate states which determine the barrier height and therefore the rate of translocation.

  10. Antioxidant Vitamin Status in the Serum and Amniotic Fluid of Women with Premature Rupture of the Fetal Membranes. (United States)

    Barrett, Bridget M.

    The purpose of this study was to examine the status of antioxidant vitamins in women with premature rupture of the fetal membranes. Specimens of blood and amniotic fluid were obtained from 80 pregnant subjects included both smokers and non-smokers during the third trimester. The concentrations of ascorbic acid (ASA), beta -carotene, retinol and alpha -tocopherol in serum and amniotic fluid were determined. The experimental group consisted of those subjects with PROM while the control subjects were those with normal pregnancy. No statistical differences were found between the PROM and control groups in retinol and vitamin E concentrations in amniotic fluid and serum. Serum ASA concentrations of PROM subjects were not different from controls, but the PROM subjects had significantly lower amniotic fluid ASA concentrations. However, in a study with fewer subjects a lower serum ASA concentration in the PROM subjects was observed. The ratio of amniotic fluid ASA concentration to ASA serum concentration was significantly lower in PROM patients than in controls in both studies. This suggests that low levels of ASA in the amniotic fluid, but not in serum is better associated with PROM. A low amniotic fluid concentration of ASA may reflect an inefficient transfer and/or increased fetal utilization. Alterations in ASA concentration in the amniotic fluid may affect the integrity of the chorioamnion leading to PROM. beta -Carotene was not found in the amniotic fluid. Serum beta-carotene levels were significantly lower in the PROM group compared to the control group. Low concentrations of beta-carotene in maternal serum in smokers not only associated with poor maternal outcome (PROM) but also compromised the fetal outcome (decreased birth weight). Maintenance of adequate serum beta-carotene concentration and amniotic fluid ASA in smokers may result in better maternal and fetal outcome. This study demonstrated that nutrition is an important factor in the prevention of PROM.

  11. Regulation of sodium channel function by bilayer elasticity

    DEFF Research Database (Denmark)

    Lundbaek, Jens A; Birn, Pia; Hansen, Anker J


    be a general mechanism regulating membrane protein function, we examined whether voltage-dependent skeletal-muscle sodium channels, expressed in HEK293 cells, are regulated by bilayer elasticity, as monitored using gramicidin A (gA) channels. Nonphysiological amphiphiles (beta-octyl-glucoside, Genapol X-100......, Triton X-100, and reduced Triton X-100) that make lipid bilayers less "stiff", as measured using gA channels, shift the voltage dependence of sodium channel inactivation toward more hyperpolarized potentials. At low amphiphile concentration, the magnitude of the shift is linearly correlated to the change...... in gA channel lifetime. Cholesterol-depletion, which also reduces bilayer stiffness, causes a similar shift in sodium channel inactivation. These results provide strong support for the notion that bilayer-protein hydrophobic coupling allows the bilayer elastic properties to regulate membrane protein...

  12. Actomyosin dynamics drive local membrane component organization in an in vitro active composite layer. (United States)

    Köster, Darius Vasco; Husain, Kabir; Iljazi, Elda; Bhat, Abrar; Bieling, Peter; Mullins, R Dyche; Rao, Madan; Mayor, Satyajit


    The surface of a living cell provides a platform for receptor signaling, protein sorting, transport, and endocytosis, whose regulation requires the local control of membrane organization. Previous work has revealed a role for dynamic actomyosin in membrane protein and lipid organization, suggesting that the cell surface behaves as an active composite composed of a fluid bilayer and a thin film of active actomyosin. We reconstitute an analogous system in vitro that consists of a fluid lipid bilayer coupled via membrane-associated actin-binding proteins to dynamic actin filaments and myosin motors. Upon complete consumption of ATP, this system settles into distinct phases of actin organization, namely bundled filaments, linked apolar asters, and a lattice of polar asters. These depend on actin concentration, filament length, and actin/myosin ratio. During formation of the polar aster phase, advection of the self-organizing actomyosin network drives transient clustering of actin-associated membrane components. Regeneration of ATP supports a constitutively remodeling actomyosin state, which in turn drives active fluctuations of coupled membrane components, resembling those observed at the cell surface. In a multicomponent membrane bilayer, this remodeling actomyosin layer contributes to changes in the extent and dynamics of phase-segregating domains. These results show how local membrane composition can be driven by active processes arising from actomyosin, highlighting the fundamental basis of the active composite model of the cell surface, and indicate its relevance to the study of membrane organization.

  13. Probing the position of resveratrol in lipid bilayers

    DEFF Research Database (Denmark)

    de Ghellinck, Alexis; Shen, Chen; Fragneto, Giovanna


    The effect of the natural antioxidant resveratrol on the structure of solid supported di-palmitoyl-phosphatidyl-choline (DPPC) bilayers in their fluid state was investigated by neutron reflectometry. Results reveal an accumulation of resveratrol (up to 25%, mol/mol) inside the headgroups...... and they exclude its presence in the hydrophobic core. The presence of resveratrol induces an increase of the average thickness and of the interfacial roughness of the headgroup layer. This may be due to a change of the tilt angle of the phosphocholine headgroups residing next to the resveratrol to a more upright...... orientation and leading to a reduction of the projected area per headgroup. This effect is propagated into the hydrophobic core, where the chain packing is modified despite the absence of resveratrol. When interacting with a DPPC/cholesterol membrane, resveratrol has a similar effect on the neighboring PC...

  14. Dynamic Morphologies of Microscale Droplet Interface Bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Mruetusatorn, Prachya [ORNL; Boreyko, Jonathan B [ORNL; Sarles, Stephen A [ORNL; Venkatesan, Guru [The University of Tennessee; Hayes, Douglas G [ORNL; Collier, Pat [ORNL


    Droplet interface bilayers (DIBs) are a powerful platform for studying the dynamics of synthetic cellular membranes; however, very little has been done to exploit the unique dynamical features of DIBs. Here, we generate microscale droplet interface bilayers ( DIBs) by bringing together femtoliter-volume water droplets in a microfluidic oil channel, and characterize morphological changes of the DIBs as the droplets shrink due to evaporation. By varying the initial conditions of the system, we identify three distinct classes of dynamic morphology. (1) Buckling and Fission: When forming DIBs using the lipid-out method (lipids in oil phase), lipids in the shrinking monolayers continually pair together and slide into the bilayer to conserve their mass. As the bilayer continues to grow, it becomes confined, buckles, and eventually fissions one or more vesicles. (2) Uniform Shrinking: When using the lipid-in method (lipids in water phase) to form DIBs, lipids uniformly transfer from the monolayers and bilayer into vesicles contained inside the water droplets. (3) Stretching and Unzipping: Finally, when the droplets are pinned to the wall(s) of the microfluidic channel, the droplets become stretched during evaporation, culminating in the unzipping of the bilayer and droplet separation. These findings offer a better understanding of the dynamics of coupled lipid interfaces.

  15. Multidimensional modeling of biofilm development and fluid dynamics in a hydrogen-based, membrane biofilm reactor (MBfR). (United States)

    Martin, Kelly J; Picioreanu, Cristian; Nerenberg, Robert


    A two-dimensional, particle-based biofilm model coupled with mass transport and computational fluid dynamics was developed to simulate autotrophic denitrification in a spiral-wound membrane biofilm reactor (MBfR), where hydrogen is supplied via hollow-fiber membrane fabric. The spiral-wound configuration consists of alternating layers of plastic spacer net and membrane fabric that create rows of flow channels, with the top and bottom walls comprised of membranes. The transversal filaments of the spacer partially obstruct the channel flow, producing complex mixing and shear patterns that require multidimensional representation. This study investigated the effect of hydrogen and nitrate concentrations, as well as spacer configuration, on biofilm development and denitrification fluxes. The model results indicate that the cavity spacer filaments, which rest on the bottom membranes, cause uneven biofilm growth. Most biofilm resided on the bottom membranes, only in the wake of the filaments where low shear zones formed. In this way, filament configuration may help achieve a desired biofilm thickness. For the conditions tested in this study, the highest nitrate fluxes were attained by minimizing the filament diameter and maximizing the filament spacing. This lowered the shear stress at the top membranes, allowing for more biofilm growth. For the scenarios studied, biomass limitation at the top membranes hindered performance more significantly than diffusion limitation in the thick biofilms at the bottom membranes. The results also highlighted the importance of two-dimensional modeling to capture uneven biofilm growth on a substratum with geometrical complexity. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. 双层羊膜移植治疗蚕蚀性角膜溃疡的显微技术探讨%Microtechnique of bilayer amniotic membrane transplantation for treatment of Mooren's ulcer

    Institute of Scientific and Technical Information of China (English)

    肖璇; 赵靖; 王殿强; 谢立信


    Objective To investigate the mierotechnique of bilayer amniotic membrane transplantation for treatment of Mooren's ulcer and evaluate the efficacy. Methods Six patients (6 eyes) with Mooren's ulcer were recruited for this study. After medical treatment or lameilar keratoplasty failed to arrest progress of corneal ulcer, bilayer amniotic membrane transplantation was performed for the treatment. We investigated the integrity of corneal epithelium, the healing of corneal ulcer, the improvement of stromal edema, the atrophy of neovessels, the transformation of amniotic membrane and the occurrence of relapse. Results All patients were followed up for 24-34 months (mean 30 months). In all cases, superficial anmiotic membrane dissolved or shed on postoperative day 7-11, disconnecting now. Corneal ulcer healed within 7-15 days postoperatively. In 5 eyes, corneal stromal edema faded away within 2-3 weeks. Corneal neovessels regressed within 2-3 months. The deeper grafts were adhered into the ulcer and fused with the cornea 3 months after the operation. Corneal transparence or macula was achieved within 5-8 months. No recurrence of Moorcn's ulcer was oc-curred in 4 patients during the follow-up period, while 2 eyes relapsed for the exposure of sutures and not re-moving the stitches timely, which had been treated with lamellar keratoplasty and no recurrence again during the follow-up period. Conclusion Bilayer amniotic membrane transplantation has advantages for Mooren's ulcer treatment. Mastering the microsurgical techniques and removing the stitches timely are the key to the success of surgery. It also provides good conditions for the further conduct of keratoplasty.%目的 探讨双层羊膜移植术治疗蚕蚀性角膜溃疡的显微手术要领及临床疗效.方法 对经药物及(或)板层角膜移植术治疗无效或复发的蚕蚀性角膜溃疡6例(6眼),行双层羊膜移植术,观察术后角膜上皮及溃疡的愈合、基质水肿消退、新生血管萎缩

  17. Reconstitution of rhodopsin into polymerizable planar supported lipid bilayers: influence of dienoyl monomer structure on photoactivation. (United States)

    Subramaniam, Varuni; D'Ambruoso, Gemma D; Hall, H K; Wysocki, Ronald J; Brown, Michael F; Saavedra, S Scott


    G-protein-coupled receptors (GPCRs) play key roles in cellular signal transduction and many are pharmacologically important targets for drug discovery. GPCRs can be reconstituted in planar supported lipid bilayers (PSLBs) with retention of activity, which has led to development of GPCR-based biosensors and biochips. However, PSLBs composed of natural lipids lack the high stability desired for many technological applications. One strategy is to use synthetic lipid monomers that can be polymerized to form robust bilayers. A key question is how lipid polymerization affects GPCR structure and activity. Here we have investigated the photochemical activity of bovine rhodopsin (Rho), a model GPCR, reconstituted into PSLBs composed of lipids having one or two polymerizable dienoyl moieties located in different regions of the acyl chains. Plasmon waveguide resonance spectroscopy was used to compare the degree of Rho photoactivation in fluid and poly(lipid) PSLBs. The position of the dienoyl moiety was found to have a significant effect: polymerization near the glycerol backbone significantly attenuates Rho activity whereas polymerization near the acyl chain termini does not. Differences in cross-link density near the acyl chain termini also do not affect Rho activity. In unpolymerized PSLBs, an equimolar mixture of phosphatidylethanolamine and phosphatidylcholine (PC) lipids enhances activity relative to pure PC; however after polymerization, the enhancement is eliminated which is attributed to stabilization of the membrane lamellar phase. These results should provide guidance for the design of robust lipid bilayers functionalized with transmembrane proteins for use in membrane-based biochips and biosensors.

  18. Electron density analysis of the effects of sugars on the structure of lipid bilayers at low hydration - a preliminary study

    Energy Technology Data Exchange (ETDEWEB)

    Lenné, T.; Kent, B.; Koster, K.L.; Garvey, C.J.; Bryant, G. (ANSTO); (USD); (ANU); (RMIT)


    Small angle X-ray scattering is used to study the effects of sugars on membranes during dehydration. Previous work has shown that the bilayer and chain-chain repeat spacings of DPPC bilayers are relatively unaffected by the presence of sugars. In this work we present a preliminary analysis of the electron density profiles of DPPC in the presence of sugars at low hydration. The difficulties of determining the correct phasing are discussed. Sugars and other small solutes have been shown to have an important role in improving the tolerance of a range of species to desiccation and freezing. In particular it has been shown that sugars can stabilize membranes in the fluid membrane phase during dehydration, and in the fully dehydrated state. Equivalently, at a particular hydration, the presence of sugars lowers the transition temperature between the fluid and gel phases. There are two competing models for explaining the effects of sugars on membrane phase transition temperatures. One, designated the water replacement hypothesis (WRH) states that sugars hydrogen bond to phospholipid headgroups, thus hindering the fluid-gel phase transition. One version of this model suggests that certain sugars (such as trehalose) achieve the measured effects by inserting between the phospholipid head groups. An alternative model explains the observed effects of sugars in terms of the sugars effect on the hydration repulsion that develops between opposing membranes during dehydration. The hydration repulsion leads to a lateral compressive stress in the bilayer which squeezes adjacent lipids more closely together, resulting in a transition to the gel phase. When sugars are present, their osmotic and volumetric effects reduce the hydration repulsion, reduce the compressive stress in the membranes, and therefore tend to maintain the average lateral separation between lipids. This model is called the hydration forces explanation (HFE). We recently showed that neither mono- nor di

  19. Evaluation of prostate-specific antigen (PSA) membrane test assays for the forensic identification of seminal fluid. (United States)

    Hochmeister, M N; Budowle, B; Rudin, O; Gehrig, C; Borer, U; Thali, M; Dirnhofer, R


    Prostate specific antigen (PSA, also known as p30), a glycoprotein produced by the prostatic gland and secreted into seminal plasma, is a marker used for demonstrating the presence of seminal fluid. Methods for the detection of PSA include Ouchterlony double diffusion, crossover electrophoresis, rocket immuno-electrophoresis, radial immunodiffusion, and ELISA. The extremely sensitive ELISA technique can detect PSA in concentrations as low as approximately 4 ng/mL. However, all these techniques are cumbersome and time consuming to perform in forensic laboratories, especially when only a few samples per week are processed. Various membrane tests are currently used in clinical settings to screen a patient's serum for the presence of PSA at levels greater than 4 ng/mL. In this study we evaluated three immunochromatographic PSA membrane tests by analyzing semen stains stored at room temperature for up to 30 years, post-coital vaginal swabs taken at different time after intercourse, semen-free vaginal swabs, and various female and male body fluids, including urine. The data demonstrate that PSA membrane test assays offer the same sensitivity as ELISA-based tests and provide a rapid approach for the forensic identification of seminal fluid. Furthermore, when the supernatant from a DNA extraction is used for the assay, there is essentially no DNA consumption for determining the presence of PSA in a forensic sample.

  20. Atomistic Monte Carlo simulation of lipid membranes

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Sklenar, Heinz


    , as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential......Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction...... into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches...

  1. Retaining activity of enzymes after capture and extraction within a single-drop of biological fluid using immunoaffinity membranes. (United States)

    Shimazaki, Youji; Sato, Yuki


    The purpose of this study was the measurement of enzyme activity within a single-drop of biological fluid after micropurification. Esterase and lactate dehydrogenase (LDH) retained their enzymatic activities after being captured by membrane-immobilized antibodies, which were prepared by non-denaturing two-dimensional electrophoresis, transferred to polyvinylidene difluoride and then stained by Ponceau S. The activities of both enzymes were also measured after being captured by antibodies and biotinylated antibodies bound to membrane-immobilized protein A or avidin, respectively. After esterase and LDH were captured from biological samples by membrane-immobilized protein A or avidin, their activities were semi-quantitatively measured on the surface of the membrane using fluorescence determination. More than 51% of enzyme activities were retained even after the enzymes were captured by biotinylated antibody bound to membrane-immobilized avidin and eluted by rinsing with 5μL of 1% Triton X-100, compared with the activities of the enzyme on the immunoaffinity membrane.

  2. Characterization of mesenchymal stem cells derived from the equine synovial fluid and membrane. (United States)

    Prado, Aline Ambrogi Franco; Favaron, Phelipe Oliveira; da Silva, Luis Claudio Lopes Correia; Baccarin, Raquel Yvonne Arantes; Miglino, Maria Angelica; Maria, Durvanei Augusto


    Isolation of mesenchymal stem cells (MSCs) in equines, has been reported for different tissues including bone marrow, adipose, umbilical cord, peripheral blood, and yolk sac. In regard to the MSCs derived from synovial fluid (SF) or membrane (SM), there is data available for humans, dogs, pigs, goats and horses. Especially in equines, these cells have being considered promising candidates for articular regeneration. Herein, we established and characterized MSCs obtained from equine SF and SM. Samples were obtained during arthroscopy and cultured using MEM (Minimum Essential Medium). MSCs were characterized by morphology and expression of specific markers for stem cells, pluripotency, inflammation, and cell cycle. The medium MEM was more effective (97% ± 2) to maintain both cultures. The cultures were composed by adherent cells with fibroblast-like shape, which had a growth pattern represented by a sigmoidal curve. After the expansion, the cells were analyzed by flow cytometry for stem cells, inflammatory, and cell cycle markers, and both lineages showed significant expression of CD45, Oct3/4, Nanog, CD105, CD90, CD34, CD117, CD133, TRA-1-81, VEGF, and LY6a. In contrast, there were differences in the cell cycle phases between the lineages, which was not observed in relation to the mitochondrial electrical potential. Given the large impact that joint pathology has on the athletic performance horses, our results suggested that the SF and SM are promising sources of stem cells with satisfactory characteristics of growth and gene expression that can be used in equine regenerative medicine.

  3. Survey of Pregnancy Outcome in Preterm Premature Rupture of Membranes with Amniotic Fluid Index <5 and ≥5

    Directory of Open Access Journals (Sweden)

    Fatemeh Tavassoli


    Full Text Available Objectives: Preterm premature rupture of membranes (PPROM is among the most important causes of perinatal morbidity and mortality. The aim of this study is to survey the pregnancy outcomes in preterm premature rupture of membranes with an amniotic fluid index of 5.Methods: This prospective cohort study was performed on 137 pregnant women complicated by preterm premature rupture of membranes (PPROM with a gestational age of 28-34 weeks during October 2006 to October 2008. The patients were divided in two groups according to their amniotic fluid index; AFI<5 (77cases, AFI≥5 (60cases. The Chi-squared test for qualitative variables and T-student test for quantitative variables were used to analyze the results.Results: The results showed that there was no significant difference in terms of the number of pregnancies, gestational age at rupture of membranes and birthweight between the two groups. However, the results demonstrated that the patients with AFI<5 exhibited a significantly shorter latency period (p=0.049, a higher rate of cesarean due to fetal distress (p=0.008, a lower neonatal Apgar score in the first minute (p=0.0127 and a higher rate of neonatal death during the first week (p=0.045.Conclusion: Overall, PPROM with oligohydroamnios is associated with shorter latency, higher rate of C/S, higher rate of early neonatal death and lower neonatal Apgar.

  4. Superdiffusion in supported lipid bilayers

    CERN Document Server

    Campagnola, Grace; Schroder, Bryce W; Peersen, Olve B; Krapf, Diego


    We study the diffusion of membrane-targeting C2 domains using single-molecule tracking in supported lipid bilayers. The ensemble-averaged mean square displacement (MSD) exhibits superdiffusive behavior. However, the time-averaged MSD of individual trajectories is found to be linear with respect to lag time, as in Brownian diffusion. These observations are explained in terms of bulk excursions that introduce jumps with a heavy-tail distribution. Our experimental results are shown to agree with analytical models of bulk-mediated diffusion and with numerical simulations.

  5. Lipids, membrane proteins and natural membranes studied by neutron scattering and diffraction: A review (United States)

    Zaccai, Giuseppe


    Diffraction first observed from myelin 50 years ago was correctly attributed to a fluid crystal of lipids, because similar patterns were observed from extracted lipid preparations. Following on more recent X-ray work which characterized a variety of lipid-water structures, neutron diffraction experiments have provided detailed descriptions of the molecular conformations in lipid bilayers. For a long time, however, the molecular structure of membrane proteins remained elusive and the development of detergents for the extraction of active membrane proteins, and the discovery of naturally crystalline purple membrane were important breakthroughs in this field. Structural parameters of membrane proteins solubilised in detergent have been measured by neutron scattering with contrast variation techniques. Purple membrane has been studied extensively by neutron diffraction. It is an excellent illustration of the use of deuterium labeling by different approaches to address specific questions of molecular structure. These studies are reviewed with a special emphasis on aspects which are applicable to membranes in general.

  6. Effects of fluid flow on elution of hydrophilic modifier from dialysis membrane surfaces. (United States)

    Matsuda, Masato; Sato, Mika; Sakata, Hiroki; Ogawa, Takahisa; Yamamoto, Ken-ichiro; Yakushiji, Taiji; Fukuda, Makoto; Miyasaka, Takehiro; Sakai, Kiyotaka


    When uremic blood flows through dialyzers during hemodialysis, dialysis membrane surfaces are exposed to shear stress and internal filtration, which may affect the surface characteristics of the dialysis membranes. In the present study, we evaluated changes in the characteristics of membrane surfaces caused by shear stress and internal filtration using blood substitutes: water purified by reverse osmosis and 6.7 wt% dextran70 solution. We focused on the levels of a hydrophilic modifier, polyvinylpyrrolidone (PVP), on the membrane surface measured by attenuated total reflectance Fourier transform infrared spectroscopy. Experiments involving 4 h dialysis, 0-144 h shear-stress loading, and 4 h dead-end filtration were performed using polyester-polymer alloy (PEPA) and polysulfone (PS) membranes. After the dialysis experiments with accompanying internal filtration, average PVP retention on the PEPA membrane surface was 93.7% in all areas, whereas that on the PS membrane surface was 98.9% in all areas. After the shear-stress loading experiments, PVP retention on the PEPA membrane surface decreased as shear-stress loading time and the magnitude of shear stress increased. However, with the PS membrane, PVP retention scarcely changed. After the dead-end filtration experiments, PVP retention decreased in all areas for both PEPA and PS membranes, but PVP retention on the PEPA membrane surface was lower than that on the PS membrane surface. PVP on the PEPA membrane surface was eluted by both shear stress and internal filtration, while that on the PS membrane surface was eluted only by internal filtration.

  7. Comparative Study of Pregnancy Outcome in Premature Rupture of Membranes With Amniotic Fluid Indices of less and more than 5

    Directory of Open Access Journals (Sweden)

    F Tavassoli


    Full Text Available Introduction: Preterm premature rupture of the membranes (PPROM is one of the most important causes of perinatal morbidity and mortality. The aim of this study was to evaluate pregnancy outcome in preterm premature rupture of the membranes with amniotic fluid index less than 5 and more than 5. Methods: This prospective study was performed on 137 pregnant women with gestational age of 28-34 weeks with PPROM from Oct 2006 to Oct 2008. The patients were divided in two groups according to amniotic fluid index: AFI<5 (77 cases, AFI≥5 (60 cases. Chi-square test was used for qualitative variables, while T-student test was used for quantitative test. Results: Both groups were similar with respect to number of pregnancies, gestational age at rupture of the membranes and birth weight. In the group with AFI<5, applied latency was significantly shorter (PV=0.049, rate of cesarean section was higher due to fetal distress (PV=0.008, neonatal Apgar score in first minute was lower during the first week (PV=0.0127 and the rate of neonatal death was higher (PV=0.045 during the first week. Conclusion: An AFI<5 cm after PPROM is associated with earlier delivery, higher rate of cesarean due to fetal distress, higher rate of neonatal death, and lower neonatal Apgar score in first minute during the first week

  8. Function curve of the membranes that regulate amniotic fluid volume in sheep. (United States)

    Faber, Job; Anderson, Debra; Hohimer, Roger; Yang, Qin; Giraud, George; Davis, Lowell


    Seven singleton 120-day fetal lambs were prepared with a shunt from the lung to the gastric end of the esophagus, a bladder catheter, and multiple amniotic fluid and vascular catheters. The urachus was ligated. Beginning 7 days later, amniotic fluid volumes were determined by drainage, followed by replacement with 1 liter of lactated Ringer (LR) solution. Urine flow into the amnion was measured continuously. In 14 of 27 experiments, amniotic fluid volumes were determined again 2 days after the inflow into the amnion had consisted of urine only and in 13 experiments after the inflow of urine had been supplemented by an intraamniotic infusion of LR solution. Intramembranous absorption was calculated from the inflows and the changes in volume between the beginning and end of each experiment. The relations between absorption rate and amniotic fluid volume, the "function curves," were highly individual. Urine production during the infusion of LR solution did not decrease, fetal plasma renin activity decreased (P amniotic fluid volume increased by 140% [SE (27%), P amniotic fluid per day. During the infusion of LR solution, the increase in the rate of absorption matched the rate of infusion (both in ml/h), with a regression coefficient of 0.75 (P amniotic fluid volumes, volume is not limited by the absorptive capacity of the amniochorion, and, at least in these preparations, the position of the function curve and not the natural rate of inflow was the major determinant of resting amniotic fluid volume.

  9. Assessing the nature of lipid raft membranes.

    Directory of Open Access Journals (Sweden)

    Perttu S Niemelä


    Full Text Available The paradigm of biological membranes has recently gone through a major update. Instead of being fluid and homogeneous, recent studies suggest that membranes are characterized by transient domains with varying fluidity. In particular, a number of experimental studies have revealed the existence of highly ordered lateral domains rich in sphingomyelin and cholesterol (CHOL. These domains, called functional lipid rafts, have been suggested to take part in a variety of dynamic cellular processes such as membrane trafficking, signal transduction, and regulation of the activity of membrane proteins. However, despite the proposed importance of these domains, their properties, and even the precise nature of the lipid phases, have remained open issues mainly because the associated short time and length scales have posed a major challenge to experiments. In this work, we employ extensive atom-scale simulations to elucidate the properties of ternary raft mixtures with CHOL, palmitoylsphingomyelin (PSM, and palmitoyloleoylphosphatidylcholine. We simulate two bilayers of 1,024 lipids for 100 ns in the liquid-ordered phase and one system of the same size in the liquid-disordered phase. The studies provide evidence that the presence of PSM and CHOL in raft-like membranes leads to strongly packed and rigid bilayers. We also find that the simulated raft bilayers are characterized by nanoscale lateral heterogeneity, though the slow lateral diffusion renders the interpretation of the observed lateral heterogeneity more difficult. The findings reveal aspects of the role of favored (specific lipid-lipid interactions within rafts and clarify the prominent role of CHOL in altering the properties of the membrane locally in its neighborhood. Also, we show that the presence of PSM and CHOL in rafts leads to intriguing lateral pressure profiles that are distinctly different from corresponding profiles in nonraft-like membranes. The results propose that the functioning of

  10. Membraner

    DEFF Research Database (Denmark)

    Bach, Finn


    Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner......Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner...

  11. On the damped oscillations of an elastic quasi-circular membrane in a two-dimensional incompressible fluid

    CERN Document Server

    Afonso, Marco Martins; Nicoud, Franck


    We propose a procedure - partly analytical and partly numerical - to find the frequency and the damping rate of the small-amplitude oscillations of a massless elastic capsule immersed in a two-dimensional viscous incompressible fluid. The unsteady Stokes equations for the stream function are decomposed onto normal modes for the angular and temporal variables, leading to a fourth-order linear ordinary differential equation in the radial variable. The forcing terms are dictated by the properties of the membrane, and result into jump conditions at the interface between the internal and external media. The equation can be solved numerically, and an excellent agreement is found with a fully-computational approach we developed in parallel. Comparisons are also shown with the results available in the scientific literature for drops, and a model based on the concept of embarked fluid is presented, which allows for a good representation of the results and a consistent interpretation of the underlying physics.


    Khanafer, Khalil; Cook, Keith; Marafie, Alia


    A numerical study was conducted to analyze fluid flow within hollow fiber membranes of the artificial lungs. The hollow fiber bundle was approximated using a porous media model. In addition, the transport equations were solved using the finite-element formulation based on the Galerkin method of weighted residuals. Comparisons with previously published work on the basis of special cases were performed and found to be in excellent agreement. A Newtonian viscous fluid model for the blood was used. Different flow models for porous media, such as the Brinkman-extended Darcy model, Darcy's law model, and the generalized flow model, were considered. Results were obtained in terms of streamlines, velocity vectors, and pressure distribution for various Reynolds numbers and Darcy numbers. The results from this investigation showed that the pressure drop across the artificial lung device increased with an increase in the Reynolds number. In addition, the pressure drop was found to increase significantly for small Darcy numbers.

  13. Asymmetric heat transfer from nanoparticles in lipid bilayers (United States)

    Potdar, Dipti; Sammalkorpi, Maria


    Here, we use molecular dynamics simulations to characterize the heat transfer properties of lipid bilayer - gold nanoparticle systems in which the nanoparticle acts as a heat source. The focus is on dipalmitoylphosphatidylcholine (DPPC) lipid bilayers and thiolated alcohol and alkyl functionalized nanoparticles as prototype hydrophilic and hydrophobic nanoparticles. We find hydrophilic nanoparticles which are partly in contact with the surrounding water environment are more efficient in transferring heat to the system than hydrophobic ones which reside surrounded by the membrane. This is because of the hydrogen bonding capability of the hydroxy pentanethiol and the more efficient heat conductivity through water than the lipid bilayer. Additionally, we find the heat conductance is strongly asymmetric and has a discontinuity between the bilayer leaflets. In total, the findings provide understanding on heat transport from localized heat sources in lipid bilayers and could bear significance, e.g., in engineering and controlling photoactivated triggering of liposomal systems.

  14. Differential dynamics of the serotonin1A receptor in membrane bilayers of varying cholesterol content revealed by all atom molecular dynamics simulation. (United States)

    Patra, Swarna M; Chakraborty, Sudip; Shahane, Ganesh; Prasanna, Xavier; Sengupta, Durba; Maiti, Prabal K; Chattopadhyay, Amitabha


    The serotonin1A receptor belongs to the superfamily of G protein-coupled receptors (GPCRs) and is a potential drug target in neuropsychiatric disorders. The receptor has been shown to require membrane cholesterol for its organization, dynamics and function. Although recent work suggests a close interaction of cholesterol with the receptor, the structural integrity of the serotonin1A receptor in the presence of cholesterol has not been explored. In this work, we have carried out all atom molecular dynamics simulations, totaling to 3 μs, to analyze the effect of cholesterol on the structure and dynamics of the serotonin1A receptor. Our results show that the presence of physiologically relevant concentration of membrane cholesterol alters conformational dynamics of the serotonin1A receptor and, on an average lowers conformational fluctuations. Our results show that, in general, transmembrane helix VII is most affected by the absence of membrane cholesterol. These results are in overall agreement with experimental data showing enhancement of GPCR stability in the presence of membrane cholesterol. Our results constitute a molecular level understanding of GPCR-cholesterol interaction, and represent an important step in our overall understanding of GPCR function in health and disease.

  15. Pair collisions of fluid-filled elastic capsules in shear flow: Effects of membrane properties and polymer additives (United States)

    Pranay, Pratik; Anekal, Samartha G.; Hernandez-Ortiz, Juan P.; Graham, Michael D.


    The dynamics and pair collisions of fluid-filled elastic capsules during Couette flow in Newtonian fluids and dilute solutions of high-molecular weight (drag-reducing) polymers are investigated via direct simulation. Capsule membranes are modeled using either a neo-Hookean constitutive model or a model introduced by Skalak et al. ["Strain energy function of red blood-cell membranes," Biophys. J. 13, 245 (1973)], which includes an energy penalty for area changes. This model was developed to capture the elastic properties of red blood cells. Polymer molecules are modeled as bead-spring trimers with finitely extensible nonlinearly elastic springs; parameters were chosen to loosely approximate 4000 kDa poly(ethylene oxide). Simulations are performed with a novel Stokes flow formulation of the immersed boundary method for the capsules, combined with Brownian dynamics for the polymer molecules. The results for isolated capsules in shear indicate that at the very low concentrations considered here, polymers have a little effect on the capsule shape. In the case of pair collisions, the effect of polymer is strongly dependent on the elastic properties of the capsules' membranes. For neo-Hookean capsules or for Skalak capsules with only a small penalty for area change, the net displacement in the gradient direction after collision is virtually unaffected by the polymer. For Skalak capsules with a large penalty for area change, polymers substantially decrease the net displacement when compared to the Newtonian case and the effect is enhanced upon increasing the polymer concentration. The differences between the polymer effects in the various cases are associated with the extensional flow generated in the region between the capsules as they leave the collision. The extension rate is highest when there is a strong resistance to a change in the membrane area and is substantially decreased in the presence of polymer.

  16. Renormalization of the tension and area expansion modulus in fluid membranes. (United States)

    Marsh, D


    Renormalization of the membrane tension and elastic area expansion modulus by thermally induced bending fluctuations is treated in terms of the formalism of Brochard, De Gennes, and Pfeuty (J. de Phys. (France). 37:1099-1104, 1976). The dependence of the renormalized tension on the bare membrane tension parallels the dependence on the fractional area extension of giant vesicles found experimentally by Evans and Rawicz (Physiol. Rev. Lett. 64:2094-2097, 1990), and suggests conditions for molecular dynamics simulations with membrane patches of limited size that might best represent the properties of macroscopic vesicles.

  17. The geometry of the ionic chànnel lumen formed by alpha-latroinsectotoxin from black widow spider venom in the bilayer lipid membranes. (United States)

    Shatursky, Oleg Ya; Volkova, Tatyana M; Himmelreich, Nina H; Grishin, Eugene V


    The dependence of single channel conductance formed by alpha-latroinsectotoxin (alpha-LIT) from black widow spider venom in the planar phospholipid membrane on the hydrodynamic radii of different nonelectrolytes allowed to determine the geometry of alpha-LIT water lumen. It was found that the cis- and trans-entrances of alpha-LIT channel had the same effective radii of 0.55-0.58 nm. Relatively small conductance of alpha-LIT channel (23.5+3.7 pS) in a symmetrical membrane bathing solution of 100 mM KCl (pH 7.4) may result from the constriction inside the channel with apparent radius of 0.37 nm located 32.5% of channel length away from the cis-entrance.

  18. Physics of biological membranes (United States)

    Mouritsen, Ole G.

    The biological membrane is a complex system consisting of an aqueous biomolecular planar aggregate of predominantly lipid and protein molecules. At physiological temperatures, the membrane may be considered a thin (˜50Å) slab of anisotropic fluid characterized by a high lateral mobility of the various molecular components. A substantial fraction of biological activity takes place in association with membranes. As a very lively piece of condensed matter, the biological membrane is a challenging research topic for both the experimental and theoretical physicists who are facing a number of fundamental physical problems including molecular self-organization, macromolecular structure and dynamics, inter-macromolecular interactions, structure-function relationships, transport of energy and matter, and interfacial forces. This paper will present a brief review of recent theoretical and experimental progress on such problems, with special emphasis on lipid bilayer structure and dynamics, lipid phase transitions, lipid-protein and lipid-cholesterol interactions, intermembrane forces, and the physical constraints imposed on biomembrane function and evolution. The paper advocates the dual point of view that there are a number of interesting physics problems in membranology and, at the same time, that the physical properties of biomembranes are important regulators of membrane function.

  19. Alpha-tocopherol inhibits pore formation in the oxidized bilayers

    CERN Document Server

    Boonnoy, Phansiri; Wong-ekkabut, Jirasak


    In biological membranes, alpha-tocopherols ({\\alpha}-toc; vitamin E) protect polyunsaturated lipids from free radicals. Although the interactions of {\\alpha}-toc with non-oxidized lipid bilayers have been studied, their on oxidized bilayers remain unknown. In this study, atomistic molecular dynamics (MD) simulations of oxidized lipid bilayers were performed with varying concentrations of {\\alpha}-toc. Bilayers with 1-palmitoyl-2-lauroyl-sn-glycero-3-phosphocholine (PLPC) lipids and its aldehyde derivatives at 1:1 ratio were studied. Our simulations show that oxidized lipids self-assemble into aggregates with a water pore rapidly developing across the lipid bilayer. The free energy of transporting an {\\alpha}-toc molecule in a lipid bilayer suggests that {\\alpha}-tocs can passively adsorb into the bilayer. When {\\alpha}-toc molecules were present at low concentrations in bilayers containing oxidized lipids, the formation of water pores was slowed down. At high {\\alpha}-toc concentra-tions, no pores were observ...

  20. A molecular dynamics model of rhodamine-labeled phospholipid incorporated into a lipid bilayer (United States)

    Kyrychenko, Alexander


    Phospholipids, labeled covalently by a fluorescent dye, are commonly applied in membrane biophysics. In this work, a molecular dynamics model of sulforhodamine attached covalently to a headgroup of 1,2-dipalmitoyl- sn-glycero-3-phosphoethanolamine is developed. It is found that the incorporation of rhodamine-labeled phospholipids into a DPPC bilayer at the low concentration results in small perturbation of the bilayer. In the dye-labeled membrane, the sulforhodamine moiety binds favorably to a polar membrane interface, forming the tilt angle 44° ± 8° to the bilayer normal. The deep location and binding of a bulk sulforhodamine fluorophore lead, therefore, to some 'softening' of the membrane structure.

  1. Pseudomonas aeruginosa outer membrane vesicles triggered by human mucosal fluid and lysozyme can prime host tissue surfaces for bacterial adhesion

    Directory of Open Access Journals (Sweden)

    Matteo Maria Emiliano Metruccio


    Full Text Available Pseudomonas aeruginosa is a leading cause of human morbidity and mortality that often targets epithelial surfaces. Host immunocompromise, or the presence of indwelling medical devices, including contact lenses, can predispose to infection. While medical devices are known to accumulate bacterial biofilms, it is not well understood why resistant epithelial surfaces become susceptible to P. aeruginosa. Many bacteria, including P. aeruginosa, release Outer Membrane Vesicles (OMVs in response to stress that can fuse with host cells to alter their function. Here, we tested the hypothesis that mucosal fluid can trigger OMV release to compromise an epithelial barrier. This was tested using tear fluid and corneal epithelial cells in vitro and in vivo. After 1 h both human tear fluid, and the tear component lysozyme, greatly enhanced OMV release from P. aeruginosa strain PAO1 compared to PBS controls (~100 fold. TEM and SDS-PAGE showed tear fluid and lysozyme-induced OMVs were similar in size and protein composition, but differed from biofilm-harvested OMVs, the latter smaller with fewer proteins. Lysozyme-induced OMVs were cytotoxic to human corneal epithelial cells in vitro and murine corneal epithelium in vivo. OMV exposure in vivo enhanced Ly6G/C expression at the corneal surface, suggesting myeloid cell recruitment, and primed the cornea for bacterial adhesion (~4-fold, P < 0.01. Sonication disrupted OMVs retained cytotoxic activity, but did not promote adhesion, suggesting the latter required OMV-mediated events beyond cell killing. These data suggest that mucosal fluid induced P. aeruginosa OMVs could contribute to loss of epithelial barrier function during medical device-related infections.

  2. SAXS investigations on lipid membranes under osmotic stress

    Energy Technology Data Exchange (ETDEWEB)

    Rubim, R.L.; Vieira, V.; Gerbelli, B.B.; Teixeira da Silva, E.R.; Oliveira, C.L.P.; Oliveira, E.A. [Universidade de Sao Paulo (USP), Sao Paulo, SP (Brazil)


    Full text: In this work we, experimentally, investigate the interactions between lipid bilayers. A structural characterization is performed by small angle x-ray scattering (SAXS) on multilamellar systems under known osmotic pressure. Changes in the composition of membranes can modify their mechanical properties and structural parameters, like the flexibility of these membranes, which plays a key role on the determination of the tridimensional organization of bilayers. The membranes are composed of soya lecithin, where the major component is DPPC (Dipalmitoylphosphatidylcholine), and fatty acids are incorporated to the membrane in different concentrations, in order to turn the membrane more fluid. The membranes are inserted in a solution of PVP [poly(vinyl-pyrrolidone) - 40000] and the polymer will apply an osmotic pressure on them. The osmotic pressure is controlled by preparing PVP solutions of desired composition and, as we know the concentration of polymer in solution, we can obtain the intensity of the osmotic pressure. SAXS experiments were done in order to determine the distance between the bilayer. From the position of the Bragg peaks, the lamellar periodicity (the thickness of the membranes plus their distance of separation) was determined. Using theoretical model for the form and structure factors we fitted those experimental data and determined the thickness of the membranes. The distance between the membranes was controlled by the osmotic pressure (P) applied to the membranes and, for a given pressure, we determine the distance between the bilayers (a) on equilibrium. The experimental curve P(a) is theoretically described by the different contributions from van der Waals, hydration and fluctuation forces. From the fitting of experimental curves, relevant parameters characterizing the strength of the different interactions are obtained, such as Hamaker and rigidity constant [2, 3]. We observe that the separation between the bilayers on equilibrium is

  3. Reator de membrana enzimático e fluidos supercríticos: associação de processos Enzymatic membrane reactor and supercritical fluids: process association

    Directory of Open Access Journals (Sweden)

    Juliana Maria Leite Nóbrega de Moura


    Full Text Available The aim of this work is to present the new applications of membrane technology in fat and oil processing, with emphasis on development and applications of the enzymatic membrane reactor and its association with extraction and purification technology by supercritical fluids (SCF. Combining the extraction by SCFs and the separation by membranes allows the integration of extractions reactions with selective separation by membranes through filtration of the supercritical mixture (SCF + extracted solutes. This association provides important energy savings regarding the SCF recompression costs.

  4. Fluid and solute transfer characteristics in a dialyzer with a high-performance membrane. (United States)

    Mineshima, Michio


    To date, many types of high-flux dialyzers with high-performance membranes have been developed. They seem to have a higher internal filtration flow rate (Q(IF)) because of a higher ultrafiltration coefficient. In the health reimbursement system in Japan, commercially available dialyzers are categorized into five types based on their β(2)-microglobulin clearance (K(BMG)) values. Although the K(BMG) value was not correlated strongly with the Q(IF) value for 26 types of commercially available dialyzers, almost all the dialyzers with a higher Q(IF) value had a higher KBMG value. These dialyzers seem to have a higher convective transport because of internal filtration in addition to diffusive transport. We measured the blood flow velocity in a cross-sectional plane of the dialyzer using pulse Doppler ultra-sonography to evaluate QIF. It is a useful method for the bedside monitoring because it is noninvasive to the patient and produces reliable data with a higher reproducibility. On the other hand, membrane fouling occurs more easily in higher Q(IF) dialyzers, compared with conventional dialyzers, because of the higher degree of membrane fouling. Internal filtration-enhanced hemodialysis (IFEHD) using these dialyzers, therefore, has the advantage of increasing solute removal efficiency by enhancing convective transport and the simultaneous disadvantage of decreasing solute removal efficiency by causing membrane fouling. Thus, IFEHD treatment should be performed using a dialyzer with a high-performance membrane to ensure that the advantage is superior to the disadvantage.

  5. The Fluid/Gravity Correspondence: a new perspective on the Membrane Paradigm

    CERN Document Server

    Hubeny, Veronika E


    This talk gives an overview of the recently-formulated Fluid/Gravity correspondence, which was developed in the context of gauge/gravity duality. Mathematically, it posits that Einstein's equations (with negative cosmological constant) in d+1 dimensions capture the (generalized) Navier-Stokes equations in d dimensions. Given an arbitrary fluid dynamical solution, we can systematically construct a corresponding asymptotically AdS black hole spacetime with a regular horizon whose properties mimic that of the fluid flow. Apart from an overview of this construction, we describe some of its applications. The presentation is intended for a broad audience of relativists, and does not assume prior knowledge of string theory or gauge/gravity duality.

  6. Amoeboid Swimming: A Generic Self-Propulsion of Cells in Fluids by Means of Membrane Deformations (United States)

    Farutin, Alexander; Rafaï, Salima; Dysthe, Dag Kristian; Duperray, Alain; Peyla, Philippe; Misbah, Chaouqi


    Microorganisms, such as bacteria, algae, or spermatozoa, are able to propel themselves forward thanks to flagella or cilia activity. By contrast, other organisms employ pronounced changes of the membrane shape to achieve propulsion, a prototypical example being the Eutreptiella gymnastica. Cells of the immune system as well as dictyostelium amoebas, traditionally believed to crawl on a substratum, can also swim in a similar way. We develop a model for these organisms: the swimmer is mimicked by a closed incompressible membrane with force density distribution (with zero total force and torque). It is shown that fast propulsion can be achieved with adequate shape adaptations. This swimming is found to consist of an entangled pusher-puller state. The autopropulsion distance over one cycle is a universal linear function of a simple geometrical dimensionless quantity A/V2/3 (V and A are the cell volume and its membrane area). This study captures the peculiar motion of Eutreptiella gymnastica with simple force distribution.

  7. Primordial membranes

    DEFF Research Database (Denmark)

    Hanczyc, Martin M; Monnard, Pierre-Alain


    Cellular membranes, which are self-assembled bilayer structures mainly composed of lipids, proteins and conjugated polysaccharides, are the defining feature of cell physiology. It is likely that the complexity of contemporary cells was preceded by simpler chemical systems or protocells during the...

  8. In-situ assessment of biofilm formation in submerged membrane system using optical coherence tomography and computational fluid dynamics

    KAUST Repository

    Fortunato, Luca


    This paper introduces a novel approach to study the biofouling development on gravity driven submerged membrane bioreactor (SMBR). The on-line monitoring of biofilm formation on a flat sheet membrane was conducted non-destructively using optical coherence tomography (OCT), allowing the in-situ investigation of the biofilm structure for 43 d. The OCT enabled to obtain a time-lapse of biofilm development on the membrane under the continuous operation. Acquired real-time information on the biofilm structure related to the change in the flux profile confirming the successful monitoring of the dynamic evolution of the biofouling layer. Four different phases were observed linking the permeate flux with the change of biofilm morphology. In particular, a stable flux of 2.1±0.1 L/m2 h was achieved with the achievement of steady biofilm morphology after 30 d of operation. Biofilm descriptors, such as thickness, biofilm area, macro-porosity and roughness (absolute and relative), were calculated for each OCT acquired scans. Interestingly, relative roughness was correlated with the flux decrease. Furthermore, the precise biofilm morphology obtained from the OCT scans was used in computational fluid dynamics (CFD) simulation to better understand the role of biofilm structure on the filtration mechanism. © 2016 Elsevier B.V.

  9. Simulation of the Effect of a Baffle Structure on Membrane Efficiency Using Computational Fluid Dynamics during the Clarification of Pomegranate Juice

    Directory of Open Access Journals (Sweden)

    Reza Sharifanfar


    Full Text Available Background and Objectives: Pomegranate juice (PJ contains large particles that stick to evaporator walls causing off flavors in the concentrate due to burning. Microfiltration is used to clarify PJ. Fouling is a limiting phenomenon that can prevent the industrialization of membrane clarification. Changes in the geometry of the membrane module such as using baffles are useful to decrease this problem. Computational fluid dynamics (CFD is a powerful numerical tool used in modeling membrane processing. Materials and Methods: The effect of baffle geometry on the efficiency of membrane clarification of pomegranate juice in a flat-sheet module was simulated using computational fluid dynamics (CFD. The geometry of the membrane unit was plotted and meshed with Gambit software, and was solved using FLUENT software. A two-dimensional double-precision method at steady state was selected to simulate the membrane process. The convective terms were discretized with a standard first-order upwind scheme in computational solution. The RNG k-  model was used due to its high accuracy in eddy flows with a low Reynolds number. The effects on the process performance of the number of baffles, their angle and the distance between the baffles and the membrane surface were evaluated. Results: The results showed that the configuration with the feed-channel height of 2 cm, the baffle angle of 90o and the distance between the membrane surface and baffles of 2 mm had maximum permeate flux. Conclusions: Reducing the distance between the baffles and the membrane surface increased the permeate flux due to create an eddy flow near the membrane surface in the flat-sheet module and reduced the total and cake-layer resistances. Keywords: Baffle, Computational fluid dynamics, Juice, Membrane, Pomegranate

  10. Meet me on the other side: trans-bilayer modulation of a model voltage-gated ion channel activity by membrane electrostatics asymmetry.

    Directory of Open Access Journals (Sweden)

    Loredana Mereuta

    Full Text Available While it is accepted that biomembrane asymmetry is generated by proteins and phospholipids distribution, little is known about how electric changes manifested in a monolayer influence functional properties of proteins localized on the opposite leaflet. Herein we used single-molecule electrophysiology and investigated how asymmetric changes in the electrostatics of an artificial lipid membrane monolayer, generated oppositely from where alamethicin--a model voltage-gated ion channel--was added, altered peptide activity. We found that phlorizin, a membrane dipole potential lowering amphiphile, augmented alamethicin activity and transport features, whereas the opposite occurred with RH-421, which enhances the monolayer dipole potential. Further, the monolayer surface potential was decreased via adsorption of sodium dodecyl sulfate, and demonstrated that vectorial modification of it also affected the alamethicin activity in a predictive manner. A new paradigm is suggested according to which asymmetric changes in the monolayer dipole and surface potential extend their effects spatially by altering the intramembrane potential, whose gradient is sensed by distantly located peptides.

  11. Fluid domain patterns in free-standing membranes captured on a solid support

    DEFF Research Database (Denmark)

    Bhartia, Tripta; Husen, Peter Rasmussen; Ipsen, John Hjort


    membrane, where lateral domain structures are kinetically trapped. High-resolution images of domain patterns in the liquid-ordered (lo) and liquid-disordered (ld) co-existence region in the phase-diagram of ternary lipid mixtures are revealed by atomic force microscopy (AFM) scans of the patches...

  12. Structural Effects of Small Molecules on Phospholipid Bilayers Investigated by Molecular Simulations

    CERN Document Server

    Lee, B W; Sum, A K; Vattulainen, I; Patra, M; Karttunen, M; Lee, Bryan W; Faller, Roland; Sum, Amadeu K; Vattulainen, Ilpo; Patra, Michael; Karttunen, Mikko


    We summarize and compare recent Molecular Dynamics simulations on the interactions of dipalmitoylphosphatidylcholine (DPPC) bilayers in the liquid crystalline phase with a number of small molecules including trehalose, a disaccharide of glucose, alcohols, and dimethylsulfoxide (DMSO). The sugar molecules tend to stabilize the structure of the bilayer as they bridge adjacent lipid headgroups. They do not strongly change the structure of the bilayer. Alcohols and DMSO destabilize the bilayer as they increase its area per molecule in the bilayer plane and decrease the order parameter. Alcohols have a stronger detrimental effect than DMSO. The observables which we compare are the area per molecule in the plane of the bilayer, the membrane thickness, and the NMR order parameter of DPPC hydrocarbon tails. The area per molecule and the order parameter are very well correlated whereas the bilayer thickness is not necessarily correlated with them.

  13. Evaporation-Induced Buckling and Fission of Microscale Droplet Interface Bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Boreyko, Jonathan B [ORNL; Mruetusatorn, Prachya [ORNL; Sarles, Stephen A [ORNL; Retterer, Scott T [ORNL; Collier, Pat [ORNL


    Droplet interface bilayers (DIBs) are a robust platform for studying synthetic cellular membranes; however, to date no DIBs have been produced at cellular length scales. Here, we create microscale droplet interface bilayers ( DIBs) at the interface between aqueous femtoliter-volume droplets within an oil-filled microfluidic channel. The uniquely large area-to-volume ratio of the droplets results in strong evaporation effects, causing the system to transition through three distinct regimes. First, the two adjacent droplets shrink into the shape of a single spherical droplet, where an augmented lipid bilayer partitions two hemi-spherical volumes. In the second regime, the combined effects of the shrinking monolayers and growing bilayer force the confined bilayer to buckle to conserve its mass. Finally, at a bending moment corresponding to a critical shear stress, the buckling bilayer fissions a vesicle to regulate its shape and stress. The DIBs produced here enable evaporation-induced bilayer dynamics reminiscent of endo- and exocytosis in cells.

  14. Less is more: removing membrane attachments stiffens the RBC cytoskeleton

    Energy Technology Data Exchange (ETDEWEB)

    Gov, Nir S [Department of Chemical Physics, The Weizmann Institute of Science, PO Box 26, Rehovot 76100 (Israel)


    The polymerized network of the cytoskeleton of the red-blood cell (RBC) contains different protein components that maintain its overall integrity and attachment to the lipid bilayer. One of these key components is the band 3-ankyrin complex that attaches the spectrin filaments to the fluid bilayer. Defects in this particular component result in the shape transformation called spherocytosis, through the shedding of membrane nano-vesicles. We show here that this transition and membrane shedding can be explained through the increased stiffness of the network when the band 3-ankyrin complexes are removed. ATP-induced transient dissociations lead to network softening, which offsets the stiffening to some extent, and causes increased fragility of these mutant cells, as is observed.

  15. Modeling and simulation of membrane separation process using computational fluid dynamics

    Directory of Open Access Journals (Sweden)

    Kambiz Tahvildari


    Full Text Available Separation of CO2 from air was simulated in this work. The considered process for removal of CO2 was a hollow-fiber membrane contactor and an aqueous solution of 2-amino-2-metyl-1-propanol (AMP as absorbent. The model was developed based on mass transfer as well as chemical reaction for CO2 and solvent in the contactor. The equations of model were solved using finite element method. Simulation results were compared with experimental data, and good agreement was observed. The results revealed that increasing solvent velocity enhances removal of CO2 in the hollow-fiber membrane contactor. Moreover, it was found that counter-current process mode is more favorable to achieve the highest separation efficiency.

  16. Computational fluid dynamics modelling of a polymer electrolyte membrane fuel cell under transient automotive operations


    Choopanya, Pattarapong


    A polymer electrolyte membrane (PEM) fuel cell is probably the most promising technology that will replace conventional internal combustion engines in the near future. As a primary power source for an automobile, the transient performance of a PEM fuel cell is of prime importance. In this thesis, a comprehensive, three-dimensional, two-phase, multi-species computational fuel cell dynamics model is developed in order to investigate the effect of flow-field design on the magnitude of current ov...

  17. Vaginal Fluid hCG Levels for Detecting Premature Rupture of Membranes

    Directory of Open Access Journals (Sweden)

    T. Artimani


    Full Text Available The aim of this study was to determine whether there is any different in hCG levels of vaginal fluids from normal pregnant women, women with confirmed PROM and with suspected PROM. After irrigating the posterior vaginal fornix with 3 ml of sterile saline and procuring vaginal washing , we measured hCG levels. Samples were analyzed from 38 normal pregnant women , 31 women with confirmed PROM , 28 women with suspected PROM. The mean of vaginal fluid hCG levels of normal pregnant women were 11.2 (0.00-98.6 mIu/ml. That of women with PROM was 140.2 (0.5 , 390.1 and that of women with suspected PROM was 21.5 (0.0 , 143.9. Sensivity was 80%, specility 83% , positive predictive value 73.7% , negative predictive value 88.1% , using a cut-value 20. It was concluded that the hCG level in vaginal fluid is a marker of PROM during third trimester.

  18. A single-channel method for evaluation of very magnitudes of Ca2+ ion fluxes through epsilon4/zeta1 N-methyl-D-aspartate receptor channels in bilayer lipid membranes. (United States)

    Wakabayashi, M; Hirano, A; Sugawara, M; Uchino, S; Nakajima-Iijima, S


    A single-channel method for evaluating agonist selectivity in terms of the very number of Ca2+ ions passed through the epsilon4/zeta1 N-methyl-D-aspartate (NMDA) receptor ion channel in bilayer lipid membranes (BLMs) is described. The number of Ca2+ passed through the single-channel was obtained from single-channel recordings in a medium where the primary permeant ion is Ca2+. The recombinant epsilon4/zeta1 NMDA channel was partially purified from Chinese hamster ovary cells expressing the channel and incorporated in BLMs formed by the tip-dip method. It was found that the epsilon4/zeta1 channel in BLMs is permeable to Ca2+ and Na+, but the number of Ca2+ passed through the channel is much fewer than that of Na+. The integrated Ca2+ currents induced by three typical agonists NMDA, L-glutamate and L-CCG-IV were obtained at concentration of 50 microM, where the integrated currents for all the agonists reached their saturated values. The integrated Ca2+ currents obtained are (3.1+/-0.21) x 10(-13) C/s for NMDA, (4.6+/-0.31) x 10(-13) C/s for L-glutamate and (5.7+/-0.25) x 10(-13) C/s for L-CCG-IV, respectively, suggesting that the three kinds of agonists have different efficacies to induce permeation of Ca2+. The range of the agonist selectivity thus obtained is much narrower than that of binding affinities for the NMDA receptors from rat brain. The present method is able to detect Ca2+ permeation with a detection limit of approximately 10(5) Ca2+ ions/s.

  19. Removal of the mechanoprotective influence of the cytoskeleton reveals PIEZO1 is gated by bilayer tension (United States)

    Cox, Charles D.; Bae, Chilman; Ziegler, Lynn; Hartley, Silas; Nikolova-Krstevski, Vesna; Rohde, Paul R.; Ng, Chai-Ann; Sachs, Frederick; Gottlieb, Philip A.; Martinac, Boris


    Mechanosensitive ion channels are force-transducing enzymes that couple mechanical stimuli to ion flux. Understanding the gating mechanism of mechanosensitive channels is challenging because the stimulus seen by the channel reflects forces shared between the membrane, cytoskeleton and extracellular matrix. Here we examine whether the mechanosensitive channel PIEZO1 is activated by force-transmission through the bilayer. To achieve this, we generate HEK293 cell membrane blebs largely free of cytoskeleton. Using the bacterial channel MscL, we calibrate the bilayer tension demonstrating that activation of MscL in blebs is identical to that in reconstituted bilayers. Utilizing a novel PIEZO1-GFP fusion, we then show PIEZO1 is activated by bilayer tension in bleb membranes, gating at lower pressures indicative of removal of the cortical cytoskeleton and the mechanoprotection it provides. Thus, PIEZO1 channels must sense force directly transmitted through the bilayer.

  20. Focal junctions retard lateral movement and disrupt fluid phase connectivity in the plasma membrane

    DEFF Research Database (Denmark)

    Vind-Kezunovic, D.; Wojewodzka, U.; Gniadecki, R.


    containing liquid-ordered (L-o) lipids. Indeed, values of maximal fluorescence recovery after photobleaching revealed that the long-range mobility of cholera toxin B subunit (CTB, marker of L-o) was similar to 1.5-fold retarded within the focal junctions compared to the surrounding membrane. However, 1......,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI-C-18:0), which specifically partitions to the liquid-disordered (L-d), non-raft phase, was also enriched in focal junctions and its mobility was slightly retarded. Cross-linking of GM(1) by CTB or raft aggregation by methyl...

  1. Direct computation of two-phase icosahedral equilibria of lipid bilayer vesicles (United States)

    Zhao, Siming; Healey, Timothy; Li, Qingdu


    Correctly formulated continuum models for lipid-bilayer membranes present a significant challenge to computational mechanics. In particular, the mid-surface behavior is that of a 2-dimensional fluid, while the membrane resists bending much like an elastic shell. Here we consider a well-known Helfrich-Cahn-Hilliard model for two-phase lipid-bilayer vesicles, incorporating mid-surface fluidity, curvature elasticity and a phase field. We present a systematic approach to the direct computation of vesical configurations possessing icosahedral symmetry, which have been observed in experiment and whose mathematical existence has recently been established. We first introduce a radial-graph formulation to overcome the difficulties associated with fluidity within a conventional Lagrangian description. We use the so-called subdivision surface finite element method combined with an icosahedral-symmetric mesh. The resulting discrete equations are well-conditioned and inherit equivariance properties under a representation of the icosahedral group. We use group-theoretic methods to obtain a reduced problem that captures all icosahedral-symmetric solutions of the full problem. Finally we explore the behavior of our reduced model, varying numerous physical parameters present in the mathematical model.

  2. Functional diversity and dynamics of bacterial communities in a membrane bioreactor for the treatment of metal-working fluid wastewater. (United States)

    Grijalbo, Lucía; Garbisu, Carlos; Martín, Iker; Etxebarria, Javier; Gutierrez-Mañero, F Javier; Lucas Garcia, Jose Antonio


    An extensive microbiological study has been carried out in a membrane bioreactor fed with activated sludge and metal-working fluids. Functional diversity and dynamics of bacterial communities were studied with different approaches. Functional diversity of culturable bacterial communities was studied with different Biolog™ plates. Structure and dynamics of bacterial communities were studied in culturable and in non-culturable fractions using a 16S rRNA analysis. Among the culturable bacteria, Alphaproteobacteria and Gammaproteobacteria were the predominant classes. However, changes in microbial community structure were detected over time. Culture-independent analysis showed that Betaproteobacteria was the most frequently detected class in the membrane bioreactor (MBR) community with Zoogloea and Acidovorax as dominant genera. Also, among non-culturable bacteria, a process of succession was observed. Longitudinal structural shifts observed were more marked for non-culturable than for culturable bacteria, pointing towards an important role in the MBR performance. Microbial community metabolic abilities assessed with Biolog™ Gram negative, Gram positive and anaerobic plates also showed differences over time for Shannon's diversity index, kinetics of average well colour development, and the intensely used substrates by bacterial community in each plate.

  3. The nanoscience behind the art of in-meso crystallization of membrane proteins. (United States)

    Zabara, Alexandru; Meikle, Thomas G; Newman, Janet; Peat, Thomas S; Conn, Charlotte E; Drummond, Calum J


    The structural changes occurring at the nanoscale level within the lipid bilayer and driving the in-meso formation of large well-diffracting membrane protein crystals have been uniquely characterized for a model membrane protein, intimin. Importantly, the order to order transitions taking place within the bilayer and the lipidic nanostructures required for crystal growth have been shown to be general, occurring for both the cubic and the sponge mesophase crystallization pathways. For the first time, a transient fluid lamellar phase has been observed and unambiguously assigned for both crystallization pathways, present at the earliest stages of protein crystallogenesis but no longer observed once the crystals surpass the size of the average lyotropic liquid crystalline domain. The reported time-resolved structural investigation provides a significantly improved and general understanding of the nanostructural changes taking place within the mesophase during in-meso crystallization which is a fundamental advance in the enabling area of membrane protein structural biology.

  4. The correlation between histologic placentitis and amnionitis and the amnioniotic fluid's inflammatory cytokines in case of spontaneous pre-term labor with intact membrane

    Directory of Open Access Journals (Sweden)

    Agus Abadi


    Full Text Available Pre-term labor is presumed to result from spreading of lower genital infection to upper part, subsequently to decidual and choioamniotic tissues. Host response to this injury include the expression of protein which is responsible to the inflammatory reactions. The expression of the inflammatory cytokines such as IL-1β, IL-6, IL-8 and TNF-α increase in case of infection.These cytokines may play an essential role in the pathophysiology of spontaneous pretem labor with intact membrane.An observational analytic cohort study was caried out on cases of spontaneous pre-tefln labor with intact membrane. The objectives of this study are to examine the relationship between l the histologic amnionitis and placentitis and the incidence of preterm delivery,2 the expression of amniotic fluid's IL-1β, IL-6, IL-8 and TNF-α and the incidence of preterm delivery, 3 the level of amniotic fluid's IL-1β, IL-6, IL-8 and TNF-α and the grade of histologic amnionitis and placentitis in case of pre-term labor with intact membrane. Cases of spontaneous Pre'teftn labor with intact membrane which underwent transabdominal amniocentesis at admission and managed as standard procedure for pre-term labor with intact membrane. Atl of the cases were observed until the delivery of the baby, eithir preterm or term. The membrane and the placentawere cut postnatally and then the histologic acute inflammation eyaluated based on the criteria of Salafia.The level of amniotic fluid IL-1β, IL-6, IL-8 and TNF-α were analyzed quantitatively by Elisa method. This study showed thet the degree of histologic amnionitis and placentitis, and the level of amniotic fluid's IL-1β, IL-6, IL-8 and TNF-α were significantly higher in pre-term compared to terrn deliveries (p<0.05 and lhere were a positive correlation between the grade of histoLogic inflammation and the level of amniotic fluid's cytokines (Spearmann Rank Conelation test; p<0,05 in cases of preterm labor with intact membrane. The

  5. Cholesterol enhances surface water diffusion of phospholipid bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Chi-Yuan; Kausik, Ravinath; Han, Songi, E-mail: [Department of Chemistry and Biochemistry and Materials Research Laboratory, University of California, Santa Barbara, California 93106 (United States); Olijve, Luuk L. C. [Laboratory of Macromolecular and Organic Chemistry and Institute for Complex Molecular Systems, Eindhoven University of Technology, P.O. Box 513, 5600 MB, Eindhoven (Netherlands)


    Elucidating the physical effect of cholesterol (Chol) on biological membranes is necessary towards rationalizing their structural and functional role in cell membranes. One of the debated questions is the role of hydration water in Chol-embedding lipid membranes, for which only little direct experimental data are available. Here, we study the hydration dynamics in a series of Chol-rich and depleted bilayer systems using an approach termed {sup 1}H Overhauser dynamic nuclear polarization (ODNP) NMR relaxometry that enables the sensitive and selective determination of water diffusion within 5–10 Å of a nitroxide-based spin label, positioned off the surface of the polar headgroups or within the nonpolar core of lipid membranes. The Chol-rich membrane systems were prepared from mixtures of Chol, dipalmitoyl phosphatidylcholine and/or dioctadecyl phosphatidylcholine lipid that are known to form liquid-ordered, raft-like, domains. Our data reveal that the translational diffusion of local water on the surface and within the hydrocarbon volume of the bilayer is significantly altered, but in opposite directions: accelerated on the membrane surface and dramatically slowed in the bilayer interior with increasing Chol content. Electron paramagnetic resonance (EPR) lineshape analysis shows looser packing of lipid headgroups and concurrently tighter packing in the bilayer core with increasing Chol content, with the effects peaking at lipid compositions reported to form lipid rafts. The complementary capability of ODNP and EPR to site-specifically probe the hydration dynamics and lipid ordering in lipid membrane systems extends the current understanding of how Chol may regulate biological processes. One possible role of Chol is the facilitation of interactions between biological constituents and the lipid membrane through the weakening or disruption of strong hydrogen-bond networks of the surface hydration layers that otherwise exert stronger repulsive forces, as reflected in

  6. Investigating Hydrophilic Pores in Model Lipid Bilayers Using Molecular Simulations: Correlating Bilayer Properties with Pore-Formation Thermodynamics. (United States)

    Hu, Yuan; Sinha, Sudipta Kumar; Patel, Sandeep


    Cell-penetrating and antimicrobial peptides show a remarkable ability to translocate across physiological membranes. Along with factors such as electric-potential-induced perturbations of membrane structure and surface tension effects, experiments invoke porelike membrane configurations during the solute transfer process into vesicles and cells. The initiation and formation of pores are associated with a nontrivial free-energy cost, thus necessitating a consideration of the factors associated with pore formation and the attendant free energies. Because of experimental and modeling challenges related to the long time scales of the translocation process, we use umbrella sampling molecular dynamics simulations with a lipid-density-based order parameter to investigate membrane-pore-formation free energy employing Martini coarse-grained models. We investigate structure and thermodynamic features of the pore in 18 lipids spanning a range of headgroups, charge states, acyl chain lengths, and saturation. We probe the dependence of pore-formation barriers on the area per lipid, lipid bilayer thickness, and membrane bending rigidities in three different lipid classes. The pore-formation free energy in pure bilayers and peptide translocating scenarios are significantly coupled with bilayer thickness. Thicker bilayers require more reversible work to create pores. The pore-formation free energy is higher in peptide-lipid systems than in peptide-free lipid systems due to penalties to maintain the solvation of charged hydrophilic solutes within the membrane environment.

  7. The use of virtual ground to control transmembrane voltages and measure bilayer currents in serial arrays of droplet interface bilayers (United States)

    Sarles, Stephen A.


    The droplet interface bilayer (DIB) is a simple technique for constructing a stable lipid bilayer at the interface of two lipid-encased water droplets submerged in oil. Networks of DIBs formed by connecting more than two droplets constitute a new form of modular biomolecular smart material, where the transduction properties of a single lipid bilayer can affect the actions performed at other interface bilayers in the network via diffusion through the aqueous environments of shared droplet connections. The passive electrical properties of a lipid bilayer and the arrangement of droplets that determine the paths for transport in the network require specific electrical control to stimulate and interrogate each bilayer. Here, we explore the use of virtual ground for electrodes inserted into specific droplets in the network and employ a multichannel patch clamp amplifier to characterize bilayer formation and ion-channel activity in a serial DIB array. Analysis of serial connections of DIBs is discussed to understand how assigning electrode connections to the measurement device can be used to measure activity across all lipid membranes within a network. Serial arrays of DIBs are assembled using the regulated attachment method within a multi-compartment flexible substrate, and wire-type electrodes inserted into each droplet compartment of the substrate enable the application of voltage and measurement of current in each droplet in the array.

  8. Atomistic Monte Carlo simulation of lipid membranes

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Sklenar, Heinz


    Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction......, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential...... of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol....

  9. Single-Sided Contacting of Out-of-Plane Polarized Piezo Films for Fluid Membrane Lenses

    CERN Document Server

    Stürmer, Moritz; Wallrabe, Ulrike


    We present an adaptive lens which tunes its focal length with a glass membrane that is deformed by an integrated piezo bending actuator. A particular challenge in the fabrication of this type of actuator is the need to contact the electrodes on both sides of the piezo element, i.e. also on the glass surface. Therefore, we show a novel contacting method where the top electrode on the piezo is segmented and the backside electrode is left at a floating potential. With this setup we achieve tuning capabilities of more than 13 dpt and response times in the range of less than 4 ms at a usable aperture of 4.5 mm.

  10. Dynamics of bolaamphiphilic fluorescent polyenes in lipid bilayers from polarization emission spectroscopy. (United States)

    Acuña, A Ulises; Amat-Guerri, Francisco; Quesada, Ernesto; Vélez, Marisela


    The rotational motions of the biamphiphilic polyenes (bolapolyenes) dimethyl all-(E)-octacosa-10,12,14,16,18-pentaenedioate (DE28:5) and dimethyl all-(E)-tetratriaconta-13,15,17,19,21-pentaenedioate (DE34:5), with head-to-head distances of 34 and 42A, respectively, have been examined by fluorescence anisotropy methods. The membrane-spanning bolapolyenes, which contain a central emitting pentaene group tethered to two methoxycarbonyl opposite polar heads by symmetric C(8) (DE28:5) and C(11) (DE34:5) polymethylene chains, were dispersed in lipid bilayers of DPPC or DMPC, and the stationary and picosecond-resolved emission was recorded as a function of temperature. In fluid-phase DMPC bilayers, three relaxation times could be determined, assigned to fast (0.2 and 2ns) single-bond isomerization processes localized on the alkyl chains, and to whole-molecule oscillations ( approximately 11ns), respectively. The anisotropy decay parameters were further analyzed in terms of a diffusive model for wobbling in a Gaussian ordering potential, to assess the anchoring effect of the symmetric polar heads. In this way, the average rotational diffusion constant of the bolapolyenes, D( perpendicular), could be estimated as 0.022-0.026rad(2) ns(-1) (DMPC bilayers, 35 degrees Celsius), a value that is only 1/3 of that corresponding to the related pentaene fatty acid spanning a single membrane monolayer. In contrast, the amplitude of the equilibrium orientational distribution (theta(half-cone) approximately 50 degrees ) is very similar for both the transmembrane and the single-headed polyenes. The reorientational oscillations of the central emitting group in the bolapolyenes necessarily would produce large-amplitude (2-5A) and very fast (ns) translational motions of the polar heads.

  11. Membrane biology: fission behind BARs. (United States)

    Haucke, Volker


    Membrane bending is accomplished in part by amphipathic helix insertion into the bilayer and the assembly of BAR domain scaffolds preparing the membrane for fission. Two recent studies highlight the roles of amphipathic helices and BAR scaffolds in membrane fission and establish the structural basis of membrane bending by the N-BAR protein endophilin.

  12. Effects of Dimethyl Sulfoxide on Surface Water near Phospholipid Bilayers. (United States)

    Lee, Yuno; Pincus, Philip A; Hyeon, Changbong


    Despite much effort to probe the properties of dimethyl sulfoxide (DMSO) solution, the effects of DMSO on water, especially near plasma membrane surfaces, still remain elusive. By performing molecular dynamics simulations at varying DMSO concentrations (XDMSO), we study how DMSO affects structural and dynamical properties of water in the vicinity of phospholipid bilayers. As proposed by a number of experiments, our simulations confirm that DMSO induces dehydration from bilayer surfaces and disrupts the H-bond structure of water. However, DMSO-enhanced water diffusivity at solvent-bilayer interfaces, an intriguing discovery reported by a spin-label measurement, is not confirmed in our simulations. To resolve this discrepancy, we examine the location of the spin label (Tempo) relative to the solvent-bilayer interface. In accord with the evidence in the literature, our simulations, which explicitly model Tempo-phosphatidylcholine, find that the Tempo moiety is equilibrated at ∼8-10 Å below the bilayer surface. Furthermore, the DMSO-enhanced surface-water diffusion is confirmed only when water diffusion is analyzed around the Tempo moiety that is immersed below the bilayer surface, which implies that the experimentally detected signal of water using Tempo stems from the interior of bilayers, not from the interface. Our analysis finds that the increase of water diffusion below the bilayer surface is coupled to the increase of area per lipid with an increasing XDMSO(≲10mol%). Underscoring the hydrophobic nature of the Tempo moiety, our study calls for careful re-evaluation of the use of Tempo in measurements on lipid bilayer surfaces. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  13. Coarse-Grained Simulations of Membranes under Tension

    CERN Document Server

    Neder, Jörg; Nielaba, Peter; Schmid, Friederike


    We investigate the properties of membranes under tension by Monte-Carlo simulations of a generic coarse-grained model for lipid bilayers. We give a comprising overview of the behavior of several membrane characteristics, such as the area per lipid, the monolayer overlap, the nematic order, and pressure profiles. Both the low-temperature regime, where the membranes are in a gel phase, and the high-temperature regime, where they are in the fluid phase, are considered. In the gel state, the membrane is hardly influenced by tension. In the fluid state, high tensions lead to structural changes in the membrane, which result in different compressibility regimes. The ripple state, which is found at tension zero in the transition regime between the fluid and the gel phase, disappears under tension and gives way to an interdigitated phase. We also study the membrane fluctuations in the fluid phase. In the low tension regime the data can be fitted nicely to a suitably extended elastic theory. At higher tensions the elas...

  14. Cholesterol effect on water permeability through DPPC and PSM lipid bilayers: a molecular dynamics study. (United States)

    Saito, Hiroaki; Shinoda, Wataru


    Water permeability of two different lipid bilayers of dipalmitoylphosphatidylcholine (DPPC) and palmitoylsphingomyelin (PSM) in the absence and presence of cholesterol (0-50 mol %) have been studied by molecular dynamics simulations to elucidate the molecular mechanism of the reduction in water leakage across the membranes by the addition of cholesterol. An enhanced free energy barrier was observed in these membranes with increased cholesterol concentration, and this was explained by the reduced cavity density around the cholesterol in the hydrophobic membrane core. There was an increase of trans conformers in the hydrophobic lipid chains adjacent to the cholesterol, which reduced the cavity density. The enhanced free energy barrier was found to be the main reason to reduce the water permeability with increased cholesterol concentration. At low cholesterol concentrations the PSM bilayer exhibited a higher free energy barrier than the DPPC bilayer for water permeation, while at greater than 30 mol % of cholesterol the difference became minor. This tendency for the PSM and DPPC bilayers to resemble each other at higher cholesterol concentrations was similar to commonly observed trends in several structural properties, such as order parameters, cross-sectional area per molecule, and cavity density profiles in the hydrophobic regions of bilayer membranes. These results demonstrate that DPPC and PSM bilayers with high cholesterol contents possess similar physical properties, which suggests that the solubility of cholesterol in these lipid bilayers has importance for an understanding of multicomponent lipid membranes with cholesterol.

  15. Chain elongation of diacylphosphatidylcholine induces fully bilayer interdigitation under atmospheric pressure. (United States)

    Goto, Masaki; Wilk, Agnieszka; Kazama, Akira; Chodankar, Shirish; Kohlbrecher, Joachim; Matsuki, Hitoshi


    The phase transitions of dibehenoylphosphatidylcholine (C22PC) bilayer membrane were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The constructed temperature-pressure phase diagram suggests that the gel phase at low temperatures is the interdigitated gel phase. To confirm the phase state, we performed small-angle neutron scattering and fluorescence measurements using a polarity-sensitive probe Prodan for the C22PC bilayer membrane under atmospheric pressure. The peaks obtained in both measurements clearly showed the characteristic patterns of the fully interdigitated gel phase. Taking into account of previous studies on the gel phase for long-chain PC bilayers under atmospheric pressure and our studies on the pressure-induced bilayer interdigitaion of diacyl-PCs, it turned out that the interdigitation of diacyl-PC bilayer membranes occurs when the carbon number of acyl chain reaches at least 22. The present study revealed that the interdigitation of PC bilayer membranes occurs not only by weakening the attractive force of polar head groups but also by strengthening the cohesive force of acyl chains. When dominating the force of acyl chains, the interdigitation can be induced even in a diacyl-PC bilayer membrane by only hydration under atmospheric pressure.

  16. Neutron diffraction studies of amphipathic helices in phospholipid bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Bradshaw, J.P.; Gilchrist, P.J. [Univ. of Edinburgh (United Kingdom); Duff, K.C. [Univ. of Edinburgh Medical School (United Kingdom); Saxena, A.M. [Brookhaven National Laboratory, Upton, NY (United States)


    The structural feature which is thought to facilitate the interaction of many peptides with phospholipid bilayers is the ability to fold into an amphipathic helix. In most cases the exact location and orientation of this helix with respect to the membrane is not known, and may vary with factors such as pH and phospholipid content of the bilayer. The growing interest in this area is stimulated by indications that similar interactions can contribute to the binding of certain hormones to their cell-surface receptors. We have been using the techniques of neutron diffraction from stacked phospholipid bilayers in an attempt to investigate this phenomenon with a number of membrane-active peptides. Here we report some of our findings with three of these: the bee venom melittin; the hormone calcitonin; and a synthetic peptide representing the ion channel fragment of influenza A M2 protein.

  17. The dependence of induced polarization on fluid salinity and pH, studied with an extended model of membrane polarization (United States)

    Hördt, Andreas; Bairlein, Katharina; Bielefeld, Anja; Bücker, Matthias; Kuhn, Eva; Nordsiek, Sven; Stebner, Hermann


    The estimation of hydraulic parameters from spectral induced polarization (SIP) measurements is difficult partly because the electrical impedance of sediments depends on several parameters that are not related to the texture. Important parameters that influence the spectral response are fluid salinity and pH. In order to understand the behaviour of SIP spectra from a mechanistic point of view, we carry out simulations with a membrane polarization model. The geometry consists of a sequence of wide and narrow pores with finite radii. The charge distribution at the mineral surface is described by a triple layer model, characterized by the zeta potential and the partition coefficient. We extended an existing model by incorporating known dependencies of the zeta potential and the partition coefficient on fluid salinity and pH. Our simulation results predict a decrease of the maximum phase shift of the complex electrical conductivity with increasing salinity, consistent with experimental observations. For very small pore radii, the phase shift may also show the opposite behaviour and increase with salinity. The imaginary conductivity at 1 Hz increases with increasing salinity, followed by a peak and a decrease at high salinities. The fact that our model predicts a decrease of the imaginary conductivity at high salinities is particularly important, because strong experimental evidence was recently found for such a decrease, which was theoretically unexplained so far. Both the maximum phase shift and the imaginary conductivity at 1 Hz decrease when pH decreases. The reason is that at low pH, the zeta potential and the partition coefficient both decrease, corresponding to a smaller charge density at the mineral surface, resulting in a weaker impact of the electrical double layer. The few existing experimental studies on pH dependence are qualitatively consistent with our simulation results.

  18. Interactions between HIV-1 Neutralizing Antibodies and Model Lipid Membranes imaged with AFM (United States)

    Zauscher, Stefan; Hardy, Gregory; Alam, Munir; Shapter, Joseph


    Lipid membrane interactions with rare, broadly neutralizing antibodies (NAbs), 2F5 and 4E10, play a critical role in HIV-1 neutralization. Our research is motivated by recent immunization studies that have shown that induction of antibodies that avidly bind the gp41-MPER antigen is not sufficient for neutralization. Rather, it is required that antigen designs induce polyreactive antibodies that recognize MPER antigens as well as the viral lipid membrane. However, the mechanistic details of how membrane properties influence NAb-lipid and NAb-antigen interactions remain unknown. Furthermore, it is well established that the native viral membrane is heterogeneous, representing a mosaic of lipid rafts and protein clustering. However, the size, physical properties, and dynamics of these regions are poorly characterized and their potential roles in HIV-1 neutralization are also unknown. To understand how membrane properties contribute to 2F5/4E10 membrane interactions, we have engineered biomimetic supported lipid bilayers (SLBs) and use atomic force microscopy to visualize membrane domains, antigen clustering, and antibody-membrane interactions at sub-nanometer z-resolution. Our results show that localized binding of HIV-1 antigens and NAbs occur preferentially with the most fluid membrane domain. This supports the theory that NAbs may interact with regions of low lateral lipid forces that allow antibody insertion into the bilayer.

  19. 固定在脂质双层膜内的细胞色素c氧化酶电化学研究%Electrochemical Study on Cytochrome C Oxidase Immobilized into Lipid Bilayer Membrane

    Institute of Scientific and Technical Information of China (English)



    含有单体牛细胞色素c氧化酶的脂质双层膜,被成功地固定在金石英晶体微平衡电极上.在较宽的pH值、温度和缓冲浓度范围内观察到了氧化酶修饰电极上的直接电子迁移.氧化酶修饰电极在80℃以上时保持直接电子迁移性质,将电极冷却到室温时,氧化酶仍保持电子迁移能力.在22~80℃范围内,温度变化可引起细胞色素c氧化酶的相转移,估计出了相转移前后的相应反应活化能通过射流分析,研究了在溶液含有亚铁细胞色素c氧化酶的电氧化过程中,乙腈同细胞色素c氧化酶的结合.结果显示:浓度1.3M具有常数Ki时,细胞色素c氧化酶和乙腈形成配合物,每个细胞色素c氧化酶配一个乙腈分子,乙腈同细胞色素氧化酶的结合过程是可逆过程.%Lipid bilayer membrane containing monomeric bovine cytochrome c oxidase was successfully immobilized in gold quartz crystal microbalance electrodes. Direct electron transfer on the oxidase modified electrode was observed over a wide range of pH, temperature and buffer concentration. The oxidase modified electrode maintains direct electron transfer properties at the temperature in excess of 80 ℃. Upon cooling down the electrode to room temperature, the oxidase still retained its electron transfer capabilities. Temperature can cause phase transition of cytochrome c oxidase in the temperature range of 22 to 80 ℃. The corresponding reaction activation energies before and after the transition were estimated.Acetonitrile binding to cytochrome c oxidase during the electroxidative process of solution-resident ferrocytochrome c was also investigated by flow injection analysis. The results showed that cytochrome c oxidase forms a complex with acetonitrile having a Ki of 1.3 M with a stoichiometry of 1 acetonitrile molecule per cytochrome c oxidase. The binding of acetonitrile to cytochrome oxidase is a reversible process.

  20. Effect of anesthetics on bending elasticity of lipid membranes (United States)

    Yi, Zheng; Michihiro, Nagao; Bossev, Dobrin


    Change in physical and chemical properties of bio-membranes is of great interest for understanding the mechanism of anesthetic action on membranes. Hypothetically the anesthetic alters the lipid membrane structure (promoting pore formation across membranes or at least switching transmembrane channels) and therefore the biophysical properties of the membrane. We have used neutron spin echo (NSE) spectroscopy to study the effect of anesthetic molecule, lidocaine, on the bending elasticity (BE) of lipid membranes. BE of lipid bilayers made of (1,2-Dimyristoyl-sn-Glycero-3-Phosphocholine) DMPC and 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC) have been measured at different temperatures and different in the fluid (Lα) phase. Using Zilman-Granek theory the BE were obtained from the decay of the NSE intermediate scattering function. We have found that in the presence of lidocaine the BE of DMPC and DPPC bilayers increases. The results were correlated with those from differential scanning calorimetry. Increase in the lidocaine concentration leads to decrease in the liquid/crystalline transition temperature.

  1. A Molecular Dynamics Study of the Structural and Dynamical Properties of Putative Arsenic Substituted Lipid Bilayers

    Directory of Open Access Journals (Sweden)

    Ratna Juwita


    Full Text Available Cell membranes are composed mainly of phospholipids which are in turn, composed of five major chemical elements: carbon, hydrogen, nitrogen, oxygen, and phosphorus. Recent studies have suggested the possibility of sustaining life if the phosphorus is substituted by arsenic. Although this issue is still controversial, it is of interest to investigate the properties of arsenated-lipid bilayers to evaluate this possibility. In this study, we simulated arsenated-lipid, 1-palmitoyl-2-oleoyl-sn-glycero-3-arsenocholine (POAC, lipid bilayers using all-atom molecular dynamics to understand basic structural and dynamical properties, in particular, the differences from analogous 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, (POPC lipid bilayers. Our simulations showed that POAC lipid bilayers have distinct structural and dynamical properties from those of native POPC lipid bilayers. Relative to POPC lipid bilayers, POAC lipid bilayers have a more compact structure with smaller lateral areas and greater order. The compact structure of POAC lipid bilayers is due to the fact that more inter-lipid salt bridges are formed with arsenate-choline compared to the phosphate-choline of POPC lipid bilayers. These inter-lipid salt bridges bind POAC lipids together and also slow down the head group rotation and lateral diffusion of POAC lipids. Thus, it would be anticipated that POAC and POPC lipid bilayers would have different biological implications.

  2. Tracking membrane protein association in model membranes.

    Directory of Open Access Journals (Sweden)

    Myriam Reffay

    Full Text Available Membrane proteins are essential in the exchange processes of cells. In spite of great breakthrough in soluble proteins studies, membrane proteins structures, functions and interactions are still a challenge because of the difficulties related to their hydrophobic properties. Most of the experiments are performed with detergent-solubilized membrane proteins. However widely used micellar systems are far from the biological two-dimensions membrane. The development of new biomimetic membrane systems is fundamental to tackle this issue.We present an original approach that combines the Fluorescence Recovery After fringe Pattern Photobleaching technique and the use of a versatile sponge phase that makes it possible to extract crucial informations about interactions between membrane proteins embedded in the bilayers of a sponge phase. The clear advantage lies in the ability to adjust at will the spacing between two adjacent bilayers. When the membranes are far apart, the only possible interactions occur laterally between proteins embedded within the same bilayer, whereas when membranes get closer to each other, interactions between proteins embedded in facing membranes may occur as well.After validating our approach on the streptavidin-biotinylated peptide complex, we study the interactions between two membrane proteins, MexA and OprM, from a Pseudomonas aeruginosa efflux pump. The mode of interaction, the size of the protein complex and its potential stoichiometry are determined. In particular, we demonstrate that: MexA is effectively embedded in the bilayer; MexA and OprM do not interact laterally but can form a complex if they are embedded in opposite bilayers; the population of bound proteins is at its maximum for bilayers separated by a distance of about 200 A, which is the periplasmic thickness of Pseudomonas aeruginosa. We also show that the MexA-OprM association is enhanced when the position and orientation of the protein is restricted by the

  3. Water reclamation from shale gas drilling flow-back fluid using a novel forward osmosis-vacuum membrane distillation hybrid system. (United States)

    Li, Xue-Mei; Zhao, Baolong; Wang, Zhouwei; Xie, Ming; Song, Jianfeng; Nghiem, Long D; He, Tao; Yang, Chi; Li, Chunxia; Chen, Gang


    This study examined the performance of a novel hybrid system of forward osmosis (FO) combined with vacuum membrane distillation (VMD) for reclaiming water from shale gas drilling flow-back fluid (SGDF). In the hybrid FO-VMD system, water permeated through the FO membrane into a draw solution reservoir, and the VMD process was used for draw solute recovery and clean water production. Using a SGDF sample obtained from a drilling site in China, the hybrid system could achieve almost 90% water recovery. Quality of the reclaimed water was comparable to that of bottled water. In the hybrid FO-VMD system, FO functions as a pre-treatment step to remove most contaminants and constituents that may foul or scale the membrane distillation (MD) membrane, whereas MD produces high quality water. It is envisioned that the FO-VMD system can recover high quality water not only from SGDF but also other wastewaters with high salinity and complex compositions.

  4. Micromachined glass apertures for artificial lipid bilayer formation in a microfluidic system


    Sandison, M.E.; Zagnoni, M.; Abu-Hantash, M.; Morgan, H


    The use of spark assisted chemical engraving (SACE) to produce glass apertures that are suitable for the formation of artificial bilayer lipid membranes is described. Prior to use, the glass apertures were rendered hydrophobic by a silanization process and were then incorporated into a simple microfluidic device. Successful bilayer lipid membrane (BLM) formation and the subsequent acquisition of single-channel recordings are demonstrated. Due to the simplicity and rapidity of the SACE process...

  5. Importance of phospholipid bilayer integrity in the analysis of protein–lipid interactions

    Energy Technology Data Exchange (ETDEWEB)

    Drücker, Patrick [Institute of Biochemistry, University of Münster, Wilhelm-Klemm-Str. 2, D-48149 Münster (Germany); Gerke, Volker [Institute of Medical Biochemistry, ZMBE, University of Münster, Von-Esmarch-Str. 56, D-48149 Münster (Germany); Galla, Hans-Joachim, E-mail: [Institute of Biochemistry, University of Münster, Wilhelm-Klemm-Str. 2, D-48149 Münster (Germany)


    Highlights: • We show long-term mechanical stabilization of solid supported bilayers. • Bilayer integrity is essential for the investigation of protein–lipid interactions. • Protein adsorption to a bilayer containing defects causes membrane destruction. - Abstract: The integrity of supported phospholipid bilayer membranes is of crucial importance for the investigation of lipid–protein interactions. Therefore we recorded the formation of supported membranes on SiO{sub 2} and mica by quartz crystal microbalance and controlled the integrity by atomic force microscopy. This study aims to analyze how membrane defects affect protein–lipid interactions. The experiments focused on a lipid mixture of POPC/DOPC/Chol/POPS/PI(4,5)P{sub 2} (37:20:20:20:3) and the binding of the peripheral membrane associated protein annexin A2. We found that formation of a continuous undisturbed bilayer is an indispensable precondition for a reliable determination and quantification of lipid–protein-interactions. If membrane defects were present, protein adsorption causes membrane disruption and lipid detachment on a support thus leading to false determination of binding constants. Our results obtained for PI(4,5)P{sub 2} and cholesterol containing supported membranes yield new knowledge to construct functional surfaces that may cover nanoporous substrates, form free standing membranes or may be used for lab-on-a-chip applications.

  6. Membrane invagination induced by Shiga toxin B-subunit

    DEFF Research Database (Denmark)

    Pezeshkian, W.; Hansen, Allan Grønhøj; Johannes, Ludger


    The bacterial Shiga toxin is composed of an enzymatically active A-subunit, and a receptor-binding homopentameric B-subunit (STxB) that mediates intracellular toxin trafficking. Upon STxB-mediated binding to the glycolipid globotriaosylceramide (Gb3) at the plasma membrane of target cells, Shiga...... toxin is internalized by clathrin-dependent and independent endocytosis. The formation of tubular membrane invaginations is an essential step in the clathrin-independent STxB uptake process. However, the mechanism by which STxB induces these invaginations has remained unclear. Using a combination of all...... toxin molecules then creates a tubular membrane invagination that drives toxin entry into the cell. This mechanism requires: (1) a precise molecular architecture of the STxB binding sites; (2) a fluid bilayer in order for the tubular invagination to form. Although, STxB binding to the membrane requires...

  7. Modeling constrained sintering of bi-layered tubular structures

    DEFF Research Database (Denmark)

    Tadesse Molla, Tesfaye; Kothanda Ramachandran, Dhavanesan; Ni, De Wei;


    . Furthermore, the model is validated using densification results from sintering of bi-layered tubular ceramic oxygen membrane based on porous MgO and Ce0.9Gd0.1O1.95-d layers. Model input parameters, such as the shrinkage kinetics and viscous parameters are obtained experimentally using optical dilatometry...

  8. Supported lipid bilayers with controlled curvature via colloidal lithography

    DEFF Research Database (Denmark)

    Sundh, Maria; Manandhar, Michal; Svedhem, Sofia


    Supported lipid bilayers (SLBs) at surfaces provide a route to quantitatively study molecular interactions with and at lipid membranes via different surface-based analytical techniques. Here, a method to fabricate SLBs with controlled curvatures, in the nanometer regime over large areas, is prese...

  9. On The Equivalence of Local and Global Area-constraint Formulations for Lipid Bilayer Vesicles

    CERN Document Server

    Dharmavaram, Sanjay


    Lipid bilayer membranes are commonly modeled as area-preserving fluid surfaces that resist bending. There appear to be two schools of thought in the literature concerning the actual area constraint. In some works the total or global area (GA) of the vesicle is a prescribed constant, while in others the local area ratio is assigned to unity. In this work we demonstrate the equivalence of these ostensibly distinct approaches in the specific case when the equilibrium configuration is a smooth, closed surface of genus zero. We accomplish this in the context of the Euler-Lagrange equilibrium equations, constraint equations and the second-variation with admissibility conditions, for a broad class of models - including the phase-field type.

  10. Thermodynamic study of benzocaine insertion into different lipid bilayers (United States)

    Cascales, J. J. López; Costa, S. D. Oliveira; Porasso, R. D.


    Despite the general consensus concerning the role played by sodium channels in the molecular mechanism of local anesthetics, the potency of anaesthetic drugs also seems to be related with their solubility in lipid bilayers. In this respect, this work represents a thermodynamic study of benzocaine insertion into lipid bilayers of different compositions by means of molecular dynamics simulation. Thus, the free energy profiles associated with benzocaine insertion into symmetric lipid bilayers composed of different proportions of dipalmitoylphosphatidylcholine and dipalmitoylphosphatidylserine were studied. From the simulation results, a maximum in the free energy (ΔG) profile was measured in the region of the lipid/solution interface. This free energy barrier appears to be very much dependent on the lipid composition of the membrane. On the other hand, the minimum free energy (ΔG) within the bilayer remained almost independent of the lipid composition of the bilayer. By repeating the study at different temperatures, it was seen how the spontaneity of benzocaine insertion into the lipid bilayer is due to an increase in the entropy associated with the process.

  11. Microporous device for local electric recordings on model lipid bilayers (United States)

    Kaufeld, Theresa; Steinem, Claudia; Schmidt, Christoph F.


    A powerful approach for characterizing lipid membranes and embedded proteins is the reconstitution of model lipid bilayers. The extreme fragility of 5 nm thick bilayers is a challenge for device design and requires a trade off of stability against accessibility. We here present a microporous lab-on-chip device that allows us to form stable, solvent-free lipid bilayers from giant unilamellar vesicles (GUVs) in a geometry that provides a unique set of access possibilities. The device is constructed around a micro-fabricated silicon chip with clusters of 1 µm-diameter pores and provides optical access to the lipid bilayers for high-NA epifluorescence imaging. At the same time, solvent exchange is possible on both sides of the lipid bilayer. Complete coverage can be achieved with GUVs, so that voltages can be applied across the lipid bilayer and single-channel currents can be measured using external or integrated silver/silver chloride electrodes. We describe the micro-fabrication by standard cleanroom techniques and the characterization of the device by atomic force microscopy, scanning electron microscopy and impedance spectroscopy. In proof-of-concept experiments we demonstrate that the device is capable of low-noise, single-ion-channel recordings. Electronic Supplementary Information (ESI) available: See DOI: 10.1039/b000000x/

  12. Different oxidized phospholipid molecules unequally affect bilayer packing. (United States)

    Megli, Francesco M; Russo, Luciana


    The aim of this study was to gain more detailed knowledge about the effect of the presence of defined oxidized phospholipid molecules in phospholipid bilayers. After chromatographic and mass spectrometry analysis, the previously used product of the Fenton reaction with unsaturated lecithins proved to consist of a plethora of oxidatively modified lecithins, useless either for the detailed study of the effects brought about in the bilayer or as the source of defined oxidized phospholipid molecules. The latter, particularly 2-(omega-carboxyacyl)- and 2-(n-hydroperoxyacyl)-lecithins, can be more conveniently prepared by chemical or enzymatic synthesis rather than by chemical or physical oxidation. The effect of those molecules and of commercially available 12-hydroxy-stearic and dodecanedioic acid was studied in planar supported phospholipid bilayers (SPBs) by use of EPR spectrometry. The SPBs also contained 2-(5-doxylstearoyl)-lecithin as the spin probe, and the EPR spectral anisotropy loss, indicative of bilayer disordering, was measured as a function of the molar percentage of oxidized lipid. Most oxidized lipid molecules examined in this study were able to induce bilayer disordering, while hydroperoxyl group-bearing acyl chains appeared to be much less effective. It is concluded that the effects of different oxidized phospholipids on phospholipid bilayer structure cannot be generalized, as happens with batch-oxidized phospholipids, and that the use of defined oxidized phospholipid molecular species for membrane oxidative stress guarantees a more reliable and detailed response.

  13. Polyglutamine expansion in huntingtin increases its insertion into lipid bilayers. (United States)

    Kegel, Kimberly B; Schewkunow, Vitali; Sapp, Ellen; Masso, Nicholas; Wanker, Erich E; DiFiglia, Marian; Goldmann, Wolfgang H


    An expanded polyglutamine (Q) tract (>37Q) in huntingtin (htt) causes Huntington disease. Htt associates with membranes and polyglutamine expansion in htt may alter membrane function in Huntington disease through a mechanism that is not known. Here we used differential scanning calorimetry to examine the effects of polyQ expansion in htt on its insertion into lipid bilayers. We prepared synthetic lipid vesicles composed of phosphatidylcholine and phosphatidylethanolamine and tested interactions of htt amino acids 1-89 with 20Q, 32Q or 53Q with the vesicles. GST-htt1-89 with 53Q inserted into synthetic lipid vesicles significantly more than GST-htt1-89 with 20Q or 32Q. We speculate that by inserting more into cell membranes, mutant huntingtin could increase disorder within the lipid bilayer and thereby disturb cellular membrane function.

  14. The potential of mesenchymal stem cells derived from amniotic membrane and amniotic fluid for neuronal regenerative therapy. (United States)

    Kim, Eun Young; Lee, Kyung-Bon; Kim, Min Kyu


    The mesenchymal stem cells (MSCs), which are derived from the mesoderm, are considered as a readily available source for tissue engineering. They have multipotent differentiation capacity and can be differentiated into various cell types. Many studies have demonstrated that the MSCs identified from amniotic membrane (AM-MSCs) and amniotic fluid (AF-MSCs) are shows advantages for many reasons, including the possibility of noninvasive isolation, multipotency, self-renewal, low immunogenicity, anti-inflammatory and nontumorigenicity properties, and minimal ethical problem. The AF-MSCs and AM-MSCs may be appropriate sources of mesenchymal stem cells for regenerative medicine, as an alternative to embryonic stem cells (ESCs). Recently, regenerative treatments such as tissue engineering and cell transplantation have shown potential in clinical applications for degenerative diseases. Therefore, amnion and MSCs derived from amnion can be applied to cell therapy in neuro-degeneration diseases. In this review, we will describe the potential of AM-MSCs and AF-MSCs, with particular focus on cures for neuronal degenerative diseases.

  15. Integrated microfluidic platform for bilayer studies and experimentation on single pore-forming species

    NARCIS (Netherlands)

    Stimberg, V.C.; Berg, van den A.; Le Gac, S.


    The combination of microfluidics with BLMs (bilayer lipid membranes) consists of a powerful tool for multiplexed and high-throughput studies on membrane properties and membrane proteins. Here, we report an integrated device for BLM ex-perimentation using both optical and electrical measurements. Thi

  16. Poly(aniline) nanowires in sol-gel coated ITO: a pH-responsive substrate for planar supported lipid bilayers. (United States)

    Ge, Chenhao; Orosz, Kristina S; Armstrong, Neal R; Saavedra, S Scott


    Facilitated ion transport across an artificial lipid bilayer coupled to a solid substrate is a function common to several types of bioelectronic devices based on supported membranes, including biomimetic fuel cells and ion channel biosensors. Described here is fabrication of a pH-sensitive transducer composed of a porous sol-gel layer derivatized with poly(aniline) (PANI) nanowires grown from an underlying planar indium-tin oxide (ITO) electrode. The upper sol-gel surface is hydrophilic, smooth, and compatible with deposition of a planar supported lipid bilayer (PSLB) formed via vesicle fusion. Conducting tip AFM was used to show that the PANI wires are connected to the ITO, which convert this electrode into a potentiometric pH sensor. The response to changes in the pH of the buffer contacting the PANI nanowire/sol-gel/ITO electrode is blocked by the very low ion permeability of the overlying fluid PSLB. The feasibility of using this assembly to monitor facilitated proton transport across the PSLB was demonstrated by doping the membrane with lipophilic ionophores that respond to a transmembrane pH gradient, which produced an apparent proton permeability several orders of magnitude greater than values measured for undoped lipid bilayers.

  17. Aqueous solutions at the interface with phospholipid bilayers. (United States)

    Berkowitz, Max L; Vácha, Robert


    In a sense, life is defined by membranes, because they delineate the barrier between the living cell and its surroundings. Membranes are also essential for regulating the machinery of life throughout many interfaces within the cell's interior. A large number of experimental, computational, and theoretical studies have demonstrated how the properties of water and ionic aqueous solutions change due to the vicinity of membranes and, in turn, how the properties of membranes depend on the presence of aqueous solutions. Consequently, understanding the character of aqueous solutions at their interface with biological membranes is critical to research progress on many fronts. The importance of incorporating a molecular-level description of water into the study of biomembrane surfaces was demonstrated by an examination of the interaction between phospholipid bilayers that can serve as model biological membranes. The results showed that, in addition to well-known forces, such as van der Waals and screened Coulomb, one has to consider a repulsion force due to the removal of water between surfaces. It was also known that physicochemical properties of biological membranes are strongly influenced by the specific character of the ions in the surrounding aqueous solutions because of the observation that different anions produce different effects on muscle twitch tension. In this Account, we describe the interaction of pure water, and also of aqueous ionic solutions, with model membranes. We show that a symbiosis of experimental and computational work over the past few years has resulted in substantial progress in the field. We now better understand the origin of the hydration force, the structural properties of water at the interface with phospholipid bilayers, and the influence of phospholipid headgroups on the dynamics of water. We also improved our knowledge of the ion-specific effect, which is observed at the interface of the phospholipid bilayer and aqueous solution, and its

  18. Influence of silybin on biophysical properties of phospholipid bilayers

    Institute of Scientific and Technical Information of China (English)

    Olga WESO(L)OWSKA; Krystyna MICHALAK; Barbara (L)ANIA-PIETRZAK; Micha(l) KU(Z)D(Z)A(L); Kamila STA(N)CZAK; Daniela MOSI(A)DZ; Piotr DOBRYSZYCKI; Andrzej O(Z)YHAR; Ma(l)gorzata KOMOROWSKA; Andrzej B HENDRICH


    Aim: Silybin (silibinin)is major biologically active flavonolignan extracted from milk thistle (Sylibum marianum). Its biological activities include hepato-protection, anticancer properties, and antioxidant- and membrane-stabilizing functions. Al-though membranes are postulated to be one of the cellular targets for silybin, little is known about its interaction with phospholipid bilayers. Methods: In the present work, the interactions of silybin with phosphatidylcholine bilayers were studied in detail using fluorescence spectroscopy, microcalorimetry and electron spin resonance techniques. Results: The results showed that silybin interacted with the surface of lipid bilayers. It affected the generalized polarization of the fluores-cent probe Prodan, while not influencing the more deeplylocated Laurdan. Silybin lowered the main phospholipid phase transition temperature as judged by microcalorimetry, and caused the immobilization of spin probe Tempo-palmitate located on the surface of membranes. The mobility of spin probes 5-and 16-doxylstearic acid was not affected by silybin. Silybin-induced quenching of 1,6-diphe-nyl-1,3,5-hexatriene fluorescence indicated that some flavonoid molecules parti-tioned into the hydrophobic region of membranes, which did not change signifi-cantly the biophysical properties of the deeper membrane regions. Conclusion: Such a behavior of silybin in membranes is in accordance with its postulated biological functions and neglectable side effects of therapies using silybin.

  19. Independent mobility of proteins and lipids in the plasma membrane of Escherichia coli. (United States)

    Nenninger, Anja; Mastroianni, Giulia; Robson, Alexander; Lenn, Tchern; Xue, Quan; Leake, Mark C; Mullineaux, Conrad W


    Fluidity is essential for many biological membrane functions. The basis for understanding membrane structure remains the classic Singer-Nicolson model, in which proteins are embedded within a fluid lipid bilayer and able to diffuse laterally within a sea of lipid. Here we report lipid and protein diffusion in the plasma membrane of live cells of the bacterium Escherichia coli, using Fluorescence Recovery after Photobleaching (FRAP) and Total Internal Reflection Fluorescence (TIRF) microscopy to measure lateral diffusion coefficients. Lipid and protein mobility within the membrane were probed by visualizing an artificial fluorescent lipid and a simple model membrane protein consisting of a single membrane-spanning alpha-helix with a Green Fluorescent Protein (GFP) tag on the cytoplasmic side. The effective viscosity of the lipid bilayer is strongly temperature-dependent, as indicated by changes in the lipid diffusion coefficient. Surprisingly, the mobility of the model protein was unaffected by changes in the effective viscosity of the bulk lipid, and TIRF microscopy indicates that it clusters in segregated, mobile domains. We suggest that this segregation profoundly influences the physical behaviour of the protein in the membrane, with strong implications for bacterial membrane function and bacterial physiology.

  20. Atomistic Monte Carlo simulation of lipid membranes. (United States)

    Wüstner, Daniel; Sklenar, Heinz


    Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches. We use our recently devised chain breakage/closure (CBC) local move set in the bond-/torsion angle space with the constant-bond-length approximation (CBLA) for the phospholipid dipalmitoylphosphatidylcholine (DPPC). We demonstrate rapid conformational equilibration for a single DPPC molecule, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol.

  1. Atomistic Monte Carlo Simulation of Lipid Membranes

    Directory of Open Access Journals (Sweden)

    Daniel Wüstner


    Full Text Available Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC simulation of lipid membranes. We provide an introduction into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches. We use our recently devised chain breakage/closure (CBC local move set in the bond-/torsion angle space with the constant-bond-length approximation (CBLA for the phospholipid dipalmitoylphosphatidylcholine (DPPC. We demonstrate rapid conformational equilibration for a single DPPC molecule, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol.

  2. Viscoelastic properties of actin-coated membranes (United States)

    Helfer, E.; Harlepp, S.; Bourdieu, L.; Robert, J.; Mackintosh, F. C.; Chatenay, D.


    In living cells, cytoskeletal filaments interact with the plasma membrane to form structures that play a key role in cell shape and mechanical properties. To study the interaction between these basic components, we designed an in vitro self-assembled network of actin filaments attached to the outer surface of giant unilamellar vesicles. Optical tweezers and single-particle tracking experiments are used to study the rich dynamics of these actin-coated membranes (ACM). We show that microrheology studies can be carried out on such an individual microscopic object. The principle of the experiment consists in measuring the thermally excited position fluctuations of a probe bead attached biochemically to the membrane. We propose a model that relates the power spectrum of these thermal fluctuations to the viscoelastic properties of the membrane. The presence of the actin network modifies strongly the membrane dynamics with respect to a fluid, lipid bilayer one. It induces first a finite (ω=0) two-dimensional (2D) shear modulus G02D~0.5 to 5 μN/m in the membrane plane. Moreover, the frequency dependence at high frequency of the shear modulus [G'2D(f )~f0.85+/-0.07] and of the bending modulus (κACM(f)~f0.55+/-0.21) demonstrate the viscoelastic behavior of the composite membrane. These results are consistent with a common exponent of 0.75 for both moduli as expected from our model and from prior measurements on actin solutions.

  3. Formation, Stability, and Mobility of One-Dimensional Lipid Bilayer on High Curvature Substrates

    Energy Technology Data Exchange (ETDEWEB)

    Huang, J; Martinez, J; Artyukhin, A; Sirbuly, D; Wang, Y; Ju, J W; Stroeve, P; Noy, A


    Curved lipid membranes are ubiquitous in living systems and play an important role in many biological processes. To understand how curvature and lipid composition affect membrane formation and fluidity we have assembled and studied mixed 1,2-Dioleoyl-sn-Glycero-3-Phosphocholine (DOPC) and 1,2-Dioleoyl-sn-Glycero-3-Phosphoethanolamine (DOPE) supported lipid bilayers on amorphous silicon nanowires with controlled diameters ranging from 20 nm to 200 nm. Addition of cone-shaped DOPE molecules to cylindrical DOPC molecules promotes vesicle fusion and bilayer formation on smaller diameter nanowires. Our experiments demonstrate that nanowire-supported bilayers are mobile, exhibit fast recovery after photobleaching, and have low concentration of defects. Lipid diffusion coefficients in these high-curvature tubular membranes are comparable to the values reported for flat supported bilayers and increase with decreasing nanowire diameter.

  4. Lipid-Bilayer Dynamics Probed by a Carbon Dot-Phospholipid Conjugate. (United States)

    Nandi, Sukhendu; Malishev, Ravit; Bhunia, Susanta Kumar; Kolusheva, Sofiya; Jopp, Jürgen; Jelinek, Raz


    Elucidating the dynamic properties of membranes is important for understanding fundamental cellular processes and for shedding light on the interactions of proteins, drugs, and viruses with the cell surface. Dynamic studies of lipid bilayers have been constrained, however, by the relatively small number of pertinent molecular probes and the limited physicochemical properties of the probes. We show that a lipid conjugate comprised of a fluorescent carbon dot (C-dot) covalently attached to a phospholipid constitutes a versatile and effective vehicle for studying bilayer dynamics. The C-dot-modified phospholipids readily incorporated within biomimetic membranes, including solid-supported bilayers and small and giant vesicles, and inserted into actual cellular membranes. We employed the C-dot-phospholipid probe to elucidate the effects of polymyxin-B (a cytolytic peptide), valproic acid (a lipophilic drug), and amyloid-β (a peptide associated with Alzheimer's disease) upon bilayer fluidity and lipid dynamics through the application of various biophysical techniques.

  5. Electrostatic interactions at the microscale modulate dynamics and distribution of lipids in bilayers. (United States)

    Mangiarotti, Agustín; Wilke, Natalia


    For decades, it has been assumed that electrostatic long-range (micron distances) repulsions in lipid bilayers are negligible due to screening from the aqueous milieu. This concept, mostly derived from theoretical calculations, is broadly accepted in the biophysical community. Here we present experimental evidence showing that domain-domain electrostatic repulsions in charged and also in neutral lipid bilayers regulate the diffusion, in-plane structuring and merging of lipid domains in the micron range. All the experiments were performed on both, lipid monolayers and bilayers, and the remarkable similarity in the results found in bilayers compared to monolayers led us to propose that inter-domain repulsions occur mainly within the plane of the membrane. Finally, our results indicate that electrostatic interactions between the species inserted in a cell membrane are not negligible, not only at nanometric but also at larger distances, suggesting another manner for regulating the membrane properties.

  6. Formation, Stability, and Mobility of One-Dimensional Lipid Bilayer on High Curvature Substrates

    Energy Technology Data Exchange (ETDEWEB)

    Huang, J; Martinez, J; Artyukhin, A; Sirbuly, D; Wang, Y; Ju, J W; Stroeve, P; Noy, A


    Curved lipid membranes are ubiquitous in living systems and play an important role in many biological processes. To understand how curvature and lipid composition affect membrane formation and fluidity we have assembled and studied mixed 1,2-Dioleoyl-sn-Glycero-3-Phosphocholine (DOPC) and 1,2-Dioleoyl-sn-Glycero-3-Phosphoethanolamine (DOPE) supported lipid bilayers on amorphous silicon nanowires with controlled diameters ranging from 20 nm to 200 nm. Addition of cone-shaped DOPE molecules to cylindrical DOPC molecules promotes vesicle fusion and bilayer formation on smaller diameter nanowires. Our experiments demonstrate that nanowire-supported bilayers are mobile, exhibit fast recovery after photobleaching, and have low concentration of defects. Lipid diffusion coefficients in these high-curvature tubular membranes are comparable to the values reported for flat supported bilayers and increase with decreasing nanowire diameter.

  7. Effects on lipid bilayer and nitrogen distribution induced by lateral pressure. (United States)

    Wang, Yu; Chen, Liang; Wang, Xiaogang; Dai, Chaoqing; Chen, Junlang


    The lateral pressure exerted on cell membrane is of great importance to signal transduction. Here, we perform molecular dynamics simulation to explore how lateral pressure affects the biophysical properties of lipid bilayer as well as nitrogen distribution in the membrane. Our results show that both physical properties of cell membrane and nitrogen distribution are highly sensitive to the lateral pressure. With the increasing lateral pressure, area per lipid drops and thickness of membrane increases obviously, while nitrogen molecules are more congested in the center of lipid bilayer than those under lower lateral pressure. These results suggest that the mechanism of nitrogen narcosis may be related to the lateral pressure.

  8. Amniotic Fluid Glucose Concentration: A Marker for Infection in Preterm Labor and Preterm Premature Rupture of Membranes

    Directory of Open Access Journals (Sweden)

    Gary A. Dildy


    Full Text Available Amniotic fluid Gram stain and culture have been utilized as laboratory tests of microbial invasion of the amniotic cavity. The Gram stain of amniotic fluid has a low sensitivity in the detection of clinical infection or microbial invasion of the amniotic cavity, and amniotic fluid culture results are not immediately available for management decisions. Glucose concentration is used to diagnose infection in other sites such as cerebrospinal fluid.

  9. Comparison of Four Types of Membrane Bioreactor Systems in Terms of Shear Stress over the Membrane Surface using Computational Fluid Dynamics

    DEFF Research Database (Denmark)

    Ratkovich, Nicolas Rios; Bentzen, Thomas Ruby


    Membrane bioreactors (MBRs) have been used successfully in biological wastewater treatment to solve the perennial problem of effective solids–liquid separation. A common problem with MBR systems is clogging of the modules and fouling of the membrane, resulting in frequent cleaning and replacement...

  10. Clinical significance of histologic chorioamnionitis with a negative amniotic fluid culture in patients with preterm labor and premature membrane rupture (United States)

    Park, Jeong Woo; Park, Kyo Hoon; Jung, Eun Young


    Objective To evaluate the effect of histological chorioamnionitis (HCA) with a negative amniotic fluid (AF) culture on adverse pregnancy and neonatal outcomes and inflammatory status in the AF compartment in women with preterm labor or preterm premature rupture of membranes (PPROM). Methods This is a retrospective cohort study of 153 women diagnosed as having a preterm labor or PPROM (20–34 weeks) who delivered singleton gestations within 48 hours of amniocentesis. AF obtained through amniocentesis was cultured, and interleukin (IL)-6, IL-8, and metalloproteinase-9 (MMP-9) levels were determined. The placentas were examined histologically. Results The prevalence of HCA with negative AF culture was 23.5% (36/153). The women with HCA but with a negative AF culture (group 2) and those with a positive AF culture (group 3) had a significantly lower mean gestational age at amniocentesis and delivery than those with a negative AF culture and without HCA (group 1). Women in group 3 had the highest levels of AF IL-6, IL-8, and MMP-9, followed by those in group 2, and those in group 1. Composite neonatal morbidity was significantly higher in groups 2 and 3 than in group 1, but this was no longer significant after adjusting for confounders caused mainly by the impact of gestational age. Discussion In the women who delivered preterm neonates, HCA with a negative AF culture was associated with increased risks of preterm birth, intense intra-amniotic inflammatory response, and prematurity-associated composite neonatal morbidity, and its risks are similar to the risk posed by positive AF culture. PMID:28278303

  11. Super-Sensitive and Robust Biosensors from Supported Polymer Bilayers

    Energy Technology Data Exchange (ETDEWEB)

    Paxton, Walter F. [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States)


    Biological organisms are potentially the most sensitive and selective biological detection systems known, yet we are currently severely limited in our ability to exploit biological interactions in sensory devices, due in part to the limited stability of biological systems and derived materials. This proposal addresses an important aspect of integrating biological sensory materials in a solid state device. If successful, such technology could enable entirely new classes of robust biosensors that could be miniaturized and deployed in the field. The critical aims of the proposed work were 1) the calibration of a more versatile approach to measuring pH, 2) the use of this method to monitor pH changes caused by the light-induced pumping of protons across vesicles with bacteriorhodopsin integrated into the membranes (either polymer or lipid); 3) the preparation of bilayer assemblies on platinum surfaces; 4) the enhanced detection of lightinduced pH changes driven by bR-loaded supported bilayers. I have developed a methodology that may enable that at interfaces and developed a methodology to characterize the functionality of bilayer membranes with reconstituted membrane proteins. The integrity of the supported bilayer films however must be optimized prior to the full realization of the work originally envisioned in the original proposal. Nevertheless, the work performed on this project and the encouraging results it has produced has demonstrated that these goals are challenging yet within reach.

  12. Evolutionary origins of membrane proteins (United States)

    Mulkidjanian, Armen Y.; Galperin, Michael Y.

    Although the genes that encode membrane proteins make about 30% of the sequenced genomes, the evolution of membrane proteins and their origins are still poorly understood. Here we address this topic by taking a closer look at those membrane proteins the ancestors of which were present in the Last Universal Common Ancestor, and in particular, the F/V-type rotating ATPases. Reconstruction of their evolutionary history provides hints for understanding not only the origin of membrane proteins, but also of membranes themselves. We argue that the evolution of biological membranes could occur as a process of coevolution of lipid bilayers and membrane proteins, where the increase in the ion-tightness of the membrane bilayer may have been accompanied by a transition from amphiphilic, pore-forming membrane proteins to highly hydrophobic integral membrane complexes.

  13. Permeation of halide anions through phospholipid bilayers occurs by the solubility-diffusion mechanism (United States)

    Paula, S.; Volkov, A. G.; Deamer, D. W.


    Two alternative mechanisms are frequently used to describe ionic permeation of lipid bilayers. In the first, ions partition into the hydrophobic phase and then diffuse across (the solubility-diffusion mechanism). The second mechanism assumes that ions traverse the bilayer through transient hydrophilic defects caused by thermal fluctuations (the pore mechanism). The theoretical predictions made by both models were tested for halide anions by measuring the permeability coefficients for chloride, bromide, and iodide as a function of bilayer thickness, ionic radius, and sign of charge. To vary the bilayer thickness systematically, liposomes were prepared from monounsaturated phosphatidylcholines (PC) with chain lengths between 16 and 24 carbon atoms. The fluorescent dye MQAE (N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide) served as an indicator for halide concentration inside the liposomes and was used to follow the kinetics of halide flux across the bilayer membranes. The observed permeability coefficients ranged from 10(-9) to 10(-7) cm/s and i