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Sample records for lipid membrane investigation

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

    PubMed

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

    2014-12-01

    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.

  2. Direct affinity of dopamine to lipid membranes investigated by Nuclear Magnetic Resonance spectroscopy.

    PubMed

    Matam, Yashasvi; Ray, Bruce D; Petrache, Horia I

    2016-04-08

    Dopamine, a naturally occurring neurotransmitter, plays an important role in the brain's reward system and acts on sensory receptors in the brain. Neurotransmitters are contained in lipid membraned vesicles and are released by exocytosis. All neurotransmitters interact with transport and receptor proteins in glial cells, on neuronal dendrites, and at the axonal button, and also must interact with membrane lipids. However, the extent of direct interaction between lipid membranes in the absence of receptors and transport proteins has not been extensively investigated. In this report, we use UV and NMR spectroscopy to determine the affinity and the orientation of dopamine interacting with lipid vesicles made of either phosphatidylcholine (PC) or phosphatidylserine (PS) lipids which are primary lipid components of synaptic vesicles. We quantify the interaction of dopamine's aromatic ring with lipid membranes using our newly developed method that involves reference spectra in hydrophobic environments. Our measurements show that dopamine interacts with lipid membranes primarily through the aromatic side opposite to the hydroxyl groups, with this aromatic side penetrating deeper into the hydrophobic region of the membrane. Since dopamine's activity involves its release into extracellular space, we have used our method to also investigate dopamine's release from lipid vesicles. We find that dopamine trapped inside PC and PS vesicles is released into the external solution despite its affinity to membranes. This result suggests that dopamine's interaction with lipid membranes is complex and involves both binding as well as permeation through lipid bilayers, a combination that could be an effective trigger for apoptosis of dopamine-generating cells.

  3. Cell-sized asymmetric lipid vesicles facilitate the investigation of asymmetric membranes

    NASA Astrophysics Data System (ADS)

    Kamiya, Koki; Kawano, Ryuji; Osaki, Toshihisa; Akiyoshi, Kazunari; Takeuchi, Shoji

    2016-09-01

    Asymmetric lipid giant vesicles have been used to model the biochemical reactions in cell membranes. However, methods for producing asymmetric giant vesicles lead to the inclusion of an organic solvent layer that affects the mechanical and physical characteristics of the membrane. Here we describe the formation of asymmetric giant vesicles that include little organic solvent, and use them to investigate the dynamic responses of lipid molecules in the vesicle membrane. We formed the giant vesicles via the inhomogeneous break-up of a lipid microtube generated by applying a jet flow to an asymmetric planar lipid bilayer. The asymmetric giant vesicles showed a lipid flip-flop behaviour in the membrane, superficially similar to the lipid flip-flop activity observed in apoptotic cells. In vitro synthesis of membrane proteins into the asymmetric giant vesicles revealed that the lipid asymmetry in bilayer membranes improves the reconstitution ratio of membrane proteins. Our asymmetric giant vesicles will be useful in elucidating lipid-lipid and lipid-membrane protein interactions involved in the regulation of cellular functions.

  4. UV-Visible and Infrared Methods for Investigating Lipid-Rhodopsin Membrane Interactions

    PubMed Central

    Brown, Michael F.

    2017-01-01

    Summary Experimental UV-visible and Fourier transform infrared (FTIR) spectroscopic methods are described for characterizing lipid-protein interactions for the example of rhodopsin in a membrane bilayer environment. The combined use of FTIR and UV-visible difference spectroscopy monitors the structural and functional changes during rhodopsin activation. Such studies investigate how membrane lipids stabilize the various rhodopsin photoproducts, analogous to mutating the protein. Interpretation of the results entails a non-specific flexible surface model for explaining the role of membrane lipid-protein interactions in biological functions. PMID:22976026

  5. Introduction to membrane lipids.

    PubMed

    Epand, Richard M

    2015-01-01

    Biological membranes are composed largely of lipids and proteins. The most common arrangement of lipids in biological membranes is as a bilayer. This arrangement spontaneously forms a barrier for the passage of polar materials. The bilayer is thin but can have a large area in the dimension perpendicular to its thickness. The physical nature of the bilayer membrane will vary according to the conditions of the environment as well as the chemical structure of the lipid constituents of the bilayer. These physical properties determine the function of the membrane together with specific structural features of the lipids that allow them to have signaling properties. The lipids of the membrane are not uniformly distributed. There is an intrinsic asymmetry between the two monolayers that constitute the bilayer. In addition, some lipids tend to be enriched in particular regions of the membrane, termed domains. There is evidence that certain domains recruit specific proteins into that domain. This has been suggested to be important for allowing interaction among different proteins involved in certain signal transduction pathways. Membrane lipids have important roles in determining the physical properties of the membrane, in modulating the activity of membrane-bound proteins and in certain cases being specific secondary messengers that can interact with specific proteins. A large variety of lipids present in biological membranes result in them possessing many functions.

  6. A new Monte Carlo method for investigating geometrical structures of lipid membranes with atomistic detail

    NASA Astrophysics Data System (ADS)

    Cheng, Sara; Qiu, Liming; Cheng, K.; Vaughn, Mark

    2011-10-01

    The distribution statistics of the surface area, volume and voids of lipid molecules are important parameters to characterize the structures of self-assembling lipid membranes. Traditional methods are mostly based on various assumptions of the thickness of the lipid membrane and the volumes of certain types of lipid molecules. However, those methods usually lead to an over- or underestimation of the average surface area of lipid molecules when compared to the experimental results of the pure lipid systems. We developed a new Monte Carlo method that is able to estimate the distributions and averages of surface area, volume and void space of the lipid molecules in the absence and presence of proteins of the MD simulation results of lipid membranes at the atomistic scale. We successfully validated our new method on an ordered hard-sphere system and on a phospholipid/cholesterol binary lipid system, all with known structural parameters. Using this new method, the structural perturbation of the conformal annular lipids in close proximity to the embedded protein in a lipid/protein system will also be presented.

  7. Computational Investigation of the Effect of Lipid Membranes on Ion Permeation in Gramicidin A

    PubMed Central

    Setiadi, Jeffry; Kuyucak, Serdar

    2016-01-01

    Membrane proteins are embedded in a lipid bilayer and interact with the lipid molecules in subtle ways. This can be studied experimentally by examining the effect of different lipid bilayers on the function of membrane proteins. Understanding the causes of the functional effects of lipids is difficult to dissect experimentally but more amenable to a computational approach. Here we perform molecular dynamics simulations and free energy calculations to study the effect of two lipid types (POPC and NODS) on the conductance of the gramicidin A (gA) channel. A larger energy barrier is found for the K+ potential of mean force in gA embedded in POPC compared to that in NODS, which is consistent with the enhanced experimental conductance of cations in gA embedded in NODS. Further analysis of the contributions to the potential energy of K+ reveals that gA and water molecules in gA make similar contributions in both bilayers but there are significant differences between the two bilayers when the lipid molecules and interfacial waters are considered. It is shown that the stronger dipole moments of the POPC head groups create a thicker layer of interfacial waters with better orientation, which ultimately is responsible for the larger energy barrier in the K+ PMF in POPC. PMID:26999229

  8. Molecular dynamics simulations of model trans-membrane peptides in lipid bilayers: a systematic investigation of hydrophobic mismatch.

    PubMed

    Kandasamy, Senthil K; Larson, Ronald G

    2006-04-01

    Hydrophobic mismatch, which is the difference between the hydrophobic length of trans-membrane segments of a protein and the hydrophobic width of the surrounding lipid bilayer, is known to play a role in membrane protein function. We have performed molecular dynamics simulations of trans-membrane KALP peptides (sequence: GKK(LA)nLKKA) in phospholipid bilayers to investigate hydrophobic mismatch alleviation mechanisms. By varying systematically the length of the peptide (KALP15, KALP19, KALP23, KALP27, and KALP31) and the lipid hydrophobic length (DLPC, DMPC, and DPPC), a wide range of mismatch conditions were studied. Simulations of durations of 50-200 ns show that under positive mismatch, the system alleviates the mismatch predominantly by tilting the peptide and to a smaller extent by increased lipid ordering in the immediate vicinity of the peptide. Under negative mismatch, alleviation takes place by a combination of local bilayer bending and the snorkeling of the lysine residues of the peptide. Simulations performed at a higher peptide/lipid molar ratio (1:25) reveal slower dynamics of both the peptide and lipid relative to those at a lower peptide/lipid ratio (1:128). The lysine residues have favorable interactions with specific oxygen atoms of the phospholipid headgroups, indicating the preferred localization of these residues at the lipid/water interface.

  9. Molecular Dynamics Simulations of Model Trans-Membrane Peptides in Lipid Bilayers: A Systematic Investigation of Hydrophobic Mismatch

    PubMed Central

    Kandasamy, Senthil K.; Larson, Ronald G.

    2006-01-01

    Hydrophobic mismatch, which is the difference between the hydrophobic length of trans-membrane segments of a protein and the hydrophobic width of the surrounding lipid bilayer, is known to play a role in membrane protein function. We have performed molecular dynamics simulations of trans-membrane KALP peptides (sequence: GKK(LA)nLKKA) in phospholipid bilayers to investigate hydrophobic mismatch alleviation mechanisms. By varying systematically the length of the peptide (KALP15, KALP19, KALP23, KALP27, and KALP31) and the lipid hydrophobic length (DLPC, DMPC, and DPPC), a wide range of mismatch conditions were studied. Simulations of durations of 50–200 ns show that under positive mismatch, the system alleviates the mismatch predominantly by tilting the peptide and to a smaller extent by increased lipid ordering in the immediate vicinity of the peptide. Under negative mismatch, alleviation takes place by a combination of local bilayer bending and the snorkeling of the lysine residues of the peptide. Simulations performed at a higher peptide/lipid molar ratio (1:25) reveal slower dynamics of both the peptide and lipid relative to those at a lower peptide/lipid ratio (1:128). The lysine residues have favorable interactions with specific oxygen atoms of the phospholipid headgroups, indicating the preferred localization of these residues at the lipid/water interface. PMID:16428278

  10. FT-IR Investigations Into The Fluidity Of Lipopolysaccharide And Lipid A Membrane Systems

    NASA Astrophysics Data System (ADS)

    Brandenburg, K.; Seydel, U.

    1989-12-01

    Lipopolysaccharides (LPS) are the major amphiphilic components of the outer membrane of Gram-negative bacteria. They are composed of a poly- or oligosaccharide portion being covalently linked to a lipid moiety called lipid A. LPS are also called endotoxins because of their ability to induce harmful effects in organisms. However, they also exhibit beneficial ('adjuvant') activity for example by stimulating the immune system and inducing the production of e.g. the tumor necrosis factor and interleukin 1. The mechanisms leading to a stimulation of the cells of the immune system should mainly be governed by the fluidity and/or the supramolecular structure of the LPS and lipid A assemblies interacting with the cell membrane. In this contribution we report on FT-IR measurements of the fluidity of various LPS and of some lipid A's, and on the influence of the concentration of divalent cations, pH and water content on this parameter. The LPS differ in the headgroup conformation/composition, i.e in the length of the sugar moiety. The lipid A's show variation in their acylation patterns.

  11. An investigation of the effect of membrane curvature on transmembrane-domain dependent protein sorting in lipid bilayers

    PubMed Central

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

    2014-01-01

    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

  12. Investigation of the interaction of dimethyl sulfoxide with lipid membranes by small-angle neutron scattering

    SciTech Connect

    Gorshkova, J. E. Gordeliy, V. I.

    2007-05-15

    The influence of dimethyl sulfoxide (CH{sub 3}){sub 2}SO (DMSO) on the structure of membranes of 1,2-dimiristoyl-sn-glycero-3-phosphatidylcholine (DMPC) in an excess of a water-DMSO solvent is investigated over a wide range of DMSO molar concentrations 0.0 {<=} X{sub DMSO} {<=} 1.0 at temperatures T = 12.5 and 55 deg. C. The dependences of the repeat distance d of multilamellar membranes and the thickness d{sub b} of single vesicles on the molar concentration X{sub DMSO} in the L{sub {beta}}{sub '} gel and L{sub {alpha}} liquid-crystalline phases are determined by small-angle neutron scattering. The intermembrane distance d{sub s} is determined from the repeat distance d and the membrane thickness d{sub b}. It is shown that an increase in the molar concentration X{sub DMSO} leads to a considerable decrease in the intermembrane distance and that, at X{sub DMSO} = 0.4, the neighboring membranes are virtually in steric contact with each other. The use of the deuterated phospholipid (DMSO-D6) and the contrast variation method makes it possible, for the first time, to determine the number of DMSO molecules strongly bound to the membrane.

  13. Investigation of molecular mechanisms of action of chelating drugs on protein-lipid model membranes by X-ray fluorescence

    SciTech Connect

    Novikova, N. N.; Zheludeva, S. I.; Koval'chuk, M. V.; Stepina, N. D.; Erko, A. I.; Yur'eva, E. A.

    2009-12-15

    Protein-lipid films based on the enzyme alkaline phosphatase were subjected to the action of chelating drugs, which are used for accelerating the removal of heavy metals from the human body, and the elemental composition of the resulting films was investigated. Total-reflection X-ray fluorescence measurements were performed at the Berlin Electron Storage Ring Company for Synchrotron Radiation (BESSY) in Germany. A comparative estimation of the protective effect of four drugs (EDTA, succimer, xydiphone, and mediphon) on membrane-bound enzymes damaged by lead ions was made. The changes in the elemental composition of the protein-lipid films caused by high doses of chelating drugs were investigated. It was shown that state-of-the-art X-ray techniques can, in principle, be used to develop new methods for the in vitro evaluation of the efficiency of drugs, providing differential data on their actions.

  14. Experimental Investigations of Direct and Converse Flexoelectric Effect in Bilayer Lipid Membranes.

    NASA Astrophysics Data System (ADS)

    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 (<300 Hz), associated with 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

  15. Lipids and Membrane Lateral Organization

    PubMed Central

    Sonnino, Sandro; Prinetti, Alessandro

    2010-01-01

    Shortly after the elucidation of the very basic structure and properties of cellular membranes, it became evident that cellular membranes are highly organized structures with multiple and multi-dimensional levels of order. Very early observations suggested that the lipid components of biological membranes might be active players in the creation of these levels of order. In the late 1980s, several different and diverse experimental pieces of evidence coalesced together giving rise to the lipid raft hypothesis. Lipid rafts became enormously (and, in the opinion of these authors, sometimes acritically) popular, surprisingly not just within the lipidologist community (who is supposed to be naturally sensitive to the fascination of lipid rafts). Today, a PubMed search using the key word “lipid rafts” returned a list of 3767 papers, including 690 reviews (as a term of comparison, searching over the same time span for a very hot lipid-related key word, “ceramide” returned 6187 hits with 799 reviews), and a tremendous number of different cellular functions have been described as “lipid raft-dependent.” However, a clear consensus definition of lipid raft has been proposed only in recent times, and the basic properties, the ruling forces, and even the existence of lipid rafts in living cells has been recently matter of intense debate. The scenario that is gradually emerging from the controversies elicited by the lipid raft hypothesis emphasizes multiple roles for membrane lipids in determining membrane order, that encompass their tendency to phase separation but are clearly not limited to this. In this review, we would like to re-focus the attention of the readers on the importance of lipids in organizing the fine structure of cellular membranes. PMID:21423393

  16. Dynamics of multicomponent lipid membranes

    NASA Astrophysics Data System (ADS)

    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.

  17. Electronic polymers in lipid membranes.

    PubMed

    Johansson, Patrik K; Jullesson, David; Elfwing, Anders; Liin, Sara I; Musumeci, Chiara; Zeglio, Erica; Elinder, Fredrik; Solin, Niclas; Inganäs, Olle

    2015-06-10

    Electrical interfaces between biological cells and man-made electrical devices exist in many forms, but it remains a challenge to bridge the different mechanical and chemical environments of electronic conductors (metals, semiconductors) and biosystems. Here we demonstrate soft electrical interfaces, by integrating the metallic polymer PEDOT-S into lipid membranes. By preparing complexes between alkyl-ammonium salts and PEDOT-S we were able to integrate PEDOT-S into both liposomes and in lipid bilayers on solid surfaces. This is a step towards efficient electronic conduction within lipid membranes. We also demonstrate that the PEDOT-S@alkyl-ammonium:lipid hybrid structures created in this work affect ion channels in the membrane of Xenopus oocytes, which shows the possibility to access and control cell membrane structures with conductive polyelectrolytes.

  18. Lipid Polymorphisms and Membrane Shape

    PubMed Central

    Frolov, Vadim A.; Shnyrova, Anna V.; Zimmerberg, Joshua

    2011-01-01

    Morphological plasticity of biological membrane is critical for cellular life, as cells need to quickly rearrange their membranes. Yet, these rearrangements are constrained in two ways. First, membrane transformations may not lead to undesirable mixing of, or leakage from, the participating cellular compartments. Second, membrane systems should be metastable at large length scales, ensuring the correct function of the particular organelle and its turnover during cellular division. Lipids, through their ability to exist with many shapes (polymorphism), provide an adequate construction material for cellular membranes. They can self-assemble into shells that are very flexible, albeit hardly stretchable, which allows for their far-reaching morphological and topological behaviors. In this article, we will discuss the importance of lipid polymorphisms in the shaping of membranes and its role in controlling cellular membrane morphology. PMID:21646378

  19. A Raman spectroscopic investigation of the lipid state in acetylcholine receptor-rich membranes from Torpedo marmorata.

    PubMed Central

    Aslanian, D; Négrerie, M

    1985-01-01

    The lipid state in acetylcholine receptor (AcChR)-rich membranes purified from electric organ of Torpedo marmorata was studied in the temperature interval from 0 degrees C to 35 degrees C using the (C-H) stretching and (C-C) skeletal optical vibrations. The Raman spectra of AcChR-rich membranes, recorded immediately after preparation of the samples, indicate that the lipids are in a predominant triclinic crystalline lattice and do not undergo a phase transition when the temperature increases up to 35 degrees C. However, the polar groups of the lipids appear subject to temperature-induced variations. After extraction of 43-kd and other non-receptor proteins, spectra indicate an order-disorder phase transition of lipids at approximately 21 degrees C. This transition appears less cooperative than the transition of the membrane lipid extract. The role of the proteins in preservation of the crystalline state of lipids in AcChR-rich membranes is discussed. PMID:4018037

  20. Lipid membranes on nanostructured silicon.

    SciTech Connect

    Slade, Andrea Lynn; Lopez, Gabriel P.; Ista, Linnea K.; O'Brien, Michael J.; Sasaki, Darryl Yoshio; Bisong, Paul; Zeineldin, Reema R.; Last, Julie A.; Brueck, Stephen R. J.

    2004-12-01

    A unique composite nanoscale architecture that combines the self-organization and molecular dynamics of lipid membranes with a corrugated nanotextured silicon wafer was prepared and characterized with fluorescence microscopy and scanning probe microscopy. The goal of this project was to understand how such structures can be assembled for supported membrane research and how the interfacial interactions between the solid substrate and the soft, self-assembled material create unique physical and mechanical behavior through the confinement of phases in the membrane. The nanometer scale structure of the silicon wafer was produced through interference lithography followed by anisotropic wet etching. For the present study, a line pattern with 100 nm line widths, 200 nm depth and a pitch of 360 nm pitch was fabricated. Lipid membranes were successfully adsorbed on the structured silicon surface via membrane fusion techniques. The surface topology of the bilayer-Si structure was imaged using in situ tapping mode atomic force microscopy (AFM). The membrane was observed to drape over the silicon structure producing an undulated topology with amplitude of 40 nm that matched the 360 nm pitch of the silicon structure. Fluorescence recovery after photobleaching (FRAP) experiments found that on the microscale those same structures exhibit anisotropic lipid mobility that was coincident with the silicon substructure. The results showed that while the lipid membrane maintains much of its self-assembled structure in the composite architecture, the silicon substructure indeed influences the dynamics of the molecular motion within the membrane.

  1. Ethanol perturbs lipid organization in models of stratum corneum membranes: An investigation combining differential scanning calorimetry, infrared and (2)H NMR spectroscopy.

    PubMed

    Kwak, Sungjong; Brief, Elana; Langlais, Denis; Kitson, Neil; Lafleur, Michel; Thewalt, Jenifer

    2012-05-01

    Ethanol is used in a variety of topical products. It is known to enhance the permeability of the skin by altering the ability of the stratum corneum (SC) intercellular membranes to form an effective barrier. In addition, ethanol and other alcohols are key components of antiseptic gels currently used for hand wash. Using infrared and deuterium NMR spectroscopy as well as calorimetry, we have investigated the effect of ethanol on a model membrane composed of lipids representing the three classes of SC lipids, an equimolar mixture of N-palmitoylsphingosine (ceramide), palmitic acid and cholesterol. Ethanol is found to influence the membrane in a dose dependent manner, disrupting packing and increasing lipid motion at low concentrations and selectively extracting lipids at moderate concentrations.

  2. Mechanics of Lipid Bilayer Membranes

    NASA Astrophysics Data System (ADS)

    Powers, Thomas R.

    All cells have membranes. The plasma membrane encapsulates the cell's interior, acting as a barrier against the outside world. In cells with nuclei (eukaryotic cells), membranes also form internal compartments (organelles) which carry out specialized tasks, such as protein modification and sorting in the case of the Golgi apparatus, and ATP production in the case of mitochondria. The main components of membranes are lipids and proteins. The proteins can be channels, carriers, receptors, catalysts, signaling molecules, or structural elements, and typically contribute a substantial fraction of the total membrane dry weight. The equilibrium properties of pure lipid membranes are relatively well-understood, and will be the main focus of this article. The framework of elasticity theory and statistical mechanics that we will develop will serve as the foundation for understanding biological phenomena such as the nonequilibrium behavior of membranes laden with ion pumps, the role of membrane elasticity in ion channel gating, and the dynamics of vesicle fission and fusion. Understanding the mechanics of lipid membranes is also important for drug encapsulation and delivery.

  3. Interactions of the baicalin and baicalein with bilayer lipid membranes investigated by cyclic voltammetry and UV-Vis spectroscopy.

    PubMed

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

    2014-02-01

    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.

  4. How Membrane-Active Peptides Get into Lipid Membranes.

    PubMed

    Sani, Marc-Antoine; Separovic, Frances

    2016-06-21

    The structure-function relationship for a family of antimicrobial peptides (AMPs) from the skin of Australian tree frogs is discussed and compared with that of peptide toxins from bee and Australian scorpion venoms. Although these membrane-active peptides induce a similar cellular fate by disrupting the lipid bilayer integrity, their lytic activity is achieved via different modes of action, which are investigated in relation to amino acid sequence, secondary structure, and membrane lipid composition. In order to better understand what structural features govern the interaction between peptides and lipid membranes, cell-penetrating peptides (CPPs), which translocate through the membrane without compromising its integrity, are also discussed. AMPs possess membrane lytic activities that are naturally designed to target the cellular membrane of pathogens or competitors. They are extremely diverse in amino acid composition and often show specificity against a particular strain of microbe. Since our antibiotic arsenal is declining precariously in the face of the rise in multiantibiotic resistance, AMPs increasingly are seen as a promising alternative. In an effort to understand their molecular mechanism, biophysical studies of a myriad of AMPs have been reported, yet no unifying mechanism has emerged, rendering difficult the rational design of drug leads. Similarly, a wide variety of cytotoxic peptides are found in venoms, the best known being melittin, yet again, predicting their activity based on a particular amino acid composition or secondary structure remains elusive. A common feature of these membrane-active peptides is their preference for the lipid environment. Indeed, they are mainly unstructured in solution and, in the presence of lipid membranes, quickly adsorb onto the surface, change their secondary structure, eventually insert into the hydrophobic core of the membrane bilayer, and finally disrupt the bilayer integrity. These steps define the molecular

  5. TOPICAL REVIEW: The microcalorimetry of lipid membranes

    NASA Astrophysics Data System (ADS)

    Heerklotz, Heiko

    2004-04-01

    Insight into the forces governing a system is essential for understanding its behaviour and function. Calorimetric investigations provide a wealth of information that is not, or is hardly, available by other methods. This paper reviews calorimetric approaches and assays for the study of lipid vesicles (liposomes) and biological membranes. With respect to the instrumentation, differential scanning calorimetry (DSC), pressure perturbation calorimetry (PPC), isothermal titration calorimetry (ITC) and water sorption calorimetry are considered. Applications of these techniques to lipid systems include the measurement of thermodynamic parameters and a detailed characterization of the thermotropic, barotropic, and lyotropic phase behaviour. The membrane binding or partitioning of solutes (proteins, peptides, drugs, surfactants, ions, etc) can also be quantified. Many calorimetric assays are available for studying the effect of proteins and other additives on membranes, characterizing non-ideal mixing, domain formation, stability, curvature strain, permeability, solubilization, and fusion. Studies of membrane proteins in lipid environments elucidate lipid-protein interactions in membranes. The systems are described in terms of enthalpic and entropic forces, equilibrium constants, heat capacities, partial volume changes etc, shedding light also on the stability of structures and the molecular origin and mechanism of structural changes.

  6. Lipid membrane domains in the brain.

    PubMed

    Aureli, Massimo; Grassi, Sara; Prioni, Simona; Sonnino, Sandro; Prinetti, Alessandro

    2015-08-01

    The brain is characterized by the presence of cell types with very different functional specialization, but with the common trait of a very high complexity of structures originated by their plasma membranes. Brain cells bear evident membrane polarization with the creation of different morphological and functional subcompartments, whose formation, stabilization and function require a very high level of lateral order within the membrane. In other words, the membrane specialization of brain cells implies the presence of distinct membrane domains. The brain is the organ with the highest enrichment in lipids like cholesterol, glycosphingolipids, and the most recently discovered brain membrane lipid, phosphatidylglucoside, whose collective behavior strongly favors segregation within the membrane leading to the formation of lipid-driven membrane domains. Lipid-driven membrane domains function as dynamic platforms for signal transduction, protein processing, and membrane turnover. Essential events involved in the development and in the maintenance of the functional integrity of the brain depend on the organization of lipid-driven membrane domains, and alterations in lipid homeostasis, leading to deranged lipid-driven membrane organization, are common in several major brain diseases. In this review, we summarize the forces behind the formation of lipid membrane domains and their biological roles in different brain cells. This article is part of a Special Issue entitled Brain Lipids.

  7. Theoretical and simulation study of lipid membranes

    NASA Astrophysics Data System (ADS)

    Khelashvili, George

    It has been established that a proper functioning of biological lipid membranes is in large part due to cholesterol's ability to regulate fluidity of a lipid bilayer. In particular, a growing body of evidence suggested that cholesterol participates in the formation of cholesterol- and sphingolipid-enriched phase-separated domains known as "rafts" in the plasma and other membranes of animal cells. Rafts have been identified as important membrane structural components in signal transduction, protein transport and sorting of membrane components. At a molecular level, the detailed, localized behavior of lipid-cholesterol bilayers is unclear. In order to better understand how cholesterols function in lipid membranes it is desirable to built theoretical models. The goal of the present research is to model lipid-cholesterol bilayers on the different length and timescales. In the first part of the work, mixtures of sphingomyelin (SM) lipid and cholesterol at different temperatures and cholesterol concentrations were investigated using Molecular Dynamics and Monte-Carlo simulation techniques. The objective was to study the properties of cholesterol- and SM-enriched raft-like domains at the atomic level. The simulations revealed that, addition of 31% cholesterol induced intermediate degree of organization in the model SM-cholesterol bilayers at temperatures below and above the main phase transition temperature of pure SM bilayer. This intermediate state of fluidity may be necessary for the binding of proteins and other molecules that associate with raft domains. In the second part of the work, dynamical self-consistent mean-field model based on atomistic simulations was developed to investigate phase properties of lipid-cholesterol bilayers on the length and timescales currently unreachable with traditional atomistic level simulation methods. This new technique allows studying systems consisting of 104 or more number of molecules, on microsecond timescales. The model was

  8. Polymer Diffusion in Lipid Membranes

    NASA Astrophysics Data System (ADS)

    Prasad, Ashok

    2005-03-01

    Motivated by experiments on fluorescently labeled DNA molecules on a supported lipid bilayer, we have examined theoretically diffusion of polymers in two dimensions. The key experimental finding we focus on is the scaling of the diffusion constant of the center of mass, D˜1/N. This implies that no effective hydrodynamic coupling exists between the diffusing DNA segments in the membrane. We construct our theoretical model using the phenomenological hydrodynamic model of supported membranes proposed by Evans and Sackmann. Our model is based on the pre-averaged Oseen tensor, and is similar to the model of Komura and Seki, but elaborated and extended to take explicit account of self-avoidance. We find that the 1/N scaling of D can be understood as a consequence of membrane hydrodynamics in the presence of a supporting surface. Further experimental consequences of the model, in particular the diffusion constant for DNA in free standing membranes, will also be discussed. This work was supported by the NSF through grants DMR-9984471 and DMR-0403997. JK is a Cottrell Scholar of Research Corporation.

  9. Electrodiffusion of lipids on membrane surfaces

    NASA Astrophysics Data System (ADS)

    Zhou, Y. C.

    2012-05-01

    Lateral translocation of lipids and proteins is a universal process on membrane surfaces. Local aggregation or organization of lipids and proteins can be induced when the random lateral motion is mediated by the electrostatic interactions and membrane curvature. Although the lateral diffusion rates of lipids on membranes of various compositions are measured and the electrostatic free energies of predetermined protein-membrane-lipid systems can be computed, the process of the aggregation and the evolution to the electrostatically favorable states remain largely undetermined. Here we propose an electrodiffusion model, based on the variational principle of the free energy functional, for the self-consistent lateral drift-diffusion of multiple species of charged lipids on membrane surfaces. Finite sizes of lipids are modeled to enforce the geometrical constraint of the lipid concentration on membrane surfaces. A surface finite element method is developed to appropriate the Laplace-Beltrami operators in the partial differential equations of the model. Our model properly describes the saturation of lipids on membrane surfaces, and correctly predicts that the MARCKS peptide can consistently sequester three multivalent phosphatidylinositol 4,5-bisphosphate lipids through its basic amino acid residues, regardless of a wide range of the percentage of monovalent phosphatidylserine in the membrane.

  10. [Germ cell membrane lipids in spermatogenesis].

    PubMed

    Wang, Ting; Shi, Xiao; Quan, Song

    2016-05-01

    Spermatogenesis is a complex developmental process in which a diploid progenitor germ cell transforms into highly specialized spermatozoa. During spermatogenesis, membrane remodeling takes place, and cell membrane permeability and liquidity undergo phase-specific changes, which are all associated with the alteration of membrane lipids. Lipids are important components of the germ cell membrane, whose volume and ratio fluctuate in different phases of spermatogenesis. Abnormal lipid metabolism can cause spermatogenic dysfunction and consequently male infertility. Germ cell membrane lipids are mainly composed of cholesterol, phospholipids and glycolipids, which play critical roles in cell adhesion and signal transduction during spermatogenesis. An insight into the correlation of membrane lipids with spermatogenesis helps us to better understand the mechanisms of spermatogenesis and provide new approaches to the diagnosis and treatment of male infertility.

  11. Pore formation in lipid membranes by alamethicin.

    PubMed Central

    Fringeli, U P; Fringeli, M

    1979-01-01

    The conformation of the linear peptide antibiotic alamethicin in dipalmitoyl phosphatidylcholine multilayers was investigated in the absence of an electric field by means of infrared attenuated total reflection spectroscopy. Alamethicin was found to be incorporated into the lipid membrane not only in the dry state but also in an aqueous environment. Its molecular conformation, however, changed from a helix when dry to an extended chain when aqueous. The extended chain aggregated to di- and multimers spanning the lipid bilayer. The equilibrium concentration of alamethicin in the surrounding water was 90 nM, which is in the range of concentrations used in black film experiments. The corresponding molar ratio of lipid to peptide was 80:1. Concerning the molecular mechanism of electric field-induced pore formation, one has to conclude that the dipole model proposed by several authors is very unlikely because it is based on the assumption that the major part of alamethicin is adsorbed on the membrane surface, from which small amounts flip into the membrane under the influence of an electric field. An alternative mechanism is proposed, based on a field-induced conformational change of the peptide from the extended state to a helix. This transition is favored by the resulting dipole moment of the alamethicin helix. PMID:291045

  12. Lipid-lipid and lipid-drug interactions in biological membranes

    NASA Astrophysics Data System (ADS)

    Martynowycz, Michael W.

    Interactions between lipids and drug molecules in biological membranes help govern proper biological function in organisms. The mechanisms responsible for hydrophobic drug permeation remain elusive. Many small molecule drugs are hydrophobic. These drugs inhibit proteins in the cellular interior. The rise of antibiotic resistance in bacteria is thought to be caused by mutations in protein structure, changing drug kinetics to favor growth. However, small molecule drugs have been shown to have different mechanisms depending in the structure of the lipid membrane of the target cell. Biological membranes are investigated using Langmuir monolayers at the air-liquid interface. These offer the highest level of control in the mimetic system and allow them to be investigated using complementary techniques. Langmuir isotherms and insertion assays are used to determine the area occupied by each lipid in the membrane and the change in area caused by the introduction of a drug molecule, respectively. Specular X-ray reflectivity is used to determine the electron density of the monolayer, and grazing incidence X-ray diffraction is used to determine the in-plane order of the monolayer. These methods determine the affinity of the drug and the mechanism of action. Studies are presented on hydrophobic drugs with mammalian membrane mimics using warfarin along with modified analogues, called superwarfarins. Data shows that toxicity of these modified drugs are modulated by the membrane cholesterol content in cells; explaining several previously unexplained effects of the drugs. Membrane mimics of bacteria are investigated along with their interactions with a hydrophobic antibiotic, novobiocin. Data suggests that permeation of the drug is mediated by modifications to the membrane lipids, and completely ceases translocation under certain circumstances. Circumventing deficiencies in small, hydrophobic drugs is approached by using biologically mimetic oligomers. Peptoids, mimetic of host

  13. Characterization of lipid domains in erythrocyte membranes.

    PubMed

    Rodgers, W; Glaser, M

    1991-02-15

    Fluorescence digital imaging microscopy was used to study the lateral distribution of the lipid components in erythrocyte membranes. Intact erythrocytes labeled with phospholipids containing a fluorophore attached to one fatty acid chain showed an uneven distribution of the phospholipids in the membrane thereby demonstrating the presence of membrane domains. The enrichment of the lipotropic compound chlor-promazine in domains in intact erythrocytes also suggested that the domains are lipid-enriched regions. Similar membrane domains were present in erythrocyte ghosts. The phospholipid enrichment was increased in the domains by inducing membrane protein aggregation. Double-labeling experiments were done to determine the relative distributions of different phospholipids in the membrane. Vesicles made from extracted lipids did not show the presence of domains consistent with the conclusion that membrane proteins were responsible for creating the domains. Overall, it was found that large domains exist in the red blood cell membrane with unequal enrichment of the different phospholipid species.

  14. Control of lipid membrane stability by cholesterol content.

    PubMed Central

    Raffy, S; Teissié, J

    1999-01-01

    Cholesterol has a concentration-dependent effect on membrane organization. It is able to control the membrane permeability by inducing conformational ordering of the lipid chains. A systematic investigation of lipid bilayer permeability is described in the present work. It takes advantage of the transmembrane potential difference modulation induced in vesicles when an external electric field is applied. The magnitude of this modulation is under the control of the membrane electrical permeability. When brought to a critical value by the external field, the membrane potential difference induces a new membrane organization. The membrane is then permeable and prone to solubilized membrane protein back-insertion. This is obtained for an external field strength, which depends on membrane native permeability. This approach was used to study the cholesterol effect on phosphatidylcholine bilayers. Studies have been performed with lipids in gel and in fluid states. When cholesterol is present, it does not affect electropermeabilization and electroinsertion in lipids in the fluid state. When lipids are in the gel state, cholesterol has a dose-dependent effect. When present at 6% (mol/mol), cholesterol prevents electropermeabilization and electroinsertion. When cholesterol is present at more than 12%, electropermeabilization and electroinsertion are obtained under milder field conditions. This is tentatively explained by a cholesterol-induced alteration of the hydrophobic barrier of the bilayer core. Our results indicate that lipid membrane permeability is affected by the cholesterol content. PMID:10096902

  15. DMSO induces dehydration near lipid membrane surfaces.

    PubMed

    Cheng, Chi-Yuan; Song, Jinsuk; Pas, Jolien; Meijer, Lenny H H; Han, Songi

    2015-07-21

    Dimethyl sulfoxide (DMSO) has been broadly used in biology as a cosolvent, a cryoprotectant, and an enhancer of membrane permeability, leading to the general assumption that DMSO-induced structural changes in cell membranes and their hydration water play important functional roles. Although the effects of DMSO on the membrane structure and the headgroup dehydration have been extensively studied, the mechanism by which DMSO invokes its effect on lipid membranes and the direct role of water in this process are unresolved. By directly probing the translational water diffusivity near unconfined lipid vesicle surfaces, the lipid headgroup mobility, and the repeat distances in multilamellar vesicles, we found that DMSO exclusively weakens the surface water network near the lipid membrane at a bulk DMSO mole fraction (XDMSO) of <0.1, regardless of the lipid composition and the lipid phase. Specifically, DMSO was found to effectively destabilize the hydration water structure at the lipid membrane surface at XDMSO <0.1, lower the energetic barrier to dehydrate this surface water, whose displacement otherwise requires a higher activation energy, consequently yielding compressed interbilayer distances in multilamellar vesicles at equilibrium with unaltered bilayer thicknesses. At XDMSO >0.1, DMSO enters the lipid interface and restricts the lipid headgroup motion. We postulate that DMSO acts as an efficient cryoprotectant even at low concentrations by exclusively disrupting the water network near the lipid membrane surface, weakening the cohesion between water and adhesion of water to the lipid headgroups, and so mitigating the stress induced by the volume change of water during freeze-thaw.

  16. Finite element modeling of lipid bilayer membranes

    NASA Astrophysics Data System (ADS)

    Feng, Feng; Klug, William S.

    2006-12-01

    A numerical simulation framework is presented for the study of biological membranes composed of lipid bilayers based on the finite element method. The classic model for these membranes employs a two-dimensional-fluid-like elastic constitutive law which is sensitive to curvature, and subjects vesicles to physically imposed constraints on surface area and volume. This model is implemented numerically via the use of C1-conforming triangular Loop subdivision finite elements. The validity of the framework is tested by computing equilibrium shapes from previously-determined axisymmetric shape-phase diagram of lipid bilayer vesicles with homogeneous material properties. Some of the benefits and challenges of finite element modeling of lipid bilayer systems are discussed, and it is indicated how this framework is natural for future investigation of biologically realistic bilayer structures involving nonaxisymmetric geometries, binding and adhesive interactions, heterogeneous mechanical properties, cytoskeletal interactions, and complex loading arrangements. These biologically relevant features have important consequences for the shape mechanics of nonidealized vesicles and cells, and their study requires not simply advances in theory, but also advances in numerical simulation techniques, such as those presented here.

  17. Characterization of Titratable Amphiphiles in Lipid Membranes by Fluorescence Spectroscopy.

    PubMed

    Pierrat, Philippe; Lebeau, Luc

    2015-11-17

    Understanding the ionization behavior of lipid membranes is a key parameter for successful development of lipid-based drug delivery systems. Accurate determination of the ionization state of a titratable species incorporated in a lipid bilayer however requires special care. Herein we investigated the behavior of titratable lipids in liposomes by fluorescence spectroscopy and determined which extrinsic parameters-i.e., besides those directly related to their molecular structure-determine their ionization state. Two fluorescent dyes, TNS and R18, have been used to investigate basic and acidic titratable lipids, respectively. Our results suggest that the titration behavior of the ionizable lipid in the membrane is more sensitive to the composition of the membrane and to its physical state than to the presence of solutes in the aqueous phase. Essentially overlooked in earlier studies on ionizable lipid assemblies, the concentration of the titratable lipid in the membrane was found to have a major effect on the ionization state of the lipid polar head. This may result in a shift in the apparent pKa value which may be as large as two pKa units and cannot be satisfactorily predicted.

  18. Self-segregation of myelin membrane lipids in model membranes.

    PubMed

    Yurlova, Larisa; Kahya, Nicoletta; Aggarwal, Shweta; Kaiser, Hermann-Josef; Chiantia, Salvatore; Bakhti, Mostafa; Pewzner-Jung, Yael; Ben-David, Oshrit; Futerman, Anthony H; Brügger, Britta; Simons, Mikael

    2011-12-07

    Rapid conduction of nerve impulses requires coating of axons by myelin sheaths, which are multilamellar, lipid-rich membranes produced by oligodendrocytes in the central nervous system. To act as an insulator, myelin has to form a stable and firm membrane structure. In this study, we have analyzed the biophysical properties of myelin membranes prepared from wild-type mice and from mouse mutants that are unable to form stable myelin. Using C-Laurdan and fluorescence correlation spectroscopy, we find that lipids are tightly organized and highly ordered in myelin isolated from wild-type mice, but not from shiverer and ceramide synthase 2 null mice. Furthermore, only myelin lipids from wild-type mice laterally segregate into physically distinct lipid phases in giant unilamellar vesicles in a process that requires very long chain glycosphingolipids. Taken together, our findings suggest that oligodendrocytes exploit the potential of lipids to self-segregate to generate a highly ordered membrane for electrical insulation of axons.

  19. Membrane Structure: Lipid-Protein Interactions in Microsomal Membranes*

    PubMed Central

    Trump, Benjamin F.; Duttera, Sue M.; Byrne, William L.; Arstila, Antti U.

    1970-01-01

    The relationships of phospholipid to membrane structure and function were examined in hepatic microsomes. Findings indicate that normal microsomal membrane structure is dependent on lipid-protein interactions and that it correlates closely with glucose-6-phosphatase activity. Modification of most phospholipid with phospholipase-C is associated with widening of the membrane which can be reversed following readdition of phospholipid. Images PMID:4317915

  20. Biosynthesis of archaeal membrane ether lipids

    PubMed Central

    Jain, Samta; Caforio, Antonella; Driessen, Arnold J. M.

    2014-01-01

    A vital function of the cell membrane in all living organism is to maintain the membrane permeability barrier and fluidity. The composition of the phospholipid bilayer is distinct in archaea when compared to bacteria and eukarya. In archaea, isoprenoid hydrocarbon side chains are linked via an ether bond to the sn-glycerol-1-phosphate backbone. In bacteria and eukarya on the other hand, fatty acid side chains are linked via an ester bond to the sn-glycerol-3-phosphate backbone. The polar head groups are globally shared in the three domains of life. The unique membrane lipids of archaea have been implicated not only in the survival and adaptation of the organisms to extreme environments but also to form the basis of the membrane composition of the last universal common ancestor (LUCA). In nature, a diverse range of archaeal lipids is found, the most common are the diether (or archaeol) and the tetraether (or caldarchaeol) lipids that form a monolayer. Variations in chain length, cyclization and other modifications lead to diversification of these lipids. The biosynthesis of these lipids is not yet well understood however progress in the last decade has led to a comprehensive understanding of the biosynthesis of archaeol. This review describes the current knowledge of the biosynthetic pathway of archaeal ether lipids; insights on the stability and robustness of archaeal lipid membranes; and evolutionary aspects of the lipid divide and the LUCA. It examines recent advances made in the field of pathway reconstruction in bacteria. PMID:25505460

  1. Human milk fat globules: polar lipid composition and in situ structural investigations revealing the heterogeneous distribution of proteins and the lateral segregation of sphingomyelin in the biological membrane.

    PubMed

    Lopez, Christelle; Ménard, Olivia

    2011-03-01

    Although human milk fat globules (MFG) are of primary importance since they are the exclusive lipid delivery carriers in the gastrointestinal tract of breast-fed infants, they remain the poorly understood aspect of milk. The objectives of this study were to investigate these unique colloidal assemblies and their interfacial properties, i.e. composition and structure of their biological membrane. In mature breast milk, MFG have a mean diameter of 4-5 microm, a surface area of about 2m(2)/g fat and an apparent zeta potential ζ=-6.7 ± 0.5 mV at 37°C. Human MFG contain 3-4mg polar lipids/g fat as quantified by HPLC/ELSD. The main polar lipids are sphingomyelin (SM; 36-45%, w/w), phosphatidylcholine (19-23%, w/w) and phosphatidylethanolamine (10-15%, w/w). In situ structural investigations of human MFG have been performed using light and confocal microscopy with adapted fluorescent probes, i.e. Nile Red, the extrinsic phospholipid Rh-DOPE, Fast Green and the lectin WGA-488. This study revealed a spatial heterogeneity in the human milk fat globule membrane (MFGM), with the lateral segregation of SM in liquid-ordered phase domains of various shapes and sizes surrounded by a liquid-disordered phase composed of the glycerophospholipids in which the proteins are dispersed. The glycocalyx formed by glycoproteins and cytoplasmic remnents have also been characterised around human MFG. A new model for the structure of the human MFGM is proposed and discussed. The unique composition and lateral organisation of the human MFGM components could be of metabolic significance and have health impact for the infants that need to be further explored.

  2. Liquid immiscibility in model bilayer lipid membranes

    NASA Astrophysics Data System (ADS)

    Veatch, Sarah L.

    There is growing evidence that cell plasma membranes are laterally organized into "raft" regions in which particular lipids and proteins are concentrated. These domains have sub-micron dimensions and have been implicated in vital cell functions. Similar liquid domains are observed in model bilayer membrane mixtures that mimick cellular lipid compositions. In model membranes, domains can be large (microns) and can readily form in the absence of proteins. This thesis presents studies of liquid immiscibility in model membrane systems using two experimental methods. By fluorescence microscopy, this thesis documents that miscibility transitions occur in a wide variety of ternary lipid mixtures containing high melting temperature (saturated) lipids, low melting temperature (usually unsaturated) lipids, and cholesterol. I have constructed detailed miscibility phase diagrams for three separate ternary lipid mixtures (DOPC/DPPC/Chol, DOPC/PSM/Chol, and POPC/PSM/Chol). Phase separation is also observed in membranes of lipids extracted from human erythrocytes. NMR experiments probe lipid order and verify the coexistence of a saturated lipid and cholesterol rich liquid ordered (Lo) phase with a more disordered, unsaturated lipid rich liquid crystalline (Lalpha) phase at low temperatures. These experiments also find multiple thermodynamic transitions and lipid organization on different length-scales. This complexity is revealed because fluorescence microscopy and NMR probe lipid order at different length-scales (>1mum vs. ˜100nm). NMR detects small domains (˜80nm) at temperatures just below the miscibility transition, even though micron-scale domains are observed by fluorescent microscopy. NMR does detect large-scale ("100nm) demixing, but at a lower temperature. In addition, it has long been known that >10nm length-scale structure is present in many lipid mixtures containing cholesterol and at least one additional lipid species, though it is shown here that only a subset of

  3. Crystallizing Membrane Proteins in Lipidic Mesophases. A Host Lipid Screen

    SciTech Connect

    Li, Dianfan; Lee, Jean; Caffrey, Martin

    2011-11-30

    The default lipid for the bulk of the crystallogenesis studies performed to date using the cubic mesophase method is monoolein. There is no good reason, however, why this 18-carbon, cis-monounsaturated monoacylglycerol should be the preferred lipid for all target membrane proteins. The latter come from an array of biomembrane types with varying properties that include hydrophobic thickness, intrinsic curvature, lateral pressure profile, lipid and protein makeup, and compositional asymmetry. Thus, it seems reasonable that screening for crystallizability based on the identity of the lipid creating the hosting mesophase would be worthwhile. For this, monoacylglycerols with differing acyl chain characteristics, such as length and olefinic bond position, must be available. A lipid synthesis and purification program is in place in the author's laboratory to serve this need. In the current study with the outer membrane sugar transporter, OprB, we demonstrate the utility of host lipid screening as a means for generating diffraction-quality crystals. Host lipid screening is likely to prove a generally useful strategy for mesophase-based crystallization of membrane proteins.

  4. Aspirin Increases the Solubility of Cholesterol in Lipid Membranes

    NASA Astrophysics Data System (ADS)

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

    2014-03-01

    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.

  5. Lipid partitioning at the nuclear envelope controls membrane biogenesis

    PubMed Central

    Barbosa, Antonio Daniel; Sembongi, Hiroshi; Su, Wen-Min; Abreu, Susana; Reggiori, Fulvio; Carman, George M.; Siniossoglou, Symeon

    2015-01-01

    Partitioning of lipid precursors between membranes and storage is crucial for cell growth, and its disruption underlies pathologies such as cancer, obesity, and type 2 diabetes. However, the mechanisms and signals that regulate this process are largely unknown. In yeast, lipid precursors are mainly used for phospholipid synthesis in nutrient-rich conditions in order to sustain rapid proliferation but are redirected to triacylglycerol (TAG) stored in lipid droplets during starvation. Here we investigate how cells reprogram lipid metabolism in the endoplasmic reticulum. We show that the conserved phosphatidate (PA) phosphatase Pah1, which generates diacylglycerol from PA, targets a nuclear membrane subdomain that is in contact with growing lipid droplets and mediates TAG synthesis. We find that cytosol acidification activates the master regulator of Pah1, the Nem1-Spo7 complex, thus linking Pah1 activity to cellular metabolic status. In the absence of TAG storage capacity, Pah1 still binds the nuclear membrane, but lipid precursors are redirected toward phospholipids, resulting in nuclear deformation and a proliferation of endoplasmic reticulum membrane. We propose that, in response to growth signals, activation of Pah1 at the nuclear envelope acts as a switch to control the balance between membrane biogenesis and lipid storage. PMID:26269581

  6. Pushing the lipid envelope: using bio-inspired nanocomposites to understand and exploit lipid membrane limitations

    NASA Astrophysics Data System (ADS)

    Montano, Gabriel

    Lipids serve as the organizing matrix material for biological membranes, the site of interaction of cells with the external environment. . As such, lipids play a critical role in structure/function relationships of an extraordinary number of critical biological processes. In this talk, we will look at bio-inspired membrane assemblies to better understand the roles of lipids in biological systems as well as attempt to generate materials that can mimic and potentially advance upon biological membrane processes. First, we will investigate the response of lipids to adverse conditions. In particular, I will present data that demonstrates the response of lipids to harsh conditions and how such responses can be exploited to generate nanocomposite rearrangements. I will also show the effect of adding the endotoxin lipopolysaccharide (LPS) to lipid bilayer assemblies and describe implications on our understanding of LPS organization in biological systems as well as describe induced lipid modifications that can be exploited to organize membrane composites with precise, two-dimensional geometric control. Lastly, I will describe the use of amphiphilic block copolymers to create membrane nanocomposites capable of mimicking biological systems. In particular, I will describe the use of our polymer-based membranes in creating artificial photosynthetic assemblies that rival biological systems in function in a more flexible, dynamic matrix.

  7. Unconventional membrane lipid biosynthesis in Xanthomonas campestris.

    PubMed

    Aktas, Meriyem; Narberhaus, Franz

    2015-09-01

    All bacteria are surrounded by at least one bilayer membrane mainly composed of phospholipids (PLs). Biosynthesis of the most abundant PLs phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and cardiolipin (CL) is well understood in model bacteria such as Escherichia coli. It recently emerged, however, that the diversity of bacterial membrane lipids is huge and that not yet explored biosynthesis pathways exist, even for the common PLs. A good example is the plant pathogen Xanthomonas campestris pv. campestris. It contains PE, PG and CL as major lipids and small amounts of the N-methylated PE derivatives monomethyl PE and phosphatidylcholine (PC = trimethylated PE). Xanthomonas campestris uses a repertoire of canonical and non-canonical enzymes for the synthesis of its membrane lipids. In this minireview, we briefly recapitulate standard pathways and integrate three recently discovered pathways into the overall picture of bacterial membrane biosynthesis.

  8. Pore dynamics in lipid membranes

    NASA Astrophysics Data System (ADS)

    Gozen, I.; Dommersnes, P.

    2014-09-01

    Transient circular pores can open in plasma membrane of cells due to mechanical stress, and failure to repair such pores lead to cell death. Similar pores in the form of defects also exist among smectic membranes, such as in myelin sheaths or mitochondrial membranes. The formation and growth of membrane defects are associated with diseases, for example multiple sclerosis. A deeper understanding of membrane pore dynamics can provide a more refined picture of membrane integrity-related disease development, and possibly also treatment options and strategies. Pore dynamics is also of great importance regarding healthcare applications such as drug delivery, gene or as recently been implied, cancer therapy. The dynamics of pores significantly differ in stacks which are confined in 2D compared to those in cells or vesicles. In this short review, we will summarize the dynamics of different types of pores that can be observed in biological membranes, which include circular transient, fusion and hemi-fusion pores. We will dedicate a section to floral and fractal pores which were discovered a few years ago and have highly peculiar characteristics. Finally, we will discuss the repair mechanisms of large area pores in conjunction with the current cell membrane repair hypotheses.

  9. Probing the Lipid Membrane Dipole Potential by Atomic Force Microscopy

    PubMed Central

    Yang, Yi; Mayer, Kathryn M.; Wickremasinghe, Nissanka S.; Hafner, Jason H.

    2008-01-01

    The electrostatic properties of biological membranes can be described by three parameters: the transmembrane potential, the membrane surface potential, and the membrane dipole potential. The first two are well characterized in terms of their magnitudes and biological effects. The dipole potential, however, is not well characterized. Various methods to measure the membrane dipole potential indirectly yield different values, and there is not even agreement on the source of the membrane dipole moment. This ambiguity impedes investigations into the biological effects of the membrane dipole moment, which should be substantial considering the large interfacial fields with which it is associated. Electrostatic analysis of phosphatidylcholine lipid membranes with the atomic force microscope reveals a repulsive force between the negatively charged probe tips and the zwitterionic lipids. This unexpected interaction has been analyzed quantitatively to reveal that the repulsion is due to a weak external field created by the internal membrane dipole potential. The analysis yields a dipole moment of 1.5 Debye per lipid with a dipole potential of +275 mV for supported phosphatidylcholine membranes. This new ability to quantitatively measure the membrane dipole moment in a noninvasive manner with nanometer scale spatial resolution will be useful in identifying the biological effects of the dipole potential. PMID:18805919

  10. Membrane-spanning lipids for an uncompromised monitoring of membrane fusion and intermembrane lipid transfer.

    PubMed

    Schwarzmann, Günter; Breiden, Bernadette; Sandhoff, Konrad

    2015-10-01

    A Förster resonance energy transfer-based fusion and transfer assay was developed to study, in model membranes, protein-mediated membrane fusion and intermembrane lipid transfer of fluorescent sphingolipid analogs. For this assay, it became necessary to apply labeled reporter molecules that are resistant to spontaneous as well as protein-mediated intermembrane transfer. The novelty of this assay is the use of nonextractable fluorescent membrane-spanning bipolar lipids. Starting from the tetraether lipid caldarchaeol, we synthesized fluorescent analogs with fluorophores at both polar ends. In addition, we synthesized radioactive glycosylated caldarchaeols. These labeled lipids were shown to stretch through bilayer membranes rather than to loop within a single lipid layer of liposomes. More important, the membrane-spanning lipids (MSLs) in contrast to phosphoglycerides proved to be nonextractable by proteins. We could show that the GM2 activator protein (GM2AP) is promiscuous with respect to glycero- and sphingolipid transfer. Saposin (Sap) B also transferred sphingolipids albeit with kinetics different from GM2AP. In addition, we could unambiguously show that the recombinant activator protein Sap C x His6 induced membrane fusion rather than intermembrane lipid transfer. These findings showed that these novel MSLs, in contrast with fluorescent phosphoglycerolipids, are well suited for an uncompromised monitoring of membrane fusion and intermembrane lipid transfer.

  11. Membrane-spanning lipids for an uncompromised monitoring of membrane fusion and intermembrane lipid transfer

    PubMed Central

    Schwarzmann, Günter; Breiden, Bernadette; Sandhoff, Konrad

    2015-01-01

    A Förster resonance energy transfer-based fusion and transfer assay was developed to study, in model membranes, protein-mediated membrane fusion and intermembrane lipid transfer of fluorescent sphingolipid analogs. For this assay, it became necessary to apply labeled reporter molecules that are resistant to spontaneous as well as protein-mediated intermembrane transfer. The novelty of this assay is the use of nonextractable fluorescent membrane-spanning bipolar lipids. Starting from the tetraether lipid caldarchaeol, we synthesized fluorescent analogs with fluorophores at both polar ends. In addition, we synthesized radioactive glycosylated caldarchaeols. These labeled lipids were shown to stretch through bilayer membranes rather than to loop within a single lipid layer of liposomes. More important, the membrane-spanning lipids (MSLs) in contrast to phosphoglycerides proved to be nonextractable by proteins. We could show that the GM2 activator protein (GM2AP) is promiscuous with respect to glycero- and sphingolipid transfer. Saposin (Sap) B also transferred sphingolipids albeit with kinetics different from GM2AP. In addition, we could unambiguously show that the recombinant activator protein Sap C x His6 induced membrane fusion rather than intermembrane lipid transfer. These findings showed that these novel MSLs, in contrast with fluorescent phosphoglycerolipids, are well suited for an uncompromised monitoring of membrane fusion and intermembrane lipid transfer. PMID:26269359

  12. The adrenal specific toxicant mitotane directly interacts with lipid membranes and alters membrane properties depending on lipid composition.

    PubMed

    Scheidt, Holger A; Haralampiev, Ivan; Theisgen, Stephan; Schirbel, Andreas; Sbiera, Silviu; Huster, Daniel; Kroiss, Matthias; Müller, Peter

    2016-06-15

    Mitotane (o,p'.-DDD) is an orphan drug approved for the treatment of adrenocortical carcinoma. The mechanisms, which are responsible for this activity of the drug, are not completely understood. It can be hypothesized that an impact of mitotane is mediated by the interaction with cellular membranes. However, an interaction of mitotane with (lipid) membranes has not yet been investigated in detail. Here, we characterized the interaction of mitotane and its main metabolite o,p'-dichlorodiphenyldichloroacetic acid (o,p'-DDA) with lipid membranes by applying a variety of biophysical approaches of nuclear magnetic resonance, electron spin resonance, and fluorescence spectroscopy. We found that mitotane and o,p'-DDA bind to lipid membranes by inserting into the lipid-water interface of the bilayer. Mitotane but not o,p'-DDA directly causes a disturbance of bilayer structure leading to an increased permeability of the membrane for polar molecules. Mitotane induced alterations of the membrane integrity required the presence of phosphatidylethanolamine and/or cholesterol. Collectively, our data for the first time characterize the impact of mitotane on the lipid membrane structure and dynamics, which may contribute to a better understanding of specific mitotane effects and side effects.

  13. Single-molecule investigation of the interactions between reconstituted planar lipid membranes and an analogue of the HP(2-20) antimicrobial peptide.

    PubMed

    Mereuta, Loredana; Luchian, Tudor; Park, Yoonkyung; Hahm, Kyung-Soo

    2008-09-05

    In this study, we employed electrophysiology experiments carried out at the single-molecule level to study the mechanism of action of the HPA3 peptide, an analogue of the linear antimicrobial peptide, HP(2-20), isolated from the N-terminal region of the Helicobacter pylori ribosomal protein. Amplitude analysis of currents fluctuations induced by HPA3 peptide at various potentials in zwitterionic lipid membranes reveal the existence of reproducible conductive states in the stochastic behavior of such events, which directly supports the existence of transmembrane pores induced the peptide. From our data recorded both at the single-pore and macroscopic levels, we propose that the HPA3 pore formation is electrophoretically facilitated by trans-negative transmembrane potentials, and HPA3 peptides translocate into the trans monolayers after forming the pores. We present evidence according to which the decrease in the membrane dipole potential of a reconstituted lipid membranes leads to an augmentation of the membrane activity of HPA3 peptides, and propose that a lower electric dipole field of the interfacial region of the membrane caused by phloretin facilitates the surface-bound HPA3 peptides to break free from one leaflet of the membrane, insert into the membrane and contribute to pore formation spanning the entire thickness of the membrane.

  14. Single-molecule investigation of the interactions between reconstituted planar lipid membranes and an analogue of the HP(2-20) antimicrobial peptide

    SciTech Connect

    Mereuta, Loredana; Luchian, Tudor Park, Yoonkyung; Hahm, Kyung-Soo

    2008-09-05

    In this study, we employed electrophysiology experiments carried out at the single-molecule level to study the mechanism of action of the HPA3 peptide, an analogue of the linear antimicrobial peptide, HP(2-20), isolated from the N-terminal region of the Helicobacter pylori ribosomal protein. Amplitude analysis of currents fluctuations induced by HPA3 peptide at various potentials in zwitterionic lipid membranes reveal the existence of reproducible conductive states in the stochastic behavior of such events, which directly supports the existence of transmembrane pores induced the peptide. From our data recorded both at the single-pore and macroscopic levels, we propose that the HPA3 pore formation is electrophoretically facilitated by trans-negative transmembrane potentials, and HPA3 peptides translocate into the trans monolayers after forming the pores. We present evidence according to which the decrease in the membrane dipole potential of a reconstituted lipid membranes leads to an augmentation of the membrane activity of HPA3 peptides, and propose that a lower electric dipole field of the interfacial region of the membrane caused by phloretin facilitates the surface-bound HPA3 peptides to break free from one leaflet of the membrane, insert into the membrane and contribute to pore formation spanning the entire thickness of the membrane.

  15. Why Hydrophilic Water can Permeate Hydrophobic Interior of Lipid Membranes

    NASA Astrophysics Data System (ADS)

    Qiao, Baofu; Olvera de La Cruz, Monica

    2014-03-01

    Water molecules as well as some small molecules have long been found to be able to diffuse across lipid membranes. Such permeation is of significant biological and biotechnological importance. For instance, the permeation of water across lipid membrane plays a important role in regulating ionic concentrations inside of cells. Such water permeation without the assistance of proteins embedded in membranes has been found to be a energetically unfavorable process. We, for the first time, explicitly depict the driving force for such an energetically unfavorable process. Atomistic molecular dynamics simulations are employed to investigate water diffusion in both liquid-crystalline and ordered gel phases of membranes containing zwitterionic DPPC or anionic DLPS lipid. The membrane conformation is calculated to have a critical role in water permeation, regardless of the type of lipid. The fluctuations in the potential energy are found to have a significant, if not the exclusive, role in the transportation of water across lipid membranes. Our results are also informative for the diffusion of small molecules of CO2, O2 and drug molecules, the absence of diffusion of ions, and the diffusion of water into the hydrophobic pores of carbon nanotubes. The authors acknowledge the support from the Office of the Director of Defense Research and Engineering (DDR & E) under Award No. FA9550-10-1-0167.

  16. Nanosecond Lipid Dynamics in Membranes Containing Cholesterol

    SciTech Connect

    Armstrong, Clare L; Haeussler, Wolfgang; Seydel, Tilo; Katsaras, John; Rheinstadter, Maikel C

    2014-01-01

    Lipid dynamics in the cholesterol-rich (40 mol%) liquid-ordered (lo) phase of dimyristoylphosphatidylcholine membranes were studied using neutron spin-echo and neutron backscattering. Recent theoretical and experimental evidence supports the notion of the liquid-ordered phase in phospholipid membranes as a locally structured liquid, with small ordered domains of a highly dynamic nature in equilibrium with a disordered matrix [S. Meinhardt, R. L. C. Vink and F. Schmid, Proc. Natl. Acad. Sci. U. S. A., 2013, 110(12), 4476 4481, C. L. Armstrong et al., PLoS One, 2013, 8(6), e66162]. This local structure was found to have a pronounced impact on the membranes' dynamical properties. We found that the long-wavelength dynamics in the liquid-ordered phase, associated with the elastic properties of the membranes, were faster by two orders of magnitude as compared to the liquid disordered phase. At the same time, collective nanoscale diffusion was significantly slower. The presence of a soft-mode (a slowing down) in the longwavelength dispersion relationship suggests an upper size limit for the ordered lipid domain of ~220 A. Moreover, from the relaxation rate of the collective lipid diffusion of lipid lipid distances, the lifetime of these domains was estimated to be about 100 nanoseconds.

  17. Engineering Lipid Bilayer Membranes for Protein Studies

    PubMed Central

    Khan, Muhammad Shuja; Dosoky, Noura Sayed; Williams, John Dalton

    2013-01-01

    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. PMID:24185908

  18. Inducing morphological changes in lipid bilayer membranes with microfabricated substrates

    NASA Astrophysics Data System (ADS)

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

    2016-11-01

    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.

  19. Photoinduced Fusion of Lipid Bilayer Membranes.

    PubMed

    Suzuki, Yui; Nagai, Ken H; Zinchenko, Anatoly; Hamada, Tsutomu

    2017-03-14

    We have developed a novel system for photocontrol of the fusion of lipid vesicles through the use of a photosensitive surfactant containing an azobenzene moiety (AzoTAB). Real-time microscopic observations clarified a change in both the surface area and internal volume of vesicles during fusion. We also determined the optimal cholesterol concentrations and temperature for inducing fusion. The mechanism of fusion can be attributed to a change in membrane tension, which is caused by the solubilization of lipids through the isomerization of AzoTAB. We used a micropipet technique to estimate membrane tension and discuss the mechanism of fusion in terms of membrane elastic energy. The obtained results regarding this novel photoinduced fusion could lead to a better understanding of the mechanism of membrane fusion in living cells and may also see wider applications, such as in drug delivery and biomimetic material design.

  20. Preparation of supported lipid membranes for aquaporin Z incorporation.

    PubMed

    Li, Xuesong; Wang, Rong; Tang, Chuyang; Vararattanavech, Ardcharaporn; Zhao, Yang; Torres, Jaume; Fane, Tony

    2012-06-01

    There has been a recent surge of interest to mimic the performance of natural cellular membranes by incorporating water channel proteins-aquaporins (AQPs) into various ultrathin films for water filtration applications. To make biomimetic membranes one of the most crucial steps is preparing a defect-free platform for AQPs incorporation on a suitable substrate. In this study two methods were used to prepare supported lipid membranes on NF membrane surfaces under a benign pH condition of 7.8. One method was direct vesicle fusion on a hydrophilic membrane NF-270; the other was vesicle fusion facilitated by hydraulic pressure on a modified hydrophilic NF-270 membrane whose surface has been spin-coated with positively charged lipids. Experiments revealed that the supported lipid membrane without AQPs prepared by the spin coating plus vesicle fusion had a much lower defect density than that prepared by vesicle fusion alone. It appears that the surface roughness and charge are the main factors determining the quality of the supported lipid membrane. Aquaporin Z (AqpZ) proteins were successfully incorporated into 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes and its permeability was measured by the stopped-flow experimental procedure. However, after the proteoliposomes have been fused onto the modified substrate, the AqpZ function in the resultant membrane was not observed and AFM images showed distinct aggregations of unfused proteoliposomes or AqpZ proteins on the substrate surface. It is speculated that the inhibition of AqpZ function may be caused by the low lipid mobility on the NF membrane surface. Further investigations to evaluate and optimize the structure-performance relationship are required.

  1. Applications of Mass Spectrometry to Lipids and Membranes

    PubMed Central

    Harkewicz, Richard; Dennis, Edward A.

    2012-01-01

    Lipidomics, a major part of metabolomics, constitutes the detailed analysis and global characterization, both spatial and temporal, of the structure and function of lipids (the lipidome) within a living system. As with proteomics, mass spectrometry has earned a central analytical role in lipidomics, and this role will continue to grow with technological developments. Currently, there exist two mass spectrometry-based lipidomics approaches, one based on a division of lipids into categories and classes prior to analysis, the “comprehensive lipidomics analysis by separation simplification” (CLASS), and the other in which all lipid species are analyzed together without prior separation, shotgun. In exploring the lipidome of various living systems, novel lipids are being discovered, and mass spectrometry is helping characterize their chemical structure. Deuterium exchange mass spectrometry (DXMS) is being used to investigate the association of lipids and membranes with proteins and enzymes, and imaging mass spectrometry (IMS) is being applied to the in situ analysis of lipids in tissues. PMID:21469951

  2. Super-resolution optical microscopy of lipid plasma membrane dynamics.

    PubMed

    Eggeling, Christian

    2015-01-01

    Plasma membrane dynamics are an important ruler of cellular activity, particularly through the interaction and diffusion dynamics of membrane-embedded proteins and lipids. FCS (fluorescence correlation spectroscopy) on an optical (confocal) microscope is a popular tool for investigating such dynamics. Unfortunately, its full applicability is constrained by the limited spatial resolution of a conventional optical microscope. The present chapter depicts the combination of optical super-resolution STED (stimulated emission depletion) microscopy with FCS, and why it is an important tool for investigating molecular membrane dynamics in living cells. Compared with conventional FCS, the STED-FCS approach demonstrates an improved possibility to distinguish free from anomalous molecular diffusion, and thus to give new insights into lipid-protein interactions and the traditional lipid 'raft' theory.

  3. Nonlinear adhesion dynamics of confined lipid membranes

    NASA Astrophysics Data System (ADS)

    To, Tung; Le Goff, Thomas; Pierre-Louis, Olivier

    Lipid membranes, which are ubiquitous objects in biological environments are often confined. For example, they can be sandwiched between a substrate and the cytoskeleton between cell adhesion, or between other membranes in stacks, or in the Golgi apparatus. We present a study of the nonlinear dynamics of membranes in a model system, where the membrane is confined between two flat walls. The dynamics derived from the lubrication approximation is highly nonlinear and nonlocal. The solution of this model in one dimension exhibits frozen states due to oscillatory interactions between membranes caused by the bending rigidity. We develope a kink model for these phenomena based on the historical work of Kawasaki and Otha. In two dimensions, the dynamics is more complex, and depends strongly on the amount of excess area in the system. We discuss the relevance of our findings for experiments on model membranes, and for biological systems. Supported by the grand ANR Biolub.

  4. Bacterial membrane lipids: diversity in structures and pathways.

    PubMed

    Sohlenkamp, Christian; Geiger, Otto

    2016-01-01

    For many decades, Escherichia coli was the main model organism for the study of bacterial membrane lipids. The results obtained served as a blueprint for membrane lipid biochemistry, but it is clear now that there is no such thing as a typical bacterial membrane lipid composition. Different bacterial species display different membrane compositions and even the membrane composition of cells belonging to a single species is not constant, but depends on the environmental conditions to which the cells are exposed. Bacterial membranes present a large diversity of amphiphilic lipids, including the common phospholipids phosphatidylglycerol, phosphatidylethanolamine and cardiolipin, the less frequent phospholipids phosphatidylcholine, and phosphatidylinositol and a variety of other membrane lipids, such as for example ornithine lipids, glycolipids, sphingolipids or hopanoids among others. In this review, we give an overview about the membrane lipid structures known in bacteria, the different metabolic pathways involved in their formation, and the distribution of membrane lipids and metabolic pathways across taxonomical groups.

  5. Atomistic Monte Carlo Simulation of Lipid Membranes

    PubMed Central

    Wüstner, Daniel; Sklenar, Heinz

    2014-01-01

    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. PMID:24469314

  6. Interaction of tau protein with model lipid membranes induces tau structural compaction and membrane disruption

    PubMed Central

    Jones, Emmalee M.; Dubey, Manish; Camp, Phillip J.; Vernon, Briana C.; Biernat, Jacek; Mandelkow, Eckhard; Majewski, Jaroslaw; Chi, Eva Y.

    2012-01-01

    The misfolding and aggregation of the intrinsically disordered, microtubule-associated tau protein into neurofibrillary tangles is implicated in the pathogenesis of Alzheimer's disease. However, the mechanisms of tau aggregation and toxicity remain unknown. Recent work has shown that lipid membrane can induce tau aggregation and that membrane permeabilization may serve as a pathway by which protein aggregates exert toxicity, suggesting that the plasma membrane may play dual roles in tau pathology. This prompted our investigation to assess tau's propensity to interact with membranes and to elucidate the mutually disruptive structural perturbations the interactions induce in both tau and the membrane. We show that although highly charged and soluble, the full-length tau (hTau40) is also highly surface active, selectively inserts into anionic DMPG lipid monolayers and induces membrane morphological changes. To resolve molecular-scale structural details of hTau40 associated with lipid membranes, X-ray and neutron scattering techniques are utilized. X-ray reflectivity indicates hTau40's presence underneath a DMPG monolayer and penetration into the lipid headgroups and tailgroups, whereas grazing incidence X-ray diffraction shows that hTau40 insertion disrupts lipid packing. Moreover, both air/water and DMPG lipid membrane interfaces induce the disordered hTau40 to partially adopt a more compact conformation with density similar to that of a folded protein. Neutron reflectivity shows that tau completely disrupts supported DMPG bilayers while leaving the neutral DPPC bilayer intact. Our results show that hTau40's strong interaction with anionic lipids induces tau structural compaction and membrane disruption, suggesting possible membrane-based mechanisms of tau aggregation and toxicity in neurodegenerative diseases. PMID:22401494

  7. Targeting Membrane Lipid a Potential Cancer Cure?

    PubMed Central

    Tan, Loh Teng-Hern; Chan, Kok-Gan; Pusparajah, Priyia; Lee, Wai-Leng; Chuah, Lay-Hong; Khan, Tahir Mehmood; Lee, Learn-Han; Goh, Bey-Hing

    2017-01-01

    Cancer mortality and morbidity is projected to increase significantly over the next few decades. Current chemotherapeutic strategies have significant limitations, and there is great interest in seeking novel therapies which are capable of specifically targeting cancer cells. Given that fundamental differences exist between the cellular membranes of healthy cells and tumor cells, novel therapies based on targeting membrane lipids in cancer cells is a promising approach that deserves attention in the field of anticancer drug development. Phosphatidylethanolamine (PE), a lipid membrane component which exists only in the inner leaflet of cell membrane under normal circumstances, has increased surface representation on the outer membrane of tumor cells with disrupted membrane asymmetry. PE thus represents a potential chemotherapeutic target as the higher exposure of PE on the membrane surface of cancer cells. This feature as well as a high degree of expression of PE on endothelial cells in tumor vasculature, makes PE an attractive molecular target for future cancer interventions. There have already been several small molecules and membrane-active peptides identified which bind specifically to the PE molecules on the cancer cell membrane, subsequently inducing membrane disruption leading to cell lysis. This approach opens up a new front in the battle against cancer, and is of particular interest as it may be a strategy that may be prove effective against tumors that respond poorly to current chemotherapeutic agents. We aim to highlight the evidence suggesting that PE is a strong candidate to be explored as a potential molecular target for membrane targeted novel anticancer therapy. PMID:28167913

  8. Dynamic Response of Model Lipid Membranes to Ultrasonic Radiation Force

    PubMed Central

    Prieto, Martin Loynaz; Oralkan, Ömer; Khuri-Yakub, Butrus T.; Maduke, Merritt C.

    2013-01-01

    Low-intensity ultrasound can modulate action potential firing in neurons in vitro and in vivo. It has been suggested that this effect is mediated by mechanical interactions of ultrasound with neural cell membranes. We investigated whether these proposed interactions could be reproduced for further study in a synthetic lipid bilayer system. We measured the response of protein-free model membranes to low-intensity ultrasound using electrophysiology and laser Doppler vibrometry. We find that ultrasonic radiation force causes oscillation and displacement of lipid membranes, resulting in small (<1%) changes in membrane area and capacitance. Under voltage-clamp, the changes in capacitance manifest as capacitive currents with an exponentially decaying sinusoidal time course. The membrane oscillation can be modeled as a fluid dynamic response to a step change in pressure caused by ultrasonic radiation force, which disrupts the balance of forces between bilayer tension and hydrostatic pressure. We also investigated the origin of the radiation force acting on the bilayer. Part of the radiation force results from the reflection of the ultrasound from the solution/air interface above the bilayer (an effect that is specific to our experimental configuration) but part appears to reflect a direct interaction of ultrasound with the bilayer, related to either acoustic streaming or scattering of sound by the bilayer. Based on these results, we conclude that synthetic lipid bilayers can be used to study the effects of ultrasound on cell membranes and membrane proteins. PMID:24194863

  9. Self-assembled tethered bimolecular lipid membranes.

    PubMed

    Sinner, Eva-Kathrin; Ritz, Sandra; Naumann, Renate; Schiller, Stefan; Knoll, Wolfgang

    2009-01-01

    This chapter describes some of the strategies developed in our group for designing, constructing and structurally and functionally characterizing tethered bimolecular lipid membranes (tBLM). We introduce this platform as a novel model membrane system that complements the existing ones, for example, Langmuir monolayers, vesicular liposomal dispersions and bimolecular ("black") lipid membranes. Moreover, it offers the additional advantage of allowing for studies of the influence of membrane structure and order on the function of integral proteins, for example, on how the composition and organization of lipids in a mixed membrane influence the ion translocation activity of integral channel proteins. The first strategy that we introduce concerns the preparation of tethered monolayers by the self-assembly of telechelics. Their molecular architecture with a headgroup, a spacer unit (the "tether") and the amphiphile that mimics the lipid molecule allows them to bind specifically to the solid support thus forming the proximal layer of the final architecture. After fusion of vesicles that could contain reconstituted proteins from a liposomal dispersion in contact to this monolayer the tethered bimolecular lipid membrane is obtained. This can then be characterized by a broad range of surface analytical techniques, including surface plasmon spectroscopies, the quartz crystal microbalance, fluorescence and IR spectroscopies, and electrochemical techniques, to mention a few. It is shown that this concept allows for the construction of tethered lipid bilayers with outstanding electrical properties including resistivities in excess of 10 MOmega cm2. A modified strategy uses the assembly of peptides as spacers that couple covalently via their engineered sulfhydryl or lipoic acid groups at the N-terminus to the employed gold substrate, while their C-terminus is being activated afterward for the coupling of, for example, dimyristoylphosphatidylethanol amine (DMPE) lipid molecules

  10. Anionic lipids and the maintenance of membrane electrostatics in eukaryotes

    PubMed Central

    Platre, Matthieu Pierre

    2017-01-01

    ABSTRACT A wide range of signaling processes occurs at the cell surface through the reversible association of proteins from the cytosol to the plasma membrane. Some low abundant lipids are enriched at the membrane of specific compartments and thereby contribute to the identity of cell organelles by acting as biochemical landmarks. Lipids also influence membrane biophysical properties, which emerge as an important feature in specifying cellular territories. Such parameters are crucial for signal transduction and include lipid packing, membrane curvature and electrostatics. In particular, membrane electrostatics specifies the identity of the plasma membrane inner leaflet. Membrane surface charges are carried by anionic phospholipids, however the exact nature of the lipid(s) that powers the plasma membrane electrostatic field varies among eukaryotes and has been hotly debated during the last decade. Herein, we discuss the role of anionic lipids in setting up plasma membrane electrostatics and we compare similarities and differences that were found in different eukaryotic cells. PMID:28102755

  11. Membrane lipids and the origin of life

    NASA Technical Reports Server (NTRS)

    Oro, J.; Holzer, G.; Rao, M.; Tornabene, T. G.

    1981-01-01

    The current state of knowledge regarding the development of biological systems is briefly reviewed. At a crucial stage concerning the evolution of such systems, the mechanisms leading to more complex structures must have evolved within the confines of a protected microenvironment, similar to those provided by the contemporary cell membranes. The major components found normally in biomembranes are phospholipids. The structure of the biomembrane is examined, and attention is given to questions concerning the availability of the structural components which are necessary in the formation of primitive lipid membranes. Two approaches regarding the study of protomembranes are discussed. The probability of obtaining ether lipids under prebiotic conditions is considered, taking into account the formation of cyclic and acyclic isoprenoids by the irradiation of isoprene with UV.

  12. Atomic force microscopy of model lipid membranes.

    PubMed

    Morandat, Sandrine; Azouzi, Slim; Beauvais, Estelle; Mastouri, Amira; El Kirat, Karim

    2013-02-01

    Supported lipid bilayers (SLBs) are biomimetic model systems that are now widely used to address the biophysical and biochemical properties of biological membranes. Two main methods are usually employed to form SLBs: the transfer of two successive monolayers by Langmuir-Blodgett or Langmuir-Schaefer techniques, and the fusion of preformed lipid vesicles. The transfer of lipid films on flat solid substrates offers the possibility to apply a wide range of surface analytical techniques that are very sensitive. Among them, atomic force microscopy (AFM) has opened new opportunities for determining the nanoscale organization of SLBs under physiological conditions. In this review, we first focus on the different protocols generally employed to prepare SLBs. Then, we describe AFM studies on the nanoscale lateral organization and mechanical properties of SLBs. Lastly, we survey recent developments in the AFM monitoring of bilayer alteration, remodeling, or digestion, by incubation with exogenous agents such as drugs, proteins, peptides, and nanoparticles.

  13. Cholesterol effect on the dipole potential of lipid membranes.

    PubMed

    Starke-Peterkovic, Thomas; Turner, Nigel; Vitha, Mark F; Waller, Mark P; Hibbs, David E; Clarke, Ronald J

    2006-06-01

    The effect of cholesterol removal by methyl-beta-cyclodextrin on the dipole potential, psi(d), of membrane vesicles composed of natural membrane lipids extracted from the kidney and brain of eight vertebrate species was investigated using the voltage-sensitive fluorescent probe di-8-ANEPPS. Cyclodextrin treatment reduced cholesterol levels by on average 80% and this was associated with an average reduction in psi(d) of 50 mV. Measurements of the effect of a range of cholesterol derivatives on the psi(d) of DMPC lipid vesicles showed that the magnitude of the effect correlated with the component of the sterol's dipole moment perpendicular to the membrane surface. The changes in psi(d) observed could not be accounted for solely by the electric field originating from the sterols' dipole moments. Additional factors must arise from sterol-induced changes in lipid packing, which changes the density of dipoles in the membrane, and changes in water penetration into the membrane, which changes the effective dielectric constant of the interfacial region. In DMPC membranes, the cholesterol-induced change in psi(d) was biphasic, i.e., a maximum in psi(d) was observed at approximately 35-45 mol %, after which psi(d) started to decrease. We suggest that this could be associated with a maximum in the strength of DMPC-cholesterol intermolecular forces at this composition.

  14. Multichannel taste sensors with lipid, lipid like polymer membranes

    NASA Astrophysics Data System (ADS)

    Szpakowska, M.; Szwacki, J.; Marjańska, E.

    2008-08-01

    The elaboration of a sensitive taste sensor for discrimination of different soft drinks is very important in food industry. The short review of taste sensors described in the literature is presented. Two types of potentiometric taste sensors, one with lipophilic compound-polymer membranes (ISE) and the other with lipid polymer membrane and a conducting polymer film (All solid state electrode, ASSE) were tested in appropriate taste solutions. Five channel ISE sensor was examined in acid, sour and sweet solutions. This sensor was sensitive to bitter and sour substances and not too sensitive to sucrose concentration. It was successfully used for discrimination of different kind of soft drinks. Four channel ASSE sensor was examined in sour solutions. It was found that stability and sensitivity of ASSE are lower than ISE. Therefore, it seems that the previous one cannot be applied in taste sensor.

  15. Influence of membrane lipid composition on flavonoid-membrane interactions: Implications on their biological activity.

    PubMed

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

    2015-04-01

    The membrane interactions and localization of flavonoids play a vital role in altering membrane-mediated cell signaling cascades as well as influence the pharmacological activities such as anti-tumour, anti-microbial and anti-oxidant properties of flavonoids. Various techniques have been used to investigate the membrane interaction of flavonoids. These include partition coefficient, fluorescence anisotropy, differential scanning calorimetry, NMR spectroscopy, electrophysiological methods and molecular dynamics simulations. Each technique will provide specific information about either alteration of membrane fluidity or localization of flavonoids within the lipid bilayer. Apart from the diverse techniques employed, the concentrations of flavonoids and lipid membrane composition employed in various studies reported in literature also are different and together these variables contribute to diverse findings that sometimes contradict each other. This review highlights different techniques employed to investigate the membrane interaction of flavonoids with special emphasis on erythrocyte model membrane systems and their significance in understanding the nature and extent of flavonoid-membrane interactions. We also attempt to correlate the membrane localization and alteration in membrane fluidity with the biological activities of flavonoids such as anti-oxidant, anti-cancer and anti-microbial properties.

  16. Lipid Gymnastics: Tethers and Fingers in membrane

    NASA Astrophysics Data System (ADS)

    Tayebi, Lobat; Miller, Gregory; Parikh, Atul

    2009-03-01

    A significant body of evidence now links local mesoscopic structure (e.g., shape and composition) of the cell membrane with its function; the mechanisms by which cellular membranes adopt the specific shapes remain poorly understood. Among all the different structures adopted by cellular membranes, the tubular shape is one of the most surprising one. While their formation is typically attributed to the reorganization of membrane cytoskeleton, many exceptions exist. We report the instantaneous formation of tubular membrane mesophases following the hydration under specific thermal conditions. The shapes emerge in a bimodal way where we have two distinct diameter ranges for tubes, ˜20μm and ˜1μm, namely fat fingers and narrow tethers. We study the roughening of hydrated drops of 3 lipids in 3 different spontaneous curvatures at various temp. and ionic strength to figure out the dominant effect in selection of tethers and fingers. Dynamics of the tubes are of particular interest where we observe four distinct steps of birth, coiling, uncoiling and retraction with different lifetime on different thermal condition. These dynamics appear to reflect interplay between membrane elasticity, surface adhesion, and thermal or hydrodynamic gradient.

  17. Effect of Amphotericin B antibiotic on the properties of model lipid membrane

    NASA Astrophysics Data System (ADS)

    Kiryakova, S.; Dencheva-Zarkova, M.; Genova, J.

    2014-12-01

    Model membranes formed from natural and synthetic lipids are an interesting object for scientific investigations due to their similarity to biological cell membrane and their simple structure with controlled composition and properties. Amphotericin B is an important polyene antifungal antibiotic, used for treatment of systemic fungal infections. It is known from the literature that the studied antibiotic has a substantial effect on the transmembrane ionic channel structures. When applied to the lipid membranes it has the tendency to create pores and in this way to affect the structure and the properties of the membrane lipid bilayer. In this work the thermally induced shape fluctuations of giant quasi-spherical liposomes have been used to study the influence of polyene antibiotic amphotericin B on the elastic properties of model lipid membranes. It have been shown experimentally that the presence of 3 mol % of AmB in the lipid membrane reduces the bending elasticity of the lipid membrane for both studied cases: pure SOPC membrane and mixed SOPC-Cholesterol membrane. Interaction of the amphotericin B with bilayer lipid membranes containing channels have been studied in this work. Model membranes were self-assembled using the patch-clamp and tip-dip patch clamp technique. We have found that amphotericin B is an ionophore and reduces the resistance of the lipid bilayer.

  18. Carotenoid binding to proteins: Modeling pigment transport to lipid membranes.

    PubMed

    Reszczynska, Emilia; Welc, Renata; Grudzinski, Wojciech; Trebacz, Kazimierz; Gruszecki, Wieslaw I

    2015-10-15

    Carotenoid pigments play numerous important physiological functions in human organism. Very special is a role of lutein and zeaxanthin in the retina of an eye and in particular in its central part, the macula lutea. In the retina, carotenoids can be directly present in the lipid phase of the membranes or remain bound to the protein-pigment complexes. In this work we address a problem of binding of carotenoids to proteins and possible role of such structures in pigment transport to lipid membranes. Interaction of three carotenoids, beta-carotene, lutein and zeaxanthin with two proteins: bovine serum albumin and glutathione S-transferase (GST) was investigated with application of molecular spectroscopy techniques: UV-Vis absorption, circular dichroism and Fourier transform infrared spectroscopy (FTIR). Interaction of pigment-protein complexes with model lipid bilayers formed with egg yolk phosphatidylcholine was investigated with application of FTIR, Raman imaging of liposomes and electrophysiological technique, in the planar lipid bilayer models. The results show that in all the cases of protein and pigment studied, carotenoids bind to protein and that the complexes formed can interact with membranes. This means that protein-carotenoid complexes are capable of playing physiological role in pigment transport to biomembranes.

  19. Three-Phase Coexistence in Lipid Membranes.

    PubMed

    Aufderhorst-Roberts, Anders; Chandra, Udayan; Connell, Simon D

    2017-01-24

    Phospholipid ternary systems are useful model systems for understanding lipid-lipid interactions and their influence on biological properties such as cell signaling and protein translocation. Despite extensive studies, there are still open questions relating to membrane phase behavior, particularly relating to a proposed state of three-phase coexistence, due to the difficulty in clearly distinguishing the three phases. We look in and around the region of the phase diagram where three phases are expected and use a combination of different atomic force microscopy (AFM) modes to present the first images of three coexisting lipid phases in biomimetic cell lipid membranes. Domains form through either nucleation or spinodal decomposition dependent upon composition, with some exhibiting both mechanisms in different domains simultaneously. Slow cooling rates are necessary to sufficiently separate mixtures with high proportions of lo and lβ phase. We probe domain heights and mechanical properties and demonstrate that the gel (lβ) domains have unusually low structural integrity in the three-phase region. This finding supports the hypothesis of a "disordered gel" state that has been proposed from NMR studies of similar systems, where the addition of small amounts of cholesterol was shown to disrupt the regular packing of the lβ state. We use NMR data from the literature on chain disorder in different mixtures and estimate an expected step height that is in excellent agreement with the AFM data. Alternatively, the disordered solid phase observed here and in the wider literature could be explained by the lβ phase being out of equilibrium, in a surface kinetically trapped state. This view is supported by the observation of unusual growth of nucleated domains, which we term "tree-ring growth," reflecting compositional heterogeneity in large disordered lβ phase domains.

  20. Hydrostatic pressure decreases membrane fluidity and lipid desaturase expression in chondrocyte progenitor cells.

    PubMed

    Montagne, Kevin; Uchiyama, Hiroki; Furukawa, Katsuko S; Ushida, Takashi

    2014-01-22

    Membrane biomechanical properties are critical in modulating nutrient and metabolite exchange as well as signal transduction. Biological membranes are predominantly composed of lipids, cholesterol and proteins, and their fluidity is tightly regulated by cholesterol and lipid desaturases. To determine whether such membrane fluidity regulation occurred in mammalian cells under pressure, we investigated the effects of pressure on membrane lipid order of mouse chondrogenic ATDC5 cells and desaturase gene expression. Hydrostatic pressure linearly increased membrane lipid packing and simultaneously repressed lipid desaturase gene expression. We also showed that cholesterol mimicked and cholesterol depletion reversed those effects, suggesting that desaturase gene expression was controlled by the membrane physical state itself. This study demonstrates a new effect of hydrostatic pressure on mammalian cells and may help to identify the molecular mechanisms involved in hydrostatic pressure sensing in chondrocytes.

  1. A sliding selectivity scale for lipid binding to membrane proteins

    PubMed Central

    Landreh, Michael; Marty, Michael T.; Gault, Joseph; Robinson, Carol V.

    2017-01-01

    Biological membranes form barriers that are essential for cellular integrity and compartmentalisation. Proteins that reside in the membrane have co-evolved with their hydrophobic lipid environment which serves as a solvent for proteins with very diverse requirements. As a result, membrane protein-lipid interactions range from completely non-selective to highly discriminating. Mass spectrometry (MS), in combination with X-ray crystallography and molecular dynamics simulations, enables us to monitor how lipids interact with intact membrane protein complexes and assess their effects on structure and dynamics. Recent studies illustrate the ability to differentiate specific lipid binding, preferential interactions with lipid subsets, and nonselective annular contacts. In this review, we consider the biological implications of different lipid-binding scenarios and propose that binding occurs on a sliding selectivity scale, in line with the view of biological membranes as facilitators of dynamic protein and lipid organization. PMID:27155089

  2. Force Field Development for Lipid Membrane Simulations.

    PubMed

    Lyubartsev, Alexander P; Rabinovich, Alexander L

    2016-10-01

    With the rapid development of computer power and wide availability of modelling software computer simulations of realistic models of lipid membranes, including their interactions with various molecular species, polypeptides and membrane proteins have become feasible for many research groups. The crucial issue of the reliability of such simulations is the quality of the force field, and many efforts, especially in the latest several years, have been devoted to parametrization and optimization of the force fields for biomembrane modelling. In this review, we give account of the recent development in this area, covering different classes of force fields, principles of the force field parametrization, comparison of the force fields, and their experimental validation. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg.

  3. Membrane Contact Sites: Complex Zones for Membrane Association and Lipid Exchange

    PubMed Central

    Quon, Evan; Beh, Christopher T.

    2015-01-01

    Lipid transport between membranes within cells involves vesicle and protein carriers, but as agents of nonvesicular lipid transfer, the role of membrane contact sites has received increasing attention. As zones for lipid metabolism and exchange, various membrane contact sites mediate direct associations between different organelles. In particular, membrane contact sites linking the plasma membrane (PM) and the endoplasmic reticulum (ER) represent important regulators of lipid and ion transfer. In yeast, cortical ER is stapled to the PM through membrane-tethering proteins, which establish a direct connection between the membranes. In this review, we consider passive and facilitated models for lipid transfer at PM–ER contact sites. Besides the tethering proteins, we examine the roles of an additional repertoire of lipid and protein regulators that prime and propagate PM–ER membrane association. We conclude that instead of being simple mediators of membrane association, regulatory components of membrane contact sites have complex and multilayered functions. PMID:26949334

  4. Tuning Membrane Thickness Fluctuations in Model Lipid Bilayers

    PubMed Central

    Ashkar, Rana; Nagao, Michihiro; Butler, Paul D.; Woodka, Andrea C.; Sen, Mani K.; Koga, Tadanori

    2015-01-01

    Membrane thickness fluctuations have been associated with a variety of critical membrane phenomena, such as cellular exchange, pore formation, and protein binding, which are intimately related to cell functionality and effective pharmaceuticals. Therefore, understanding how these fluctuations are controlled can remarkably impact medical applications involving selective macromolecule binding and efficient cellular drug intake. Interestingly, previous reports on single-component bilayers show almost identical thickness fluctuation patterns for all investigated lipid tail-lengths, with similar temperature-independent membrane thickness fluctuation amplitude in the fluid phase and a rapid suppression of fluctuations upon transition to the gel phase. Presumably, in vivo functions require a tunability of these parameters, suggesting that more complex model systems are necessary. In this study, we explore lipid tail-length mismatch as a regulator for membrane fluctuations. Unilamellar vesicles of an equimolar mixture of dimyristoylphosphatidylcholine and distearoylphosphatidylcholine molecules, with different tail-lengths and melting transition temperatures, are used as a model system for this next level of complexity. Indeed, this binary system exhibits a significant response of membrane dynamics to thermal variations. The system also suggests a decoupling of the amplitude and the relaxation time of the membrane thickness fluctuations, implying a potential for independent control of these two key parameters. PMID:26153707

  5. Reconstitution of a Kv Channel into Lipid Membranes for Structural and Functional Studies

    PubMed Central

    Shi, Liang; Jiang, Qiu-Xing

    2013-01-01

    To study the lipid-protein interaction in a reductionistic fashion, it is necessary to incorporate the membrane proteins into membranes of well-defined lipid composition. We are studying the lipid-dependent gating effects in a prototype voltage-gated potassium (Kv) channel, and have worked out detailed procedures to reconstitute the channels into different membrane systems. Our reconstitution procedures take consideration of both detergent-induced fusion of vesicles and the fusion of protein/detergent micelles with the lipid/detergent mixed micelles as well as the importance of reaching an equilibrium distribution of lipids among the protein/detergent/lipid and the detergent/lipid mixed micelles. Our data suggested that the insertion of the channels in the lipid vesicles is relatively random in orientations, and the reconstitution efficiency is so high that no detectable protein aggregates were seen in fractionation experiments. We have utilized the reconstituted channels to determine the conformational states of the channels in different lipids, record electrical activities of a small number of channels incorporated in planar lipid bilayers, screen for conformation-specific ligands from a phage-displayed peptide library, and support the growth of 2D crystals of the channels in membranes. The reconstitution procedures described here may be adapted for studying other membrane proteins in lipid bilayers, especially for the investigation of the lipid effects on the eukaryotic voltage-gated ion channels. PMID:23892292

  6. Crystallizing Membrane Proteins Using Lipidic Mesophases

    PubMed Central

    Caffrey, Martin; Cherezov, Vadim

    2009-01-01

    A detailed protocol for crystallizing membrane proteins that makes use of lipidic mesophases is described. This has variously been referred to as the lipid cubic phase or in meso method. The method has been shown to be quite general in that it has been used to solve X-ray crystallographic structures of prokaryotic and eukaryotic proteins, proteins that are monomeric, homo- and hetero-multimeric, chromophore-containing and chromophore-free, and α-helical and β-barrel proteins. Its most recent successes are the human engineered β2-adrenergic and adenosine A2A G protein-coupled receptors. Protocols are provided for preparing and characterizing the lipidic mesophase, for reconstituting the protein into the monoolein-based mesophase, for functional assay of the protein in the mesophase, and for setting up crystallizations in manual mode. Methods for harvesting micro-crystals are also described. The time required to prepare the protein-loaded mesophase and to set up a crystallization plate manually is about one hour. PMID:19390528

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

    NASA Astrophysics Data System (ADS)

    Rønnest, A. K.; Peters, G. H.; Hansen, F. Y.; Taub, H.; Miskowiec, A.

    2016-04-01

    Molecular dynamics simulations have been used to investigate the influence of the valency of counter-ions on the structure of freestanding bilayer membranes of the anionic 1,2-dimyristoyl-sn-glycero-3-phosphoglycerol (DMPG) lipid at 310 K and 1 atm. At this temperature, the membrane is in the fluid phase with a monovalent counter-ion and in the gel phase with a divalent counter-ion. The diffusion constant of water as a function of its depth in the membrane has been determined from mean-square-displacement calculations. Also, calculated incoherent quasielastic neutron scattering functions have been 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 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 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 for DMPG bilayers and found ˜1 V (˜2 ṡ 108 V m-1) when in the fluid phase with a monovalent counter-ion and ˜1.4 V (˜2.8 ṡ 108 V m-1) when in the gel phase with a divalent counter-ion. The number of water molecules for a fully hydrated DMPG membrane has been estimated to be 9.7 molecules per lipid 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

  8. Supported lipid bilayers as models for studying membrane domains.

    PubMed

    Kiessling, Volker; Yang, Sung-Tae; Tamm, Lukas K

    2015-01-01

    Supported lipid bilayers have been in use for over 30 years. They have been employed to study the structure, composition, and dynamics of lipid bilayer phases, the binding and distribution of soluble, integral, and lipidated proteins in membranes, membrane fusion, and interactions of membranes with elements of the cytoskeleton. This review focuses on the unique ability of supported lipid bilayers to study liquid-ordered and liquid-disordered domains in membranes. We highlight methods to produce asymmetric lipid bilayers with lipid compositions that mimic those of the extracellular and cytoplasmic leaflets of cell membranes and the functional reconstitution of membrane proteins into such systems. Questions related to interleaflet domain coupling and membrane protein activation have been addressed and answered using advanced reconstitution and imaging procedures in symmetric and asymmetric supported membranes with and without coexisting lipid phase domains. Previously controversial topics regarding anomalous and anisotropic diffusion in membranes have been resolved by using supported membrane approaches showing that the propensity of certain lipid compositions to form "rafts" are important but overlaid with "picket-fence" interactions that are imposed by a subtended cytoskeletal network.

  9. Film Balance Studies of Membrane Lipids and Related Molecules

    ERIC Educational Resources Information Center

    Cadenhead, D. A.

    1972-01-01

    Discusses apparatus, techniques, and measurements used to determine cell membrane composition. The use of a film balance to study monolayer membranes of selected lipids is described and results reported. (TS)

  10. [Role of membrane lipids in myocardial cytoprotection

    NASA Technical Reports Server (NTRS)

    Grynberg, A.

    2000-01-01

    The cardiomyocyte capacity to regulate ATP production to face any change in energy demand is a major determinant of cardiac function. This process is based on a balanced fatty acid (FA) metabolism, because FA is the main fuel of the heart, although the most expensive one in oxygen. The pathway is, however, weakly controlled by the cardiac myocyte which can well regulate FA mitochondrial entry but not cell FA uptake. For this reason, several pathological situations often result from either harmful accumulation of FA and derivatives or excess FA-oxidation. Control of the FA/glucose balance by decreased energy production from FA would thus offer an alternative strategy in the treatment of ischaemia, providing the cardiomyocytes weak ability in handling the non-metabolised FA is controlled. The initiation and the regulation of cardiac contraction both result from membrane activity; the other major role of lipids in the heart is their contribution to membrane homeostasis through phospholipid synthesis pathways and phospholipases. The anti-anginal activity of Trimetazidine, reported as a cytoprotective effect without a haemo-dynamic component; is associated with reduced use of FA for energy. However, accumulation of FA and derivatives has never been observed. Trimetazidine is reported to increase significantly the synthesis of phospholipids without influencing the other lipid classes, thus increasing the incorporation of FA in membrane structures. This cytoprotection appears to be based on the redirection of the use of FA to phospholipid synthesis, which would decrease their availability for energy production. This class of compounds, with the same properties as Trimetazidine, offers a metabolic approach to the treatment of ischaemia.

  11. Aspirin inhibits formation of cholesterol rafts in fluid lipid membranes.

    PubMed

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

    2015-03-01

    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.

  12. Analysis of membrane lipid biogenesis pathways using yeast genetics.

    PubMed

    Gsell, Martina; Daum, Günther

    2013-01-01

    The yeast Saccharomyces cerevisiae has become a valuable eukaryotic model organism to study biochemical and cellular processes at a molecular basis. A common strategy for such studies is the use of single and multiple mutants constructed by genetic manipulation which are compromised in individual enzymatic steps or certain metabolic pathways. Here, we describe selected examples of yeast research on phospholipid metabolism with emphasis on our own work dealing with investigations of phosphatidylethanolamine synthesis. Such studies start with the selection and construction of appropriate mutants and lead to phenotype analysis, lipid profiling, enzymatic analysis, and in vivo experiments. Comparing results obtained with wild-type and mutant strains allows us to understand the role of gene products and metabolic processes in more detail. Such studies are valuable not only for contributing to our knowledge of the complex network of lipid metabolism, but also of effects of lipids on structure and function of cellular membranes.

  13. Photopolymerization of Dienoyl Lipids Creates Planar Supported Poly(lipid) Membranes with Retained Fluidity.

    PubMed

    Orosz, Kristina S; Jones, Ian W; Keogh, John P; Smith, Christopher M; Griffin, Kaitlyn R; Xu, Juhua; Comi, Troy J; Hall, H K; Saavedra, S Scott

    2016-02-16

    Polymerization of substrate-supported bilayers composed of dienoylphosphatidylcholine (PC) lipids is known to greatly enhance their chemical and mechanical stability; however, the effects of polymerization on membrane fluidity have not been investigated. Here planar supported lipid bilayers (PSLBs) composed of dienoyl PCs on glass substrates were examined to assess the degree to which UV-initiated polymerization affects lateral lipid mobility. Fluorescence recovery after photobleaching (FRAP) was used to measure the diffusion coefficients (D) and mobile fractions of rhodamine-DOPE in unpolymerized and polymerized PSLBs composed of bis-sorbyl phosphatidylcholine (bis-SorbPC), mono-sorbyl-phosphatidylcholine (mono-SorbPC), bis-dienoyl-phosphatidylcholine (bis-DenPC), and mono-dienoyl phosphatidylcholine (mono-DenPC). Polymerization was performed in both the Lα and Lβ phase for each lipid. In all cases, polymerization reduced membrane fluidity; however, measurable lateral diffusion was retained which is attributed to a low degree of polymerization. The D values for sorbyl lipids were less than those of the denoyl lipids; this may be a consequence of the distal location of polymerizable group in the sorbyl lipids which may facilitate interleaflet bonding. The D values measured after polymerization were 0.1-0.8 of those measured before polymerization, a range that corresponds to fluidity intermediate between that of a Lα phase and a Lβ phase. This D range is comparable to ratios of D values reported for liquid-disordered (Ld) and liquid-ordered (Lo) lipid phases and indicates that the effect of UV polymerization on lateral diffusion in a dienoyl PSLB is similar to the transition from a Ld phase to a Lo phase. The partial retention of fluidity in UV-polymerized PSLBs, their enhanced stability, and the activity of incorporated transmembrane proteins and peptides is discussed.

  14. Photopolymerization of dienoyl lipids creates planar supported poly(lipid) membranes with retained fluidity

    PubMed Central

    Orosz, Kristina S.; Jones, Ian W.; Keogh, John P.; Smith, Christopher M.; Griffin, Kaitlyn R.; Xu, Juhua; Comi, Troy J.; Hall, H. K.

    2016-01-01

    Polymerization of substrate-supported bilayers composed of dienoyl phosphatidylcholine (PC) lipids is known to greatly enhance their chemical and mechanical stability, however the effects of polymerization on membrane fluidity have not been investigated. Here planar supported lipid bilayers (PSLBs) composed of dienoyl PCs on glass substrates were examined to assess the degree to which UV-initiated polymerization affects lateral lipid mobility. Fluorescence recovery after photobleaching (FRAP) was used to measure the diffusion coefficients (D) and mobile fractions of rhodamine-DOPE in unpolymerized and polymerized PSLBs composed of bis-sorbyl phosphatidylcholine (bis-SorbPC), mono-sorbyl phosphatidylcholine (mono-SorbPC), bis-dienoyl phosphatidylcholine (bis-DenPC) and mono-dienoyl phosphatidylcholine (mono-DenPC). Polymerization was performed in both the Lα and Lβ phase for each lipid. In all cases, polymerization reduced membrane fluidity, however measurable lateral diffusion was retained which is attributed to a low degree of polymerization. The D values for sorbyl lipids were less than those of the denoyl lipids; this may be a consequence of the distal location of polymerizable group in the sorbyl lipids which may facilitate inter-leaflet bonding. The D values measured after polymerization were 0.1 to 0.8 of those measured before polymerization, a range that corresponds to fluidity intermediate between that of a Lα phase and a Lβ phase. This D range is comparable to ratios of D values reported for liquid-disordered (Ld) and liquid-ordered (Lo) lipid phases, and indicates that the effect of UV polymerization on lateral diffusion in a dienoyl PSLB is similar to the transition from a Ld phase to a Lo phase. The partial retention of fluidity in UV polymerized PSLBs, their enhanced stability, and the activity of incorporated transmembrane proteins and peptides is discussed. PMID:26794208

  15. Lipid exchange between membranes: effects of membrane surface charge, composition, and curvature.

    PubMed

    Zhu, Tao; Jiang, Zhongying; Ma, Yuqiang

    2012-09-01

    Intermembrane lipid exchange is critical to membrane functions and pharmaceutical applications. The exchange process is not fully understood and it is explored by quartz crystal microbalance with dissipation monitor method in this research. It is found that intermembrane lipid exchange is accelerated with the decrease of vesicle size and the increase of charge and liquid crystalline lipid composition ratio. Vesicle adsorption rate, membrane lateral pressure gradient, and lipid lateral diffusion coefficient are inferred to be critical in deciding the lipid exchange kinetics between membranes. Besides that, the membrane contact situation during lipid exchange is also studied. The maximum total membrane contact area is found to increase with the decrease of vesicle size, charged and liquid crystalline lipid composition ratio. A competition mechanism between the vesicle adsorption rate and the intermembrane lipid exchange rate was proposed to control the maximum total membrane contact area.

  16. Lipids: architects and regulators of membrane dynamics and trafficking.

    PubMed

    Moreau, Patrick

    2007-05-01

    We have recently shown that an inhibition of sterol synthesis by fenpropimorph leads to an accumulation of sterol precursors, hydroxypalmitic acid-containing glucosylceramides and detergent resistant membranes in the Golgi bodies instead of the plasma membrane, suggesting that the individual molecules or the microdomains were blocked in the Golgi. These results and others from several eukaryotic models link lipid metabolism with membrane morphodynamics that are involved in membrane trafficking. Focus has been expanded to other lipid families, and numerous evidences are given showing lipids and lipid-modifying enzymes as key regulators of membrane homeostasis which can strongly regulate membrane morphodynamics and therefore trafficking. Beside protein-based machineries, lipid-based machineries are also shown as crucial regulatory forces involved in protein transport and sorting.

  17. Engineering lipid structure for recognition of the liquid ordered membrane phase

    DOE PAGES

    Bordovsky, Stefan S.; Wong, Christopher S.; Bachand, George D.; ...

    2016-08-26

    The selective partitioning of lipid components in phase-separated membranes is essential for domain formation involved in cellular processes. Identifying and tracking the movement of lipids in cellular systems would be improved if we understood how to achieve selective affinity between fluorophore-labeled lipids and membrane assemblies. Furthermore, we investigated the structure and chemistry of membrane lipids to evaluate lipid designs that partition to the liquid ordered (Lo) phase. A range of fluorophores at the headgroup position and lengths of PEG spacer between the lipid backbone and fluorophore were examined. On a lipid body with saturated palmityl or palmitoyl tails, we foundmore » that although the lipid tails can direct selective partitioning to the Lo phase through favorable packing interactions, headgroup hydrophobicity can override the partitioning behavior and direct the lipid to the disordered membrane phase (Ld). The PEG spacer can serve as a buffer to mute headgroup–membrane interactions and thus improve Lo phase partitioning, but its effect is limited with strongly hydrophobic fluorophore headgroups. We present a series of lipid designs leading to the development of novel fluorescently labeled lipids with selective affinity for the Lo phase.« less

  18. Engineering lipid structure for recognition of the liquid ordered membrane phase

    SciTech Connect

    Bordovsky, Stefan S.; Wong, Christopher S.; Bachand, George D.; Stachowiak, Jeanne C.; Sasaki, Darryl Y.

    2016-08-26

    The selective partitioning of lipid components in phase-separated membranes is essential for domain formation involved in cellular processes. Identifying and tracking the movement of lipids in cellular systems would be improved if we understood how to achieve selective affinity between fluorophore-labeled lipids and membrane assemblies. Furthermore, we investigated the structure and chemistry of membrane lipids to evaluate lipid designs that partition to the liquid ordered (Lo) phase. A range of fluorophores at the headgroup position and lengths of PEG spacer between the lipid backbone and fluorophore were examined. On a lipid body with saturated palmityl or palmitoyl tails, we found that although the lipid tails can direct selective partitioning to the Lo phase through favorable packing interactions, headgroup hydrophobicity can override the partitioning behavior and direct the lipid to the disordered membrane phase (Ld). The PEG spacer can serve as a buffer to mute headgroup–membrane interactions and thus improve Lo phase partitioning, but its effect is limited with strongly hydrophobic fluorophore headgroups. We present a series of lipid designs leading to the development of novel fluorescently labeled lipids with selective affinity for the Lo phase.

  19. The role of ceramide chain length distribution on the barrier properties of the skin lipid membranes.

    PubMed

    Mojumdar, E H; Kariman, Z; van Kerckhove, L; Gooris, G S; Bouwstra, J A

    2014-10-01

    The skin barrier function is provided by the stratum corneum (SC). The lipids in the SC are composed of three lipid classes: ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs) which form two crystalline lamellar structures. In the present study, we investigate the effect of CER chain length distribution on the barrier properties of model lipid membranes mimicking the lipid composition and organization of SC. The membranes were prepared with either isolated pig CERs (PCERs) or synthetic CERs. While PCERs have a wide chain length distribution, the synthetic CERs are quite uniform in chain length. The barrier properties were examined by means of permeation studies using hydrocortisone as a model drug. Our studies revealed a reduced barrier in lipid membranes prepared with PCERs compared to synthetic CERs. Additional studies revealed that a wider chain length distribution of PCERs results in an enhanced hexagonal packing and increased conformational disordering of the lipid tails compared to synthetic CERs, while the lamellar phases did not change. This demonstrates that the chain length distribution affects the lipid barrier by reducing the lipid ordering and density within the lipid lamellae. In subsequent studies, the effect of increased levels of FFAs or CERs with a long acyl chain in the PCERs membranes was also studied. These changes in lipid composition enhanced the level of orthorhombic packing, reduced the conformational disordering and increased the barrier of the lipid membranes. In conclusion, the CER chain length distribution is an important key factor for maintaining a proper barrier.

  20. Electrophoretic separation method for membrane pore-forming proteins in multilayer lipid membranes.

    PubMed

    Okamoto, Yukihiro; Tsujimoto, Yusuke; Umakoshi, Hiroshi

    2016-03-01

    In this paper, we report on a novel electrophoretic separation and analysis method for membrane pore-forming proteins in multilayer lipid membranes (MLMs) in order to overcome the problems related to current separation and analysis methods of membrane proteins, and to obtain a high-performance separation method on the basis of specific properties of the lipid membranes. We constructed MLMs, and subsequently characterized membrane pore-forming protein behavior in MLMs. Through the use of these MLMs, we were able to successfully separate and analyze membrane pore-forming proteins in MLMs. To the best of our knowledge, this research is the first example of membrane pore-forming protein separation in lipid membranes. Our method can be expected to be applied for the separation and analysis of other membrane proteins including intrinsic membrane proteins and to result in high-performance by utilizing the specific properties of lipid membranes.

  1. Brain membrane lipids in major depression and anxiety disorders.

    PubMed

    Müller, Christian P; Reichel, Martin; Mühle, Christiane; Rhein, Cosima; Gulbins, Erich; Kornhuber, Johannes

    2015-08-01

    Major depression and anxiety disorders have high prevalence rates and are frequently comorbid. The neurobiological bases for these disorders are not fully understood, and available treatments are not always effective. Current models assume that dysfunctions in neuronal proteins and peptide activities are the primary causes of these disorders. Brain lipids determine the localization and function of proteins in the cell membrane and in doing so regulate synaptic throughput in neurons. Lipids may also leave the membrane as transmitters and relay signals from the membrane to intracellular compartments or to other cells. Here we review how membrane lipids, which play roles in the membrane's function as a barrier and a signaling medium for classical transmitter signaling, contribute to depression and anxiety disorders and how this role may provide targets for lipid-based treatment approaches. Preclinical findings have suggested a crucial role for the membrane-forming n-3 polyunsaturated fatty acids, glycerolipids, glycerophospholipids, and sphingolipids in the induction of depression- and anxiety-related behaviors. These polyunsaturated fatty acids also offer new treatment options such as targeted dietary supplementation or pharmacological interference with lipid-regulating enzymes. While clinical trials support this view, effective lipid-based therapies may need more individualized approaches. Altogether, accumulating evidence suggests a crucial role for membrane lipids in the pathogenesis of depression and anxiety disorders; these lipids could be exploited for improved prevention and treatment. This article is part of a Special Issue entitled Brain Lipids.

  2. On the Importance of Hydrodynamic Interactions in Lipid Membrane Formation

    PubMed Central

    Ando, Tadashi; Skolnick, Jeffrey

    2013-01-01

    Hydrodynamic interactions (HI) give rise to collective motions between molecules, which are known to be important in the dynamics of random coil polymers and colloids. However, their role in the biological self-assembly of many molecule systems has not been investigated. Here, using Brownian dynamics simulations, we evaluate the importance of HI on the kinetics of self-assembly of lipid membranes. One-thousand coarse-grained lipid molecules in periodic simulation boxes were allowed to assemble into stable bilayers in the presence and absence of intermolecular HI. Hydrodynamic interactions reduce the monomer-monomer association rate by 50%. In contrast, the rate of association of lipid clusters is much faster in the presence of intermolecular HI. In fact, with intermolecular HI, the membrane self-assembly rate is 3–10 times faster than that without intermolecular HI. We introduce an analytical model to describe the size dependence of the diffusive encounter rate of particle clusters, which can qualitatively explain our simulation results for the early stage of the membrane self-assembly process. These results clearly suggest that HI greatly affects the kinetics of self-assembly and that simulations without HI will significantly underestimate the kinetic parameters of such processes. PMID:23332062

  3. MemProtMD: Automated Insertion of Membrane Protein Structures into Explicit Lipid Membranes

    PubMed Central

    Stansfeld, Phillip J.; Goose, Joseph E.; Caffrey, Martin; Carpenter, Elisabeth P.; Parker, Joanne L.; Newstead, Simon; Sansom, Mark S.P.

    2015-01-01

    Summary There has been exponential growth in the number of membrane protein structures determined. Nevertheless, these structures are usually resolved in the absence of their lipid environment. Coarse-grained molecular dynamics (CGMD) simulations enable insertion of membrane proteins into explicit models of lipid bilayers. We have automated the CGMD methodology, enabling membrane protein structures to be identified upon their release into the PDB and embedded into a membrane. The simulations are analyzed for protein-lipid interactions, identifying lipid binding sites, and revealing local bilayer deformations plus molecular access pathways within the membrane. The coarse-grained models of membrane protein/bilayer complexes are transformed to atomistic resolution for further analysis and simulation. Using this automated simulation pipeline, we have analyzed a number of recently determined membrane protein structures to predict their locations within a membrane, their lipid/protein interactions, and the functional implications of an enhanced understanding of the local membrane environment of each protein. PMID:26073602

  4. Critical point fluctuations in supported lipid membranes.

    PubMed

    Connell, Simon D; Heath, George; Olmsted, Peter D; Kisil, Anastasia

    2013-01-01

    In this paper, we demonstrate that it is possible to observe many aspects of critical phenomena in supported lipid bilayers using atomic force microscopy (AFM) with the aid of stable and precise temperature control. The regions of criticality were determined by accurately measuring and calculating phase diagrams for the 2 phase L(d)-L(o) region, and tracking how it moves with temperature, then increasing the sampling density around the estimated critical regions. Compositional fluctuations were observed above the critical temperature (T(c)) and characterised using a spatial correlation function. From this analysis, the phase transition was found to be most closely described by the 2D Ising model, showing it is a critical transition. Below T(c) roughening of the domain boundaries occurred due to the reduction in line tension close to the critical point. Smaller scale density fluctuations were also detected just below T(c). At T(c), we believe we have observed fluctuations on length scales greater than 10 microm. The region of critically fluctuating 10-100 nm nanodomains has been found to extend a considerable distance above T(c) to temperatures within the biological range, and seem to be an ideal candidate for the actual structure of lipid rafts in cell membranes. Although evidence for this idea has recently emerged, this is the first direct evidence for nanoscale domains in the critical region.

  5. Membrane proteins, lipids and detergents: not just a soap opera.

    PubMed

    Seddon, Annela M; Curnow, Paul; Booth, Paula J

    2004-11-03

    Studying membrane proteins represents a major challenge in protein biochemistry, with one of the major difficulties being the problems encountered when working outside the natural lipid environment. In vitro studies such as crystallization are reliant on the successful solubilization or reconstitution of membrane proteins, which generally involves the careful selection of solubilizing detergents and mixed lipid/detergent systems. This review will concentrate on the methods currently available for efficient reconstitution and solubilization of membrane proteins through the use of detergent micelles, mixed lipid/detergent micelles and bicelles or liposomes. We focus on the relevant molecular properties of the detergents and lipids that aid understanding of these processes. A significant barrier to membrane protein research is retaining the stability and function of the protein during solubilization, reconstitution and crystallization. We highlight some of the lessons learnt from studies of membrane protein folding in vitro and give an overview of the role that lipids can play in stabilizing the proteins.

  6. Monoolein lipid phases as incorporation and enrichment materials for membrane protein crystallization.

    SciTech Connect

    Wallace, E.; Dranow, D.; Laible, P. D.; Christensen, J.; Nollert, P.

    2011-01-01

    The crystallization of membrane proteins in amphiphile-rich materials such as lipidic cubic phases is an established methodology in many structural biology laboratories. The standard procedure employed with this methodology requires the generation of a highly viscous lipidic material by mixing lipid, for instance monoolein, with a solution of the detergent solubilized membrane protein. This preparation is often carried out with specialized mixing tools that allow handling of the highly viscous materials while minimizing dead volume to save precious membrane protein sample. The processes that occur during the initial mixing of the lipid with the membrane protein are not well understood. Here we show that the formation of the lipidic phases and the incorporation of the membrane protein into such materials can be separated experimentally. Specifically, we have investigated the effect of different initial monoolein-based lipid phase states on the crystallization behavior of the colored photosynthetic reaction center from Rhodobacter sphaeroides. We find that the detergent solubilized photosynthetic reaction center spontaneously inserts into and concentrates in the lipid matrix without any mixing, and that the initial lipid material phase state is irrelevant for productive crystallization. A substantial in-situ enrichment of the membrane protein to concentration levels that are otherwise unobtainable occurs in a thin layer on the surface of the lipidic material. These results have important practical applications and hence we suggest a simplified protocol for membrane protein crystallization within amphiphile rich materials, eliminating any specialized mixing tools to prepare crystallization experiments within lipidic cubic phases. Furthermore, by virtue of sampling a membrane protein concentration gradient within a single crystallization experiment, this crystallization technique is more robust and increases the efficiency of identifying productive crystallization

  7. Alteration of macrophage membrane lipids following processing of bacterial peptidoglycan

    SciTech Connect

    Polanski, M.; Gray, G.R.

    1986-03-01

    As part of the continuing investigation into the role played by macrophages in antigen presentation and bacterial adjuvant activation, the authors have examined the metabolites produced by macrophages after encounter with peptidoglycan. Peptidoglycan was chosen because it contains N-acetyl-muramyl-L-alanyl-D-isoglutamine (muramyl dipeptide), a known adjuvant whose primary target cell is the macrophage. In previous work, the authors established that a series of muramyl dipeptide-like glycopeptides was released into the medium following phagocytosis of peptidoglycan by a macrophage cell line. Here the authors report on the finding that, additionally, a membrane lipid has been covalently altered by the addition of a peptidoglycan fragment. Bacillus subtilis cell walls which had been radiolabeled in their muramic acid, glucosamine and alanine residues, were incubated with the murine macrophage cell line RAW264. Using standard lipid extraction procedures, a lipid was isolated and found to contain equal molar ratios of alanine, glutamic acid and diaminopimelic acid. Since lipidated peptidoglycan peptides have been shown to be immunoactivators, the isolated lipid derivative may serve as a signal for interactions with other lymphocytes.

  8. Biophysical interactions with model lipid membranes: applications in drug discovery and drug delivery

    PubMed Central

    Peetla, Chiranjeevi; Stine, Andrew; Labhasetwar, Vinod

    2009-01-01

    The transport of drugs or drug delivery systems across the cell membrane is a complex biological process, often difficult to understand because of its dynamic nature. In this regard, model lipid membranes, which mimic many aspects of cell-membrane lipids, have been very useful in helping investigators to discern the roles of lipids in cellular interactions. One can use drug-lipid interactions to predict pharmacokinetic properties of drugs, such as their transport, biodistribution, accumulation, and hence efficacy. These interactions can also be used to study the mechanisms of transport, based on the structure and hydrophilicity/hydrophobicity of drug molecules. In recent years, model lipid membranes have also been explored to understand their mechanisms of interactions with peptides, polymers, and nanocarriers. These interaction studies can be used to design and develop efficient drug delivery systems. Changes in the lipid composition of cells and tissue in certain disease conditions may alter biophysical interactions, which could be explored to develop target-specific drugs and drug delivery systems. In this review, we discuss different model membranes, drug-lipid interactions and their significance, studies of model membrane interactions with nanocarriers, and how biophysical interaction studies with lipid model membranes could play an important role in drug discovery and drug delivery. PMID:19432455

  9. Membrane humidity control investigation

    NASA Technical Reports Server (NTRS)

    Elam, J.; Ruder, J.; Strumpf, H.

    1974-01-01

    The basic performance data on a hollow fiber membrane unit that removes water from a breathing gas loop by diffusion is presented. Using available permeability data for cellulose acetate, a preliminary design was made of a dehumidifier unit that would meet the problem statement.

  10. Counterion-mediated pattern formation in membranes containing anionic lipids

    PubMed Central

    Slochower, David R.; Wang, Yu-Hsiu; Tourdot, Richard W.; Radhakrishnan, Ravi; Janmey, Paul A.

    2014-01-01

    Most lipid components of cell membranes are either neutral, like cholesterol, or zwitterionic, like phosphatidylcholine and sphingomyelin. Very few lipids, such as sphingosine, are cationic at physiological pH. These generally interact only transiently with the lipid bilayer, and their synthetic analogs are often designed to destabilize the membrane for drug or DNA delivery. However, anionic lipids are common in both eukaryotic and prokaryotic cell membranes. The net charge per anionic phospholipid ranges from −1 for the most abundant anionic lipids such has phosphatidylserine, to near −7 for phosphatidylinositol 3,4,5 trisphosphate, although the effective charge depends on many environmental factors. Anionic phospholipids and other negatively charged lipids such as lipopolysaccharides are not randomly distributed in the lipid bilayer, but are highly restricted to specific leaflets of the bilayer and to regions near transmembrane proteins or other organized structures within the plane of the membrane. This review highlights some recent evidence that counterions, in the form of monovalent or divalent metal ions, polyamines, or cationic protein domains, have a large influence of the lateral distribution of anionic lipids within the membrane, and that lateral demixing of anionic lipids has effects on membrane curvature and protein function that are important for biological control. PMID:24556233

  11. Thermal Adaptation of the Archaeal and Bacterial Lipid Membranes

    PubMed Central

    Koga, Yosuke

    2012-01-01

    The physiological characteristics that distinguish archaeal and bacterial lipids, as well as those that define thermophilic lipids, are discussed from three points of view that (1) the role of the chemical stability of lipids in the heat tolerance of thermophilic organisms: (2) the relevance of the increase in the proportion of certain lipids as the growth temperature increases: (3) the lipid bilayer membrane properties that enable membranes to function at high temperatures. It is concluded that no single, chemically stable lipid by itself was responsible for the adaptation of surviving at high temperatures. Lipid membranes that function effectively require the two properties of a high permeability barrier and a liquid crystalline state. Archaeal membranes realize these two properties throughout the whole biological temperature range by means of their isoprenoid chains. Bacterial membranes meet these requirements only at or just above the phase-transition temperature, and therefore their fatty acid composition must be elaborately regulated. A recent hypothesis sketched a scenario of the evolution of lipids in which the “lipid divide” emerged concomitantly with the differentiation of archaea and bacteria. The two modes of thermal adaptation were established concurrently with the “lipid divide.” PMID:22927779

  12. Curvature forces in membrane lipid-protein interactions.

    PubMed

    Brown, Michael F

    2012-12-11

    Membrane biochemists are becoming increasingly aware of the role of lipid-protein interactions in diverse cellular functions. This review describes how conformational changes in membrane proteins, involving folding, stability, and membrane shape transitions, potentially involve elastic remodeling of the lipid bilayer. Evidence suggests that membrane lipids affect proteins through interactions of a relatively long-range nature, extending beyond a single annulus of next-neighbor boundary lipids. It is assumed the distance scale of the forces is large compared to the molecular range of action. Application of the theory of elasticity to flexible soft surfaces derives from classical physics and explains the polymorphism of both detergents and membrane phospholipids. A flexible surface model (FSM) describes the balance of curvature and hydrophobic forces in lipid-protein interactions. Chemically nonspecific properties of the lipid bilayer modulate the conformational energetics of membrane proteins. The new biomembrane model challenges the standard model (the fluid mosaic model) found in biochemistry texts. The idea of a curvature force field based on data first introduced for rhodopsin gives a bridge between theory and experiment. Influences of bilayer thickness, nonlamellar-forming lipids, detergents, and osmotic stress are all explained by the FSM. An increased awareness of curvature forces suggests that research will accelerate as structural biology becomes more closely entwined with the physical chemistry of lipids in explaining membrane structure and function.

  13. Curvature Forces in Membrane Lipid-Protein Interactions

    PubMed Central

    Brown, Michael F.

    2012-01-01

    Membrane biochemists are becoming increasingly aware of the role of lipid-protein interactions in diverse cellular functions. This review describes how conformational changes of membrane proteins—involving folding, stability, and membrane shape transitions—potentially involve elastic remodeling of the lipid bilayer. Evidence suggests that membrane lipids affect proteins through interactions of a relatively long-range nature, extending beyond a single annulus of next-neighbor boundary lipids. It is assumed the distance scale of the forces is large compared to the molecular range of action. Application of the theory of elasticity to flexible soft surfaces derives from classical physics, and explains the polymorphism of both detergents and membrane phospholipids. A flexible surface model (FSM) describes the balance of curvature and hydrophobic forces in lipid-protein interactions. Chemically nonspecific properties of the lipid bilayer modulate the conformational energetics of membrane proteins. The new biomembrane model challenges the standard model (the fluid mosaic model) found in biochemistry texts. The idea of a curvature force field based on data first introduced for rhodopsin gives a bridge between theory and experiment. Influences of bilayer thickness, nonlamellar-forming lipids, detergents, and osmotic stress are all explained by the FSM. An increased awareness of curvature forces suggests that research will accelerate as structural biology becomes more closely entwined with the physical chemistry of lipids in explaining membrane structure and function. PMID:23163284

  14. Probing Peptide and Protein Insertion in a Biomimetic S-Layer Supported Lipid Membrane Platform

    PubMed Central

    Damiati, Samar; Schrems, Angelika; Sinner, Eva-Kathrin; Sleytr, Uwe B.; Schuster, Bernhard

    2015-01-01

    The most important aspect of synthetic lipid membrane architectures is their ability to study functional membrane-active peptides and membrane proteins in an environment close to nature. Here, we report on the generation and performance of a biomimetic platform, the S-layer supported lipid membrane (SsLM), to investigate the structural and electrical characteristics of the membrane-active peptide gramicidin and the transmembrane protein α-hemolysin in real-time using a quartz crystal microbalance with dissipation monitoring in combination with electrochemical impedance spectroscopy. A shift in membrane resistance is caused by the interaction of α-hemolysin and gramicidin with SsLMs, even if only an attachment onto, or functional channels through the lipid membrane, respectively, are formed. Moreover, the obtained results did not indicate the formation of functional α-hemolysin pores, but evidence for functional incorporation of gramicidin into this biomimetic architecture is provided. PMID:25633104

  15. The influence of membrane lipid structure on plasma membrane Ca2+ -ATPase activity.

    PubMed

    Tang, Daxin; Dean, William L; Borchman, Douglas; Paterson, Christopher A

    2006-03-01

    Lipid composition and Ca(2+)-ATPase activity both change with age and disease in many tissues. We explored relationships between lipid composition/structure and plasma membrane Ca(2+)-ATPase (PMCA) activity. PMCA was purified from human erythrocytes and was reconstituted into liposomes prepared from human ocular lens membrane lipids and synthetic lipids. Lens lipids were used in this study as a model for naturally ordered lipids, but the influence of lens lipids on PMCA function is especially relevant to the lens since calcium homeostasis is vital to lens clarity. Compared to fiber cell lipids, epithelial lipids exhibited an ordered to disordered phase transition temperature that was 12 degrees C lower. Reconstitution of PMCA into lipids was essential for maximal activity. PMCA activity was two to three times higher when the surrounding phosphatidylcholine molecules contained acyl chains that were ordered (stiff) compared to disordered (fluid) acyl chains. In a completely ordered lipid hydrocarbon chain environment, PMCA associates more strongly with the acidic lipid phosphatidylserine in comparison to phosphatidylcholine. PMCA associates much more strongly with phosphatidylcholine containing disordered hydrocarbon chains than ordered hydrocarbon chains. PMCA activity is influenced by membrane lipid composition and structure. The naturally high degree of lipid order in plasma membranes such as those found in the human lens may serve to support PMCA activity. The absence of PMCA activity in the cortical region of human lenses is apparently not due to a different lipid environment. Changes in lipid composition such as those observed with age or disease could potentially influence PMCA function.

  16. The Interaction of Polyene Antibiotics with Thin Lipid Membranes

    PubMed Central

    Andreoli, Thomas E.; Monahan, Marcia

    1968-01-01

    Optically black, thin lipid membranes prepared from sheep erythrocyte lipids have a high dc resistance (Rm ≅ 108 ohm-cm2) when the bathing solutions contain NaCl or KCl. The ionic transference numbers (Ti) indicate that these membranes are cation-selective (TNa ≅ 0.85; TCl ≅ 0.15). These electrical properties are independent of the cholesterol content of the lipid solutions from which the membranes are formed. Nystatin, and probably amphotericin B, are cyclic polyene antibiotics containing ≈36 ring atoms and a free amino and carboxyl group. When the lipid solutions used to form membranes contained equimolar amounts of cholesterol and phospholipid, these antibiotics reduced Rm to ≈102 ohm-cm2; concomitantly, TCl became ≅0.92. The slope of the line relating log Rm and log antibiotic concentration was ≅4.5. Neither nystatin (2 x 10-5 M) nor amphotericin B (2 x 10-7 M) had any effect on membrane stability. The antibiotics had no effect on Rm or membrane permselectivity when the lipids used to form membranes were cholesterol-depleted. Filipin (10-5 M), an uncharged polyene with 28 ring atoms, produced striking membrane instability, but did not affect Rm or membrane ionic selectivity. These data suggest that amphotericin B or nystatin may interact with membrane-bound sterols to produce multimolecular complexes which greatly enhance the permeability of such membranes for anions (Cl-, acetate), and, to a lesser degree, cations (Na+, K+, Li+). PMID:5672005

  17. Membranes: a meeting point for lipids, proteins and therapies

    PubMed Central

    Escribá, Pablo V; González-Ros, José M; Goñi, Félix M; Kinnunen, Paavo K J; Vigh, Lászlo; Sánchez-Magraner, Lissete; Fernández, Asia M; Busquets, Xavier; Horváth, Ibolya; Barceló-Coblijn, Gwendolyn

    2008-01-01

    Abstract Membranes constitute a meeting point for lipids and proteins. Not only do they define the entity of cells and cytosolic organelles but they also display a wide variety of important functions previously ascribed to the activity of proteins alone. Indeed, lipids have commonly been considered a mere support for the transient or permanent association of membrane proteins, while acting as a selective cell/organelle barrier. However, mounting evidence demonstrates that lipids themselves regulate the location and activity of many membrane proteins, as well as defining membrane microdomains that serve as spatio-temporal platforms for interacting signalling proteins. Membrane lipids are crucial in the fission and fusion of lipid bilayers and they also act as sensors to control environmental or physiological conditions. Lipids and lipid structures participate directly as messengers or regulators of signal transduction. Moreover, their alteration has been associated with the development of numerous diseases. Proteins can interact with membranes through lipid co-/post-translational modifications, and electrostatic and hydrophobic interactions, van der Waals forces and hydrogen bonding are all involved in the associations among membrane proteins and lipids. The present study reviews these interactions from the molecular and biomedical point of view, and the effects of their modulation on the physiological activity of cells, the aetiology of human diseases and the design of clinical drugs. In fact, the influence of lipids on protein function is reflected in the possibility to use these molecular species as targets for therapies against cancer, obesity, neurodegenerative disorders, cardiovascular pathologies and other diseases, using a new approach called membrane-lipid therapy. PMID:18266954

  18. Membranes: a meeting point for lipids, proteins and therapies.

    PubMed

    Escribá, Pablo V; González-Ros, José M; Goñi, Félix M; Kinnunen, Paavo K J; Vigh, Lászlo; Sánchez-Magraner, Lissete; Fernández, Asia M; Busquets, Xavier; Horváth, Ibolya; Barceló-Coblijn, Gwendolyn

    2008-06-01

    Membranes constitute a meeting point for lipids and proteins. Not only do they define the entity of cells and cytosolic organelles but they also display a wide variety of important functions previously ascribed to the activity of proteins alone. Indeed, lipids have commonly been considered a mere support for the transient or permanent association of membrane proteins, while acting as a selective cell/organelle barrier. However, mounting evidence demonstrates that lipids themselves regulate the location and activity of many membrane proteins, as well as defining membrane microdomains that serve as spatio-temporal platforms for interacting signalling proteins. Membrane lipids are crucial in the fission and fusion of lipid bilayers and they also act as sensors to control environmental or physiological conditions. Lipids and lipid structures participate directly as messengers or regulators of signal transduction. Moreover, their alteration has been associated with the development of numerous diseases. Proteins can interact with membranes through lipid co-/post-translational modifications, and electrostatic and hydrophobic interactions, van der Waals forces and hydrogen bonding are all involved in the associations among membrane proteins and lipids. The present study reviews these interactions from the molecular and biomedical point of view, and the effects of their modulation on the physiological activity of cells, the aetiology of human diseases and the design of clinical drugs. In fact, the influence of lipids on protein function is reflected in the possibility to use these molecular species as targets for therapies against cancer, obesity, neurodegenerative disorders, cardiovascular pathologies and other diseases, using a new approach called membrane-lipid therapy.

  19. Elucidating how bamboo salt interacts with supported lipid membranes: influence of alkalinity on membrane fluidity.

    PubMed

    Jeong, Jong Hee; Choi, Jae-Hyeok; Kim, Min Chul; Park, Jae Hyeon; Herrin, Jason Scott; Kim, Seung Hyun; Lee, Haiwon; Cho, Nam-Joon

    2015-07-01

    Bamboo salt is a traditional medicine produced from sea salt. It is widely used in Oriental medicine and is an alkalizing agent with reported antiinflammatory, antimicrobial and chemotherapeutic properties. Notwithstanding, linking specific molecular mechanisms with these properties has been challenging to establish in biological systems. In part, this issue may be related to bamboo salt eliciting nonspecific effects on components found within these systems. Herein, we investigated the effects of bamboo salt solution on supported lipid bilayers as a model system to characterize the interaction between lipid membranes and bamboo salt. The atomic composition of unprocessed and processed bamboo salts was first analyzed by mass spectrometry, and we identified several elements that have not been previously reported in other bamboo salt preparations. The alkalinity of hydrated samples was also measured and determined to be between pH 10 and 11 for bamboo salts. The effect of processed bamboo salt solutions on the fluidic properties of a supported lipid bilayer on glass was next investigated by fluorescence recovery after photobleaching (FRAP) analysis. It was demonstrated that, with increasing ionic strength of the bamboo salt solution, the fluidity of a lipid bilayer increased. On the contrary, increasing the ionic strength of near-neutral buffer solutions with sodium chloride salt diminished fluidity. To reconcile these two observations, we identified that solution alkalinity is critical for the effects of bamboo salt on membrane fluidity, as confirmed using three additional commercial bamboo salt preparations. Extended-DLVO model calculations support that the effects of bamboo salt on lipid membranes are due to the alkalinity imparting a stronger hydration force. Collectively, the results of this work demonstrate that processing of bamboo salt strongly affects its atomic composition and that the alkalinity of bamboo salt solutions contributes to its effect on membrane

  20. Lipid membrane-assisted condensation and assembly of amphiphilic Janus particles

    SciTech Connect

    Chambers, Mariah; Mallory, Stewart Anthony; Malone, Heather; Gao, Yuan; Anthony, Stephen M.; Yi, Yi; Cacciuto, Angelo; Yu, Yan

    2016-01-01

    Amphiphilic Janus particles self-assemble into complex metastructures, but little is known about how their assembly might be modified by weak interactions with a nearby biological membrane surface. Here, we report an integrated experimental and molecular dynamics simulation study to investigate the self-assembly of amphiphilic Janus particles on a lipid membrane. We created an experimental system in which Janus particles are allowed to self-assemble in the same medium where zwitterionic lipids form giant unilamellar vesicles (GUVs). Janus particles spontaneously concentrated on the inner leaflet of the GUVs. They exhibited biased orientation and heterogeneous rotational dynamics as revealed by single particle rotational tracking. The combined experimental and simulation results show that Janus particles concentrate on the lipid membranes due to weak particle–lipid attraction, whereas the biased orientation of particles is driven predominantly by inter-particle interactions. Furthermore, this study demonstrates the potential of using lipid membranes to influence the self-assembly of Janus particles.

  1. Isolation and analysis of membrane lipids and lipid rafts in common carp (Cyprinus carpio L.).

    PubMed

    Brogden, Graham; Propsting, Marcus; Adamek, Mikolaj; Naim, Hassan Y; Steinhagen, Dieter

    2014-03-01

    Cell membranes act as an interface between the interior of the cell and the exterior environment and facilitate a range of essential functions including cell signalling, cell structure, nutrient uptake and protection. It is composed of a lipid bilayer with integrated proteins, and the inner leaflet of the lipid bilayer comprises of liquid ordered (Lo) and liquid disordered (Ld) domains. Lo microdomains, also named as lipid rafts are enriched in cholesterol, sphingomyelin and certain types of proteins, which facilitate cell signalling and nutrient uptake. Lipid rafts have been extensively researched in mammals and the presence of functional lipid rafts was recently demonstrated in goldfish, but there is currently very little knowledge about their composition and function in fish. Therefore a protocol was established for the analysis of lipid rafts and membranous lipids in common carp (Cyprinus carpio L.) tissues. Twelve lipids were identified and analysed in the Ld domain of the membrane with the most predominant lipids found in all tissues being; triglycerides, cholesterol, phosphoethanolamine and phosphatidylcholine. Four lipids were identified in lipid rafts in all tissues analysed, triglycerides (33-62%) always found in the highest concentration followed by cholesterol (24-32%), phosphatidylcholine and sphingomyelin. Isolation of lipid rafts was confirmed by identifying the presence of the lipid raft associated protein flotillin, present at higher concentrations in the detergent resistant fraction. The data provided here build a lipid library of important carp tissues as a baseline for further studies into virus entry, protein trafficking or environmental stress analysis.

  2. Measuring lipid packing of model and cellular membranes with environment sensitive probes.

    PubMed

    Sezgin, Erdinc; Sadowski, Tomasz; Simons, Kai

    2014-07-15

    The extent of lipid packing is one of the key physicochemical features of biological membranes and is involved in many membrane processes. Polarity sensitive fluorescent probes are commonly used tools to measure membrane lipid packing in both artificial and biological membranes. In this paper, we have systematically compared eight different probes to measure membrane lipid ordering. We investigated how these probes behave in small unilamellar liposomes, phase-separated giant unilamellar vesicles, cell-derived giant plasma membrane vesicles, and live cells. We have tested the order sensitivity of a variety of measurable parameters, including generalized polarization, peak shift, or intensity shift. We also investigated internalization and photostability of the probes to assess probe potential for time-lapse live cell imaging. These results provide a catalogue of properties to facilitate the choice of probe according to need.

  3. Lipid Clustering Correlates with Membrane Curvature as Revealed by Molecular Simulations of Complex Lipid Bilayers

    PubMed Central

    Koldsø, Heidi; Shorthouse, David; Hélie, Jean; Sansom, Mark S. P.

    2014-01-01

    Cell membranes are complex multicomponent systems, which are highly heterogeneous in the lipid distribution and composition. To date, most molecular simulations have focussed on relatively simple lipid compositions, helping to inform our understanding of in vitro experimental studies. Here we describe on simulations of complex asymmetric plasma membrane model, which contains seven different lipids species including the glycolipid GM3 in the outer leaflet and the anionic lipid, phosphatidylinositol 4,5-bisphophate (PIP2), in the inner leaflet. Plasma membrane models consisting of 1500 lipids and resembling the in vivo composition were constructed and simulations were run for 5 µs. In these simulations the most striking feature was the formation of nano-clusters of GM3 within the outer leaflet. In simulations of protein interactions within a plasma membrane model, GM3, PIP2, and cholesterol all formed favorable interactions with the model α-helical protein. A larger scale simulation of a model plasma membrane containing 6000 lipid molecules revealed correlations between curvature of the bilayer surface and clustering of lipid molecules. In particular, the concave (when viewed from the extracellular side) regions of the bilayer surface were locally enriched in GM3. In summary, these simulations explore the nanoscale dynamics of model bilayers which mimic the in vivo lipid composition of mammalian plasma membranes, revealing emergent nanoscale membrane organization which may be coupled both to fluctuations in local membrane geometry and to interactions with proteins. PMID:25340788

  4. Lipid clustering correlates with membrane curvature as revealed by molecular simulations of complex lipid bilayers.

    PubMed

    Koldsø, Heidi; Shorthouse, David; Hélie, Jean; Sansom, Mark S P

    2014-10-01

    Cell membranes are complex multicomponent systems, which are highly heterogeneous in the lipid distribution and composition. To date, most molecular simulations have focussed on relatively simple lipid compositions, helping to inform our understanding of in vitro experimental studies. Here we describe on simulations of complex asymmetric plasma membrane model, which contains seven different lipids species including the glycolipid GM3 in the outer leaflet and the anionic lipid, phosphatidylinositol 4,5-bisphophate (PIP2), in the inner leaflet. Plasma membrane models consisting of 1500 lipids and resembling the in vivo composition were constructed and simulations were run for 5 µs. In these simulations the most striking feature was the formation of nano-clusters of GM3 within the outer leaflet. In simulations of protein interactions within a plasma membrane model, GM3, PIP2, and cholesterol all formed favorable interactions with the model α-helical protein. A larger scale simulation of a model plasma membrane containing 6000 lipid molecules revealed correlations between curvature of the bilayer surface and clustering of lipid molecules. In particular, the concave (when viewed from the extracellular side) regions of the bilayer surface were locally enriched in GM3. In summary, these simulations explore the nanoscale dynamics of model bilayers which mimic the in vivo lipid composition of mammalian plasma membranes, revealing emergent nanoscale membrane organization which may be coupled both to fluctuations in local membrane geometry and to interactions with proteins.

  5. How Lipid Membranes Affect Pore Forming Toxin Activity.

    PubMed

    Rojko, Nejc; Anderluh, Gregor

    2015-12-15

    Pore forming toxins (PFTs) evolved to permeate the plasma membrane of target cells. This is achieved in a multistep mechanism that usually involves binding of soluble protein monomer to the lipid membrane, oligomerization at the plane of the membrane, and insertion of part of the polypeptide chain across the lipid membrane to form a conductive channel. Introduced pores allow uncontrolled transport of solutes across the membrane, inflicting damage to the target cell. PFTs are usually studied from the perspective of structure-function relationships, often neglecting the important role of the bulk membrane properties on the PFT mechanism of action. In this Account, we discuss how membrane lateral heterogeneity, thickness, and fluidity influence the pore forming process of PFTs. In general, lipid molecules are more accessible for binding in fluid membranes due to steric reasons. When PFT specifically binds ordered domains, it usually recognizes a specific lipid distribution pattern, like sphingomyelin (SM) clusters or SM/cholesterol complexes, and not individual lipid species. Lipid domains were also suggested to act as an additional concentration platform facilitating PFT oligomerization, but this is yet to be shown. The last stage in PFT action is the insertion of the transmembrane segment across the membranes to build the transmembrane pore walls. Conformational changes are a spontaneous process, and sufficient free energy has to be available for efficient membrane penetration. Therefore, fluid bilayers are permeabilized more readily in comparison to highly ordered and thicker liquid ordered lipid phase (Lo). Energetically more costly insertion into the Lo phase can be driven by the hydrophobic mismatch between the thinner liquid disordered phase (Ld) and large protein complexes, which are unable to tilt like single transmembrane segments. In the case of proteolipid pores, membrane properties can directly modulate pore size, stability, and even selectivity. Finally

  6. Photon correlation spectroscopy of bilayer lipid membranes.

    PubMed Central

    Crilly, J F; Earnshaw, J C

    1983-01-01

    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. PMID:6838962

  7. The superlattice model of lateral organization of membranes and its implications on membrane lipid homeostasis.

    PubMed

    Somerharju, Pentti; Virtanen, Jorma A; Cheng, Kwan H; Hermansson, Martin

    2009-01-01

    Most biological membranes are extremely complex structures consisting of hundreds of different lipid and protein molecules. According to the famous fluid-mosaic model lipids and many proteins are free to diffuse very rapidly in the plane of the membrane. While such fast diffusion implies that different membrane lipids would be laterally randomly distributed, accumulating evidence indicates that in model and natural membranes the lipid components tend to adopt regular (superlattice-like) distributions. The superlattice model, put forward based on such evidence, is intriguing because it predicts that 1) there is a limited number of allowed compositions representing local minima in membrane free energy and 2) those energy minima could provide set-points for enzymes regulating membrane lipid compositions. Furthermore, the existence of a discrete number of allowed compositions could help to maintain organelle identity in the face of rapid inter-organelle membrane traffic.

  8. Effects of dimethyl sulfoxide on lipid membrane electroporation.

    PubMed

    Fernández, M Laura; Reigada, Ramon

    2014-08-07

    Pores can be generated in lipid membranes by the application of an external electric field or by the addition of particular chemicals such as dimethyl sulfoxide (DMSO). Molecular dynamics (MD) has been shown to be a useful tool for unveiling many aspects of pore formation in lipid membranes in both situations. By means of MD simulations, we address the formation of electropores in cholesterol-containing lipid bilayers under the influence of DMSO. We show how a combination of physical and chemical mechanisms leads to more favorable conditions for generating membrane pores and, in particular, how the addition of DMSO to the medium significantly reduces the minimum electric field required to electroporate a lipid membrane. The strong alteration of membrane transversal properties and the energetic stabilization of the hydrophobic pore stage by DMSO provide the physicochemical mechanisms that explain this effect.

  9. Dynamic sorting of lipids and proteins in membrane tubes with a moving phase boundary

    PubMed Central

    Heinrich, Michael; Tian, Aiwei; Esposito, Cinzia; Baumgart, Tobias

    2010-01-01

    Cellular organelle membranes maintain their integrity, global shape, and composition despite vigorous exchange among compartments of lipids and proteins during trafficking and signaling. Organelle homeostasis involves dynamic molecular sorting mechanisms that are far from being understood. In contrast, equilibrium thermodynamics of membrane mixing and sorting, particularly the phase behavior of binary and ternary model membrane mixtures and its coupling to membrane mechanics, is relatively well characterized. Elucidating the continuous turnover of live cell membranes, however, calls for experimental and theoretical membrane models enabling manipulation and investigation of directional mass transport. Here we introduce the phenomenon of curvature-induced domain nucleation and growth in membrane mixtures with fluid phase coexistence. Membrane domains were consistently observed to nucleate precisely at the junction between a strongly curved cylindrical (tube) membrane and a pipette-aspirated giant unilamellar vesicle. This experimental geometry mimics intracellular sorting compartments, because they often show tubular-vesicular membrane regions. Nucleated domains at tube necks were observed to present diffusion barriers to the transport of lipids and proteins. We find that curvature-nucleated domains grow with characteristic parabolic time dependence that is strongly curvature-dependent. We derive an analytical model that reflects the observed growth dynamics. Numerically calculated membrane shapes furthermore allow us to elucidate mechanical details underlying curvature-dependent directed lipid transport. Our observations suggest a novel dynamic membrane sorting principle that may contribute to intracellular protein and lipid sorting and trafficking. PMID:20368457

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

    PubMed Central

    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

    2012-01-01

    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. PMID:23269904

  11. Nucleic acid-lipid membrane interactions studied by DSC.

    PubMed

    Giatrellis, Sarantis; Nounesis, George

    2011-01-01

    The interactions of nucleic acids with lipid membranes are of great importance for biological mechanisms as well as for biotechnological applications in gene delivery and drug carriers. The optimization of liposomal vectors for clinical use is absolutely dependent upon the formation mechanisms, the morphology, and the molecular organization of the lipoplexes, that is, the complexes of lipid membranes with DNA. Differential scanning calorimetry (DSC) has emerged as an efficient and relatively easy-to-operate experimental technique that can straightforwardly provide data related to the thermodynamics and the kinetics of the DNA-lipid complexation and especially to the lipid organization and phase transitions within the membrane. In this review, we summarize DSC studies considering nucleic acid-membrane systems, accentuating DSC capabilities, and data analysis. Published work involving cationic, anionic, and zwitterionic lipids as well as lipid mixtures interacting with RNA and DNA of different sizes and conformations are included. It is shown that despite limitations, issues such as DNA- or RNA-induced phase separation and microdomain lipid segregation, liposomal aggregation and fusion, alterations of the lipid long-range molecular order, as well as membrane-induced structural changes of the nucleic acids can be efficiently treated by systematic high-sensitivity DSC studies.

  12. Thermodynamics of sodium dodecyl sulfate partitioning into lipid membranes.

    PubMed

    Tan, Anmin; Ziegler, André; Steinbauer, Bernhard; Seelig, Joachim

    2002-09-01

    The partition equilibria of sodium dodecyl sulfate (SDS) and lithium dodecyl sulfate between water and bilayer membranes were investigated with isothermal titration calorimetry and spectroscopic methods (light scattering, (31)P-nuclear magnetic resonance) in the temperature range of 28 degrees C to 56 degrees C. The partitioning of the dodecyl sulfate anion (DS(-)) into the bilayer membrane is energetically favored by an exothermic partition enthalpy of Delta H(O)(D) = -6.0 kcal/mol at 28 degrees C. This is in contrast to nonionic detergents where Delta H(O)(D) is usually positive. The partition enthalpy decreases linearly with increasing temperature and the molar heat capacity is Delta C(O)(P) = -50 +/- 3 cal mol(-1) K(-1). The partition isotherm is nonlinear if the bound detergent is plotted versus the free detergent concentration in bulk solution. This is caused by the electrostatic repulsion between the DS(-) ions inserted into the membrane and those free in solution near the membrane surface. The surface concentration of DS(-) immediately above the plane of binding was hence calculated with the Gouy-Chapman theory, and a strictly linear relationship was obtained between the surface concentration and the extent of DS(-) partitioning. The surface partition constant K describes the chemical equilibrium in the absence of electrostatic effects. For the SDS-membrane equilibrium K was found to be 1.2 x 10(4) M(-1) to 6 x 10(4) M(-1) for the various systems and conditions investigated, very similar to data available for nonionic detergents of the same chain length. The membrane-micelle phase diagram was also studied. Complete membrane solubilization requires a ratio of 2.2 mol SDS bound per mole of total lipid at 56 degrees C. The corresponding equilibrium concentration of SDS free in solution is C (sat)(D,F) approximately 1.7 mM and is slightly below the critical micelles concentration (CMC) = 2.1 mM (at 56 degrees C and 0.11 M buffer). Membrane saturation occurs at

  13. Electro-Optical Imaging Microscopy of Dye-Doped Artificial Lipidic Membranes

    PubMed Central

    Hajj, Bassam; De Reguardati, Sophie; Hugonin, Loïc; Le Pioufle, Bruno; Osaki, Toshihisa; Suzuki, Hiroaki; Takeuchi, Shoji; Mojzisova, Halina; Chauvat, Dominique; Zyss, Joseph

    2009-01-01

    Artificial lipidic bilayers are widely used as a model for the lipid matrix in biological cell membranes. We use the Pockels electro-optical effect to investigate the properties of an artificial lipidic membrane doped with nonlinear molecules in the outer layer. We report here what is believed to be the first electro-optical Pockels signal and image from such a membrane. The electro-optical dephasing distribution within the membrane is imaged and the signal is shown to be linear as a function of the applied voltage. A theoretical analysis taking into account the statistical orientation distribution of the inserted dye molecules allows us to estimate the doped membrane nonlinearity. Ongoing extensions of this work to living cell membranes are discussed. PMID:19948120

  14. Differential Effect of Plant Lipids on Membrane Organization

    PubMed Central

    Grosjean, Kevin; Mongrand, Sébastien; Beney, Laurent; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

    2015-01-01

    The high diversity of the plant lipid mixture raises the question of their respective involvement in the definition of membrane organization. This is particularly the case for plant plasma membrane, which is enriched in specific lipids, such as free and conjugated forms of phytosterols and typical phytosphingolipids, such as glycosylinositolphosphoceramides. This question was here addressed extensively by characterizing the order level of membrane from vesicles prepared using various plant lipid mixtures and labeled with an environment-sensitive probe. Fluorescence spectroscopy experiments showed that among major phytosterols, campesterol exhibits a stronger ability than β-sitosterol and stigmasterol to order model membranes. Multispectral confocal microscopy, allowing spatial analysis of membrane organization, demonstrated accordingly the strong ability of campesterol to promote ordered domain formation and to organize their spatial distribution at the membrane surface. Conjugated sterol forms, alone and in synergy with free sterols, exhibit a striking ability to order membrane. Plant sphingolipids, particularly glycosylinositolphosphoceramides, enhanced the sterol-induced ordering effect, emphasizing the formation and increasing the size of sterol-dependent ordered domains. Altogether, our results support a differential involvement of free and conjugated phytosterols in the formation of ordered domains and suggest that the diversity of plant lipids, allowing various local combinations of lipid species, could be a major contributor to membrane organization in particular through the formation of sphingolipid-sterol interacting domains. PMID:25575593

  15. The electrical interplay between proteins and lipids in membranes.

    PubMed

    Richens, Joanna L; Lane, Jordan S; Bramble, Jonathan P; O'Shea, Paul

    2015-09-01

    All molecular interactions that are relevant to cellular and molecular structures are electrical in nature but manifest in a rich variety of forms that each has its own range and influences on the net effect of how molecular species interact. This article outlines how electrical interactions between the protein and lipid membrane components underlie many of the activities of membrane function. Particular emphasis is placed on spatially localised behaviour in membranes involving modulation of protein activity and microdomain structure. The interactions between membrane lipids and membrane proteins together with their role within cell biology represent an enormous body of work. Broad conclusions are not easy given the complexities of the various systems and even consensus with model membrane systems containing two or three lipid types is difficult. By defining two types of broad lipid-protein interaction, respectively Type I as specific and Type II as more non-specific and focussing on the electrical interactions mostly in the extra-membrane regions it is possible to assemble broad rules or a consensus of the dominant features of the interplay between these two fundamentally important classes of membrane component. This article is part of a special issue entitled: Lipid-protein interactions.

  16. Electro-hydrodynamic effects on lipid membranes in giant vesicles

    NASA Astrophysics Data System (ADS)

    Staykova, Margarita; Yamamoto, Tetsuya; Lipowsky, Reinhard; Dimova, Rumiana

    2009-11-01

    Electric fields are widely applied for cell manipulation in numerous micron-scale systems. Here, we show for the first time that alternating electric fields may cause pronounced flows in the membrane of giant lipid vesicles as well as in the surrounding fluid media.^ The lipid vesicles are not only biomimetic model for the cell membrane but also have many potential biotechnological applications, e.g. as drug-delivery systems and micro-reactors. The reported effects should be considered in electric micro-manipulation procedures on cells and vesicles. They might be useful for applications in microfluidic technologies, for lipid mixing, trapping and displacement, as will be demonstrated. We also believe that our method for visualization of the lipid flows by fluorescently labeled intra-membrane domains will be helpful for studies on membrane behavior in vesicles subjected to shear or mechanical stresses.

  17. Membrane proteins bind lipids selectively to modulate their structure and function

    PubMed Central

    Allison, Timothy M.; Ulmschneider, Martin B.; Degiacomi, Matteo T.; Baldwin, Andrew J.; Robinson, Carol V.

    2014-01-01

    Previous studies have established that the folding, structure and function of membrane proteins are influenced by their lipid environments1-7 and that lipids can bind to specific sites, for example in potassium channels8. Fundamental questions remain however regarding the extent of membrane protein selectivity toward lipids. Here we report a mass spectrometry (MS) approach designed to determine the selectivity of lipid binding to membrane protein complexes. We investigate the mechanosensitive channel of large conductance (MscL), aquaporin Z (AqpZ), and the ammonia channel (AmtB) using ion mobility MS (IM-MS), which reports gas-phase collision cross sections. We demonstrate that folded conformations of membrane protein complexes can exist in the gas-phase. By resolving lipid-bound states we then rank bound lipids based on their ability to resist gas phase unfolding and thereby stabilize membrane protein structure. Results show that lipids bind non-selectively and with high avidity to MscL, all imparting comparable stability, the highest-ranking lipid however is phosphatidylinositol phosphate, in line with its proposed functional role in mechanosensation9. AqpZ is also stabilized by many lipids with cardiolipin imparting the most significant resistance to unfolding. Subsequently, through functional assays, we discover that cardiolipin modulates AqpZ function. Analogous experiments identify AmtB as being highly selective for phosphatidylglycerol prompting us to obtain an X-ray structure in this lipid membrane-like environment. The 2.3Å resolution structure, when compared with others obtained without lipid bound, reveals distinct conformational changes that reposition AmtB residues to interact with the lipid bilayer. Overall our results demonstrate that resistance to unfolding correlates with specific lipid-binding events enabling distinction of lipids that merely bind from those that modulate membrane protein structure and/or function. We anticipate that these

  18. Electroporation of archaeal lipid membranes using MD simulations.

    PubMed

    Polak, Andraž; Tarek, Mounir; Tomšič, Matija; Valant, Janez; Ulrih, Nataša Poklar; Jamnik, Andrej; Kramar, Peter; Miklavčič, Damijan

    2014-12-01

    Molecular dynamics (MD) simulations were used to investigate the electroporation of archaeal lipid bilayers when subjected to high transmembrane voltages induced by a charge imbalance, mimicking therefore millisecond electric pulse experiments. The structural characteristics of the bilayer, a 9:91 mol% 2,3-di-O-sesterterpanyl-sn-glicerol-1-phospho-myo-inositol (AI) and 2,3-di-O-sesterterpanyl-sn-glicerol-1-phospho-1'(2'-O-α-D-glucosyl)-myo-inositol (AGI) were compared to small angle X-ray scattering data. A rather good agreement of the electron density profiles at temperatures of 298 and 343 K was found assessing therefore the validity of the protocols and force fields used in simulations. Compared to dipalmitoyl-phosphatidylcholine (DPPC), the electroporation threshold for the bilayer was found to increase from ~2 V to 4.3 V at 323 K, and to 5.2 V at 298 K. Comparing the electroporation thresholds of the archaeal lipids to those of simple diphytanoyl-phosphatidylcholine (DPhPC) bilayers (2.5 V at 323 K) allowed one to trace back the stability of the membranes to the structure of their lipid head groups. Addition of DPPC in amounts of 50 mol% to the archaeal lipid bilayers decreases their stability and lowers the electroporation thresholds to 3.8 V and 4.1 V at respectively 323 and 298 K. The present study therefore shows how membrane compositions can be selected to cover a wide range of responses to electric stimuli. This provides new routes for the design of liposomes that can be efficiently used as drug delivery carriers, as the selection of their composition allows one to tune in their electroporation threshold for subsequent release of their load.

  19. Chemotherapy drugs form ion pores in membranes due to physical interactions with lipids.

    PubMed

    Ashrafuzzaman, Mohammad; Tseng, Chih-Yuan; Duszyk, Marek; Tuszynski, Jack A

    2012-12-01

    We demonstrate the effects on membrane of the tubulin-binding chemotherapy drugs: thiocolchicoside and taxol. Electrophysiology recordings across lipid membranes in aqueous phases containing drugs were used to investigate the drug effects on membrane conductance. Molecular dynamics simulation of the chemotherapy drug-lipid complexes was used to elucidate the mechanism at an atomistic level. Both drugs are observed to induce stable ion-flowing pores across membranes. Discrete pore current-time plots exhibit triangular conductance events in contrast to rectangular ones found for ion channels. Molecular dynamics simulations indicate that drugs and lipids experience electrostatic and van der Waals interactions for short periods of time when found within each other's proximity. The energies from these two interactions are found to be similar to the energies derived theoretically using the screened Coulomb and the van der Waals interactions between peptides and lipids due to mainly their charge properties while forming peptide-induced ion channels in lipid bilayers. Experimental and in silico studies together suggest that the chemotherapy drugs induce ion pores inside lipid membranes due to drug-lipid physical interactions. The findings reveal cytotoxic effects of drugs on the cell membrane, which may aid in novel drug development for treatment of cancer and other diseases.

  20. Designing lipids for selective partitioning into liquid ordered membrane domains.

    PubMed

    Momin, Noor; Lee, Stacey; Gadok, Avinash K; Busch, David J; Bachand, George D; Hayden, Carl C; Stachowiak, Jeanne C; Sasaki, Darryl Y

    2015-04-28

    Self-organization of lipid molecules into specific membrane phases is key to the development of hierarchical molecular assemblies that mimic cellular structures. While the packing interaction of the lipid tails should provide the major driving force to direct lipid partitioning to ordered or disordered membrane domains, numerous examples show that the headgroup and spacer play important but undefined roles. We report here the development of several new biotinylated lipids that examine the role of spacer chemistry and structure on membrane phase partitioning. The new lipids were prepared with varying lengths of low molecular weight polyethylene glycol (EGn) spacers to examine how spacer hydrophilicity and length influence their partitioning behavior following binding with FITC-labeled streptavidin in liquid ordered (Lo) and liquid disordered (Ld) phase coexisting membranes. Partitioning coefficients (Kp Lo/Ld) of the biotinylated lipids were determined using fluorescence measurements in studies with giant unilamellar vesicles (GUVs). Compared against DPPE-biotin, DPPE-cap-biotin, and DSPE-PEG2000-biotin lipids, the new dipalmityl-EGn-biotin lipids exhibited markedly enhanced partitioning into liquid ordered domains, achieving Kp of up to 7.3 with a decaethylene glycol spacer (DP-EG10-biotin). We further demonstrated biological relevance of the lipids with selective partitioning to lipid raft-like domains observed in giant plasma membrane vesicles (GPMVs) derived from mammalian cells. Our results found that the spacer group not only plays a pivotal role for designing lipids with phase selectivity but may also influence the structural order of the domain assemblies.

  1. Adhesion and hemifusion of cytoplasmic myelin lipid membranes are highly dependent on the lipid composition

    PubMed Central

    Banquy, Xavier; Kristiansen, Kai; Lee, Dong Woog; Israelachvili, Jacob N.

    2012-01-01

    We report the effects of calcium ions on the adhesion and hemifusion mechanisms of model supported myelin lipid bilayer membranes of differing lipid composition. As in our previous studies [1, 2], the lipid compositions used mimic “healthy” and “diseased-like” (experimental autoimmune encephalomyelitis, EAE) membranes. Our results show that the interaction forces as a function of membrane separation distance are well described by a generic model that also (and in particular) includes the hydrophobic interaction arising from the hydrophobically exposed (interior) parts of the bilayers. The model is able to capture the mechanical instability that triggers the onset of the hemifusion event, and highlights the primary role of the hydrophobic interaction in membrane fusion. The effects of lipid composition on the fusion mechanism, and the adhesion forces between myelin lipid bilayers, can be summarized as follow: in calcium-free buffer, healthy membranes do not present any signs of adhesion or hemifusion, while diseased membranes hemifuse easily. Addition of 2 mM calcium favors adhesion and hemifusion of the membranes independently of their composition, but the mechanisms involved in the two processes were different: healthy bilayers systematically presented stronger adhesion forces and lower energy barriers to fusion compared to diseased bilayers. These results are of particular relevance for understanding lesion development (demyelination, swelling, vacuolization and/or vesiculation) in myelin associated diseases such as multiple sclerosis and its relationship to lipid domain formation in myelin membranes. PMID:22047743

  2. Lipid rafts and detergent-resistant membranes in epithelial keratinocytes.

    PubMed

    McGuinn, Kathleen P; Mahoney, Mỹ G

    2014-01-01

    Our understanding of the plasma membrane has markedly increased since Singer and Nicolson proposed the fluid mosaic model in 1972. While their revolutionary theory of the lipid bilayer remains largely valid, it is now known that lipids and proteins are not randomly dispersed throughout the plasma membrane but instead may be organized within membrane microdomains, commonly referred to as lipid rafts. Lipid rafts are highly dynamic, detergent resistant, and enriched with both cholesterol and glycosphingolipids. The two main types are flotillin-rich planar lipid rafts and caveolin-rich caveolae. It is proposed that flotillin and caveolin proteins regulate cell communication by compartmentalizing and interacting with signal transduction proteins within their respective lipid microdomains. Consequently, membrane rafts play an important role in vital cellular functions including migration, invasion, and signaling; thus, alterations in their microenvironment can initiate signaling pathways that affect cellular function and behavior. Therefore, the identification of lipid rafts and their associated proteins is integral to the study of transmembrane signaling. Here, we review the current standard protocols and biochemical approaches used to isolate and define raft proteins from epithelial cells and tissues. Furthermore, in Section 3 of this chapter, detailed protocols are offered for isolating lipid rafts by subjection to detergent and sucrose density centrifugation, as well as an approach for selectively isolating caveolae. Methods to manipulate rafts with treatments such as methyl-β-cyclodextrin and flotillin III are also described.

  3. Interaction of poloxamine block copolymers with lipid membranes: Role of copolymer structure and membrane cholesterol content.

    PubMed

    Sandez-Macho, Isabel; Casas, Matilde; Lage, Emilio V; Rial-Hermida, M Isabel; Concheiro, Angel; Alvarez-Lorenzo, Carmen

    2015-09-01

    Interactions of X-shaped poly(ethylene oxide)-poly(propylene oxide) (PEO-PPO) block copolymers with cell membranes were investigated recording the π-A isotherms of monolayer systems of dipalmitoylphosphatidylcholine (DPPC):cholesterol 100:0; 80:20 and 60:40 mol ratio and evaluating the capability of the copolymers to trigger haemolysis or to protect from haemolytic agents. Four varieties of poloxamine (Tetronic 904, 908, 1107 and 1307) were chosen in order to cover a wide range of EO and PO units contents and molecular weights, and compared to a variety of poloxamer (Pluronic P85). The π-A isotherms revealed that the greater the content in cholesterol, the stronger the interaction of the block copolymers with the lipids monolayer. The interactions were particularly relevant at low pressures and low lipid proportions, mimicking the conditions of damaged membranes. Relatively hydrophobic copolymers bearing short PEO blocks (e.g., T904 and P85) intercalated among the lipids expanding the surface area (ΔGexc) but not effectively sealing the pores. These varieties showed haemolytic behavior. Oppositely, highly hydrophilic copolymers bearing long PEO blocks (e.g., T908, T1107 and T1307) caused membrane contraction and outer leaflet sealing due to strong interactions of PEO with cholesterol and diamine core with phospholipids. These later varieties were not haemolytic and exerted a certain protective effect against spontaneous haemolysis for both intact erythrocytes and cholesterol-depleted erythrocytes.

  4. Lipid flow through fusion pores connecting membranes of different tensions.

    PubMed

    Chizmadzhev, Y A; Kumenko, D A; Kuzmin, P I; Chernomordik, L V; Zimmerberg, J; Cohen, F S

    1999-06-01

    When two membranes fuse, their components mix; this is usually described as a purely diffusional process. However, if the membranes are under different tensions, the material will spread predominantly by convection. We use standard fluid mechanics to rigorously calculate the steady-state convective flux of lipids. A fusion pore is modeled as a toroid shape, connecting two planar membranes. Each of the membrane monolayers is considered separately as incompressible viscous media with the same shear viscosity, etas. The two monolayers interact by sliding past each other, described by an intermonolayer viscosity, etar. Combining a continuity equation with an equation that balances the work provided by the tension difference, Deltasigma, against the energy dissipated by flow in the viscous membrane, yields expressions for lipid velocity, upsilon, and area of lipid flux, Phi. These expressions for upsilon and Phi depend on Deltasigma, etas, etar, and geometrical aspects of a toroidal pore, but the general features of the theory hold for any fusion pore that has a roughly hourglass shape. These expressions are readily applicable to data from any experiments that monitor movement of lipid dye between fused membranes under different tensions. Lipid velocity increases nonlinearly from a small value for small pore radii, rp, to a saturating value at large rp. As a result of velocity saturation, the flux increases linearly with pore radius for large pores. The calculated lipid flux is in agreement with available experimental data for both large and transient fusion pores.

  5. Lipid flow through fusion pores connecting membranes of different tensions.

    PubMed Central

    Chizmadzhev, Y A; Kumenko, D A; Kuzmin, P I; Chernomordik, L V; Zimmerberg, J; Cohen, F S

    1999-01-01

    When two membranes fuse, their components mix; this is usually described as a purely diffusional process. However, if the membranes are under different tensions, the material will spread predominantly by convection. We use standard fluid mechanics to rigorously calculate the steady-state convective flux of lipids. A fusion pore is modeled as a toroid shape, connecting two planar membranes. Each of the membrane monolayers is considered separately as incompressible viscous media with the same shear viscosity, etas. The two monolayers interact by sliding past each other, described by an intermonolayer viscosity, etar. Combining a continuity equation with an equation that balances the work provided by the tension difference, Deltasigma, against the energy dissipated by flow in the viscous membrane, yields expressions for lipid velocity, upsilon, and area of lipid flux, Phi. These expressions for upsilon and Phi depend on Deltasigma, etas, etar, and geometrical aspects of a toroidal pore, but the general features of the theory hold for any fusion pore that has a roughly hourglass shape. These expressions are readily applicable to data from any experiments that monitor movement of lipid dye between fused membranes under different tensions. Lipid velocity increases nonlinearly from a small value for small pore radii, rp, to a saturating value at large rp. As a result of velocity saturation, the flux increases linearly with pore radius for large pores. The calculated lipid flux is in agreement with available experimental data for both large and transient fusion pores. PMID:10354423

  6. Lipids and topological rules governing membrane protein assembly☆

    PubMed Central

    Bogdanov, Mikhail; Dowhan, William; Vitrac, Heidi

    2014-01-01

    Membrane protein folding and topogenesis are tuned to a given lipid profile since lipids and proteins have co-evolved to follow a set of interdependent rules governing final protein topological organization. Transmembrane domain (TMD) topology is determined via a dynamic process in which topogenic signals in the nascent protein are recognized and interpreted initially by the translocon followed by a given lipid profile in accordance with the Positive Inside Rule. The net zero charged phospholipid phosphatidylethanolamine and other neutral lipids dampen the translocation potential of negatively charged residues in favor of the cytoplasmic retention potential of positively charged residues (Charge Balance Rule). This explains why positively charged residues are more potent topological signals than negatively charged residues. Dynamic changes in orientation of TMDs during or after membrane insertion are attributed to non-sequential cooperative and collective lipid–protein charge interactions as well as long-term interactions within a protein. The proportion of dual topological conformers of a membrane protein varies in a dose responsive manner with changes in the membrane lipid composition not only in vivo but also in vitro and therefore is determined by the membrane lipid composition. Switching between two opposite TMD topologies can occur in either direction in vivo and also in liposomes (designated as fliposomes) independent of any other cellular factors. Such lipid-dependent post-insertional reversibility of TMD orientation indicates a thermodynamically driven process that can occur at any time and in any cell membrane driven by changes in the lipid composition. This dynamic view of protein topological organization influenced by the lipid environment reveals previously unrecognized possibilities for cellular regulation and understanding of disease states resulting from mis-folded proteins. This article is part of a Special Issue entitled: Protein Trafficking

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

    PubMed Central

    Cheney, Philip P.; Weisgerber, Alan W.; Feuerbach, Alec M.; Knowles, Michelle K.

    2017-01-01

    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. PMID:28294967

  8. Do local anesthetics interact preferentially with membrane lipid rafts? Comparative interactivities with raft-like membranes.

    PubMed

    Tsuchiya, Hironori; Ueno, Takahiro; Mizogami, Maki; Takakura, Ko

    2010-08-01

    Membranous lipid bilayers have been reconsidered as the site of action of local anesthetics (LAs). Recent understanding of biomembranes indicates the existence of lipid raft microdomains enriched in cholesterol and sphingolipids as potential platforms for channels and receptors. Based on the hypothesis that LAs may interact preferentially with lipid rafts over non-raft membranes, we compared their effects on raft model membranes and cardiolipin-containing biomimetic membranes. Liposomes were prepared with phospholipids, sphingomyelin, cerebroside, and cholesterol to have compositions corresponding to lipid rafts and cardiomyocyte mitochondrial membranes. After reacting LAs (50-200 microM) with the membrane preparations, their interactivities were determined by measuring fluorescence polarization with 1,6-diphenyl-1,3,5-hexatriene. Although bupivacaine and lidocaine acted on different raft-like liquid-ordered membranes to reduce polarization values, their effects on biomimetic less ordered membranes were much greater. LAs interacted with biomimetic membranes with the potency being R(+)-bupivacaine > racemic bupivacaine > S(-)-bupivacaine > ropivacaine > lidocaine > prilocaine, which is consistent with the rank order of pharmacotoxicological potency. However, raft model membranes showed neither structure-dependence nor stereoselectivity. The relevance of membrane lipid rafts to LAs is questionable at least in their effects on raft-like liquid-ordered membranes.

  9. A novel biotinylated lipid raft reporter for electron microscopic imaging of plasma membrane microdomains[S

    PubMed Central

    Krager, Kimberly J.; Sarkar, Mitul; Twait, Erik C.; Lill, Nancy L.; Koland, John G.

    2012-01-01

    The submicroscopic spatial organization of cell surface receptors and plasma membrane signaling molecules is readily characterized by electron microscopy (EM) via immunogold labeling of plasma membrane sheets. Although various signaling molecules have been seen to segregate within plasma membrane microdomains, the biochemical identity of these microdomains and the factors affecting their formation are largely unknown. Lipid rafts are envisioned as submicron membrane subdomains of liquid ordered structure with differing lipid and protein constituents that define their specific varieties. To facilitate EM investigation of inner leaflet lipid rafts and the localization of membrane proteins therein, a unique genetically encoded reporter with the dually acylated raft-targeting motif of the Lck kinase was developed. This reporter, designated Lck-BAP-GFP, incorporates green fluorescent protein (GFP) and biotin acceptor peptide (BAP) modules, with the latter allowing its single-step labeling with streptavidin-gold. Lck-BAP-GFP was metabolically biotinylated in mammalian cells, distributed into low-density detergent-resistant membrane fractions, and was readily detected with avidin-based reagents. In EM images of plasma membrane sheets, the streptavidin-gold-labeled reporter was clustered in 20–50 nm microdomains, presumably representative of inner leaflet lipid rafts. The utility of the reporter was demonstrated in an investigation of the potential lipid raft localization of the epidermal growth factor receptor. PMID:22822037

  10. A novel biotinylated lipid raft reporter for electron microscopic imaging of plasma membrane microdomains.

    PubMed

    Krager, Kimberly J; Sarkar, Mitul; Twait, Erik C; Lill, Nancy L; Koland, John G

    2012-10-01

    The submicroscopic spatial organization of cell surface receptors and plasma membrane signaling molecules is readily characterized by electron microscopy (EM) via immunogold labeling of plasma membrane sheets. Although various signaling molecules have been seen to segregate within plasma membrane microdomains, the biochemical identity of these microdomains and the factors affecting their formation are largely unknown. Lipid rafts are envisioned as submicron membrane subdomains of liquid ordered structure with differing lipid and protein constituents that define their specific varieties. To facilitate EM investigation of inner leaflet lipid rafts and the localization of membrane proteins therein, a unique genetically encoded reporter with the dually acylated raft-targeting motif of the Lck kinase was developed. This reporter, designated Lck-BAP-GFP, incorporates green fluorescent protein (GFP) and biotin acceptor peptide (BAP) modules, with the latter allowing its single-step labeling with streptavidin-gold. Lck-BAP-GFP was metabolically biotinylated in mammalian cells, distributed into low-density detergent-resistant membrane fractions, and was readily detected with avidin-based reagents. In EM images of plasma membrane sheets, the streptavidin-gold-labeled reporter was clustered in 20-50 nm microdomains, presumably representative of inner leaflet lipid rafts. The utility of the reporter was demonstrated in an investigation of the potential lipid raft localization of the epidermal growth factor receptor.

  11. Mechanisms of membrane deformation by lipid-binding domains.

    PubMed

    Itoh, Toshiki; Takenawa, Tadaomi

    2009-09-01

    Among an increasing number of lipid-binding domains, a group that not only binds to membrane lipids but also changes the shape of the membrane has been found. These domains are characterized by their strong ability to transform globular liposomes as well as flat plasma membranes into elongated membrane tubules both in vitro and in vivo. Biochemical studies on the structures of these proteins have revealed the importance of the amphipathic helix, which potentially intercalates into the lipid bilayer to induce and/or sense membrane curvature. Among such membrane-deforming domains, BAR and F-BAR/EFC domains form crescent-shaped dimers, suggesting a preference for a curved membrane, which is important for curvature sensing. Bioinformatics in combination with structural analyses has been identifying an increasing number of novel families of lipid-binding domains. This review attempts to summarize the evidence obtained by recent studies in order to gain general insights into the roles of membrane-deforming domains in a variety of biological events.

  12. Anthrax toxin-induced rupture of artificial lipid bilayer membranes

    PubMed Central

    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.

    2013-01-01

    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. PMID:23947891

  13. Anthrax toxin-induced rupture of artificial lipid bilayer membranes

    NASA Astrophysics Data System (ADS)

    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.

    2013-08-01

    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.

  14. Steady-state compartmentalization of lipid membranes by active proteins.

    PubMed Central

    Sabra, M C; Mouritsen, O G

    1998-01-01

    Using a simple microscopic model of lipid-protein interactions, based on the hydrophobic matching principle, we study some generic aspects of lipid-membrane compartmentalization controlled by a dispersion of active integral membrane proteins. The activity of the proteins is simulated by conformational excitations governed by an external drive, and the deexcitation is controlled by interaction of the protein with its lipid surroundings. In response to the flux of energy into the proteins from the environment and the subsequent dissipation of energy into the lipid bilayer, the lipid-protein assembly reorganizes into a steady-state structure with a typical length scale determined by the strength of the external drive. In the specific case of a mixed dimyristoylphosphatidylcholine-distearoylphosphatidylcholine bilayer in the gel-fluid coexistence region, it is shown explicitly by computer simulation that the activity of an integral membrane protein can lead to a compartmentalization of the lipid-bilayer membrane. The compartmentalization is related to the dynamical process of phase separation and lipid domain formation. PMID:9533687

  15. Procyanidins can interact with Caco-2 cell membrane lipid rafts: involvement of cholesterol.

    PubMed

    Verstraeten, Sandra V; Jaggers, Grayson K; Fraga, Cesar G; Oteiza, Patricia I

    2013-11-01

    Large procyanidins (more than three subunits) are not absorbed at the gastrointestinal tract but could exert local effects through their interactions with membranes. We previously showed that hexameric procyanidins (Hex), although not entering cells, interact with membranes modulating cell signaling and fate. This paper investigated if Hex, as an example of large procyanidins, can selectively interact with lipid rafts which could in part explain its biological actions. This mechanism was studied in both synthetic membranes (liposomes) and Caco-2 cells. Hex promoted Caco-2 cell membrane rigidification and dehydration, effects that were abolished upon cholesterol depletion with methyl-β-cyclodextrin (MCD). Hex prevented lipid raft structure disruption induced by cholesterol depletion/redistribution by MCD or sodium deoxycholate. Supporting the involvement of cholesterol-Hex bonding in Hex interaction with lipid rafts, the absence of cholesterol markedly decreased the capacity of Hex to prevent deoxycholate- and Triton X-100-mediated disruption of lipid raft-like liposomes. Stressing the functional relevance of this interaction, Hex mitigated lipid raft-associated activation of the extracellular signal-regulated kinases (ERK) 1/2. Results support the capacity of a large procyanidin (Hex) to interact with membrane lipid rafts mainly through Hex-cholesterol bondings. Procyanidin-lipid raft interactions can in part explain the capacity of large procyanidins to modulate cell physiology.

  16. Plasma Membrane Lipids and Their Role in Fungal Virulence

    PubMed Central

    Del Poeta, Maurizio

    2016-01-01

    There has been considerable evidence in recent years suggesting that plasma membrane lipids are important regulators of fungal pathogenicity. Various glycolipids have been shown to impart virulent properties in several fungal species, while others have been shown to play a role in host defense. In addition to their role as virulence factors, lipids also contribute to other virulence mechanisms such as drug resistance, biofilm formation, and release of extracellular vesicles. In addition, lipids also affect the mechanical properties of the plasma membrane through the formation of packed microdomains composed mainly of sphingolipids and sterols. Changes in the composition of lipid microdomains has been shown to disrupt the localization of virulence factors and affect fungal pathogenicity. This review gathers evidence on the various roles of plasma membrane lipids in fungal virulence and how lipids might contribute to the different processes that occur during infection and treatment. Insight into the role of lipids in fungal virulence can lead to an improved understanding of the process of fungal pathogenesis and the development of new lipid-mediated therapeutic strategies. PMID:26703191

  17. Interactions of Lipid Membranes with Fibrillar Protein Aggregates.

    PubMed

    Gorbenko, Galyna; Trusova, Valeriya; Girych, Mykhailo; Adachi, Emi; Mizuguchi, Chiharu; Saito, Hiroyuki

    2015-01-01

    Amyloid fibrils are an intriguing class of protein aggregates with distinct physicochemical, structural and morphological properties. They display peculiar membrane-binding behavior, thus adding complexity to the problem of protein-lipid interactions. The consensus that emerged during the past decade is that amyloid cytotoxicity arises from a continuum of cross-β-sheet assemblies including mature fibrils. Based on literature survey and our own data, in this chapter we address several aspects of fibril-lipid interactions, including (i) the effects of amyloid assemblies on molecular organization of lipid bilayer; (ii) competition between fibrillar and monomeric membrane-associating proteins for binding to the lipid surface; and (iii) the effects of lipids on the structural morphology of fibrillar aggregates. To illustrate some of the processes occurring in fibril-lipid systems, we present and analyze fluorescence data reporting on lipid bilayer interactions with fibrillar lysozyme and with the N-terminal 83-residue fragment of amyloidogenic mutant apolipoprotein A-I, 1-83/G26R/W@8. The results help understand possible mechanisms of interaction and mutual remodeling of amyloid fibers and lipid membranes, which may contribute to amyloid cytotoxicity.

  18. Imidazolium-Based Lipid Analogues and Their Interaction with Phosphatidylcholine Membranes.

    PubMed

    Wang, Da; de Jong, Djurre H; Rühling, Andreas; Lesch, Volker; Shimizu, Karina; Wulff, Stephanie; Heuer, Andreas; Glorius, Frank; Galla, Hans-Joachim

    2016-12-06

    4,5-Dialkylated imidazolium lipid salts are a new class of lipid analogues showing distinct biological activities. The potential effects of the imidazolium lipids on artificial lipid membranes and the corresponding membrane interactions was analyzed. Therefore, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) was employed to create an established lipid monolayer model and a bilayer membrane. Mixed monolayers of DPPC and 4,5-dialkylimidazolium lipids differing by their alkyl chain length (C7, C11, and C15) were characterized by surface pressure-area (π-A) isotherms using a Wilhelmy film balance in combination with epifluorescence microscopy. Monolayer hysteresis for binary mixtures was examined by recording triplicate consecutive compression-expansion cycles. The lipid miscibility and membrane stability of DPPC/imidazolium lipids were subsequently evaluated by the excess mean molecular area (ΔA(ex)) and the excess Gibbs free energy (ΔG(ex)) of mixing. Furthermore, the thermotropic behavior of mixed liposomes of DPPC/imidazolium lipids was investigated by differential scanning calorimetry (DSC). The C15-imidazolium lipid (C15-IMe·HI) forms a thermodynamically favored and kinetically reversible Langmuir monolayer with DPPC and exhibits a rigidification effect on both DPPC monolayer and bilayer structures at low molar fractions (X ≤ 0.3). However, the incorporation of the C11-imidazolium lipid (C11-IMe·HI) causes the formation of an unstable and irreversible Langmuir-Gibbs monolayer with DPPC and disordered DPPC liposomes. The C7-imidazolium lipid (C7-IMe·HI) displays negligible membrane activity. To better understand these results on a molecular level, all-atom molecular dynamics (MD) simulations were performed. The simulations yield two opposing molecular mechanisms governing the different behavior of the three imidazolium lipids: a lateral ordering effect and a free volume/stretching effect. Overall, our study provides the first evidence that the membrane

  19. Stabilization of concentration fluctuations in mixed membranes by hybrid lipids

    NASA Astrophysics Data System (ADS)

    Palmieri, Benoit; Safran, Samuel

    2012-02-01

    Finite-size domains have been observed at the surface of cells. These lipids ``rafts'' are stable nanodomains enriched in saturated lipids and cholesterol. While line tension favors macrodomains, one explanation for raft stabilization suggests that the membrane composition is tuned close to a spinodal temperature. From this point of view, rafts are long-lived concentration fluctuations in the mixed phase. We propose a ternary mixture model for the cell membrane that includes hybrid lipids which have one saturated and one unsaturated hydrocarbon chain. Finite amount of hybrid lipids reduces the packing incompatibility at the saturated/unsaturated lipid interface and stabilizes the concentration fluctuations. Hybrid-Hybrid interactions are included in the model and further increase the life-time of the rafts and decrease their length-scales. Moreover, the hybrid has extra orientational degrees of freedom that may lead to modulated phases.

  20. Edelfosine and miltefosine effects on lipid raft properties: membrane biophysics in cell death by antitumor lipids.

    PubMed

    Castro, Bruno M; Fedorov, Aleksander; Hornillos, Valentin; Delgado, Javier; Acuña, A Ulises; Mollinedo, Faustino; Prieto, Manuel

    2013-07-03

    Edelfosine (1-O-octadecyl-2-O-methyl-sn-glycero-phosphocholine) and miltefosine (hexadecylphosphocholine) are synthetic alkylphospholipids (ALPs) that are reported to selectively accumulate in tumor cell membranes, inducing Fas clustering and activation on lipid rafts, triggering apoptosis. However, the exact mechanism by which these lipids elicit these events is still not fully understood. Recent studies propose that their mode of action might be related with alterations of lipid rafts biophysical properties caused by these lipid drugs. To achieve a clear understanding of this mechanism, we studied the effects of pharmacologically relevant amounts of edelfosine and miltefosine in the properties of model and cellular membranes. The influence of these molecules on membrane order, lateral organization, and lipid rafts molar fraction and size were studied by steady-state and time-resolved fluorescence methods, Förster resonance energy transfer (FRET), confocal and fluorescence lifetime imaging microscopy (FLIM). We found that the global membrane and lipid rafts biophysical properties of both model and cellular membranes were not significantly affected by both the ALPs. Nonetheless, in model membranes, a mild increase in membrane fluidity induced by both alkyl lipids was detected, although this effect was more noticeable for edelfosine than miltefosine. This absence of drastic alterations shows for the first time that ALPs mode of action is unlikely to be directly linked to alterations of lipid rafts biophysical properties caused by these drugs. The biological implications of this result are discussed in the context of ALPs effects on lipid metabolism, mitochondria homeostasis modulation, and their relationship with tumor cell death.

  1. Lipid binding and membrane penetration of polymyxin B derivatives studied in a biomimetic vesicle system.

    PubMed Central

    Katz, Marina; Tsubery, Haim; Kolusheva, Sofiya; Shames, Alex; Fridkin, Mati; Jelinek, Raz

    2003-01-01

    Understanding membrane interactions and cell-wall permeation of Gram-negative bacteria is of great importance, owing to increasing bacterial resistance to existing drugs and therapeutic treatments. Here we use biomimetic lipid vesicles to analyse membrane association and penetration by synthetic derivatives of polymyxin B (PMB), a potent naturally occurring antibacterial cyclic peptide. The PMB analogues studied were PMB nonapeptide (PMBN), in which the hydrophobic alkyl residue was cleaved, PMBN diastereomer containing D-instead of L-amino acids within the cyclic ring (dPMBN) and PMBN where the hydrophobic alkyl chain was replaced with an Ala6 repeat (Ala6-PMBN). Peptide binding measurements, colorimetric transitions induced within the vesicles, fluorescence quenching experiments and ESR spectroscopy were applied to investigate the structural parameters underlying the different membrane-permeation profiles and biological activities of the analogues. The experiments point to the role of negatively charged lipids in membrane binding and confirm the prominence of lipopolisaccharide (LPS) in promoting membrane association and penetration by the peptides. Examination of the lipid interactions of the PMB derivatives shows that the cyclic moiety of PMB is not only implicated in lipid attachment and LPS binding, but also affects penetration into the inner bilayer core. The addition of the Ala6 peptide moiety, however, does not significantly promote peptide insertion into the hydrophobic lipid environment. The data also indicate that the extent of penetration into the lipid bilayer is not related to the overall affinity of the peptides to the membrane. PMID:12848621

  2. Mechano-capacitive properties of polarized membranes and the application to conductance measurements of lipid membrane patches

    NASA Astrophysics Data System (ADS)

    Zecchi, Karis A.; Mosgaard, Lars D.; Heimburg, Thomas

    2017-01-01

    Biological membranes are capacitors that can be charged by applying an electric field across the membrane. The charges on the capacitor exert a force on the membrane that leads to electrostriction, i.e., a thinning of the membrane. This effect is especially strong close to chain melting transitions. A consequence is voltage induced pore formation in the lipid membrane. Since the force is quadratic in voltage, negative and positive voltages have an identical influence on the physics of symmetric membranes. This is not the case for a membrane with an asymmetry leading to a permanent electric polarization. Positive and negative voltages of identical magnitude lead to different physical properties. Such an asymmetry can originate from a lipid composition that is different on the two monolayers of the membrane, or from membrane curvature. The latter effect is called flexoelectricity. It was investigated in detail by A.G. Petrov in the recent decades. As a consequence of permanent polarization, the membrane capacitor is discharged at a voltage different from zero. This leads to interesting electrical phenomena such as outward or inward rectification of membrane permeability. The changes in current-voltage relationships are consistent with the known magnitude of the flexoelectric effect.

  3. Coupling between pore formation and phase separation in charged lipid membranes

    NASA Astrophysics Data System (ADS)

    Himeno, Hiroki; Ito, Hiroaki; Higuchi, Yuji; Hamada, Tsutomu; Shimokawa, Naofumi; Takagi, Masahiro

    2015-12-01

    We investigated the effect of charge on the membrane morphology of giant unilamellar vesicles (GUVs) composed of various mixtures containing charged lipids. We observed the membrane morphologies by fluorescent and confocal laser microscopy in lipid mixtures consisting of a neutral unsaturated lipid [dioleoylphosphatidylcholine (DOPC)], a neutral saturated lipid [dipalmitoylphosphatidylcholine (DPPC)], a charged unsaturated lipid [dioleoylphosphatidylglycerol (DOP G(-)) ], a charged saturated lipid [dipalmitoylphosphatidylglycerol (DPP G(-)) ], and cholesterol (Chol). In binary mixtures of neutral DOPC-DPPC and charged DOPC -DPP G(-) , spherical vesicles were formed. On the other hand, pore formation was often observed with GUVs consisting of DOP G(-) and DPPC. In a DPPC-DPPG(-) -Chol ternary mixture, pore-formed vesicles were also frequently observed. The percentage of pore-formed vesicles increased with the DPP G(-) concentration. Moreover, when the head group charges of charged lipids were screened by the addition of salt, pore-formed vesicles were suppressed in both the binary and ternary charged lipid mixtures. We discuss the mechanisms of pore formation in charged lipid mixtures and the relationship between phase separation and the membrane morphology. Finally, we reproduce the results seen in experimental systems by using coarse-grained molecular dynamics simulations.

  4. Lipid domains in model membranes: a brief historical perspective.

    PubMed

    Mouritsen, Ole G; Bagatolli, Luis A

    2015-01-01

    All biological membranes consist of a complex composite of macromolecules and macromolecular assemblies, of which the fluid lipid-bilayer component is a core element with regard to cell encapsulation and barrier properties. The fluid lipid bilayer also supports the functional machinery of receptors, channels and pumps that are associated with the membrane. This bilayer is stabilized by weak physical and colloidal forces, and its nature is that of a self-assembled system of amphiphiles in water. Being only approximately 5 nm in thickness and still encapsulating a cell that is three orders of magnitude larger in diameter, the lipid bilayer as a material has very unusual physical properties, both in terms of structure and dynamics. Although the lipid bilayer is a fluid, it has a distinct and structured trans-bilayer profile, and in the plane of the bilayer the various molecular components, viz different lipid species and membrane proteins, have the capacity to organize laterally in terms of differentiated domains on different length and time scales. These elements of small-scale structure and order are crucial for the functioning of the membrane. It has turned out to be difficult to quantitatively study the small-scale structure of biological membranes. A major part of the insight into membrane micro- and nano-domains and the concepts used to describe them have hence come from studies of simple lipid bilayers as models of membranes, by use of a wide range of theoretical, experimental and simulational approaches. Many questions remain to be answered as to which extent the result from model studies can carry over to real biological membranes.

  5. Properties of Membranes Derived from the Total Lipids Extracted from the Human Lens Cortex and Nucleus

    PubMed Central

    Mainali, Laxman; Raguz, Marija; O’Brien, William J.; Subczynski, Witold K.

    2013-01-01

    Human lens lipid membranes prepared using a rapid solvent exchange method from the total lipids extracted from the clear lens cortex and nucleus of 41- to 60-year-old donors were investigated using electron paramagnetic resonance spin-labeling. Profiles of the phospholipid alkyl-chain order, fluidity, oxygen transport parameter, and hydrophobicity were assessed across coexisting membrane domains. Membranes prepared from the lens cortex and nucleus were found to contain two distinct lipid environments, the bulk phospholipid-cholesterol domain and the cholesterol bilayer domain (CBD). The alkyl chains of phospholipids were strongly ordered at all depths, indicating that the amplitude of the wobbling motion of alkyl chains was small. However, profiles of the membrane fluidity, which explicitly contain time (expressed as the spin-lattice relaxation rate) and depend on the rotational motion of spin labels, show relatively high fluidity of alkyl chains close to the membrane center. Profiles of the oxygen transport parameter and hydrophobicity have a rectangular shape and also indicate a high fluidity and hydrophobicity of the membrane center. The amount of CBD was greater in nuclear membranes than in cortical membranes. The presence of the CBD in lens lipid membranes, which at 37°C showed a permeability coefficient for oxygen about 60% smaller than across a water layer of the same thickness, would be expected to raise the barrier for oxygen transport across the fiber cell membrane. Properties of human membranes are compared with those obtained for membranes made of lipids extracted from cortex and nucleus of porcine and bovine eye lenses. PMID:23438364

  6. Equilibrium microphase separation in the two-leaflet model of lipid membranes

    NASA Astrophysics Data System (ADS)

    Reigada, Ramon; Mikhailov, Alexander S.

    2016-01-01

    Because of the coupling between local lipid composition and the thickness of the membrane, microphase separation in two-component lipid membranes can take place; such effects may underlie the formation of equilibrium nanoscale rafts. Using a kinetic description, this phenomenon is analytically and numerically investigated. The phase diagram is constructed through the stability analysis for linearized kinetic equations, and conditions for microphase separation are discussed. Simulations of the full kinetic model reveal the development of equilibrium membrane nanostructures with various morphologies from the initial uniform state.

  7. 3D-Membrane Stacks on Supported Membranes Composed of Diatom Lipids Induced by Long-Chain Polyamines.

    PubMed

    Gräb, Oliver; Abacilar, Maryna; Daus, Fabian; Geyer, Armin; Steinem, Claudia

    2016-10-04

    Long-chain polyamines (LCPAs) are intimately involved in the biomineralization process of diatoms taking place in silica deposition vesicles being acidic compartments surrounded by a lipid bilayer. Here, we addressed the question whether and how LCPAs interact with lipid membranes composed of glycerophospholipids and glyceroglycolipids mimicking the membranes of diatoms and higher plants. Solid supported lipid bilayers and monolayers containing the three major components that are unique in diatoms and higher plants, i.e., monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and sulfoquinovosyldiacylglycerol (SQDG), were prepared by spreading small unilamellar vesicles. The integrity of the membranes was investigated by fluorescence microscopy and atomic force microscopy showing continuous flat bilayers and monolayers with small protrusions on top of the membrane. The addition of a synthetic polyamine composed of 13 amine groups separated by a propyl spacer (C3N13) results in flat but three-dimensional membrane stacks within minutes. The membrane stacks are connected with the underlying membrane as verified by fluorescence recovery after photobleaching experiments. Membrane stack formation was found to be independent of the lipid composition; i.e., neither glyceroglycolipids nor negatively charged lipids were required. However, the formation process was strongly dependent on the chain length of the polyamine. Whereas short polyamines such as the naturally occurring spermidine, spermine, and the synthetic polyamines C3N4 and C3N5 do not induce stack formation, those containing seven and more amine groups (C3N7, C3N13, and C3N18) do form membrane stacks. The observed stack formation might have implications for the stability and expansion of the silica deposition vesicle during valve and girdle band formation in diatoms.

  8. Ceramides in the skin lipid membranes: length matters.

    PubMed

    Skolová, Barbora; Janůšová, Barbora; Zbytovská, Jarmila; Gooris, Gert; Bouwstra, Joke; Slepička, Petr; Berka, Pavel; Roh, Jaroslav; Palát, Karel; Hrabálek, Alexandr; Vávrová, Kateřina

    2013-12-17

    Ceramides are essential constituents of the skin barrier that allow humans to live on dry land. Reduced levels of ceramides have been associated with skin diseases, e.g., atopic dermatitis. However, the structural requirements and mechanisms of action of ceramides are not fully understood. Here, we report the effects of ceramide acyl chain length on the permeabilities and biophysics of lipid membranes composed of ceramides (or free sphingosine), fatty acids, cholesterol, and cholesterol sulfate. Short-chain ceramides increased the permeability of the lipid membranes compared to a long-chain ceramide with maxima at 4-6 carbons in the acyl. By a combination of differential scanning calorimetry, Fourier transform infrared spectroscopy, X-ray diffraction, Langmuir monolayers, and atomic force microscopy, we found that the reason for this effect in short ceramides was a lower proportion of tight orthorhombic packing and phase separation of continuous short ceramide-enriched domains with shorter lamellar periodicity compared to native long ceramides. Thus, long acyl chains in ceramides are essential for the formation of tightly packed impermeable lipid lamellae. Moreover, the model skin lipid membranes are a valuable tool to study the relationships between the lipid structure and composition, lipid organization, and the membrane permeability.

  9. Digital holographic microscopy of phase separation in multicomponent lipid membranes

    NASA Astrophysics Data System (ADS)

    Farzam Rad, Vahideh; Moradi, Ali-Reza; Darudi, Ahmad; Tayebi, Lobat

    2016-12-01

    Lateral in-homogeneities in lipid compositions cause microdomains formation and change in the physical properties of biological membranes. With the presence of cholesterol and mixed species of lipids, phospholipid membranes segregate into lateral domains of liquid-ordered and liquid-disordered phases. Coupling of two-dimensional intralayer phase separations and interlayer liquid-crystalline ordering in multicomponent membranes has been previously demonstrated. By the use of digital holographic microscopy (DHMicroscopy), we quantitatively analyzed the volumetric dynamical behavior of such membranes. The specimens are lipid mixtures composed of sphingomyelin, cholesterol, and unsaturated phospholipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine. DHMicroscopy in a transmission mode is an effective tool for quantitative visualization of phase objects. By deriving the associated phase changes, three-dimensional information on the morphology variation of lipid stacks at arbitrary time scales is obtained. Moreover, the thickness distribution of the object at demanded axial planes can be obtained by numerical focusing. Our results show that the volume evolution of lipid domains follows approximately the same universal growth law of previously reported area evolution. However, the thickness of the domains does not alter significantly by time; therefore, the volume evolution is mostly attributed to the changes in area dynamics. These results might be useful in the field of membrane-based functional materials.

  10. Spectral Imaging to Measure Heterogeneity in Membrane Lipid Packing

    PubMed Central

    Sezgin, Erdinc; Waithe, Dominic; Bernardino de la Serna, Jorge; Eggeling, Christian

    2015-01-01

    Physicochemical properties of the plasma membrane have been shown to play an important role in cellular functionality. Among those properties, the molecular order of the lipids, or the lipid packing, is of high importance. Changes in lipid packing are believed to compartmentalize cellular signaling by initiating coalescence and conformational changes of proteins. A common way to infer membrane lipid packing is by using membrane-embedded polarity-sensitive dyes, whose emission spectrum is dependent on the molecular order of the immediate membrane environment. Here, we report on an improved determination of such spectral shifts in the emission spectrum of the polarity-sensitive dyes. This improvement is based on the use of spectral imaging on a scanning confocal fluorescence microscope in combination with an improved analysis, which considers the whole emission spectrum instead of just single wavelength ranges. Using this approach and the polarity-sensitive dyes C-Laurdan or Di-4-ANEPPDHQ, we were able to image—with high accuracy—minute differences in the lipid packing of model and cellular membranes. PMID:25755090

  11. Effect of membrane tension on the physical properties of DOPC lipid bilayer membrane

    PubMed Central

    Reddy, A. Srinivas; Warshaviak, Dora Toledo; Chachisvilis, Mirianas

    2013-01-01

    Molecular dynamics simulations of a dioleoylphosphocholine (DOPC) lipid bilayer were performed to explore its mechanosensitivity. Variations in the bilayer properties, such as area per lipid, volume, thickness, hydration depth (HD), hydration thickness (HT), lateral diffusion coefficient, and changes in lipid structural order were computed in the membrane tension range 0 to 15 dyn/cm. We determined that an increase in membrane tension results in a decrease in the bilayer thickness and HD of ∼5% and ∼5.7% respectively, whereas area per lipid, volume, and HT/HD increased by 6.8%, 2.4%, and 5% respectively. The changes in lipid conformation and orientation were characterized using orientational (S2) and deuterium (SCD) order parameters. Upon increase of membrane tension both order parameters indicated an increase in lipid disorder by 10– 20%, mostly in the tail end region of the hydrophobic chains. The effect of membrane tension on lipid lateral diffusion in the DOPC bilayer was analyzed on three different time scales corresponding to inertial motion, anomalous diffusion and normal diffusion. The results showed that lateral diffusion of lipid molecules is anomalous in nature due to the non-exponential distribution of waiting times. The anomalous and normal diffusion coefficients increased by 20% and 52% when the membrane tension changed from 0 to 15 dyn/cm, respectively. In conclusion, our studies showed that membrane tension causes relatively significant changes in the area per lipid, volume, polarity, membrane thickness, and fluidity of the membrane suggesting multiple mechanisms by which mechanical perturbation of the membrane could trigger mechanosensitive response in cells. PMID:22588133

  12. A theoretical formalism for aggregation of peroxidized lipids and plasma membrane stability during photolysis.

    PubMed Central

    Busch, N A; Yarmush, M L; Toner, M

    1998-01-01

    The objective of this investigation was to examine, from a theoretical perspective, the mechanism underlying the lysis of plasma membranes by photoinduced, chemically mediated damage such as is found in photolysis. Toward this end, a model is presented which relates the membrane lifetime to the thermodynamic parameters of the membrane components based upon the kinetic theory of aggregate formation. The formalism includes a standard birth/death process for the formation of damaged membrane components (i.e., peroxidized lipids) as well as a terminating condensation process for the formation of aggregates of peroxidized plasma membrane lipids. Our theory predicts that 1) the membrane lifetime is inversely correlated with predicted rate of membrane damage; 2) an upper limit on the duration of membrane damage exists, above which the mean and variance of the membrane lifetime is independent of further membrane damage; and 3) both the mean and variance of the time of membrane lifetime distribution are correlated with the number of sites that may be damaged to form a single membrane defect. The model provides a framework to optimize the lysis of cell membranes by photodynamic therapy. PMID:9826616

  13. Kinetic and thermodynamic aspects of lipid translocation in biological membranes.

    PubMed Central

    Frickenhaus, S; Heinrich, R

    1999-01-01

    A theoretical analysis of the lipid translocation in cellular bilayer membranes is presented. We focus on an integrative model of active and passive transport processes determining the asymmetrical distribution of the major lipid components between the monolayers. The active translocation of the aminophospholipids phosphatidylserine and phosphatidylethanolamine is mathematically described by kinetic equations resulting from a realistic ATP-dependent transport mechanism. Concerning the passive transport of the aminophospholipids as well as of phosphatidylcholine, sphingomyelin, and cholesterol, two different approaches are used. The first treatment makes use of thermodynamic flux-force relationships. Relevant forces are transversal concentration differences of the lipids as well as differences in the mechanical states of the monolayers due to lateral compressions. Both forces, originating primarily from the operation of an aminophospholipid translocase, are expressed as functions of the lipid compositions of the two monolayers. In the case of mechanical forces, lipid-specific parameters such as different molecular surface areas and compression force constants are taken into account. Using invariance principles, it is shown how the phenomenological coefficients depend on the total lipid amounts. In a second approach, passive transport is analyzed in terms of kinetic mechanisms of carrier-mediated translocation, where mechanical effects are incorporated into the translocation rate constants. The thermodynamic as well as the kinetic approach are applied to simulate the time-dependent redistribution of the lipid components in human red blood cells. In the thermodynamic model the steady-state asymmetrical lipid distribution of erythrocyte membranes is simulated well under certain parameter restrictions: 1) the time scales of uncoupled passive transbilayer movement must be different among the lipid species; 2) positive cross-couplings of the passive lipid fluxes are

  14. Laurdan fluorescence senses mechanical strain in the lipid bilayer membrane.

    PubMed

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

    2006-09-01

    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.

  15. Measuring the composition-curvature coupling in binary lipid membranes by computer simulations

    SciTech Connect

    Barragán Vidal, I. A. Müller, M.; Rosetti, C. M.; Pastorino, C.

    2014-11-21

    The coupling between local composition fluctuations in binary lipid membranes and curvature affects the lateral membrane structure. We propose an efficient method to compute the composition-curvature coupling in molecular simulations and apply it to two coarse-grained membrane models—a minimal, implicit-solvent model and the MARTINI model. Both the weak-curvature behavior that is typical for thermal fluctuations of planar bilayer membranes as well as the strong-curvature regime corresponding to narrow cylindrical membrane tubes are studied by molecular dynamics simulation. The simulation results are analyzed by using a phenomenological model of the thermodynamics of curved, mixed bilayer membranes that accounts for the change of the monolayer area upon bending. Additionally the role of thermodynamic characteristics such as the incompatibility between the two lipid species and asymmetry of composition are investigated.

  16. DNA Release from Lipoplexes by Anionic Lipids: Correlation with Lipid Mesomorphism, Interfacial Curvature, and Membrane Fusion

    PubMed Central

    Tarahovsky, Yury S.; Koynova, Rumiana; MacDonald, Robert C.

    2004-01-01

    DNA release from lipoplexes is an essential step during lipofection and is probably a result of charge neutralization by cellular anionic lipids. As a model system to test this possibility, fluorescence resonance energy transfer between DNA and lipid covalently labeled with Cy3 and BODIPY, respectively, was used to monitor the release of DNA from lipid surfaces induced by anionic liposomes. The separation of DNA from lipid measured this way was considerably slower and less complete than that estimated with noncovalently labeled DNA, and depends on the lipid composition of both lipoplexes and anionic liposomes. This result was confirmed by centrifugal separation of released DNA and lipid. X-ray diffraction revealed a clear correlation of the DNA release capacity of the anionic lipids with the interfacial curvature of the mesomorphic structures developed when the anionic and cationic liposomes were mixed. DNA release also correlated with the rate of fusion of anionic liposomes with lipoplexes. It is concluded that the tendency to fuse and the phase preference of the mixed lipid membranes are key factors for the rate and extent of DNA release. The approach presented emphasizes the importance of the lipid composition of both lipoplexes and target membranes and suggests optimal transfection may be obtained by tailoring lipoplex composition to the lipid composition of target cells. PMID:15298910

  17. Randomization of membrane lipids in relation to transport system assembly in Escherichia coli.

    PubMed

    Thilo, L; Overath, P

    1976-01-27

    The distribution of newly synthesized lipid molecules in the pre-existing lipid phase of the membrane was studied in whole cells of the fatty acid requiring Escheria coli strain K1062. The fluorescence probe N-phenyl-1-naphthylamine revealed reversible lipid phase transitions in cells supplemented with cis-delta9-octadecenoate (transition temperature Tt = 14 degrees C; width of the transition deltaT = 13 degrees C) or trans-delta9-hexadecenoate (Tt = 27 degrees C; deltaT = 7 degrees C). Cells were first grown in the presence of cis-delta9-octadecenoate at 37 degrees C and subsequently for various periods in the presence of trans-delta9-hexadecenoate at 37 or 22 degrees C, i.e. above or below the transition of the newly formed lipids. Reproducible phase transitions with single, well-defined Tt values between 14 and 27 degrees C were observed under both conditions. Beta-Galactoside transport induced in a similar experiment before or during a change in the fatty acid composition showed a single change in activation energy at a temperature close to the lipid transition temperature, Tt. Starvation of cis-delta9-octadecenoate-supplemented cells for this fatty acid led to a gradual rise in the transition temperature, due to an increase in the percentage of saturated acyl chains in the membrane lipids. It is concluded that under all conditions investigated a mixed lipid phase composed of newly synthesized and pre-existing lipid molecules is formed in the membrane. Since conserved domains of newly synthesized lipids surrounding simultaneously formed transport proteins could not be demonstrated, the results do not support a membrane assembly mechanism proposed by N. Tsukagoshi and C. F. Fox [(1973), Biochemistry 12, 2822-2829]. It rather appears that newly formed lipid molecules are continuously released from their sites of synthesis into the lipid matrix by a rapid diffusion-controlled process.

  18. EDTA-induced membrane fluidization and destabilization: biophysical studies on artificial lipid membranes.

    PubMed

    Prachayasittikul, Virapong; Isarankura-Na-Ayudhya, Chartchalerm; Tantimongcolwat, Tanawut; Nantasenamat, Chanin; Galla, Hans-Joachim

    2007-11-01

    The molecular mechanism of ethylenediaminetetraacetic acid (EDTA)-induced membrane destabilization has been studied using a combination of four biophysical techniques on artificial lipid membranes. Data from Langmuir film balance and epifluorescence microscopy revealed the fluidization and expansion effect of EDTA on phase behavior of monolayers of either 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or mixtures of DPPC and metal-chelating lipids, such as N(alpha),N(alpha)-Bis[carboxymethyl]-N(epsilon)-[(dioctadecylamino)succinyl]-L-lysine or 1,2-dioleoyl-sn-glycero-3-[N-(5-amino-1-carboxypentyl iminodiacetic acid) succinyl]. A plausible explanation could be drawn from the electrostatic interaction between negatively charged groups of EDTA and the positively charged choline head group of DPPC. Intercalation of EDTA into the lipid membrane induced membrane curvature as elucidated by atomic force microscopy. Growth in size and shape of the membrane protrusion was found to be time-dependent upon exposure to EDTA. Further loss of material from the lipid membrane surface was monitored in real time using a quartz crystal microbalance. This indicates membrane restabilization by exclusion of the protrusions from the surface. Loss of lipid components facilitates membrane instability, leading to membrane permeabilization and lysis.

  19. Penetration of Cell Membranes and Synthetic Lipid Bilayers by Nanoprobes

    PubMed Central

    Angle, Matthew R.; Wang, Andrew; Thomas, Aman; Schaefer, Andreas T.; Melosh, Nicholas A.

    2014-01-01

    Nanoscale devices have been proposed as tools for measuring and controlling intracellular activity by providing electrical and/or chemical access to the cytosol. Unfortunately, nanostructures with diameters of 50–500 nm do not readily penetrate the cell membrane, and rationally optimizing nanoprobes for cell penetration requires real-time characterization methods that are capable of following the process of membrane penetration with nanometer resolution. Although extensive work has examined the rupture of supported synthetic lipid bilayers, little is known about the applicability of these model systems to living cell membranes with complex lipid compositions, cytoskeletal attachment, and membrane proteins. Here, we describe atomic force microscopy (AFM) membrane penetration experiments in two parallel systems: live HEK293 cells and stacks of synthetic lipid bilayers. By using the same probes in both systems, we were able to clearly identify membrane penetration in synthetic bilayers and compare these events with putative membrane penetration events in cells. We examined membrane penetration forces for three tip geometries and 18 chemical modifications of the probe surface, and in all cases the median forces required to penetrate cellular and synthetic lipid bilayers with nanoprobes were greater than 1 nN. The penetration force was sensitive to the probe's sharpness, but not its surface chemistry, and the force did not depend on cell surface or cytoskeletal properties, with cells and lipid stacks yielding similar forces. This systematic assessment of penetration under various mechanical and chemical conditions provides insights into nanoprobe-cell interactions and informs the design of future intracellular nanoprobes. PMID:25418094

  20. Structure of spin-coated lipid films and domain formation in supported membranes formed by hydration.

    PubMed

    Simonsen, Adam Cohen; Bagatolli, Luis A

    2004-10-26

    An atomic force and fluorescence microscopy investigation of the structure of spin-coated lipid films is presented. In the surface of the dry film, lipids are found to orient in a conformation where acyl chains are pointing outward while laterally the individual layers of the multilamellar film exhibit a dewetting pattern similar to what is found in polymer thin films. Hydration of the film in liquid water promotes detachment of bilayers from the surface while a single membrane remains on the mica substrate. This supported membrane is highly uniform and defect-free as compared to supported membranes prepared by conventional methods. It is further demonstrated that supported membranes of binary lipid mixtures prepared by this method exhibit gel-fluid domain coexistence in accordance with expectations from the phase diagrams.

  1. Lipid membranes in external electric fields: kinetics of large pore formation causing rupture.

    PubMed

    Winterhalter, Mathias

    2014-06-01

    About 40 years ago, Helfrich introduced an elastic model to explain shapes and shape transitions of cells (Z Naturforsch C, 1973; 28:693). This seminal article stimulated numerous theoretical as well as experimental investigations and created new research fields. In particular, the predictive power of his approach was demonstrated in a large variety of lipid model system. Here in this review, we focus on the development with respect to planar lipid membranes in external electric fields. Stimulated by the early work of Helfrich on electric field forces acting on liposomes, we extended his early approach to understand the kinetics of lipid membrane rupture. First, we revisit the main forces determining the kinetics of membrane rupture followed by an overview on various experiments. Knowledge on the kinetics of defect formation may help to design stable membranes or serve for novel mechanism for controlled release.

  2. Probing protein-lipid interactions by FRET between membrane fluorophores

    NASA Astrophysics Data System (ADS)

    Trusova, Valeriya M.; Gorbenko, Galyna P.; Deligeorgiev, Todor; Gadjev, Nikolai

    2016-09-01

    Förster resonance energy transfer (FRET) is a powerful fluorescence technique that has found numerous applications in medicine and biology. One area where FRET proved to be especially informative involves the intermolecular interactions in biological membranes. The present study was focused on developing and verifying a Monte-Carlo approach to analyzing the results of FRET between the membrane-bound fluorophores. This approach was employed to quantify FRET from benzanthrone dye ABM to squaraine dye SQ-1 in the model protein-lipid system containing a polycationic globular protein lysozyme and negatively charged lipid vesicles composed of phosphatidylcholine and phosphatidylglycerol. It was found that acceptor redistribution between the lipid bilayer and protein binding sites resulted in the decrease of FRET efficiency. Quantification of this effect in terms of the proposed methodology yielded both structural and binding parameters of lysozyme-lipid complexes.

  3. Lipid Bilayer Vesicles with Numbers of Membrane-Linking Pores

    NASA Astrophysics Data System (ADS)

    Ken-ichirou Akashi,; Hidetake Miyata,

    2010-06-01

    We report that phospholipid membranes spontaneously formed in aqueous medium giant unilamellar vesicles (GUVs) possessing many membranous wormhole-like structures (membrane-linking pores, MLPs). By phase contract microscopy and confocal fluorescence microscopy, the structures of the MLPs, consisting of lipid bilayer, were resolvable, and a variety of vesicular shapes having many MLPs (a high genus topology) were found. These vesicles were stable but easily deformed by micromanipulation with a microneedle. We also observed the size reduction of the MLPs with the increase in membrane tension, which was qualitatively consistent with a prediction from a simple dynamical model.

  4. Phosphoinositides, Major Actors in Membrane Trafficking and Lipid Signaling Pathways

    PubMed Central

    De Craene, Johan-Owen; Bertazzi, Dimitri L.; Bär, Séverine; Friant, Sylvie

    2017-01-01

    Phosphoinositides are lipids involved in the vesicular transport of proteins and lipids between the different compartments of eukaryotic cells. They act by recruiting and/or activating effector proteins and thus are involved in regulating various cellular functions, such as vesicular budding, membrane fusion and cytoskeleton dynamics. Although detected in small concentrations in membranes, their role is essential to cell function, since imbalance in their concentrations is a hallmark of many cancers. Their synthesis involves phosphorylating/dephosphorylating positions D3, D4 and/or D5 of their inositol ring by specific lipid kinases and phosphatases. This process is tightly regulated and specific to the different intracellular membranes. Most enzymes involved in phosphoinositide synthesis are conserved between yeast and human, and their loss of function leads to severe diseases (cancer, myopathy, neuropathy and ciliopathy). PMID:28294977

  5. Molecular mechanisms of protein and lipid targeting to ciliary membranes

    PubMed Central

    Emmer, Brian T.; Maric, Danijela; Engman, David M.

    2010-01-01

    Cilia are specialized surface regions of eukaryotic cells that serve a variety of functions, ranging from motility to sensation and to regulation of cell growth and differentiation. The discovery that a number of human diseases, collectively known as ciliopathies, result from defective cilium function has expanded interest in these structures. Among the many properties of cilia, motility and intraflagellar transport have been most extensively studied. The latter is the process by which multiprotein complexes associate with microtubule motors to transport structural subunits along the axoneme to and from the ciliary tip. By contrast, the mechanisms by which membrane proteins and lipids are specifically targeted to the cilium are still largely unknown. In this Commentary, we review the current knowledge of protein and lipid targeting to ciliary membranes and outline important issues for future study. We also integrate this information into a proposed model of how the cell specifically targets proteins and lipids to the specialized membrane of this unique organelle. PMID:20145001

  6. Oxygen permeability of the lipid bilayer membrane made of calf lens lipids

    PubMed Central

    Widomska, Justyna; Raguz, Marija; Subczynski, Witold K.

    2007-01-01

    The oxygen permeability coefficient across the membrane made of the total lipid extract from the plasma membrane of calf lens was estimated from the profile of the oxygen transport parameter (local oxygen diffusion-concentration product) and compared with those estimated for membranes made of an equimolar 1-palmitoyl-2-oleoylphosphatidylcholine/cholesterol (POPC/Chol) mixture and of pure POPC. Profiles of the oxygen transport parameter were obtained by observing the collision of molecular oxygen with nitroxide radical spin labels placed at different depths in the membrane using the saturation-recovery EPR technique and were published by us earlier (J. Widomska, M. Raguz, J. Dillon, E. R. Gaillard, W. K. Subczynski, Biochim. Biophys. Acta. Epub 2007 March 20). At 35°C, the estimated oxygen permeability coefficients were 51.3, 49.7, and 157.4 cm/s for lens lipid, POPC/Chol, and POPC membranes, respectively (compared with 53.3 cm/s for a water layer with the same thickness as a membrane). Membrane permeability significantly decreases at lower temperatures. In the lens lipid membrane, resistance to the oxygen transport is located in and near the polar headgroup region of the membrane to the depth of the ninth carbon, which is approximately where the steroid-ring structure of cholesterol reaches into the membrane. In the central region of the membrane, oxygen transport is enhanced, significantly exceeding that in bulk water. It is concluded that the high level of cholesterol in lens lipids is responsible for these unique membrane properties. PMID:17662231

  7. Ionic channels and nerve membrane lipids. Cholesterol-tetrodotoxin interaction.

    PubMed

    Villegas, R; Barnola, F V; Camejo, G

    1970-04-01

    Experiments were carried out to investigate possible interactions of tetrodotoxin (TTX) with lipid molecules isolated from nerve fiber plasma membranes of the squid Dosidicus gigas. TTX has a highly selective ability to block the channel normally used by Na(+) to cross the axolemma during nervous impulse conduction. In order to investigate the interaction each lipid sample was spread on 5 x 10(-7)M TTX and TTX-free 0.15 M NaCl solutions adjusted to pH 7.4 with 7 x 10(-3)M phosphate buffer. The surface pressure-area diagrams of the lipid monolayers revealed that TTX interacts only with cholesterol. The expansion of the cholesterol monolayers at 5 x 10(-7)M TTX was 2 A(2)/molecule at zero pressure for the experiments at 20 degrees C and 2.5 A(2)/molecule for those at 25 degrees C. Similar results were obtained in KCl subphases. The apparent dissociation constant of the cholesterol-TTX complex calculated from dose-response experiments is 2.6 x 10(-7)M. Experiments at pH 10.1 revealed that the zwitter ionic form of TTX is less active. Experiments with cholesterol derivatives (cholesteryl acetate, cholesterol methyl ether, cholestanol, and cholestanyl acetate) indicate that for the interaction with TTX a partial negatively charged group at C-3 and a double bond between C-5 and C-6 on the steroid nucleus are required. Tetrodonic acid, a biologically inactive derivative of TTX, does not interact with cholesterol. The results lead us to propose that cholesterol is part of the Na(+) channel.

  8. PE and PS Lipids Synergistically Enhance Membrane Poration by a Peptide with Anticancer Properties.

    PubMed

    Leite, Natália Bueno; Aufderhorst-Roberts, Anders; Palma, Mario Sergio; Connell, Simon D; Ruggiero Neto, João; Beales, Paul A

    2015-09-01

    Polybia-MP1 (MP1) is a bioactive host-defense peptide with known anticancer properties. Its activity is attributed to excess serine (phosphatidylserine (PS)) on the outer leaflet of cancer cells. Recently, higher quantities of phosphatidylethanolamine (PE) were also found at these cells' surface. We investigate the interaction of MP1 with model membranes in the presence and absence of POPS (PS) and DOPE (PE) to understand the role of lipid composition in MP1's anticancer characteristics. Indeed we find that PS lipids significantly enhance the bound concentration of peptide on the membrane by a factor of 7-8. However, through a combination of membrane permeability assays and imaging techniques we find that PE significantly increases the susceptibility of the membrane to disruption by these peptides and causes an order-of-magnitude increase in membrane permeability by facilitating the formation of larger transmembrane pores. Significantly, atomic-force microscopy imaging reveals differences in the pore formation mechanism with and without the presence of PE. Therefore, PS and PE lipids synergistically combine to enhance membrane poration by MP1, implying that the combined enrichment of both these lipids in the outer leaflet of cancer cells is highly significant for MP1's anticancer action. These mechanistic insights could aid development of novel chemotherapeutics that target pathological changes in the lipid composition of cancerous cells.

  9. PE and PS Lipids Synergistically Enhance Membrane Poration by a Peptide with Anticancer Properties

    PubMed Central

    Leite, Natália Bueno; Aufderhorst-Roberts, Anders; Palma, Mario Sergio; Connell, Simon D.; Neto, João Ruggiero; Beales, Paul A.

    2015-01-01

    Polybia-MP1 (MP1) is a bioactive host-defense peptide with known anticancer properties. Its activity is attributed to excess serine (phosphatidylserine (PS)) on the outer leaflet of cancer cells. Recently, higher quantities of phosphatidylethanolamine (PE) were also found at these cells’ surface. We investigate the interaction of MP1 with model membranes in the presence and absence of POPS (PS) and DOPE (PE) to understand the role of lipid composition in MP1’s anticancer characteristics. Indeed we find that PS lipids significantly enhance the bound concentration of peptide on the membrane by a factor of 7–8. However, through a combination of membrane permeability assays and imaging techniques we find that PE significantly increases the susceptibility of the membrane to disruption by these peptides and causes an order-of-magnitude increase in membrane permeability by facilitating the formation of larger transmembrane pores. Significantly, atomic-force microscopy imaging reveals differences in the pore formation mechanism with and without the presence of PE. Therefore, PS and PE lipids synergistically combine to enhance membrane poration by MP1, implying that the combined enrichment of both these lipids in the outer leaflet of cancer cells is highly significant for MP1’s anticancer action. These mechanistic insights could aid development of novel chemotherapeutics that target pathological changes in the lipid composition of cancerous cells. PMID:26331251

  10. Lamellar biogels: Fluid-membrane-based hydrogels containing polymer lipids

    SciTech Connect

    Warriner, H.E.; Idziak, S.H.J.; Slack, N.L.

    1996-02-16

    A class of lamellar biological hydrogels comprised of fluid membranes of lipids and surfactants with small amounts of low molecular weight poly(ethylene glycol)-derived polymer pipids (PEG-lipids) were studied by x-ray diffraction, polarized light microscopy, and rheometry. In contrast to isotropic hydrogels of polymer networks, these membrane-based birefringent liquid crystalline biogels, labeled L{sub {alpha},g,} form the gel phase when water is added to the liquid-like lamellar L{sub {alpha}} phase, which reenters a liquid-like mixed phase upon further dilution. Furthermore, gels with larger water content require less PEG-lipid to remain stable. Although concentrated ({approx}50 weight percent) mixtures of free PEG (molecular weight, 5000) and water do not gel, gelatin does occur in mixtures containing as little as 0.5 weight percent PEG lipid. A defining signature of the L{sub {alpha}, g} regime as it sets in from the fluid lamellar L{sub {alpha}} phase is the proliferation of layer-dislocation-type defects, which are stabilized by the segregation of PEG-lipids to the defect regions of high membrane curvature that connect the membranes. 32 refs., 5 figs.

  11. Two-Phase Contiguous Supported Lipid Bilayer Model for Membrane Rafts via Polymer Blotting and Stenciling.

    PubMed

    Richards, Mark J; Daniel, Susan

    2017-02-07

    The supported lipid bilayer has been portrayed as a useful model of the cell membrane compatible with many biophysical tools and techniques that demonstrate its appeal in learning about the basic features of the plasma membrane. However, some of its potential has yet to be realized, particularly in the area of bilayer patterning and phase/composition heterogeneity. In this work, we generate contiguous bilayer patterns as a model system that captures the general features of membrane domains and lipid rafts. Micropatterned polymer templates of two types are investigated for generating patterned bilayer formation: polymer blotting and polymer lift-off stenciling. While these approaches have been used previously to create bilayer arrays by corralling bilayers patches with various types of boundaries impenetrable to bilayer diffusion, unique to the methods presented here, there are no physical barriers to diffusion. In this work, interfaces between contiguous lipid phases define the pattern shapes, with continuity between them allowing transfer of membrane-bound biomolecules between the phases. We examine effectors of membrane domain stability including temperature and cholesterol content to investigate domain dynamics. Contiguous patterning of supported bilayers as a model of lipid rafts expands the application of the SLB to an area with current appeal and brings with it a useful toolset for characterization and analysis. These combined tools should be helpful to researchers investigating lipid raft dynamics and function and biomolecule partitioning studies. Additionally, this patterning technique may be useful for applications such as bioseparations that exploit differences in lipid phase partitioning or creation of membranes that bind species like viruses preferentially at lipid phase boundaries, to name a few.

  12. Membrane Lipid Metabolism in Germinating Castor Bean Endosperm 1

    PubMed Central

    Donaldson, Robert P.

    1976-01-01

    Castor bean (Ricinus communis L. var. Hale) endosperms, excised after 2 days germination at 30 C, were incubated 5 min to 8 hr with 14C-acetate and 3H-glycerol. Homogenates were fractionated by sucrose gradient centrifugation. Organelles found to be active in lipid synthesis were the lipid bodies and the endoplasmic reticulum. The products of incorporation in the lipid bodies were 3H-diglycerides containing 14C-fatty acids of more than 20 carbons. In contrast, the endoplasmic reticulum produced 3H-phospholipids as well as 3H-diglycerides rich in 14C-linoleate. The phospholipids synthesized and their acyl contents were of the types known to be the major components of organelle membranes in this tissue. Phospholipids and diglycerides containing 14C and 3H were found in the glyoxysomes and mitochondria subsequent to their appearance in the endoplasmic reticulum. The results show that germinating castor bean endosperm synthesizes membrane lipids de novo from acetate rather than reutilizing stored lipid components directly. It is also apparent that the endoplasmic reticulum is responsible for several steps in membrane lipid production. PMID:16659516

  13. Fatty Acids from Membrane Lipids Become Incorporated into Lipid Bodies during Myxococcus xanthus Differentiation

    PubMed Central

    Bhat, Swapna; Boynton, Tye O.; Pham, Dan; Shimkets, Lawrence J.

    2014-01-01

    Myxococcus xanthus responds to amino acid limitation by producing fruiting bodies containing dormant spores. During development, cells produce triacylglycerides in lipid bodies that become consumed during spore maturation. As the cells are starved to induce development, the production of triglycerides represents a counterintuitive metabolic switch. In this paper, lipid bodies were quantified in wild-type strain DK1622 and 33 developmental mutants at the cellular level by measuring the cross sectional area of the cell stained with the lipophilic dye Nile red. We provide five lines of evidence that triacylglycerides are derived from membrane phospholipids as cells shorten in length and then differentiate into myxospores. First, in wild type cells, lipid bodies appear early in development and their size increases concurrent with an 87% decline in membrane surface area. Second, developmental mutants blocked at different stages of shortening and differentiation accumulated lipid bodies proportionate with their cell length with a Pearson's correlation coefficient of 0.76. Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten. Fourth, genes for fatty acid synthesis are down-regulated while genes for fatty acid degradation are up regulated. Finally, direct movement of fatty acids from membrane lipids in growing cells to lipid bodies in developing cells was observed by pulse labeling cells with palmitate. Recycling of lipids released by Programmed Cell Death appears not to be necessary for lipid body production as a fadL mutant was defective in fatty acid uptake but proficient in lipid body production. The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation. MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes. PMID:24906161

  14. Bioavailability of endocrine disrupting chemicals (EDCs): Liposome-water partitioning and lipid membrane permeation

    NASA Astrophysics Data System (ADS)

    Kwon, Jung-Hwan

    The bioavailability of endocrine disrupting chemicals (EDCs) is a function of a number of parameters including the ability of the chemical to partition into organic tissue and reach receptor sites within an organism. In this dissertation, equilibrium partition coefficients between water and lipid membrane vesicles and artificial lipid membrane permeability were investigated for evaluating bioavailability of aqueous pollutants. Structurally diverse endocrine disrupting chemicals were chosen as model compounds for partitioning experiments and simple hydrophobic organic chemicals were used for the evaluation of a parallel artificial membrane device developed to mimic bioconcentration rates in fish. Hydrophobic interactions represented by octanol/water partition coefficients (KOWs) were not appropriate for estimating lipid membrane/water partition coefficients (Klipws) for the selected EDCs having a relatively large molar liquid volume (MLV) and containing polar functional groups. Correlations that include MLV and polar surface area (PSA) reduce the predicted value of log K lipw, suggesting that lipid membranes are less favorable than 1-octanol for a hydrophobic solute because of the changes in membrane fluidity and the amount of cholesterol in the lipid bilayers. These results suggested that KOW alone has limited potential for estimating K lipw, and MLV or PSA may be used as additional descriptors for developing quantitative structure-activity relationships (QSARs). The poor correlations between KOW and Klipw observed in this research may be due to the highly organized structure of lipid bilayers. Measured thermodynamic constants demonstrated that the entropy contribution becomes more dominant for more organized liposomes having saturated lipid tails. This implies that entropy-driven partitioning process makes Klipw different from KOW especially for more saturated lipid bilayer membranes. In the parallel artificial membrane system developed, a membrane filter

  15. Thermodynamics and mechanics of membrane curvature generation and sensing by proteins and lipids.

    PubMed

    Baumgart, Tobias; Capraro, Benjamin R; Zhu, Chen; Das, Sovan L

    2011-01-01

    Research investigating lipid membrane curvature generation and sensing is a rapidly developing frontier in membrane physical chemistry and biophysics. The fast recent progress is based on the discovery of a plethora of proteins involved in coupling membrane shape to cellular membrane function, the design of new quantitative experimental techniques to study aspects of membrane curvature, and the development of analytical theories and simulation techniques that allow a mechanistic interpretation of quantitative measurements. The present review first provides an overview of important classes of membrane proteins for which function is coupled to membrane curvature. We then survey several mechanisms that are assumed to underlie membrane curvature sensing and generation. Finally, we discuss relatively simple thermodynamic/mechanical models that allow quantitative interpretation of experimental observations.

  16. Hybrid lipids increase nanoscale fluctuation lifetimes in mixed membranes

    NASA Astrophysics Data System (ADS)

    Palmieri, Benoit; Safran, Samuel A.

    2013-09-01

    A recently proposed ternary mixture model is used to predict fluctuation domain lifetimes in the one phase region. The membrane is made of saturated, unsaturated, and hybrid lipids that have one saturated and one unsaturated hydrocarbon chain. The hybrid lipid is a natural linactant which can reduce the packing incompatibility between saturated and unsaturated lipids. The fluctuation lifetimes are predicted as a function of the hybrid lipid fraction and the fluctuation domain size. These lifetimes can be increased by up to three orders of magnitude compared to the case of no hybrids. With hybrid, small length scale fluctuations have sizable amplitudes even close to the critical temperature and, hence, benefit from enhanced critical slowing down. The increase in lifetime is particularly important for nanometer scale fluctuation domains where the hybrid orientation and the other lipids composition are highly coupled.

  17. Hybrid lipids increase nanoscale fluctuation lifetimes in mixed membranes.

    PubMed

    Palmieri, Benoit; Safran, Samuel A

    2013-09-01

    A recently proposed ternary mixture model is used to predict fluctuation domain lifetimes in the one phase region. The membrane is made of saturated, unsaturated, and hybrid lipids that have one saturated and one unsaturated hydrocarbon chain. The hybrid lipid is a natural linactant which can reduce the packing incompatibility between saturated and unsaturated lipids. The fluctuation lifetimes are predicted as a function of the hybrid lipid fraction and the fluctuation domain size. These lifetimes can be increased by up to three orders of magnitude compared to the case of no hybrids. With hybrid, small length scale fluctuations have sizable amplitudes even close to the critical temperature and, hence, benefit from enhanced critical slowing down. The increase in lifetime is particularly important for nanometer scale fluctuation domains where the hybrid orientation and the other lipids composition are highly coupled.

  18. Intermonolayer Friction and Surface Shear Viscosity of Lipid Bilayer Membranes

    PubMed Central

    den Otter, W. K.; Shkulipa, S. A.

    2007-01-01

    The flow behavior of lipid bilayer membranes is characterized by a surface viscosity for in-plane shear deformations, and an intermonolayer friction coefficient for slip between the two leaflets of the bilayer. Both properties have been studied for a variety of coarse-grained double-tailed model lipids, using equilibrium and nonequilibrium molecular dynamics simulations. For lipids with two identical tails, the surface shear viscosity rises rapidly with tail length, while the intermonolayer friction coefficient is less sensitive to the tail length. Interdigitation of lipid tails across the bilayer midsurface, as observed for lipids with two distinct tails, strongly enhances the intermonolayer friction coefficient, but hardly affects the surface shear viscosity. The simulation results are compared against the available experimental data. PMID:17468168

  19. Effect of Gold Nanoparticle on Structure and Fluidity of Lipid Membrane

    PubMed Central

    Mhashal, Anil R.; Roy, Sudip

    2014-01-01

    This paper deals with the effect of different size gold nanoparticles on the fluidity of lipid membrane at different regions of the bilayer. To investigate this, we have considered significantly large bilayer leaflets and incorporated only one nanoparticle each time, which was subjected to all atomistic molecular dynamics simulations. We have observed that, lipid molecules located near to the gold nanoparticle interact directly with it, which results in deformation of lipid structure and slower dynamics of lipid molecules. However, lipid molecules far away from the interaction site of the nanoparticle get perturbed, which gives rise to increase in local ordering of the lipid domains and decrease in fluidity. The bilayer thickness and area per head group in this region also get altered. Similar trend, but with different magnitude is also observed when different size nanoparticle interact with the bilayer. PMID:25469786

  20. Stabilization of Lipid Membranes With Dendritic Polymers

    DTIC Science & Technology

    2004-12-01

    monolayer coverage was possible using the spin - coating procedure for concentrations of 10-5 w/w dendrimers in solution or lower. Higher coverages...The fluorescence intensity increased as coverage time before spin coating increased and as the dendrimer solution concentration increased (Fig...enhanced the adsorption of lipids to the substrate. Figure 7: Fluorescence intensity as a function of the PAMAM concentration used in the spin

  1. Impact of monoolein on aquaporin1-based supported lipid bilayer membranes

    PubMed Central

    Wang, Zhining; Wang, Xida; Ding, Wande; Wang, Miaoqi; Qi, Xin; Gao, Congjie

    2015-01-01

    Aquaporin (AQP) based biomimetic membranes have attracted considerable attention for their potential water purification applications. In this paper, AQP1 incorporated biomimetic membranes were prepared and characterized. The morphology and structure of the biomimetic membranes were characterized by in situ atomic force microscopy (AFM), infrared absorption spectroscopy, fluorescence microscopy, and contact angle measurements. The nanofiltration performance of the AQP1 incorporated membranes was investigated at 4 bar by using 2 g l−1 NaCl as feed solution. Lipid mobility plays an important role in the performance of the AQP1 incorporated supported lipid bilayer (SLB) membranes. We demonstrated that the lipid mobility is successfully tuned by the addition of monoolein (MO). Through in situ AFM and fluorescence recovery after photo-bleaching (FRAP) measurements, the membrane morphology and the molecular mobility were studied. The lipid mobility increased in the sequence DPPC < DPPC/MO (RMO = 5/5) < DOPC/MO (RMO = 5/5) < DOPC, which is consistent with the flux increment and salt rejection. This study may provide some useful insights for improving the water purification performance of biomimetic membranes. PMID:27877825

  2. Impact of monoolein on aquaporin1-based supported lipid bilayer membranes

    NASA Astrophysics Data System (ADS)

    Wang, Zhining; Wang, Xida; Ding, Wande; Wang, Miaoqi; Qi, Xin; Gao, Congjie

    2015-08-01

    Aquaporin (AQP) based biomimetic membranes have attracted considerable attention for their potential water purification applications. In this paper, AQP1 incorporated biomimetic membranes were prepared and characterized. The morphology and structure of the biomimetic membranes were characterized by in situ atomic force microscopy (AFM), infrared absorption spectroscopy, fluorescence microscopy, and contact angle measurements. The nanofiltration performance of the AQP1 incorporated membranes was investigated at 4 bar by using 2 g l-1 NaCl as feed solution. Lipid mobility plays an important role in the performance of the AQP1 incorporated supported lipid bilayer (SLB) membranes. We demonstrated that the lipid mobility is successfully tuned by the addition of monoolein (MO). Through in situ AFM and fluorescence recovery after photo-bleaching (FRAP) measurements, the membrane morphology and the molecular mobility were studied. The lipid mobility increased in the sequence DPPC < DPPC/MO (RMO = 5/5) < DOPC/MO (RMO = 5/5) < DOPC, which is consistent with the flux increment and salt rejection. This study may provide some useful insights for improving the water purification performance of biomimetic membranes.

  3. Interaction of pristine and functionalized carbon nanotubes with lipid membranes.

    PubMed

    Baoukina, Svetlana; Monticelli, Luca; Tieleman, D Peter

    2013-10-10

    Carbon nanotubes are widely used in a growing number of applications. Their interactions with biological materials, cell membranes in particular, is of interest in applications including drug delivery and for understanding the toxicity of carbon nanotubes. We use extensive molecular dynamics simulations with the MARTINI model to study the interactions of model nanotubes of different thickness, length, and patterns of chemical modification with model membranes. In addition, we characterize the interactions of small bundles of carbon nanotubes with membrane models. Short pristine carbon nanotubes readily insert into membranes and adopt an orientation parallel to the plane of the membrane in the center of the membrane. Larger aggregates and functionalized nanotubes exhibit a range of possible interactions. The distribution and orientation of carbon nanotubes can be controlled by functionalizing the nanotubes. Free energy calculations provide thermodynamic insight into the preferred orientations of different nanotubes and quantify structural defects in the lipid matrix.

  4. Reversible control of current across lipid membranes by local heating

    PubMed Central

    Urban, Patrick; Kirchner, Silke R.; Mühlbauer, Christian; Lohmüller, Theobald; Feldmann, Jochen

    2016-01-01

    Lipid membranes are almost impermeable for charged molecules and ions that can pass the membrane barrier only with the help of specialized transport proteins. Here, we report how temperature manipulation at the nanoscale can be employed to reversibly control the electrical resistance and the amount of current that flows through a bilayer membrane with pA resolution. For this experiment, heating is achieved by irradiating gold nanoparticles that are attached to the bilayer membrane with laser light at their plasmon resonance frequency. We found that controlling the temperature on the nanoscale renders it possible to reproducibly regulate the current across a phospholipid membrane and the membrane of living cells in absence of any ion channels. PMID:26940847

  5. Proteomic Profiling of Detergent Resistant Membranes (Lipid Rafts) of Prostasomes.

    PubMed

    Dubois, Louise; Ronquist, Karl K Göran; Ek, Bo; Ronquist, Gunnar; Larsson, Anders

    2015-11-01

    Prostasomes are exosomes derived from prostate epithelial cells through exocytosis by multivesicular bodies. Prostasomes have a bilayered membrane and readily interact with sperm. The membrane lipid composition is unusual with a high contribution of sphingomyelin at the expense of phosphatidylcholine and saturated and monounsaturated fatty acids are dominant. Lipid rafts are liquid-ordered domains that are more tightly packed than the surrounding nonraft phase of the bilayer. Lipid rafts are proposed to be highly dynamic, submicroscopic assemblies that float freely within the liquid disordered membrane bilayer and some proteins preferentially partition into the ordered raft domains. We asked the question whether lipid rafts do exist in prostasomes and, if so, which proteins might be associated with them. Prostasomes of density range 1.13-1.19g/ml were subjected to density gradient ultracentrifugation in sucrose fabricated by phosphate buffered saline (PBS) containing 1% Triton X-100 with capacity for banding at 1.10 g/ml, i.e. the classical density of lipid rafts. Prepared prostasomal lipid rafts (by gradient ultracentrifugation) were analyzed by mass spectrometry. The clearly visible band on top of 1.10g/ml sucrose in the Triton X-100 containing gradient was subjected to liquid chromatography-tandem MS and more than 370 lipid raft associated proteins were identified. Several of them were involved in intraluminal vesicle formation, e.g. tetraspanins, ESCRTs, and Ras-related proteins. This is the first comprehensive liquid chromatography-tandem MS profiling of proteins in lipid rafts derived from exosomes. Data are available via ProteomeXchange with identifier PXD002163.

  6. Pore spanning lipid bilayers on silanised nanoporous alumina membranes

    NASA Astrophysics Data System (ADS)

    Md Jani, Abdul M.; Zhou, Jinwen; Nussio, Matthew R.; Losic, Dusan; Shapter, Joe G.; Voelcker, Nicolas H.

    2008-12-01

    The preparation of bilayer lipid membranes (BLMs) on solid surfaces is important for many studies probing various important biological phenomena including the cell barrier properties, ion-channels, biosensing, drug discovery and protein/ligand interactions. In this work we present new membrane platforms based on suspended BLMs on nanoporous anodic aluminium oxide (AAO) membranes. AAO membranes were prepared by electrochemical anodisation of aluminium foil in 0.3 M oxalic acid using a custom-built etching cell and applying voltage of 40 V, at 1oC. AAO membranes with controlled diameter of pores from 30 - 40 nm (top of membrane) and 60 -70 nm (bottom of membrane) were fabricated. Pore dimensions have been confirmed by scanning electron microscopy (SEM) and atomic force microscopy (AFM). AAO membranes were chemically functionalised with 3-aminopropyltriethoxysilane (APTES). Confirmation of the APTES attachment to the AAO membrane was achieved by means of infrared spectroscopy, X-ray photoelectron spectroscopy and contact angle measurements. The Fourier transform infrared (FTIR) spectra of functionalised membranes show several peaks from 2800 to 3000 cm-1 which were assigned to symmetric and antisymmetric CH2 bands. XPS data of the membrane showed a distinct increase in C1s (285 eV), N1s (402 eV) and Si2p (102 eV) peaks after silanisation. The water contact angle of the functionalised membrane was 80o as compared to 20o for the untreated membrane. The formation of BLMs comprising dioleoyl-phosphatidylserine (DOPS) on APTESmodified AAO membranes was carried using the vesicle spreading technique. AFM imaging and force spectroscopy was used to characterise the structural and nanomechanical properties of the suspended membrane. This technique also confirmed the stability of bilayers on the nanoporous alumina support for several days. Fabricated suspended BLMs on nanoporous AAO hold promise for the construction of biomimetic membrane architectures with embedded

  7. Stabilization of composition fluctuations in mixed membranes by hybrid lipids

    NASA Astrophysics Data System (ADS)

    Safran, Samuel; Palmieri, Benoit

    2013-03-01

    A ternary mixture model is proposed to describe composition fluctuations in mixed membranes composed of saturated, unsaturated and hybrid lipids. The asymmetric hybrid lipid has one saturated and one unsaturated hydrocarbon chain and it can reduce the packing incompatibility between saturated and unsaturated lipids. A methodology to recast the free-energy of the lattice in terms of a continuous isotropic field theory is proposed and used to analyze composition fluctuations above the critical temperature. The effect of hybrid lipids on fluctuations domains rich in saturated/unsaturated lipids is predicted. The correlation length of such fluctuations decreases significantly with increasing amounts of hybrids even if the temperature is maintained close to the critical temperature. This provides an upper bound for the domain sizes expected in rafts stabilized by hybrids, above the critical temperature. When the hybrid composition of the membrane is increased further, a crossover value is found above which ``stripe-like'' fluctuations are observed. The wavelength of these fluctuations decreases with increasing hybrid fraction and tends toward a molecular size in a membrane that contains only hybrids.

  8. Reduced Lateral Mobility of Lipids and Proteins in Crowded Membranes

    PubMed Central

    Goose, Joseph E.; Sansom, Mark S. P.

    2013-01-01

    Coarse-grained molecular dynamics simulations of the E. coli outer membrane proteins FhuA, LamB, NanC, OmpA and OmpF in a POPE/POPG (3∶1) bilayer were performed to characterise the diffusive nature of each component of the membrane. At small observation times (<10 ns) particle vibrations dominate phospholipid diffusion elevating the calculated values from the longer time-scale bulk value (>50 ns) of 8.5×10−7 cm2 s−1. The phospholipid diffusion around each protein was found to vary based on distance from protein. An asymmetry in the diffusion of annular lipids in the inner and outer leaflets was observed and correlated with an asymmetry in charged residues in the vicinity of the inner and outer leaflet head-groups. Protein rotational and translational diffusion were also found to vary with observation time and were inversely correlated with the radius of gyration of the protein in the plane of the bilayer. As the concentration of protein within the bilayer was increased, the overall mobility of the membrane decreased reflected in reduced lipid diffusion coefficients for both lipid and protein components. The increase in protein concentration also resulted in a decrease in the anomalous diffusion exponent α of the lipid. Formation of extended clusters and networks of proteins led to compartmentalisation of lipids in extreme cases. PMID:23592975

  9. Membrane lipid rafts disturbance in the response of rat skeletal muscle to short-term disuse.

    PubMed

    Petrov, Alexey M; Kravtsova, Violetta V; Matchkov, Vladimir V; Vasiliev, Alexander N; Zefirov, Andrey L; Chibalin, Alexander V; Heiny, Judith A; Krivoi, Igor I

    2017-03-08

    Marked loss of skeletal muscle mass occurs under various conditions of disuse, but the molecular and cellular mechanisms leading to atrophy are not completely understood. We investigate early molecular events which might play a role in skeletal muscle remodeling during mechanical unloading (disuse). The effects of acute (6 - 12 h) hindlimb suspension on the soleus muscles from adult rats were examined. The integrity of plasma membrane lipid rafts was tested utilizing cholera toxin B subunit, or fluorescent sterols. In addition, resting intracellular Ca(2+) level was analyzed. Acute disuse disturbed the plasma membrane lipid-ordered phase throughout the sarcolemma and was more pronounced in junctional membrane regions. Ouabain (1 µM), which specifically inhibits the Na,K-ATPase α2 isozyme in rodent skeletal muscles, produced similar lipid rafts changes in control muscles, but was ineffective in suspended muscles, which show an initial loss of α2 Na,K-ATPase activity. Lipid rafts were able to recover with cholesterol supplementation, suggesting that disturbance results from cholesterol loss. Repetitive nerve stimulation also restores lipid rafts, specifically in junctional sarcolemma region. Disuse locally lowered the resting intracellular Ca(2+) concentration only near the neuromuscular junction of muscle fibers. Our results provide the evidence to suggest that the ordering of lipid rafts strongly depends on motor nerve input and may involve interactions with the α2 Na,K-ATPase. Lipid rafts disturbance, accompanied by intracellular Ca(2+) dysregulation are among the earliest remodeling events induced by skeletal muscle disuse.

  10. Anesthetic diffusion through lipid membranes depends on the protonation rate.

    PubMed

    Pérez-Isidoro, Rosendo; Sierra-Valdez, F J; Ruiz-Suárez, J C

    2014-12-18

    Hundreds of substances possess anesthetic action. However, despite decades of research and tests, a golden rule is required to reconcile the diverse hypothesis behind anesthesia. What makes an anesthetic to be local or general in the first place? The specific targets on proteins, the solubility in lipids, the diffusivity, potency, action time? Here we show that there could be a new player equally or even more important to disentangle the riddle: the protonation rate. Indeed, such rate modulates the diffusion speed of anesthetics into lipid membranes; low protonation rates enhance the diffusion for local anesthetics while high ones reduce it. We show also that there is a pH and membrane phase dependence on the local anesthetic diffusion across multiple lipid bilayers. Based on our findings we incorporate a new clue that may advance our understanding of the anesthetic phenomenon.

  11. Cholesterol drives aβ(1-42) interaction with lipid rafts in model membranes.

    PubMed

    Seghezza, Silvia; Diaspro, Alberto; Canale, Claudio; Dante, Silvia

    2014-11-25

    The molecular mechanism at the basis of the neurodegenerative process related to Alzheimer's disease (AD) is triggered by the local composition of the neural plasma membrane. The role of cholesterol is controversial. In this investigation the interaction of the AD peptide amyloid-beta (1-42) with model membranes containing lipid rafts has been investigated by atomic force microscopy techniques. Supported lipid membranes made of phospholipids/sphingomyelin/cholesterol have been investigated as a function of the molar content of cholesterol, in a range spanning the phase diagram of the lipid system. The administration of amyloid-beta induced a phase reorganization of the lipid domains, when the cholesterol molar fraction was below 5%. At the same time, a mechanical destabilization and an appreciable thinning of the membrane induced by the peptide were detected. The major interaction was observed in the presence of the gel phase Lβ, and was enhanced by a low cholesterol amount. With the appearance of the liquid ordered phase Lo, the effect was hindered. At high cholesterol content (20% mol), no detectable effects in the bilayer morphology or in its mechanical stability were recorded. These findings give new insights on the molecular mechanism of the amyloid/membrane interaction, highlighting the peculiar role of cholesterol.

  12. Lipid-protein interactions in plasma membranes of fiber cells isolated from the human eye lens.

    PubMed

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2014-03-01

    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.

  13. Lipid-Protein Interactions in Plasma Membranes of Fiber Cells Isolated from the Human Eye Lens

    PubMed Central

    Raguz, Marija; Mainali, Laxman; O’Brien, William J.; Subczynski, Witold K.

    2014-01-01

    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. PMID:24486794

  14. Micrometer-Scale Membrane Transition of Supported Lipid Bilayer Membrane Reconstituted with Cytosol of Dictyostelium discoideum

    PubMed Central

    Takahashi, Kei; Toyota, Taro

    2017-01-01

    Background: The transformation of the supported lipid bilayer (SLB) membrane by extracted cytosol from living resources, has recently drawn much attention. It enables us to address the question of whether the purified phospholipid SLB membrane, including lipids related to amoeba locomotion, which was discussed in many previous studies, exhibits membrane deformation in the presence of cytosol extracted from amoeba; Methods: In this report, a method for reconstituting a supported lipid bilayer (SLB) membrane, composed of purified phospholipids and cytosol extracted from Dictyostelium discoideum, is described. This technique is a new reconstitution method combining the artificial constitution of membranes with the reconstitution using animate cytosol (without precise purification at a molecular level), contributing to membrane deformation analysis; Results: The morphology transition of a SLB membrane composed of phosphatidylcholines, after the addition of cytosolic extract, was traced using a confocal laser scanning fluorescence microscope. As a result, pore formation in the SLB membrane was observed and phosphatidylinositides incorporated into the SLB membrane tended to suppress pore formation and expansion; Conclusions: The current findings imply that phosphatidylinositides have the potential to control cytoplasm activity and bind to a phosphoinositide-containing SLB membrane. PMID:28272354

  15. Lipid domains control myelin basic protein adsorption and membrane interactions between model myelin lipid bilayers.

    PubMed

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

    2014-02-25

    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.

  16. Determining the pivotal plane of fluid lipid membranes in simulations

    NASA Astrophysics Data System (ADS)

    Wang, Xin; Deserno, Markus

    2015-10-01

    Each leaflet of a curved lipid membrane contains a surface at which the area strain vanishes, the so-called pivotal plane. Its distance z0 from the bilayer's midplane arises in numerous contexts, for instance the connection between monolayer and bilayer moduli, stress-profile moments, or area-difference elasticity theories. Here, we propose two precise methods for determining the location of the pivotal plane in computer simulations, both of which rely on monitoring the lipid imbalance across a curved bilayer. The first method considers the ratio of lipid number between the two leaflets of cylindrical or spherical vesicles; it hence requires lipid flip-flop for equilibration. The second method looks at the leaflet difference across local sections cut out from a buckled membrane; this observable equilibrates even in the absence of flip-flop. We apply our methods to two different coarse-grained lipid models, the generic three-bead solvent-free Cooke model and a ten-bead representation of dimyristoylphosphocholine with the explicit solvent MARTINI model. The Cooke model is amenable to both methods and gives results that agree at the percent level. Using it, we also show that the pivotal plane moves outward as lipid curvature becomes more positive. The MARTINI model can only be analyzed with the buckling method; the obtained value z0 = 0.850(11) nm lies about 0.4 nm inwards of the glycerol backbone and is hence unexpectedly small. We attribute this to limitations of the coarse-grained description, suggesting that the location of the pivotal plane might be a good indicator for how well lipid models capture the microscopic origins of curvature elasticity. Finally, we also show that the pivotal plane position itself moves as the membrane is bent. The leading correction is linear in curvature, dependent on the Poisson ratio, and can matter when analyzing experimental results obtained from highly curved inverse hexagonal phases.

  17. Cyclohexane Rings Reduce Membrane Permeability to Small Ions in Archaea-Inspired Tetraether Lipids.

    PubMed

    Koyanagi, Takaoki; Leriche, Geoffray; Onofrei, David; Holland, Gregory P; Mayer, Michael; Yang, Jerry

    2016-01-26

    Extremophile archaeal organisms overcome problems of membrane permeability by producing lipids with structural elements that putatively improve membrane integrity compared to lipids from other life forms. Herein, we describe a series of lipids that mimic some key structural features of archaeal lipids, such as: 1) single tethering of lipid tails to create fully transmembrane tetraether lipids and 2) the incorporation of small rings into these tethered segments. We found that membranes formed from pure tetraether lipids leaked small ions at a rate that was about two orders of magnitude slower than common bilayer-forming lipids. Incorporation of cyclopentane rings into the tetraether lipids did not affect membrane leakage, whereas a cyclohexane ring reduced leakage by an additional 40 %. These results show that mimicking certain structural features of natural archaeal lipids results in improved membrane integrity, which may help overcome limitations of many current lipid-based technologies.

  18. DNA-Tile Structures Induce Ionic Currents through Lipid Membranes.

    PubMed

    Göpfrich, Kerstin; Zettl, Thomas; Meijering, Anna E C; Hernández-Ainsa, Silvia; Kocabey, Samet; Liedl, Tim; Keyser, Ulrich F

    2015-05-13

    Self-assembled DNA nanostructures have been used to create man-made transmembrane channels in lipid bilayers. Here, we present a DNA-tile structure with a nominal subnanometer channel and cholesterol-tags for membrane anchoring. With an outer diameter of 5 nm and a molecular weight of 45 kDa, the dimensions of our synthetic nanostructure are comparable to biological ion channels. Because of its simple design, the structure self-assembles within a minute, making its creation scalable for applications in biology. Ionic current recordings demonstrate that the tile structures enable ion conduction through lipid bilayers and show gating and voltage-switching behavior. By demonstrating the design of DNA-based membrane channels with openings much smaller than that of the archetypical six-helix bundle, our work showcases their versatility inspired by the rich diversity of natural membrane components.

  19. Dynamical Clustering and the Origin of Raft-like Structures in a Model Lipid Membrane

    NASA Astrophysics Data System (ADS)

    Starr, Francis

    2014-03-01

    We investigate the dynamical heterogeneity of a model single-component lipid membrane using simulations of a coarse-grained representation of lipid molecules. In the liquid-ordered (LO) phase, lipid diffusion is hindered by the transient trapping of molecules by their neighbors, giving rise to two distinct mobility groups: low-mobility lipids which are temporarily ``caged'', and lipids with displacements on the scale of the intermolecular spacing. The lipid molecules within these distinct mobility states cluster, giving rise to transient ``islands'' of enhanced mobility having the size and time scale expected for lipid ``rafts''. These clusters are strikingly similar to the dynamical clusters found in glass-forming fluids, and distinct from phase-separation clusters. Such dynamic heterogeneity is ubiquitous in disordered condensed-phase systems. Thus, we hypothesize that rafts may originate from this universal mechanism, explaining why raft-like regions should arise, regardless of lipid structural or compositional details. This perspective provides a new approach to understand membrane transport.

  20. Polyunsaturation in lipid membranes: dynamic properties and lateral pressure profiles.

    PubMed

    Ollila, Samuli; Hyvönen, Marja T; Vattulainen, Ilpo

    2007-03-29

    We elucidate the influence of unsaturation on single-component membrane properties, focusing on their dynamical aspects and lateral pressure profiles across the membrane. To this end, we employ atomistic molecular dynamics simulations to study five different membrane systems with varying degrees of unsaturation, starting from saturated membranes and systematically increasing the level of unsaturation, ending up with a bilayer of phospholipids containing the docosahexaenoic acid. For an increasing level of unsaturation, we find considerable effects on dynamical properties, such as accelerated dynamics of the phosphocholine head groups and glycerol backbones and speeded up rotational dynamics of the lipid molecules. The lateral pressure profile is found to be altered by the degree of unsaturation. For an increasing number of double bonds, the peak in the middle of the bilayer decreases. This is compensated for by changes in the membrane-water interface region in terms of increasing peak heights of the lateral pressure profile. Implications of the findings are briefly discussed.

  1. Artificial black membranes from bipolar lipids of thermophilic Archaebacteria.

    PubMed Central

    Gliozzi, A; Rolandi, R; De Rosa, M; Gambacorta, A

    1982-01-01

    The membrane of thermophilic archaebacteria is characterized by the presence of unusual isoprenoid bipolar lipids. The molecular organization of these lipids is still a matter of study. Important information could come from forming artificial black membranes. Black films can be formed from n-alkane or squalene dispersions of bipolar lipids extracted from the membrane of Caldariella acidophila. Membrane formation occurred only above a critical temperature (approximately 70 degrees C) corresponding to the physiological one. At lower temperatures, special solvent systems (n-alkanes or squalene, butanol and n-alkanes or squalene, butanol chloroform) were required. To characterize the physical parameters of these membranes, conductance and capacitance measurements were performed. Conductance was in the range of 10(-8) - 10(-7) omega -1 cm -2 , where specific capacitance at T = 72 degrees C was Cs = 0.685 +/- 0.004 microF/cm2 and Cs = 0.658 +/- 0.08 microF/cm2, corresponding to a dielectric thickness of 27 and 29 A for squalene and dodecane dispersions, respectively. Capacitance was shown to vary as the square of membrane potential, as usual in lipid bilayers. Values of the proportionality constant alpha have been compared to those of solvent-containing and solvent-free bilayers. The behavior of capacitance as a function of temperature is also shown by lowering temperature; the occurrence of complex structural changes was indicated. All the experimental data suggest that the presence of solvent is very low. Two possible molecular configurations of the films are discussed. PMID:6800415

  2. Distribution of Fullerene Nanoparticles between Water and Solid Supported Lipid Membranes: Thermodynamics and Effects of Membrane Composition on Distribution.

    PubMed

    Ha, Yeonjeong; Katz, Lynn E; Liljestrand, Howard M

    2015-12-15

    The distribution coefficient (Klipw) of fullerene between solid supported lipid membranes (SSLMs) and water was examined using different lipid membrane compositions. Klipw of fullerene was significantly higher with a cationic lipid membrane compared to that with a zwitterionic or anionic lipid membrane, potentially due to the strong interactions between negative fullerene dispersions and positive lipid head groups. The higher Klipw for fullerene distribution to ternary lipid mixture membranes was attributed to an increase in the interfacial surface area of the lipid membrane resulting from phase separation. These results imply that lipid composition can be a critical factor that affects bioconcentration of fullerene. Distribution of fullerene into zwitterionic unsaturated lipid membranes was dominated by the entropy contribution (ΔS) and the process was endothermic (ΔH > 0). This result contrasts the partitioning thermodynamics of highly and moderately hydrophobic chemicals indicating that the lipid-water distribution mechanism of fullerene may be different from that of molecular level chemicals. Potential mechanisms for the distribution of fullerene that may explain these differences include adsorption on the lipid membrane surfaces and partitioning into the center of lipid membranes (i.e., absorption).

  3. Incorporation of large guest molecules into liposomes via chemical reactions in lipid membranes.

    PubMed

    Tsuchiya, Yuki; Sugikawa, Kouta; Ueda, Masafumi; Ikeda, Atsushi

    2017-02-22

    The incorporation of hydrophobic guest molecules into lipid membranes by the exchange of the guest molecule from a cyclodextrin (CDx) complex to a liposome is limited to guest molecules that can be included in CDxs. To solve this problem, large guest molecules were incorporated into liposomes by chemical reactions of guest molecules in lipid membranes. Stable lipid-membrane-incorporated fullerene derivatives with large substituent(s) were prepared by Diels-Alder reactions in lipid membranes.

  4. Lipid membrane domain formation and alamethicin aggregation studied by calorimetry, sound velocity measurements, and atomic force microscopy.

    PubMed

    Oliynyk, Vitaliy; Jäger, Markus; Heimburg, Thomas; Buckin, Vitaly; Kaatze, Udo

    2008-05-01

    An experimental study of phosphocholine membranes made from one lipid, from mixtures of DPPC and DLPC, and also from lipids and small amounts of alamethicin is presented. We used atomic force microscopy to investigate the spatial organization and structure of lipid domains and also of the defects induced by the peptide. Alamethicin was found to alter the state of lipids in the gel state in a way that domains of fluid lipids are formed close to the defects. Differential calorimetry revealed phase characteristics of the lipid mixtures and the effect of small amounts of alamethicin on the phase behavior. It was also shown that the sound velocity profiles of the membranes suspensions can be well calculated from the heat capacity traces of the samples. This result confirms the correlation between the mechanical properties and the specific heat of membrane systems.

  5. Mechanical properties that influence antimicrobial peptide activity in lipid membranes.

    PubMed

    Marín-Medina, Nathaly; Ramírez, Diego Alejandro; Trier, Steve; Leidy, Chad

    2016-12-01

    Antimicrobial peptides are small amphiphilic proteins found in animals and plants as essential components of the innate immune system and whose function is to control bacterial infectious activity. In order to accomplish their function, antimicrobial peptides use different mechanisms of action which have been deeply studied in view of their potential exploitation to treat antibiotic-resistant bacterial infections. One of the main mechanisms of action of these peptides is the disruption of the bacterial membrane through pore formation, which, in some cases, takes place via a monomer to oligomer cooperative transition. Previous studies have shown that lipid composition, and the presence of exogenous components, such as cholesterol in model membranes or carotenoids in bacteria, can affect the potency of distinct antimicrobial peptides. At the same time, considering the membrane as a two-dimensional material, it has been shown that membrane composition defines its mechanical properties which might be relevant in many membrane-related processes. Nevertheless, the correlation between the mechanical properties of the membrane and antimicrobial peptide potency has not been considered according to the importance it deserves. The relevance of these mechanical properties in membrane deformation due to peptide insertion is reviewed here for different types of pores in order to elucidate if indeed membrane composition affects antimicrobial peptide activity by modulation of the mechanical properties of the membrane. This would also provide a better understanding of the mechanisms used by bacteria to overcome antimicrobial peptide activity.

  6. Lipid domains in intact fiber-cell plasma membranes isolated from cortical and nuclear regions of human eye lenses of donors from different age groups.

    PubMed

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2015-03-01

    The results reported here clearly document changes in the properties and the organization of fiber-cell membrane lipids that occur with age, based on electron paramagnetic resonance (EPR) analysis of lens membranes of clear lenses from donors of age groups from 0 to 20, 21 to 40, and 61 to 80 years. The physical properties, including profiles of the alkyl chain order, fluidity, hydrophobicity, and oxygen transport parameter, were investigated using EPR spin-labeling methods, which also provide an opportunity to discriminate coexisting lipid domains and to evaluate the relative amounts of lipids in these domains. Fiber-cell membranes were found to contain three distinct lipid environments: bulk lipid domain, which appears minimally affected by membrane proteins, and two domains that appear due to the presence of membrane proteins, namely boundary and trapped lipid domains. In nuclear membranes the amount of boundary and trapped phospholipids as well as the amount of cholesterol in trapped lipid domains increased with the donors' age and was greater than that in cortical membranes. The difference between the amounts of lipids in domains uniquely formed due to the presence of membrane proteins in nuclear and cortical membranes increased with the donors' age. It was also shown that cholesterol was to a large degree excluded from trapped lipid domains in cortical membranes. It is evident that the rigidity of nuclear membranes was greater than that of cortical membranes for all age groups. The amount of lipids in domains of low oxygen permeability, mainly in trapped lipid domains, were greater in nuclear than cortical membranes and increased with the age of donors. These results indicate that the nuclear fiber cell plasma membranes were less permeable to oxygen than cortical membranes and become less permeable to oxygen with age. In clear lenses, age-related changes in the lens lipid and protein composition and organization appear to occur in ways that increase fiber

  7. Lipid Domains in Intact Fiber-Cell Plasma Membranes Isolated from Cortical and Nuclear Regions of Human Eye Lenses of Donors from Different Age Groups

    PubMed Central

    Raguz, Marija; Mainali, Laxman; O’Brien, William J.; Subczynski, Witold K.

    2015-01-01

    The results reported here clearly document changes in the properties and the organization of fiber-cell membrane lipids that occur with age, based on electron paramagnetic resonance (EPR) analysis of lens membranes of clear lenses from donors of age groups from 0 to 20, 21 to 40, and 61 to 80 years. The physical properties, including profiles of the alkyl chain order, fluidity, hydrophobicity, and oxygen transport parameter, were investigated using EPR spin-labeling methods, which also provide an opportunity to discriminate coexisting lipid domains and to evaluate the relative amounts of lipids in these domains. Fiber-cell membranes were found to contain three distinct lipid environments: bulk lipid domain, which appears minimally affected by membrane proteins, and two domains that appear due to the presence of membrane proteins, namely boundary and trapped lipid domains. In nuclear membranes the amount of boundary and trapped phospholipids as well as the amount of cholesterol in trapped lipid domains increased with the donors’ age and was greater than that in cortical membranes. The difference between the amounts of lipids in domains uniquely formed due to the presence of membrane proteins in nuclear and cortical membranes increased with the donors’ age. It was also shown that cholesterol was to a large degree excluded from trapped lipid domains in cortical membranes. It is evident that the rigidity of nuclear membranes was greater than that of cortical membranes for all age groups. The amount of lipids in domains of low oxygen permeability, mainly in trapped lipid domains, were greater in nuclear than cortical membranes and increased with the age of donors. These results indicate that the nuclear fiber cell plasma membranes were less permeable to oxygen than cortical membranes and become less permeable to oxygen with age. In clear lenses, age-related changes in the lens lipid and protein composition and organization appear to occur in ways that increase fiber

  8. Single Molecule Kinetics of ENTH Binding to Lipid Membranes

    SciTech Connect

    Rozovsky, Sharon; Forstner, Martin B.; Sondermann, Holger; Groves, Jay T.

    2012-04-03

    Transient recruitment of proteins to membranes is a fundamental mechanism by which the cell exerts spatial and temporal control over proteins’ localization and interactions. Thus, the specificity and the kinetics of peripheral proteins’ membrane residence are an attribute of their function. In this article, we describe the membrane interactions of the interfacial epsin N-terminal homology (ENTH) domain with its target lipid phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2). The direct visualization and quantification of interactions of single ENTH molecules with supported lipid bilayers is achieved using total internal reflection fluorescence microscopy (TIRFM) with a time resolution of 13 ms. This enables the recording of the kinetic behavior of ENTH interacting with membranes with physiologically relevant concentrations of PtdIns(4,5)P2 despite the low effective binding affinity. Subsequent single fluorophore tracking permits us to build up distributions of residence times and to measure ENTH dissociation rates as a function of membrane composition. In addition, due to the high time resolution, we are able to resolve details of the motion of ENTH associated with a simple, homogeneous membrane. In this case ENTH’s diffusive transport appears to be the result of at least three different diffusion processes.

  9. Molecular dynamics study of lipid bilayers modeling the plasma membranes of mouse hepatocytes and hepatomas

    NASA Astrophysics Data System (ADS)

    Andoh, Yoshimichi; Aoki, Noriyuki; Okazaki, Susumu

    2016-02-01

    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.

  10. Important roles for membrane lipids in haloarchaeal bioenergetics.

    PubMed

    Kellermann, Matthias Y; Yoshinaga, Marcos Y; Valentine, Raymond C; Wörmer, Lars; Valentine, David L

    2016-11-01

    Recent advances in lipidomic analysis in combination with various physiological experiments set the stage for deciphering the structure-function of haloarchaeal membrane lipids. Here we focused primarily on changes in lipid composition of Haloferax volcanii, but also performed a comparative analysis with four other haloarchaeal species (Halobacterium salinarum, Halorubrum lacusprofundi, Halorubrum sodomense and Haloplanus natans) all representing distinctive cell morphologies and behaviors (i.e., rod shape vs. pleomorphic behavior). Common to all five haloarchaea, our data reveal an extraordinary high level of menaquinone, reaching up to 72% of the total lipids. This ubiquity suggests that menaquinones may function beyond their ordinary role as electron and proton transporter, acting simultaneously as ion permeability barriers and as powerful shield against oxidative stress. In addition, we aimed at understanding the role of cations interacting with the characteristic negatively charged surface of haloarchaeal membranes. We propose for instance that by bridging the negative charges of adjacent anionic phospholipids, Mg(2+) acts as surrogate for cardiolipin, a molecule that is known to control curvature stress of membranes. This study further provides a bioenergetic perspective as to how haloarchaea evolved following oxygenation of Earth's atmosphere. The success of the aerobic lifestyle of haloarchaea includes multiple membrane-based strategies that successfully balance the need for a robust bilayer structure with the need for high rates of electron transport - collectively representing the molecular basis to inhabit hypersaline water bodies around the planet.

  11. The interaction of polyglutamine peptides with lipid membranes is regulated by flanking sequences associated with huntingtin.

    PubMed

    Burke, Kathleen A; Kauffman, Karlina J; Umbaugh, C Samuel; Frey, Shelli L; Legleiter, Justin

    2013-05-24

    Huntington disease (HD) is caused by an expanded polyglutamine (poly(Q)) repeat near the N terminus of the huntingtin (htt) protein. Expanded poly(Q) facilitates formation of htt aggregates, eventually leading to deposition of cytoplasmic and intranuclear inclusion bodies containing htt. Flanking sequences directly adjacent to the poly(Q) domain, such as the first 17 amino acids on the N terminus (Nt17) and the polyproline (poly(P)) domain on the C-terminal side of the poly(Q) domain, heavily influence aggregation. Additionally, htt interacts with a variety of membraneous structures within the cell, and Nt17 is implicated in lipid binding. To investigate the interaction between htt exon1 and lipid membranes, a combination of in situ atomic force microscopy, Langmuir trough techniques, and vesicle permeability assays were used to directly monitor the interaction of a variety of synthetic poly(Q) peptides with different combinations of flanking sequences (KK-Q35-KK, KK-Q35-P10-KK, Nt17-Q35-KK, and Nt17-Q35-P10-KK) on model membranes and surfaces. Each peptide aggregated on mica, predominately forming extended, fibrillar aggregates. In contrast, poly(Q) peptides that lacked the Nt17 domain did not appreciably aggregate on or insert into lipid membranes. Nt17 facilitated the interaction of peptides with lipid surfaces, whereas the poly(P) region enhanced this interaction. The aggregation of Nt17-Q35-P10-KK on the lipid bilayer closely resembled that of a htt exon1 construct containing 35 repeat glutamines. Collectively, this data suggests that the Nt17 domain plays a critical role in htt binding and aggregation on lipid membranes, and this lipid/htt interaction can be further modulated by the presence of the poly(P) domain.

  12. Buffers affect the bending rigidity of model lipid membranes.

    PubMed

    Bouvrais, Hélène; Duelund, Lars; Ipsen, John H

    2014-01-14

    In biophysical and biochemical studies of lipid bilayers the influence of the used buffer is often ignored or assumed to be negligible on membrane structure, elasticity, or physical properties. However, we here present experimental evidence, through bending rigidity measurements performed on giant vesicles, of a more complex behavior, where the buffering molecules may considerably affect the bending rigidity of phosphatidylcholine bilayers. Furthermore, a synergistic effect on the bending modulus is observed in the presence of both salt and buffer molecules, which serves as a warning to experimentalists in the data interpretation of their studies, since typical lipid bilayer studies contain buffer and ion molecules.

  13. Lipid membrane-assisted condensation and assembly of amphiphilic Janus particles

    DOE PAGES

    Chambers, Mariah; Mallory, Stewart Anthony; Malone, Heather; ...

    2016-01-01

    Amphiphilic Janus particles self-assemble into complex metastructures, but little is known about how their assembly might be modified by weak interactions with a nearby biological membrane surface. Here, we report an integrated experimental and molecular dynamics simulation study to investigate the self-assembly of amphiphilic Janus particles on a lipid membrane. We created an experimental system in which Janus particles are allowed to self-assemble in the same medium where zwitterionic lipids form giant unilamellar vesicles (GUVs). Janus particles spontaneously concentrated on the inner leaflet of the GUVs. They exhibited biased orientation and heterogeneous rotational dynamics as revealed by single particle rotationalmore » tracking. The combined experimental and simulation results show that Janus particles concentrate on the lipid membranes due to weak particle–lipid attraction, whereas the biased orientation of particles is driven predominantly by inter-particle interactions. Furthermore, this study demonstrates the potential of using lipid membranes to influence the self-assembly of Janus particles.« less

  14. Influence of membrane surface charge on adsorption of complement proteins onto supported lipid bilayers.

    PubMed

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

    2016-12-01

    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.

  15. Lipids and topological rules of membrane protein assembly: balance between long and short range lipid-protein interactions.

    PubMed

    Vitrac, Heidi; Bogdanov, Mikhail; Heacock, Phil; Dowhan, William

    2011-04-29

    The N-terminal six-transmembrane domain (TM) bundle of lactose permease of Escherichia coli is uniformly inverted when assembled in membranes lacking phosphatidylethanolamine (PE). Inversion is dependent on the net charge of cytoplasmically exposed protein domains containing positive and negative residues, net charge of the membrane surface, and low hydrophobicity of TM VII acting as a molecular hinge between the two halves of lactose permease (Bogdanov, M., Xie, J., Heacock, P., and Dowhan, W. (2008) J. Cell Biol. 182, 925-935). Net neutral lipids suppress the membrane translocation potential of negatively charged amino acids, thus increasing the cytoplasmic retention potential of positively charged amino acids. Herein, TM organization of sucrose permease (CscB) and phenylalanine permease (PheP) as a function of membrane lipid composition was investigated to extend these principles to other proteins. For CscB, topological dependence on PE only becomes evident after a significant increase in the net negative charge of the cytoplasmic surface of the N-terminal TM bundle. High negative charge is required to overcome the thermodynamic block to inversion due to the high hydrophobicity of TM VII. Increasing the positive charge of the cytoplasmic surface of the N-terminal TM hairpin of PheP, which is misoriented in PE-lacking cells, favors native orientation in the absence of PE. PheP and CscB also display co-existing dual topologies dependent on changes in the charge balance between protein domains and the membrane lipids. Therefore, the topology of both permeases is dependent on PE. However, CscB topology is governed by thermodynamic balance between opposing lipid-dependent electrostatic and hydrophobic interactions.

  16. Exploiting lipopolysaccharide-induced deformation of lipid bilayers to modify membrane composition and generate two-dimensional geometric membrane array patterns

    SciTech Connect

    Adams, Peter G.; Swingle, Kirstie L.; Paxton, Walter F.; Nogan, John J.; Stromberg, Loreen R.; Firestone, Millicent A.; Mukundan, Harshini; Montaño, Gabriel A.

    2015-05-27

    Supported lipid bilayers have proven effective as model membranes for investigating biophysical processes and in development of sensor and array technologies. The ability to modify lipid bilayers after their formation and in situ could greatly advance membrane technologies, but is difficult via current state-of-the-art technologies. Here we demonstrate a novel method that allows the controlled post-formation processing and modification of complex supported lipid bilayer arrangements, under aqueous conditions. We exploit the destabilization effect of lipopolysaccharide, an amphiphilic biomolecule, interacting with lipid bilayers to generate voids that can be backfilled to introduce desired membrane components. We further demonstrate that when used in combination with a single, traditional soft lithography process, it is possible to generate hierarchically-organized membrane domains and microscale 2-D array patterns of domains. Significantly, this technique can be used to repeatedly modify membranes allowing iterative control over membrane composition. This approach expands our toolkit for functional membrane design, with potential applications for enhanced materials templating, biosensing and investigating lipid-membrane processes.

  17. Exploiting lipopolysaccharide-induced deformation of lipid bilayers to modify membrane composition and generate two-dimensional geometric membrane array patterns

    PubMed Central

    Adams, Peter G.; Swingle, Kirstie L.; Paxton, Walter F.; Nogan, John J.; Stromberg, Loreen R.; Firestone, Millicent A.; Mukundan, Harshini; Montaño, Gabriel A.

    2015-01-01

    Supported lipid bilayers have proven effective as model membranes for investigating biophysical processes and in development of sensor and array technologies. The ability to modify lipid bilayers after their formation and in situ could greatly advance membrane technologies, but is difficult via current state-of-the-art technologies. Here we demonstrate a novel method that allows the controlled post-formation processing and modification of complex supported lipid bilayer arrangements, under aqueous conditions. We exploit the destabilization effect of lipopolysaccharide, an amphiphilic biomolecule, interacting with lipid bilayers to generate voids that can be backfilled to introduce desired membrane components. We further demonstrate that when used in combination with a single, traditional soft lithography process, it is possible to generate hierarchically-organized membrane domains and microscale 2-D array patterns of domains. Significantly, this technique can be used to repeatedly modify membranes allowing iterative control over membrane composition. This approach expands our toolkit for functional membrane design, with potential applications for enhanced materials templating, biosensing and investigating lipid-membrane processes. PMID:26015293

  18. Exploiting lipopolysaccharide-induced deformation of lipid bilayers to modify membrane composition and generate two-dimensional geometric membrane array patterns

    DOE PAGES

    Adams, Peter G.; Swingle, Kirstie L.; Paxton, Walter F.; ...

    2015-05-27

    Supported lipid bilayers have proven effective as model membranes for investigating biophysical processes and in development of sensor and array technologies. The ability to modify lipid bilayers after their formation and in situ could greatly advance membrane technologies, but is difficult via current state-of-the-art technologies. Here we demonstrate a novel method that allows the controlled post-formation processing and modification of complex supported lipid bilayer arrangements, under aqueous conditions. We exploit the destabilization effect of lipopolysaccharide, an amphiphilic biomolecule, interacting with lipid bilayers to generate voids that can be backfilled to introduce desired membrane components. We further demonstrate that when usedmore » in combination with a single, traditional soft lithography process, it is possible to generate hierarchically-organized membrane domains and microscale 2-D array patterns of domains. Significantly, this technique can be used to repeatedly modify membranes allowing iterative control over membrane composition. This approach expands our toolkit for functional membrane design, with potential applications for enhanced materials templating, biosensing and investigating lipid-membrane processes.« less

  19. Phytosphingosine, sphingosine and dihydrosphingosine ceramides in model skin lipid membranes: permeability and biophysics.

    PubMed

    Školová, Barbora; Kováčik, Andrej; Tesař, Ondřej; Opálka, Lukáš; Vávrová, Kateřina

    2017-05-01

    Ceramides based on phytosphingosine, sphingosine and dihydrosphingosine are essential constituents of the skin lipid barrier that protects the body from excessive water loss. The roles of the individual ceramide subclasses in regulating skin permeability and the reasons for C4-hydroxylation of these sphingolipids are not completely understood. We investigated the chain length-dependent effects of dihydroceramides, sphingosine ceramides (with C4-unsaturation) and phytoceramides (with C4-hydroxyl) on the permeability, lipid organization and thermotropic behavior of model stratum corneum lipid membranes composed of ceramide/lignoceric acid/cholesterol/cholesteryl sulfate. Phytoceramides with very long C24 acyl chains increased the permeability of the model lipid membranes compared to dihydroceramides or sphingosine ceramides with the same chain lengths. Either unsaturation or C4-hydroxylation of dihydroceramides induced chain length-dependent increases in membrane permeability. Infrared spectroscopy showed that C4-hydroxylation of the sphingoid base decreased the relative ratio of orthorhombic chain packing in the membrane and lowered the miscibility of C24 phytoceramide with lignoceric acid. The phase separation in phytoceramide membranes was confirmed by X-ray diffraction. In contrast, phytoceramides formed strong hydrogen bonds and highly thermostable domains. Thus, the large heterogeneity in ceramide structures and in their aggregation mechanisms may confer resistance towards the heterogeneous external stressors that are constantly faced by the skin barrier.

  20. Membrane lipid rafts and neurobiology: age-related changes in membrane lipids and loss of neuronal function.

    PubMed

    Egawa, Junji; Pearn, Matthew L; Lemkuil, Brian P; Patel, Piyush M; Head, Brian P

    2016-08-15

    A better understanding of the cellular physiological role that plasma membrane lipids, fatty acids and sterols play in various cellular systems may yield more insight into how cellular and whole organ function is altered during the ageing process. Membrane lipid rafts (MLRs) within the plasma membrane of most cells serve as key organizers of intracellular signalling and tethering points of cytoskeletal components. MLRs are plasmalemmal microdomains enriched in sphingolipids, cholesterol and scaffolding proteins; they serve as a platform for signal transduction, cytoskeletal organization and vesicular trafficking. Within MLRs are the scaffolding and cholesterol binding proteins named caveolin (Cav). Cavs not only organize a multitude of receptors including neurotransmitter receptors (NMDA and AMPA receptors), signalling proteins that regulate the production of cAMP (G protein-coupled receptors, adenylyl cyclases, phosphodiesterases (PDEs)), and receptor tyrosine kinases involved in growth (Trk), but also interact with components that modulate actin and tubulin cytoskeletal dynamics (e.g. RhoGTPases and actin binding proteins). MLRs are essential for the regulation of the physiology of organs such as the brain, and age-related loss of cholesterol from the plasma membrane leads to loss of MLRs, decreased presynaptic vesicle fusion, and changes in neurotransmitter release, all of which contribute to different forms of neurodegeneration. Thus, MLRs provide an active membrane domain that tethers and reorganizes the cytoskeletal machinery necessary for membrane and cellular repair, and genetic interventions that restore MLRs to normal cellular levels may be exploited as potential therapeutic means to reverse the ageing and neurodegenerative processes.

  1. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells.

    PubMed

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi

    2013-04-01

    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.

  2. Modulation of a membrane lipid lamellar-nonlamellar phase transition by cationic lipids: A measure for transfection efficiency

    SciTech Connect

    Tenchov, Boris G; Wang, Li; Koynova, Rumiana; MacDonald, Robert C

    2010-01-18

    Synthetic cationic lipids can be used as DNA carriers and are regarded to be the most promising non-viral gene carriers. For this investigation, six novel phosphatidylcholine (PC) cationic derivatives with various hydrophobic moieties were synthesized and their transfection efficiencies for human umbilical artery endothelial cells (HUAEC) were determined. Three compounds with relatively short, myristoleoyl or myristelaidoyl 14:1 chains exhibited very high activity, exceeding by {approx}10 times that of the reference cationic derivative dioleoyl ethylPC (EDOPC). Noteworthy, cationic lipids with 14:1 hydrocarbon chains have not been tested as DNA carriers in transfection assays previously. The other three lipids, which contained oleoyl 18:1 and longer chains, exhibited moderate to weak transfection activity. Transfection efficiency was found to correlate strongly with the effect of the cationic lipids on the lamellar-to-inverted hexagonal, L{sub {alpha}} {yields} H{sub II}, phase conversion in dipalmitoleoyl phosphatidylethanolamine dispersions (DPoPE). X-ray diffraction on binary DPoPE/cationic lipid mixtures showed that the superior transfection agents eliminated the direct L{sub {alpha}} {yields} H{sub II} phase transition and promoted formation of an inverted cubic phase between the L{sub {alpha}} and H{sub II} phases. In contrast, moderate and weak transfection agents retained the direct L{sub {alpha}} {yields} H{sub II} transition but shifted to higher temperatures than that of pure DPoPE, and induced cubic phase formation at a later stage. On the basis of current models of lipid membrane fusion, promotion of a cubic phase by the high-efficiency agents may be considered as an indication that their high transfection activity results from enhanced lipoplex fusion with cellular membranes. The distinct, well-expressed correlation established between transfection efficiency of a cationic lipid and the way it modulates nonlamellar phase formation of a membrane lipid

  3. Physical aspects of heterogeneities in multi-component lipid membranes.

    PubMed

    Komura, Shigeyuki; Andelman, David

    2014-06-01

    Ever since the raft model for biomembranes has been proposed, the traditional view of biomembranes based on the fluid-mosaic model has been altered. In the raft model, dynamical heterogeneities in multi-component lipid bilayers play an essential role. Focusing on the lateral phase separation of biomembranes and vesicles, we review some of the most relevant research conducted over the last decade. We mainly refer to those experimental works that are based on physical chemistry approach, and to theoretical explanations given in terms of soft matter physics. In the first part, we describe the phase behavior and the conformation of multi-component lipid bilayers. After formulating the hydrodynamics of fluid membranes in the presence of the surrounding solvent, we discuss the domain growth-law and decay rate of concentration fluctuations. Finally, we review several attempts to describe membrane rafts as two-dimensional microemulsion.

  4. Peroxidation of tobacco membrane lipids by the photosensitizing toxin, cercosporin.

    PubMed

    Daub, M E

    1982-06-01

    Cercosporin, a nonspecific toxin from Cercospora species, is a photosensitizing compound which rapidly kills plant cells in the light. Cell death appears to be due to a cercosporin-mediated peroxidation of membrane lipids. Tobacco leaf discs treated with cercosporin showed a large increase in electrolyte leakage 1 to 2 minutes after irradiation with light. All tobacco protoplasts exposed to cercosporin in the light were damaged within 45 minutes. Chloroform:methanol extracts of toxin-treated suspension cultures gave positive reactions for lipid hydroperoxides in the thiobarbituric acid test. Cercosporin-treated leaf discs emitted high concentrations of ethane 12 to 24 hours after incubation in the light. Cercosporin also oxidized solutions of methyl linolenate as determined by the thiobarbituric acid assay and the emission of ethane. alpha-Tocopherol had an inhibitory effect on the cercosporin-mediated lipid peroxidation.

  5. Electrochemical characterization of bilayer lipid membrane-semiconductor junctions

    SciTech Connect

    Zhao, Xiao Kang; Baral, S.; Fendler, J.H. )

    1990-03-08

    Three different systems of glyceryl monooleate (GMO), bilayer lipid membrane (BLM) supported semiconductor particles have been prepared and characterized. A single composition of particulate semiconductor deposited only on one side of the BLM constituted system A, two different compositions of particulate semiconductors sequentially deposited on the same side of the BLM represented system B, and two different compositions of particulate semiconductors deposited on the opposite sides of the BLM made up system C.

  6. Simulation of Nanoparticle Permeation through a Lipid Membrane

    PubMed Central

    Fiedler, Steven L.; Violi, Angela

    2010-01-01

    Abstract A metric of nanoparticle toxicity is the passive permeability rate through cellular membranes. To assess the influence of nanoparticle morphology on this process, the permeability of buckyball-sized molecules through a representative lipid bilayer was investigated by molecular-dynamics simulation. When C60 was compared with a prototypical opened C60 molecule and a representative combustion-generated particle, C68H29, the calculated free-energy profiles along the permeation coordinate revealed a sizable variation in form and depth. The orientation of the anisotropic molecules was determined by monitoring the principal axis corresponding to the largest moment of inertia, and free rotation was shown to be hindered in the bilayer interior. Diffusion constant values of the permeant molecules were calculated from a statistical average of seven to 10 trajectories at five locations along the permeation coordinate. A relatively minor variation of the values was observed in the bilayer interior; however, local resistance values spanned up to 24 orders of magnitude from the water layer to the bilayer center, due primarily to its exponential dependence on free energy. The permeability coefficient values calculated for the three similarly sized but structurally distinct nanoparticles showed a significant variance. The use of C60 to represent similarly sized carbonaceous nanoparticles for assessments of toxicity is questioned. PMID:20655842

  7. [The action of week inorganic acids and lower carboxylic acids on the conductivity of bilayer lipid membranes].

    PubMed

    Kilivnik, K E; Khmarskaia, L A; Ksenzhek, O S

    2009-01-01

    The ability of weak inorganic acids (H2S, HCN) and lower carboxylic acids to interact with bilayer lipid membranes, change their conductivity, and act as protonophores has been investigated. The mechanism of changes in BLM conductivity was studied. Factors influencing the interaction of acids with model lipid membranes were determined. Maximum changes in conductivity were observed at pH values equal to the dissociation constants of weak acids and depend on the coefficients of distribution "octanol-water".

  8. Polymer-Induced Swelling of Solid-Supported Lipid Membranes

    PubMed Central

    Kreuzer, Martin; Trapp, Marcus; Dahint, Reiner; Steitz, Roland

    2015-01-01

    In this paper, we study the interaction of charged polymers with solid-supported 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) membranes by in-situ neutron reflectivity. We observe an enormous swelling of the oligolamellar lipid bilayer stacks after incubation in solutions of poly(allylamine hydrochloride) (PAH) in D2O. The positively charged polyelectrolyte molecules interact with the lipid bilayers and induce a drastic increase in their d-spacing by a factor of ~4. Temperature, time, and pH influence the swollen interfacial lipid linings. From our study, we conclude that electrostatic interactions introduced by the adsorbed PAH are the main cause for the drastic swelling of the lipid coatings. The DMPC membrane stacks do not detach from their solid support at T > Tm. Steric interactions, also introduced by the PAH molecules, are held responsible for the stabilizing effect. We believe that this novel system offers great potential for fundamental studies of biomembrane properties, keeping the membrane’s natural fluidity and freedom, decoupled from a solid support at physiological conditions. PMID:26703746

  9. Lipid membrane-mediated attraction between curvature inducing objects

    PubMed Central

    van der Wel, Casper; Vahid, Afshin; Šarić, Anđela; Idema, Timon; Heinrich, Doris; Kraft, Daniela J.

    2016-01-01

    The interplay of membrane proteins is vital for many biological processes, such as cellular transport, cell division, and signal transduction between nerve cells. Theoretical considerations have led to the idea that the membrane itself mediates protein self-organization in these processes through minimization of membrane curvature energy. Here, we present a combined experimental and numerical study in which we quantify these interactions directly for the first time. In our experimental model system we control the deformation of a lipid membrane by adhering colloidal particles. Using confocal microscopy, we establish that these membrane deformations cause an attractive interaction force leading to reversible binding. The attraction extends over 2.5 times the particle diameter and has a strength of three times the thermal energy (−3.3 kBT). Coarse-grained Monte-Carlo simulations of the system are in excellent agreement with the experimental results and prove that the measured interaction is independent of length scale. Our combined experimental and numerical results reveal membrane curvature as a common physical origin for interactions between any membrane-deforming objects, from nanometre-sized proteins to micrometre-sized particles. PMID:27618764

  10. Lipid membrane-mediated attraction between curvature inducing objects

    NASA Astrophysics Data System (ADS)

    van der Wel, Casper; Vahid, Afshin; Šarić, Anđela; Idema, Timon; Heinrich, Doris; Kraft, Daniela J.

    2016-09-01

    The interplay of membrane proteins is vital for many biological processes, such as cellular transport, cell division, and signal transduction between nerve cells. Theoretical considerations have led to the idea that the membrane itself mediates protein self-organization in these processes through minimization of membrane curvature energy. Here, we present a combined experimental and numerical study in which we quantify these interactions directly for the first time. In our experimental model system we control the deformation of a lipid membrane by adhering colloidal particles. Using confocal microscopy, we establish that these membrane deformations cause an attractive interaction force leading to reversible binding. The attraction extends over 2.5 times the particle diameter and has a strength of three times the thermal energy (‑3.3 kBT). Coarse-grained Monte-Carlo simulations of the system are in excellent agreement with the experimental results and prove that the measured interaction is independent of length scale. Our combined experimental and numerical results reveal membrane curvature as a common physical origin for interactions between any membrane-deforming objects, from nanometre-sized proteins to micrometre-sized particles.

  11. Phosphatidylserine lipids and membrane order precisely regulate the activity of Polybia-MP1 peptide.

    PubMed

    Alvares, Dayane S; Neto, João Ruggiero; Ambroggio, Ernesto E

    2017-03-05

    Polybia-MP1 (IDWKKLLDAAKQIL-NH2) is a lytic peptide from the Brazilian wasp venom with known anti-cancer properties. Previous evidence indicates that phosphatidylserine (PS) lipids are relevant for the lytic activity of MP1. In agreement with this requirement, phosphatidylserine lipids are translocated to the outer leaflet of cells, and are available for MP1 binding, depending on the presence of liquid-ordered domains. Here, we investigated the effect of PS on MP1 activity when this lipid is reconstituted in membranes of giant or large liposomes with different lipid-phase states. By monitoring the membrane and soluble luminal content of giant unilamellar vesicles (GUVs), using fluorescence confocal microscopy, we were able to determine that MP1 has a pore-forming activity at the membrane level. Liquid-ordered domains, which were phase-separated within the membrane of GUVs, influenced the pore-forming activity of MP1. Experiments evaluating the membrane-binding and lytic activity of MP1 on large unilamellar vesicles (LUVs), with the same lipid composition as GUVs, demonstrated that there was synergy between liquid-ordered domains and PS, which enhanced both activities. Based on our findings, we propose that the physicochemical properties of cancer cell membranes, which possess a much higher concentration of PS than normal cells, renders them susceptible to MP1 binding and lytic pore formation. These results can be correlated with MP1's potent and selective anti-cancer activity and pave the way for future research to develop cancer therapies that harness and exploit the properties of MP1.

  12. Anomalous surface diffusion of protons on lipid membranes.

    PubMed

    Wolf, Maarten G; Grubmüller, Helmut; Groenhof, Gerrit

    2014-07-01

    The cellular energy machinery depends on the presence and properties of protons at or in the vicinity of lipid membranes. To asses the energetics and mobility of a proton near a membrane, we simulated an excess proton near a solvated DMPC bilayer at 323 K, using a recently developed method to include the Grotthuss proton shuttling mechanism in classical molecular dynamics simulations. We obtained a proton surface affinity of -13.0 ± 0.5 kJ mol(-1). The proton interacted strongly with both lipid headgroup and linker carbonyl oxygens. Furthermore, the surface diffusion of the proton was anomalous, with a subdiffusive regime over the first few nanoseconds, followed by a superdiffusive regime. The time- and distance dependence of the proton surface diffusion coefficient within these regimes may also resolve discrepancies between previously reported diffusion coefficients. Our simulations show that the proton anomalous surface diffusion originates from restricted diffusion in two different surface-bound states, interrupted by the occasional bulk-mediated long-range surface diffusion. Although only a DMPC membrane was considered in this work, we speculate that the restrictive character of the on-surface diffusion is highly sensitive to the specific membrane conditions, which can alter the relative contributions of the surface and bulk pathways to the overall diffusion process. Finally, we discuss the implications of our findings for the energy machinery.

  13. A Critical Reassessment of Penetratin Translocation Across Lipid Membranes

    PubMed Central

    Bárány-Wallje, Elsa; Keller, Sandro; Serowy, Steffen; Geibel, Sebastian; Pohl, Peter; Bienert, Michael; Dathe, Margitta

    2005-01-01

    Penetratin is a short, basic cell-penetrating peptide able to induce cellular uptake of a vast variety of large, hydrophilic cargos. We have reassessed the highly controversial issue of direct permeation of the strongly cationic peptide across negatively charged lipid membranes. Confocal laser scanning microscopy on rhodamine-labeled giant vesicles incubated with carboxyfluorescein-labeled penetratin yielded no evidence of transbilayer movement, in contradiction to previously reported results. Confocal fluorescence spectroscopy on black lipid membranes confirmed this finding, which was also not affected by application of a transmembrane electric potential difference. A novel dialysis assay based on tryptophan absorbance and fluorescence spectroscopy demonstrated that the permeability of small and large unilamellar vesicles to penetratin is <10−13 m/s. Taken together, the results show that penetratin is not capable of overcoming model membrane systems irrespective of the bilayer curvature or the presence of a transmembrane voltage. Thus, direct translocation across the hydrophobic core of the plasma membrane cannot account for the efficient uptake of penetratin into live cells, which is in accord with recent in vitro studies underlining the importance of endocytosis in the internalization process of cationic cell-penetrating peptides. PMID:16040762

  14. Investigating cell membrane structure and dynamics with TCSPC-FLIM

    NASA Astrophysics Data System (ADS)

    Le Marois, Alix; Owen, Dylan M.; Suhling, Klaus

    2015-03-01

    We report the use of Time-Correlated Single Photon Counting (TCSPC) in a polarization-resolved Fluorescence Lifetime Imaging (FLIM) setup for the investigation of cell membrane structural and dynamic properties. This technique allows us to study the orientation and mobility of fluorescent membrane dyes, namely di-4-ANEPPDHQ and DiO, in model bilayers of different lipid compositions. Dipole alignment and extent of rotational motion can be linked to membrane order and fluidity. Comparison of the time-resolved anisotropy decays of the two fluorescent dyes suggests that rotational motion of membrane constituents is restricted in liquid-ordered phases, and appears to be limited to the region of aliphatic tails in liquid-disordered phases. In living cells, understanding the membrane structure provides crucial information on its functional properties, such as exo- and endocytosis, cell mobility and signal transduction.

  15. Interaction measurement of particles bound to a lipid membrane

    NASA Astrophysics Data System (ADS)

    Sarfati, Raphael; Dufresne, Eric

    2015-03-01

    The local shape and dynamics of the plasma membrane play important roles in many cellular processes. Local membrane deformations are often mediated by the adsorption of proteins (notably from the BAR family), and their subsequent self-assembly. The emerging hypothesis is that self-assembly arises from long-range interactions of individual proteins through the membrane's deformation field. We study these interactions in a model system of micron-sized colloidal particles adsorbed onto a lipid bilayer. We use fluorescent microscopy, optical tweezers and particle tracking to measure dissipative and conservative forces as a function of the separation between the particles. We find that particles are driven together with forces of order 100 fN and remain bound in a potential well with a stiffness of order 100 fN/micron.

  16. Elasto-plasticity in wrinkled polymerized lipid membranes

    NASA Astrophysics Data System (ADS)

    Chaieb, Sahraoui

    2014-01-01

    Biomembranes shown to behave like elastic sheets, can also suffer plastic deformations. Neutron scattering experiments on partially polymerised wrinkled membranes revealed that when a critical degree of polymerisation is crossed, the wrinkled membranes do not resume their spherical shapes. Instead they remain wrinkled and rigid while their non-polymerised counterparts resume their spherical floppy shapes. The yield stress of these membranes, measured for the first time via the fractal dimension, is intimately related to the degree of polymerisation probably through a 2D disorder that quenches the lateral diffusion of the lipid molecules. This work might shed light on the physical reason behind the irreversible deformation of echinocytes, acanthocytes and malaria infected red blood cells.

  17. Quantitative characterization of the lateral distribution of membrane proteins within the lipid bilayer.

    PubMed Central

    Freire, E; Snyder, B

    1982-01-01

    The dependence of the lateral distribution of membrane proteins on the size, protein/lipoid molar ratio, and the magnitude of the interaction potentials has been investigated by computer modeling protein-lipid distributions with Monte Carlo calculations. These results have allowed us to develop a quantitative characterization of the distribution of membrane proteins and to correlate these distributions with experimental observables. The topological arrangement of protein domains, protein plus annular lipid domains, and free lipid domains is described in terms of radial distribution, pair connectedness, and cluster distribution functions. The radial distribution functions are used to measure the distribution of intermolecular distances between protein molecules, whereas the pair connectedness functions are used to estimate the physical extension of compositional domains. It is shown that, at characteristic protein/lipid molar ratios, previously isolated domains become connected, forming domain networks that extend over the entire membrane surface. These changes in the lateral connectivity of compositional domains are paralleled by changes in the calculated lateral diffusion coefficients and might have important implications for the regulation of diffusion controlled processes within the membrane. PMID:7074188

  18. Partitioning of exogenous delta-tocopherol between the triacylglycerol and membrane lipid fractions of chicken muscle.

    PubMed

    Sigfusson, Halldor; Hultin, Herbert O

    2002-11-20

    The partitioning of exogenous delta-tocopherol, added dissolved in ethanol, between the neutral triacylglycerols and membranes of chicken leg muscles was investigated. The two lipid fractions were separated using differential ultracentrifugation techniques. Triacylglycerols were obtained after high-speed centrifugation of the minced muscle at 130000 g for 30 min. Membranes were collected from a muscle-buffer homogenate (pH 7.5) between 10000 g for 20 min and 130000 g for 30 min. The triacylglycerols collected represented from 15 to 80% of the total triacylglycerols of the minced muscle, the yields increasing with increasing muscle triacylglycerol content. The phospholipids in the isolated membrane fraction represented from 20 to 35% of the total phospholipids of the muscle. At low muscle total lipid contents (3-5%), the added delta-tocopherol was present in approximately the same concentration in both muscle lipid fractions. At higher total lipid contents, achieved by adding exogenous triacylglycerols, the delta-tocopherol concentration in the membranes increased relative to that in the triacylglycerols.

  19. Lipid bilayer thickness determines cholesterol's location in model membranes

    SciTech Connect

    Marquardt, Drew; Heberle, Frederick A.; Greathouse, Denise V.; Koeppe, II, Roger E.; Standaert, Robert F.; Van Oosten, Brad J.; Harroun, Thad A.; Kinnun, Jacob J.; Williams, Justin A.; Wassall, Stephen R.; Katsaras, John

    2016-10-11

    Cholesterol is an essential biomolecule of animal cell membranes, and an important precursor for the biosynthesis of certain hormones and vitamins. It is also thought to play a key role in cell signaling processes associated with functional plasma membrane microdomains (domains enriched in cholesterol), commonly referred to as rafts. In all of these diverse biological phenomena, the transverse location of cholesterol in the membrane is almost certainly an important structural feature. Using a combination of neutron scattering and solid-state 2H NMR, we have determined the location and orientation of cholesterol in phosphatidylcholine (PC) model membranes having fatty acids of different lengths and degrees of unsaturation. The data establish that cholesterol reorients rapidly about the bilayer normal in all the membranes studied, but is tilted and forced to span the bilayer midplane in the very thin bilayers. The possibility that cholesterol lies flat in the middle of bilayers, including those made from PC lipids containing polyunsaturated fatty acids (PUFAs), is ruled out. Finally, these results support the notion that hydrophobic thickness is the primary determinant of cholesterol's location in membranes.

  20. Lipid bilayer thickness determines cholesterol's location in model membranes

    DOE PAGES

    Marquardt, Drew; Heberle, Frederick A.; Greathouse, Denise V.; ...

    2016-10-11

    Cholesterol is an essential biomolecule of animal cell membranes, and an important precursor for the biosynthesis of certain hormones and vitamins. It is also thought to play a key role in cell signaling processes associated with functional plasma membrane microdomains (domains enriched in cholesterol), commonly referred to as rafts. In all of these diverse biological phenomena, the transverse location of cholesterol in the membrane is almost certainly an important structural feature. Using a combination of neutron scattering and solid-state 2H NMR, we have determined the location and orientation of cholesterol in phosphatidylcholine (PC) model membranes having fatty acids of differentmore » lengths and degrees of unsaturation. The data establish that cholesterol reorients rapidly about the bilayer normal in all the membranes studied, but is tilted and forced to span the bilayer midplane in the very thin bilayers. The possibility that cholesterol lies flat in the middle of bilayers, including those made from PC lipids containing polyunsaturated fatty acids (PUFAs), is ruled out. Finally, these results support the notion that hydrophobic thickness is the primary determinant of cholesterol's location in membranes.« less

  1. Area per lipid and cholesterol interactions in membranes from separated local-field (13)C NMR spectroscopy.

    PubMed

    Leftin, Avigdor; Molugu, Trivikram R; Job, Constantin; Beyer, Klaus; Brown, Michael F

    2014-11-18

    Investigations of lipid membranes using NMR spectroscopy generally require isotopic labeling, often precluding structural studies of complex lipid systems. Solid-state (13)C magic-angle spinning NMR spectroscopy at natural isotopic abundance gives site-specific structural information that can aid in the characterization of complex biomembranes. Using the separated local-field experiment DROSS, we resolved (13)C-(1)H residual dipolar couplings that were interpreted with a statistical mean-torque model. Liquid-disordered and liquid-ordered phases were characterized according to membrane thickness and average cross-sectional area per lipid. Knowledge of such structural parameters is vital for molecular dynamics simulations, and provides information about the balance of forces in membrane lipid bilayers. Experiments were conducted with both phosphatidylcholine (dimyristoylphosphatidylcholine (DMPC) and palmitoyloleoylphosphatidylcholine (POPC)) and egg-yolk sphingomyelin (EYSM) lipids, and allowed us to extract segmental order parameters from the (13)C-(1)H residual dipolar couplings. Order parameters were used to calculate membrane structural quantities, including the area per lipid and bilayer thickness. Relative to POPC, EYSM is more ordered in the ld phase and experiences less structural perturbation upon adding 50% cholesterol to form the lo phase. The loss of configurational entropy is smaller for EYSM than for POPC, thus favoring its interaction with cholesterol in raftlike lipid systems. Our studies show that solid-state (13)C NMR spectroscopy is applicable to investigations of complex lipids and makes it possible to obtain structural parameters for biomembrane systems where isotope labeling may be prohibitive.

  2. Distinguishing Bicontinuous Lipid Cubic Phases from Isotropic Membrane Morphologies Using 31P Solid-State NMR Spectroscopy

    PubMed Central

    Yang, Yu; Yao, Hongwei

    2015-01-01

    Nonlamellar lipid membranes are frequently induced by proteins that fuse, bend, and cut membranes. Understanding the mechanism of action of these proteins requires the elucidation of the membrane morphologies that they induce. While hexagonal phases and lamellar phases are readily identified by their characteristic solid-state NMR lineshapes, bicontinuous lipid cubic phases are more difficult to discern, since the static NMR spectra of cubic-phase lipids consist of an isotropic 31P or 2H peak, indistinguishable from the spectra of isotropic membrane morphologies such as micelles and small vesicles. To date, small-angle X-ray scattering is the only method to identify bicontinuous lipid cubic phases. To explore unique NMR signatures of lipid cubic phases, we first describe the orientation distribution of lipid molecules in cubic phases and simulate the static 31P chemical shift lineshapes of oriented cubic-phase membranes in the limit of slow lateral diffusion. We then show that 31P T2 relaxation times differ significantly between isotropic micelles and cubic-phase membranes: the latter exhibit two-orders-of magnitude shorter T2 relaxation times. These differences are explained by the different timescales of lipid lateral diffusion on the cubic-phase surface versus the timescales of micelle tumbling. Using this relaxation NMR approach, we investigated a DOPE membrane containing the transmembrane domain (TMD) of a viral fusion protein. The static 31P spectrum of DOPE shows an isotropic peak, whose T2 relaxation times correspond to that of a cubic phase. Thus, the viral fusion protein TMD induces negative Gaussian curvature, which is an intrinsic characteristic of cubic phases, to the DOPE membrane. This curvature induction has important implications to the mechanism of virus-cell fusion. This study establishes a simple NMR diagnostic probe of lipid cubic phases, which is expected to be useful for studying many protein-induced membrane remodeling phenomena in biology

  3. Modeling the phase separation in binary lipid membrane under externally imposed oscillatory shear flow.

    PubMed

    Chen, Xiao-Bo; Niu, Li-Sha; Shi, Hui-Ji

    2008-09-01

    By adding external velocity terms, the two-dimensional time-dependent Ginzburg-Landau (TDGL) equations are modified. Based on this, the phase separation in binary lipid membrane under externally imposed oscillatory shear flow is numerically modeled employing the Cell Dynamical System (CDS) approach. Considering shear flows with different frequencies and amplitudes, several aspects of such a phase evolving process are studied. Firstly, visualized results are shown via snapshot figures of the membrane shape. And then, the simulated scattering patterns at typical moments are presented. Furthermore, in order to more quantitatively discuss this phase-separation process, the time growth laws of the characteristic domain sizes in both directions parallel and perpendicular to the flow are investigated for each case. Finally, the peculiar rheological properties of such binary lipid membrane system have been discussed, mainly the normal stress difference and the viscoelastic complex shear moduli.

  4. Chemical and structural investigation of lipid nanoparticles: drug-lipid interaction and molecular distribution

    NASA Astrophysics Data System (ADS)

    Anantachaisilp, Suranan; Meejoo Smith, Siwaporn; Treetong, Alongkot; Pratontep, Sirapat; Puttipipatkhachorn, Satit; Rungsardthong Ruktanonchai, Uracha

    2010-03-01

    Lipid nanoparticles are a promising alternative to existing carriers in chemical or drug delivery systems. A key challenge is to determine how chemicals are incorporated and distributed inside nanoparticles, which assists in controlling chemical retention and release characteristics. This study reports the chemical and structural investigation of γ-oryzanol loading inside a model lipid nanoparticle drug delivery system composed of cetyl palmitate as solid lipid and Miglyol 812® as liquid lipid. The lipid nanoparticles were prepared by high pressure homogenization at varying liquid lipid content, in comparison with the γ-oryzanol free systems. The size of the lipid nanoparticles, as measured by the photon correlation spectroscopy, was found to decrease with increased liquid lipid content from 200 to 160 nm. High-resolution proton nuclear magnetic resonance (1H-NMR) measurements of the medium chain triglyceride of the liquid lipid has confirmed successful incorporation of the liquid lipid in the lipid nanoparticles. Differential scanning calorimetric and powder x-ray diffraction measurements provide complementary results to the 1H-NMR, whereby the crystallinity of the lipid nanoparticles diminishes with an increase in the liquid lipid content. For the distribution of γ-oryzanol inside the lipid nanoparticles, the 1H-NMR revealed that the chemical shifts of the liquid lipid in γ-oryzanol loaded systems were found at rather higher field than those in γ-oryzanol free systems, suggesting incorporation of γ-oryzanol in the liquid lipid. In addition, the phase-separated structure was observed by atomic force microscopy for lipid nanoparticles with 0% liquid lipid, but not for lipid nanoparticles with 5 and 10% liquid lipid. Raman spectroscopic and mapping measurements further revealed preferential incorporation of γ-oryzanol in the liquid part rather than the solid part of in the lipid nanoparticles. Simple models representing the distribution of γ-oryzanol and

  5. β2-Microglobulin Amyloid Fibril-Induced Membrane Disruption Is Enhanced by Endosomal Lipids and Acidic pH

    PubMed Central

    Goodchild, Sophia C.; Sheynis, Tania; Thompson, Rebecca; Tipping, Kevin W.; Xue, Wei-Feng; Ranson, Neil A.; Beales, Paul A.; Hewitt, Eric W.; Radford, Sheena E.

    2014-01-01

    Although the molecular mechanisms underlying the pathology of amyloidoses are not well understood, the interaction between amyloid proteins and cell membranes is thought to play a role in several amyloid diseases. Amyloid fibrils of β2-microglobulin (β2m), associated with dialysis-related amyloidosis (DRA), have been shown to cause disruption of anionic lipid bilayers in vitro. However, the effect of lipid composition and the chemical environment in which β2m-lipid interactions occur have not been investigated previously. Here we examine membrane damage resulting from the interaction of β2m monomers and fibrils with lipid bilayers. Using dye release, tryptophan fluorescence quenching and fluorescence confocal microscopy assays we investigate the effect of anionic lipid composition and pH on the susceptibility of liposomes to fibril-induced membrane damage. We show that β2m fibril-induced membrane disruption is modulated by anionic lipid composition and is enhanced by acidic pH. Most strikingly, the greatest degree of membrane disruption is observed for liposomes containing bis(monoacylglycero)phosphate (BMP) at acidic pH, conditions likely to reflect those encountered in the endocytic pathway. The results suggest that the interaction between β2m fibrils and membranes of endosomal origin may play a role in the molecular mechanism of β2m amyloid-associated osteoarticular tissue destruction in DRA. PMID:25100247

  6. Raman Imaging in Cell Membranes, Lipid-Rich Organelles, and Lipid Bilayers.

    PubMed

    Syed, Aleem; Smith, Emily A

    2017-03-15

    Raman-based optical imaging is a promising analytical tool for noninvasive, label-free chemical imaging of lipid bilayers and cellular membranes. Imaging using spontaneous Raman scattering suffers from a low intensity that hinders its use in some cellular applications. However, developments in coherent Raman imaging, surface-enhanced Raman imaging, and tip-enhanced Raman imaging have enabled video-rate imaging, excellent detection limits, and nanometer spatial resolution, respectively. After a brief introduction to these commonly used Raman imaging techniques for cell membrane studies, this review discusses selected applications of these modalities for chemical imaging of membrane proteins and lipids. Finally, recent developments in chemical tags for Raman imaging and their applications in the analysis of selected cell membrane components are summarized. Ongoing developments toward improving the temporal and spatial resolution of Raman imaging and small-molecule tags with strong Raman scattering cross sections continue to expand the utility of Raman imaging for diverse cell membrane studies. Expected final online publication date for the Annual Review of Analytical Chemistry Volume 10 is June 12, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

  7. Genetic Analysis of Arabidopsis Mutants Impaired in Plastid Lipid Import Reveals a Role of Membrane Lipids in Chloroplast Division

    SciTech Connect

    Fan, J.; Xu, C.

    2011-03-01

    The biogenesis of photosynthetic membranes in plants relies largely on lipid import from the endoplasmic reticulum (ER) and this lipid transport process is mediated by TGD proteins in Arabidopsis. Such a dependency of chloroplast biogenesis on ER-to-plastid lipid transport was recently exemplified by analyzing double mutants between tgd1-1 or tgd4-3 and fad6 mutants. The fad6 mutants are defective in the desaturation of membrane lipids in chloroplasts and therefore dependent on import of polyunsaturated lipid precursors from the ER for constructing a competent thylakoid membrane system. In support of a critical role of TGD proteins in ER-to-plastid lipid trafficking, we showed that the introduction of the tgd mutations into fad6 mutant backgrounds led to drastic reductions in relative amounts of thylakoid lipids. Moreover, the tgd1-1 fad6 and tgd4-3 fad6 double mutants were deficient in polyunsaturated fatty acids in chloroplast membrane lipids, and severely compromised in the biogenesis of photosynthetic membrane systems. Here we report that these double mutants are severely impaired in chloroplast division. The possible role of membrane lipids in chloroplast division is discussed.

  8. Imaging lipid domains in cell membranes: the advent of super-resolution fluorescence microscopy

    PubMed Central

    Owen, Dylan M.; Gaus, Katharina

    2013-01-01

    The lipid bilayer of model membranes, liposomes reconstituted from cell lipids, and plasma membrane vesicles and spheres can separate into two distinct liquid phases to yield lipid domains with liquid-ordered and liquid-disordered properties. These observations are the basis of the lipid raft hypothesis that postulates the existence of cholesterol-enriched ordered-phase lipid domains in cell membranes that could regulate protein mobility, localization and interaction. Here we review the evidence that nano-scaled lipid complexes and meso-scaled lipid domains exist in cell membranes and how new fluorescence microscopy techniques that overcome the diffraction limit provide new insights into lipid organization in cell membranes. PMID:24376453

  9. Kinetics of enzymatic reactions in lipid membranes containing domains.

    PubMed

    Zhdanov, Vladimir P; Höök, Fredrik

    2015-03-06

    An appreciable part of enzymes operating in vivo is associated with lipid membranes. The function of such enzymes can be influenced by the presence of domains containing proteins and/or composed of different lipids. The corresponding experimental model-system studies can be performed under well controlled conditions, e.g., on a planar supported lipid bilayer or surface-immobilized vesicles. To clarify what may happen in such systems, we propose general kinetic equations describing the enzyme-catalyzed substrate conversion occurring via the Michaelis-Menten (MM) mechanism on a membrane with domains which do not directly participate in reaction. For two generic situations when a relatively slow reaction takes place primarily in or outside domains, we take substrate saturation and lateral substrate-substrate interactions at domains into account and scrutinize the dependence of the reaction rate on the average substrate coverage. With increasing coverage, depending on the details, the reaction rate reaches saturation via an inflection point or monotonously as in the conventional MM case. In addition, we show analytically the types of reaction kinetics occurring primarily at domain boundaries. In the physically interesting situation when the domain growth is fast on the reaction time scale, the latter kinetics are far from conventional. The opposite situation when the reaction is fast and controlled by diffusion has been studied by using the Monte Carlo technique. The corresponding results indicate that the dependence of the reaction kinetics on the domain size may be weak.

  10. Acylation of Glucagon-Like Peptide-2: Interaction with Lipid Membranes and In Vitro Intestinal Permeability

    PubMed Central

    Trier, Sofie; Linderoth, Lars; Bjerregaard, Simon; Andresen, Thomas Lars; Rahbek, Ulrik Lytt

    2014-01-01

    Background Acylation of peptide drugs with fatty acid chains has proven beneficial for prolonging systemic circulation as well as increasing enzymatic stability without disrupting biological potency. Acylation has furthermore been shown to increase interactions with the lipid membranes of mammalian cells. The extent to which such interactions hinder or benefit delivery of acylated peptide drugs across cellular barriers such as the intestinal epithelia is currently unknown. The present study investigates the effect of acylating peptide drugs from a drug delivery perspective. Purpose We hypothesize that the membrane interaction is an important parameter for intestinal translocation, which may be used to optimize the acylation chain length for intestinal permeation. This work aims to characterize acylated analogues of the intestinotrophic Glucagon-like peptide-2 by systematically increasing acyl chain length, in order to elucidate its influence on membrane interaction and intestinal cell translocation in vitro. Results Peptide self-association and binding to both model lipid and cell membranes was found to increase gradually with acyl chain length, whereas translocation across Caco-2 cells depended non-linearly on chain length. Short and medium acyl chains increased translocation compared to the native peptide, but long chain acylation displayed no improvement in translocation. Co-administration of a paracellular absorption enhancer was found to increase translocation irrespective of acyl chain length, whereas a transcellular enhancer displayed increased synergy with the long chain acylation. Conclusions These results show that membrane interactions play a prominent role during intestinal translocation of an acylated peptide. Acylation benefits permeation for shorter and medium chains due to increased membrane interactions, however, for longer chains insertion in the membrane becomes dominant and hinders translocation, i.e. the peptides get ‘stuck’ in the cell

  11. Fuel cell ion-exchange membrane investigation

    NASA Technical Reports Server (NTRS)

    Toy, M. S.

    1972-01-01

    The present deficiencies in the fluorocarbon sulfonic acid membrane used as the solid polymer electrolyte in the H2/O2 fuel cell are studied. Considered are: Adhesives selection, elastomeric formulations, scavenger exploration, and membrane characterization. The significant data are interpreted and recommendations are given for both short and long range further investigations in two of the four major areas: membrane adhesives and membrane stabilization.

  12. Redistribution of Cholesterol in Model Lipid Membranes in Response to the Membrane-Active Peptide Alamethicin

    NASA Astrophysics Data System (ADS)

    Heller, William; Qian, Shuo

    2013-03-01

    The cellular membrane is a heterogeneous, dynamic mixture of molecules and macromolecules that self-assemble into a tightly-regulated functional unit that provides a semipermeable barrier between the cell and its environment. Among the many compositional differences between mammalian and bacterial cell membranes that impact its physical properties, one key difference is cholesterol content, which is more prevalent in mammals. Cholesterol is an amphiphile that associates with membranes and serves to maintain its fluidity and permeability. Membrane-active peptides, such as the alpha-helical peptide alamethicin, interact with membranes in a concentration- and composition-dependent manner to form transmembrane pores that are responsible for the lytic action of the peptide. Through the use of small-angle neutron scattering and deuterium labeling, it was possible to observe a redistribution of the lipid and cholesterol in unilamellar vesicles in response to the presence of alamethicin at a peptide-to-lipid ratio of 1/200. The results demonstrate that the membrane remodeling powers of alamethicin reach beyond the membrane thinning effect to altering the localization of specific components in the bilayer, complementing the accepted two-state mechanism of pore formation. Research was supported by U. S. DOE-OBER (CSMB; FWP ERKP291) and the U. S. DOE-BES Scientific User Facilities Division (ORNL's SNS and HFIR).

  13. An ER protein functionally couples neutral lipid metabolism on lipid droplets to membrane lipid synthesis in the ER.

    PubMed

    Markgraf, Daniel F; Klemm, Robin W; Junker, Mirco; Hannibal-Bach, Hans K; Ejsing, Christer S; Rapoport, Tom A

    2014-01-16

    Eukaryotic cells store neutral lipids such as triacylglycerol (TAG) in lipid droplets (LDs). Here, we have addressed how LDs are functionally linked to the endoplasmic reticulum (ER). We show that, in S. cerevisiae, LD growth is sustained by LD-localized enzymes. When LDs grow in early stationary phase, the diacylglycerol acyl-transferase Dga1p moves from the ER to LDs and is responsible for all TAG synthesis from diacylglycerol (DAG). During LD breakdown in early exponential phase, an ER membrane protein (Ice2p) facilitates TAG utilization for membrane-lipid synthesis. Ice2p has a cytosolic domain with affinity for LDs and is required for the efficient utilization of LD-derived DAG in the ER. Ice2p breaks a futile cycle on LDs between TAG degradation and synthesis, promoting the rapid relocalization of Dga1p to the ER. Our results show that Ice2p functionally links LDs with the ER and explain how cells switch neutral lipid metabolism from storage to consumption.

  14. Membrane on a chip: a functional tethered lipid bilayer membrane on silicon oxide surfaces.

    PubMed

    Atanasov, Vladimir; Knorr, Nikolaus; Duran, Randolph S; Ingebrandt, Sven; Offenhäusser, Andreas; Knoll, Wolfgang; Köper, Ingo

    2005-09-01

    Tethered membranes have been proven during recent years to be a powerful and flexible biomimetic platform. We reported in a previous article on the design of a new architecture based on the self-assembly of a thiolipid on ultrasmooth gold substrates, which shows extremely good electrical sealing properties as well as functionality of a bilayer membrane. Here, we describe the synthesis of lipids for a more modular design and the adaptation of the linker part to silane chemistry. We were able to form a functional tethered bilayer lipid membrane with good electrical sealing properties covering a silicon oxide surface. We demonstrate the functional incorporation of the ion carrier valinomycin and of the ion channel gramicidin.

  15. Membrane lipid peroxidation by UV-A: Mechanism and implications

    SciTech Connect

    Bose, B.; Agarwal, S.; Chatterjee, S.N. )

    1990-10-01

    UV-A produced a dose-dependent linear increase of lipid peroxidation in liposomal membrane, as detected by the assay of (i) conjugated dienes, (ii) lipid hydroperoxides, (iii) malondialdehydes (MDA), and (iv) the fluorescent adducts formed by the reaction of MDA with glycine and also a linear dose-dependent increase of ({sup 14}C)glucose efflux from the liposomes. UV-A-induced MDA production could not be inhibited by any significant degree by sodium formate, dimethyl sulfoxide, EDTA, or superoxide dismutase but was very significantly inhibited by butylated hydroxytoluene, alpha-tocopherol, sodium azide, L-histidine, dimethylfuran, and beta-carotene. MDA formation increased with an increase in the D{sub 2}O content in water, leading to a maximal amount of nearly 50% enhancement of lipid peroxidation in 100% D{sub 2}O vis-a-vis water used as dispersion medium. The experimental findings indicate the involvement of singlet oxygen as the initiator of the UV-A-induced lipid peroxidation.

  16. Millimeter microwave effect on ion transport across lipid bilayer membranes.

    PubMed

    Alekseev, S I; Ziskin, M C

    1995-01-01

    The effects of millimeter microwaves in the frequency range of 54-76 GHz on capacitance and conductance of lipid bilayer membranes (BLM) were studied. Some of the membranes were modified by gramicidin A and amphotericin B or by tetraphenylboron anions (TPhB-). The millimeter microwaves were pulse-modulated (PW) at repetition rates ranging from 1 to 100 pps, PW at 1000 pps, or unmodulated continuous waves (CW). The maximum output power at the waveguide outlet was 20 mW. It was found that CW irradiation decreased the unmodified BLM capacitance by 1.2% +/- 0.5%. At the same time, membrane current induced by TPhB- transport increased by 5% +/- 1%. The changes in conductance of ionic channels formed by gramicidin A and amphotericin B were small (0.6% +/- 0.4%). No "resonance-like" effects of mm-wave irradiation on membrane capacitance, ionic channel currents, or TPhB- transport were detected. All changes in membrane capacitance and currents were independent of the modulation employed and were equivalent to heating by approximately 1.1 degrees C.

  17. Solubility and Permeation of Hydrogen Sulfide in Lipid Membranes

    PubMed Central

    Cuevasanta, Ernesto; Denicola, Ana; Alvarez, Beatriz; Möller, Matías N.

    2012-01-01

    Hydrogen sulfide (H2S) is mainly known for its toxicity but has recently been shown to be produced endogenously in mammalian tissues and to be associated with physiological regulatory functions. To better understand the role of biomembranes in modulating its biological distribution and effects; we measured the partition coefficient of H2S in models of biological membranes. The partition coefficients were found to be 2.1±0.2, 1.9±0.5 and 2.0±0.6 in n-octanol, hexane and dilauroylphosphatidylcholine liposome membranes relative to water, respectively (25°C). This two-fold higher concentration of H2S in the membrane translates into a rapid membrane permeability, Pm = 3 cm s−1. We used a mathematical model in three dimensions to gain insight into the diffusion of total sulfide in tissues. This model shows that the sphere of action of sulfide produced by a single cell expands to involve more than 200 neighboring cells, and that the resistance imposed by lipid membranes has a significant effect on the diffusional spread of sulfide at pH 7.4, increasing local concentrations. These results support the role of hydrogen sulfide as a paracrine signaling molecule and reveal advantageous pharmacokinetic properties for its therapeutic applications. PMID:22509322

  18. Binding of amphiphilic and triphilic block copolymers to lipid model membranes: the role of perfluorinated moieties.

    PubMed

    Schwieger, Christian; Achilles, Anja; Scholz, Sven; Rüger, Jan; Bacia, Kirsten; Saalwaechter, Kay; Kressler, Jörg; Blume, Alfred

    2014-09-07

    A novel class of symmetric amphi- and triphilic (hydrophilic, lipophilic, fluorophilic) block copolymers has been investigated with respect to their interactions with lipid membranes. The amphiphilic triblock copolymer has the structure PGMA(20)-PPO(34)-PGMA(20) (GP) and it becomes triphilic after attaching perfluoroalkyl moieties (F9) to either end which leads to F(9)-PGMA(20)-PPO(34)-PGMA(20)-F(9) (F-GP). The hydrophobic poly(propylene oxide) (PPO) block is sufficiently long to span a lipid bilayer. The poly(glycerol monomethacrylate) (PGMA) blocks have a high propensity for hydrogen bonding. The hydrophobic and lipophobic perfluoroalkyl moieties have the tendency to phase segregate in aqueous as well as in hydrocarbon environments. We performed differential scanning calorimetry (DSC) measurements on polymer bound lipid vesicles under systematic variation of the bilayer thickness, the nature of the lipid headgroup, and the polymer concentration. The vesicles were composed of phosphatidylcholines (DMPC, DPPC, DAPC, DSPC) or phosphatidylethanolamines (DMPE, DPPE, POPE). We showed that GP as well as F-GP binding have membrane stabilizing and destabilizing components. PPO and F9 blocks insert into the hydrophobic part of the membrane concomitantly with PGMA block adsorption to the lipid headgroup layer. The F9 chains act as additional membrane anchors. The insertion of the PPO blocks of both GP and F-GP could be proven by 2D-NOESY NMR spectroscopy. By fluorescence microscopy we show that F-GP binding increases the porosity of POPC giant unilamellar vesicles (GUVs), allowing the influx of water soluble dyes as well as the translocation of the complete triphilic polymer and its accumulation at the GUV surface. These results open a new route for the rational design of membrane systems with specific properties.

  19. In vitro study of interaction of synaptic vesicles with lipid membranes

    NASA Astrophysics Data System (ADS)

    Ghosh, S. K.; Castorph, S.; Konovalov, O.; Jahn, R.; Holt, M.; Salditt, T.

    2010-10-01

    The fusion of synaptic vesicles (SVs) with the plasma membrane in neurons is a crucial step in the release of neurotransmitters, which are responsible for carrying signals between nerve cells. While many of the molecular players involved in this fusion process have been identified, a precise molecular description of their roles in the process is still lacking. A case in point is the plasma membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2). Although PIP2 is known to be essential for vesicle fusion, its precise role in the process remains unclear. We have re-investigated the role of this lipid in membrane structure and function using the complementary experimental techniques of x-ray reflectivity, both on lipid monolayers at an air-water interface and bilayers on a solid support, and grazing incidence x-ray diffraction on lipid monolayers. These techniques provide unprecedented access to structural information at the molecular level, and detail the profound structural changes that occur in a membrane following PIP2 incorporation. Further, we also confirm and extend previous findings that the association of SVs with membranes is enhanced by PIP2 incorporation, and reveal the structural changes that underpin this phenomenon. Further, the association is further intensified by a physiologically relevant amount of Ca2+ ions in the subphase of the monolayer, as revealed by the increase in interfacial pressure seen with the lipid monolayer system. Finally, a theoretical calculation concerning the products arising from the fusion of these SVs with proteoliposomes is presented, with which we aim to illustrate the potential future uses of this system.

  20. Role of lipid II and membrane thickness in the mechanism of action of the lantibiotic bovicin HC5.

    PubMed

    Paiva, Aline Dias; Breukink, Eefjan; Mantovani, Hilário Cuquetto

    2011-11-01

    Lantibiotics are antimicrobial peptides produced by Gram-positive bacteria, nisin being the most well-known member. Nisin inhibits peptidoglycan synthesis and forms pores at sensitive membranes upon interaction with lipid II, the essential bacterial cell wall precursor. Bovicin HC5, a bacteriocin produced by Streptococcus bovis HC5, has the putative N-terminal lipid II binding motif, and we investigated the mode of action of bovicin HC5 using both living bacteria and model membranes, with special emphasis on the role of lipid II. Bovicin HC5 showed activity against Staphylococcus cohnii and Staphylococcus warneri, but bovicin HC5 hardly interfered with the membrane potential of S. cohnii. In model membranes, bovicin HC5 was not able to cause carboxyfluorescein release or proton influx from DOPC vesicles containing lipid II. Bovicin HC5 blocked lipid II-dependent pore formation activity of nisin, and a high-affinity interaction with lipid II was observed (apparent binding constant [K(a)] = 3.1 × 10(6) M(-1)), with a 1:1 stoichiometry. In DOPC vesicles containing lipid II, bovicin HC5 was able to assemble with lipid II into a prepore-like structure. Furthermore, we observed pore formation activity of bovicin HC5, which was stimulated by the presence of lipid II, in thin membranes. Moreover, bovicin HC5 induced the segregation of lipid II into domains in giant model membrane vesicles. In conclusion, bovicin HC5 has a primary mode of action similar to that of nisin, but some differences regarding the pore-forming capacity were demonstrated.

  1. Biological Membranes in Extreme Conditions: Simulations of Anionic Archaeal Tetraether Lipid Membranes

    PubMed Central

    Pineda De Castro, Luis Felipe; Dopson, Mark

    2016-01-01

    In contrast to the majority of organisms that have cells bound by di-ester phospholipids, archaeal membranes consist of di- and tetraether phospholipids. Originating from organisms that withstand harsh conditions (e.g., low pH and a wide range of temperatures) such membranes have physical properties that make them attractive materials for biological research and biotechnological applications. We developed force-field parameters based on the widely used Generalized Amber Force Field (GAFF) to enable the study of anionic tetraether membranes of the model archaean Sulfolobus acidocaldarius by computer simulations. The simulations reveal that the physical properties of these unique membranes depend on the number of cyclopentane rings included in each lipid unit, and on the size of cations that are used to ensure charge neutrality. This suggests that the biophysical properties of Sulfolobus acidocaldarius cells depend not only on the compositions of their membranes but also on the media in which they grow. PMID:27167213

  2. Membrane protein simulations with a united-atom lipid and all-atom protein model: lipid protein interactions, side chain transfer free energies and model proteins

    NASA Astrophysics Data System (ADS)

    Tieleman, D. Peter; MacCallum, Justin L.; Ash, Walter L.; Kandt, Christian; Xu, Zhitao; Monticelli, Luca

    2006-07-01

    We have reparameterized the dihedral parameters in a commonly used united-atom lipid force field so that they can be used with the all-atom OPLS force field for proteins implemented in the molecular dynamics simulation software GROMACS. Simulations with this new combination give stable trajectories and sensible behaviour of both lipids and protein. We have calculated the free energy of transfer of amino acid side chains between water and 'lipid-cyclohexane', made of lipid force field methylene groups, as a hydrophobic mimic of the membrane interior, for both the OPLS-AA and a modified OPLS-AA force field which gives better hydration free energies under simulation conditions close to those preferred for the lipid force field. The average error is 4.3 kJ mol-1 for water-'lipid-cyclohexane' compared to 3.2 kJ mol-1 for OPLS-AA cyclohexane and 2.4 kJ mol-1 for the modified OPLS-AA water-'lipid-cyclohexane'. We have also investigated the effect of different methods to combine parameters between the united-atom lipid force field and the united-atom protein force field ffgmx. In a widely used combination, the strength of interactions between hydrocarbon lipid tails and proteins is significantly overestimated, causing a decrease in the area per lipid and an increase in lipid ordering. Using straight combination rules improves the results. Combined, we suggest that using OPLS-AA together with the united-atom lipid force field implemented in GROMACS is a reasonable approach to membrane protein simulations. We also suggest that using partial volume information and free energies of transfer may help to improve the parameterization of lipid-protein interactions and point out the need for accurate experimental data to validate and improve force field descriptions of such interactions.

  3. Membrane protein simulations with a united-atom lipid and all-atom protein model: lipid-protein interactions, side chain transfer free energies and model proteins.

    PubMed

    Tieleman, D Peter; Maccallum, Justin L; Ash, Walter L; Kandt, Christian; Xu, Zhitao; Monticelli, Luca

    2006-07-19

    We have reparameterized the dihedral parameters in a commonly used united-atom lipid force field so that they can be used with the all-atom OPLS force field for proteins implemented in the molecular dynamics simulation software GROMACS. Simulations with this new combination give stable trajectories and sensible behaviour of both lipids and protein. We have calculated the free energy of transfer of amino acid side chains between water and 'lipid-cyclohexane', made of lipid force field methylene groups, as a hydrophobic mimic of the membrane interior, for both the OPLS-AA and a modified OPLS-AA force field which gives better hydration free energies under simulation conditions close to those preferred for the lipid force field. The average error is 4.3 kJ mol(-1) for water-'lipid-cyclohexane' compared to 3.2 kJ mol(-1) for OPLS-AA cyclohexane and 2.4 kJ mol(-1) for the modified OPLS-AA water-'lipid-cyclohexane'. We have also investigated the effect of different methods to combine parameters between the united-atom lipid force field and the united-atom protein force field ffgmx. In a widely used combination, the strength of interactions between hydrocarbon lipid tails and proteins is significantly overestimated, causing a decrease in the area per lipid and an increase in lipid ordering. Using straight combination rules improves the results. Combined, we suggest that using OPLS-AA together with the united-atom lipid force field implemented in GROMACS is a reasonable approach to membrane protein simulations. We also suggest that using partial volume information and free energies of transfer may help to improve the parameterization of lipid-protein interactions and point out the need for accurate experimental data to validate and improve force field descriptions of such interactions.

  4. A moderate change in temperature induces changes in fatty acid composition of storage and membrane lipids in a soil arthropod.

    PubMed

    van Dooremalen, Coby; Ellers, Jacintha

    2010-02-01

    A moderate change in ambient temperature can lead to vital physiological and biochemical adjustments in ectotherms, one of which is a change in fatty acid composition. When temperature decreases, the composition of membrane lipids (phospholipid fatty acids) is expected to become more unsaturated to be able to maintain homeoviscosity. Although different in function, storage lipids (triacylglycerol fatty acids) are expected to respond to temperature changes in a similar way. Age-specific differences, however, could influence this temperature response between different life stages. Here, we investigate if fatty acid composition of membrane and storage lipids responds similarly to temperature changes for two different life stages of Orchesella cincta. Juveniles and adults were cold acclimated (15 degrees C-->5 degrees C) for 28 days and then re-acclimated (5 degrees C-->15 degrees C) for another 28 days. We found adult membranes had a more unsaturated fatty acid composition than juveniles. Membrane lipids became more unsaturated during cold acclimation, and a reversed response occurred during warm acclimation. Membrane lipids, however, showed no warm acclimation, possibly due to the moderate temperature change. The ability to adjust storage lipid composition to moderate changes in ambient temperature may be an underestimated fitness component of temperature adaptation because fluidity of storage lipids permits accessibility of enzymes to energy reserves.

  5. Evidence for annexin A6-dependent plasma membrane remodelling of lipid domains

    PubMed Central

    Alvarez-Guaita, Anna; Vilà de Muga, Sandra; Owen, Dylan M; Williamson, David; Magenau, Astrid; García-Melero, Ana; Reverter, Meritxell; Hoque, Monira; Cairns, Rose; Cornely, Rhea; Tebar, Francesc; Grewal, Thomas; Gaus, Katharina; Ayala-Sanmartín, Jesús; Enrich, Carlos; Rentero, Carles

    2015-01-01

    Background and Purpose Annexin A6 (AnxA6) is a calcium-dependent phospholipid-binding protein that can be recruited to the plasma membrane to function as a scaffolding protein to regulate signal complex formation, endo- and exocytic pathways as well as distribution of cellular cholesterol. Here, we have investigated how AnxA6 influences the membrane order. Experimental Approach We used Laurdan and di-4-ANEPPDHQ staining in (i) artificial membranes; (ii) live cells to investigate membrane packing and ordered lipid phases; and (iii) a super-resolution imaging (photoactivated localization microscopy, PALM) and Ripley's K second-order point pattern analysis approach to assess how AnxA6 regulates plasma membrane order domains and protein clustering. Key Results In artificial membranes, purified AnxA6 induced a global increase in membrane order. However, confocal microscopy using di-4-ANEPPDHQ in live cells showed that cells expressing AnxA6, which reduces plasma membrane cholesterol levels and modifies the actin cytoskeleton meshwork, displayed a decrease in membrane order (∼15 and 30% in A431 and MEF cells respectively). PALM data from Lck10 and Src15 membrane raft/non-raft markers revealed that AnxA6 expression induced clustering of both raft and non-raft markers. Altered clustering of Lck10 and Src15 in cells expressing AnxA6 was also observed after cholesterol extraction with methyl-β-cyclodextrin or actin cytoskeleton disruption with latrunculin B. Conclusions and Implications AnxA6-induced plasma membrane remodelling indicated that elevated AnxA6 expression decreased membrane order through the regulation of cellular cholesterol homeostasis and the actin cytoskeleton. This study provides the first evidence from live cells that support current models of annexins as membrane organizers. PMID:25409976

  6. Elastic deformation and area per lipid of membranes: atomistic view from solid-state deuterium NMR spectroscopy.

    PubMed

    Kinnun, Jacob J; Mallikarjunaiah, K J; Petrache, Horia I; Brown, Michael F

    2015-01-01

    This article reviews the application of solid-state ²H nuclear magnetic resonance (NMR) spectroscopy for investigating the deformation of lipid bilayers at the atomistic level. For liquid-crystalline membranes, the average structure is manifested by the segmental order parameters (SCD) of the lipids. Solid-state ²H NMR yields observables directly related to the stress field of the lipid bilayer. The extent to which lipid bilayers are deformed by osmotic pressure is integral to how lipid-protein interactions affect membrane functions. Calculations of the average area per lipid and related structural properties are pertinent to bilayer remodeling and molecular dynamics (MD) simulations of membranes. To establish structural quantities, such as area per lipid and volumetric bilayer thickness, a mean-torque analysis of ²H NMR order parameters is applied. Osmotic stress is introduced by adding polymer solutions or by gravimetric dehydration, which are thermodynamically equivalent. Solid-state NMR studies of lipids under osmotic stress probe membrane interactions involving collective bilayer undulations, order-director fluctuations, and lipid molecular protrusions. Removal of water yields a reduction of the mean area per lipid, with a corresponding increase in volumetric bilayer thickness, by up to 20% in the liquid-crystalline state. Hydrophobic mismatch can shift protein states involving mechanosensation, transport, and molecular recognition by G-protein-coupled receptors. Measurements of the order parameters versus osmotic pressure yield the elastic area compressibility modulus and the corresponding bilayer thickness at an atomistic level. Solid-state ²H NMR thus reveals how membrane deformation can affect protein conformational changes within the stress field of the lipid bilayer.

  7. Cholesterol as a factor regulating the influence of natural (PAF and lysoPAF) vs synthetic (ED) ether lipids on model lipid membranes.

    PubMed

    Flasiński, Michał; Wydro, Paweł; Hąc-Wydro, Katarzyna; Dynarowicz-Łątka, Patrycja

    2013-11-01

    In this work we have performed a comparative study on the effect of antineoplastic ether lipid-edelfosine (ED), its natural analogs - Platelet Activating Factor (PAF) and its precursor (lyso-PAF), both lacking anticancer properties, on cholesterol/phosphatidylcholine (Chol/PC) monolayers, serving as model membranes. Since all the above ether lipids are membrane active, it can be expected that their effect on membranes may differentiate their biological activity. Our investigations were aimed at studying potential relationship of the effect of ED, PAF and lyso-PAF on model membranes, differing in condensation. We have modified molecular packing of Chol/PC model systems either by increasing the level of sterol in the system or changing the structure of PC, while keeping the same sterol content. Additionally, we have performed a detailed comparison of the miscibility of ED, PAF and lyso-PAF with various membrane lipids. The collected data evidenced that all the investigated ether lipids influence Chol/PC films in the same way; however, in a different magnitude. Moreover, the interactions of ED, PAF and lyso-PAF with model membranes were the strongest at the highest level of sterol in the system. A thorough analysis of the obtained results has proved that the effect of the investigated ether lipids on membranes is not dependent on the condensation of the system, but it is strongly determined by the concentration of cholesterol. Since ED was found to interact with model membranes stronger than PAF and lyso-PAF, we have suggested that this fact may contribute to differences in cytotoxicity of these compounds.

  8. Electrostatic properties of membrane lipids coupled to metarhodopsin II formation in visual transduction.

    PubMed

    Wang, Yin; Botelho, Ana Vitória; Martinez, Gary V; Brown, Michael F

    2002-07-03

    Changes in lipid composition have recently been shown to exert appreciable influences on the activities of membrane-bound proteins and peptides. We tested the hypothesis that the conformational states of rhodopsin linked to visual signal transduction are related to biophysical properties of the membrane lipid bilayer. For bovine rhodopsin, the meta I-meta II conformational transition was studied in egg phosphatidylcholine (PC) recombinants versus the native rod outer segment (ROS) membranes by means of flash photolysis. Formation of metarhodopsin II was observed by the change in absorbance at 478 nm after a single actinic flash was delivered to the sample. The meta I/meta II ratio was investigated as a function of both temperature and pH. The data clearly demonstrated thermodynamic reversibility of the transition for both the egg PC recombinants and the native ROS membranes. A significant shift of the apparent pK(a) for the acid-base equilibrium to lower values was evident in the egg PC recombinant, with little meta II produced under physiological conditions. Calculations of the membrane surface pH using a Poisson-Boltzmann model suggested the free energies of the meta I and meta II states were significantly affected by electrostatic properties of the bilayer lipids. In the ROS membranes, phosphatidylserine (PS) is needed for full formation of meta II, in combination with phosphatidylethanolamine (PE) and polyunsaturated docosahexaenoic acid (DHA; 22:6omega3) chains. We propose that the PS surface potential leads to an accumulation of hydronium ions, H(3)O(+), in the electrical double layer, which drive the reaction together with the large negative spontaneous curvature (H(0)) conferred by PE plus DHA chains. The elastic stress/strain of the bilayer arises from an interplay of the approximately zero H(0) from PS and the negative H(0) due to the PE headgroups and polyunsaturated chains. The lipid influences are further explained in terms of matching of the bilayer

  9. MODEL AND CELL MEMBRANE PARTITIONING OF PERFLUOROOCTANESULFONATE IS INDEPENDENT OF THE LIPID CHAIN LENGTH

    PubMed Central

    Xie, Wei; Ludewig, Gabriele; Wang, Kai; Lehmler, Hans-Joachim

    2009-01-01

    Perfluorooctanesulfonic acid (PFOS) is a persistent environmental pollutant that may cause adverse health effects in humans and animals by interacting with and disturbing of the normal properties of biological lipid assemblies. To gain further insights into these interactions, we investigated the effect of PFOS potassium salt on dimyristoyl- (DMPC), dipalmitoyl- (DPPC) and distearoylphosphatidylcholine (DSPC) model membranes using fluorescence anisotropy measurements and differential scanning calorimetry (DSC) and on the cell membrane of HL-60 human leukemia cells and freshly isolated rat alveolar macrophages using fluorescence anisotropy measurements. PFOS caused a concentration-dependent decrease of the main phase transition temperature (Tm) and an increased peak width (ΔTw) in both the fluorescence anisotropy and the DSC experiments, with a rank order DMPC > DPPC > DSPC. PFOS caused a fluidization of the gel phase of all phosphatidylcholines investigated, but had the opposite effect on the liquid crystalline phase. The apparent partition coefficients of PFOS between the phosphatidylcholine bilayer and the bulk aqueous phase were largely independent of the phosphatidylcholine chain length and ranged from 4.4 × 104 to 8.8 × 104. PFOS also significantly increased the fluidity of membranes of cells. These findings suggest that PFOS readily partitions into lipid assemblies, independent of their composition, and may cause adverse biological effects by altering their fluidity in a manner that depends on the membrane cooperativity and state (e.g., gel versus liquid crystalline phase) of the lipid assembly. PMID:19932010

  10. Direct observation of brownian motion of lipids in a membrane.

    PubMed Central

    Lee, G M; Ishihara, A; Jacobson, K A

    1991-01-01

    Nanovid microscopy, which uses 30- to 40-nm colloidal gold probes combined with video-enhanced contrast, can be used to examine random and directed movements of individual molecules in the plasma membrane of living cells. To validate the technique in a model system, the movements of lipid molecules were followed in a supported, planar bilayer containing fluorescein-conjugated phosphatidylethanolamine (Fl-PtdEtn) labeled with 30-nm gold anti-fluorescein (anti-Fl). Multivalent gold probes were prepared by conjugating only anti-Fl to the gold. Paucivalent probes were prepared by mixing an irrelevant antibody with the anti-Fl prior to conjugation. The membrane-bound gold particles moved in random patterns that were indistinguishable from those produced by computer simulations of two-dimensional random motion. The multivalent gold probes had an average lateral diffusion coefficient (D) of 0.26 x 10(-8) cm2/sec, and paucivalent probes had an average D of 0.73 x 10(-8) cm2/sec. Sixteen percent of the multivalent and 50% of the paucivalent probes had values for D in excess of 0.6 x 10(-8) cm2/sec, which, after allowance for stochastic variation, are consistent with the D of 1.3 x 10(-8) cm2/sec measured by fluorescence recovery after photobleaching of Fl-PtdEtn in the planar bilayer. The effect of valency on diffusion suggests that the multivalent gold binds several lipids forming a disk up to 30-40 nm in diameter, resulting in reduced diffusion with respect to the paucivalent gold, which binds one or a very few lipids. Provided the valency of the gold probe is considered in the interpretation of the results. Nanovid microscopy is a valid method for analyzing the movements of single or small groups of molecules within membranes. Images PMID:1712486

  11. New fluorescent octadecapentaenoic acids as probes of lipid membranes and protein-lipid interactions.

    PubMed Central

    Mateo, C R; Souto, A A; Amat-Guerri, F; Acuña, A U

    1996-01-01

    The chemical and spectroscopic properties of the new fluorescent acids all(E)-8, 10, 12, 14, 16-octadecapentaenoic acid (t-COPA) and its (8Z)-isomer (c-COPA) have been characterized in solvents of different polarity, synthetic lipid bilayers, and lipid/protein systems. These compounds are reasonably photostable in solution, present an intense UV absorption band (epsilon(350 nm) approximately 10(5) M(-1) cm(-1)) strongly overlapped by tryptophan fluorescence and their emission, centered at 470 nm, is strongly polarized (r(O) = 0.385 +/- 0.005) and decays with a major component (85%) of lifetime 23 ns and a faster minor one of lifetime 2 ns (D,L-alpha-dimyristoylphosphatidylcholine (DMPC), 15 degrees C). Both COPA isomers incorporate readily into vesicles and membranes (K(p) approximately 10(6)) and align parallel to the lipids. t-COPA distributes homogeneously between gel and fluid lipid domains and the changes in polarization accurately reflect the lipid T(m) values. From the decay of the fluorescence anisotropy in spherical bilayers of DMPC and POPC it is shown that t-COPA also correctly reflects the lipid order parameters, determined by 2H NMR techniques. Resonance energy transfer from tryptophan to the bound pentaenoic acid in serum albumin in solution, and from the tryptophan residues of gramicidin in lipid bilayers also containing the pentaenoic acid, show that this probe is a useful acceptor of protein tryptophan excitation, with R(O) values of 30-34 A. Images FIGURE 10 PMID:8889194

  12. Snake Cytotoxins Bind to Membranes via Interactions with Phosphatidylserine Head Groups of Lipids

    PubMed Central

    Konshina, Anastasia G.; Boldyrev, Ivan A.; Utkin, Yuri N.; Omel'kov, Anton V.; Efremov, Roman G.

    2011-01-01

    The major representatives of Elapidae snake venom, cytotoxins (CTs), share similar three-fingered fold and exert diverse range of biological activities against various cell types. CT-induced cell death starts from the membrane recognition process, whose molecular details remain unclear. It is known, however, that the presence of anionic lipids in cell membranes is one of the important factors determining CT-membrane binding. In this work, we therefore investigated specific interactions between one of the most abundant of such lipids, phosphatidylserine (PS), and CT 4 of Naja kaouthia using a combined, experimental and modeling, approach. It was shown that incorporation of PS into zwitterionic liposomes greatly increased the membrane-damaging activity of CT 4 measured by the release of the liposome-entrapped calcein fluorescent dye. The CT-induced leakage rate depends on the PS concentration with a maximum at approximately 20% PS. Interestingly, the effects observed for PS were much more pronounced than those measured for another anionic lipid, sulfatide. To delineate the potential PS binding sites on CT 4 and estimate their relative affinities, a series of computer simulations was performed for the systems containing the head group of PS and different spatial models of CT 4 in aqueous solution and in an implicit membrane. This was done using an original hybrid computational protocol implementing docking, Monte Carlo and molecular dynamics simulations. As a result, at least three putative PS-binding sites with different affinities to PS molecule were delineated. Being located in different parts of the CT molecule, these anion-binding sites can potentially facilitate and modulate the multi-step process of the toxin insertion into lipid bilayers. This feature together with the diverse binding affinities of the sites to a wide variety of anionic targets on the membrane surface appears to be functionally meaningful and may adjust CT action against different types of

  13. Membrane evaporator/sublimator investigation

    NASA Technical Reports Server (NTRS)

    Elam, J.; Ruder, J.; Strumpf, H.

    1974-01-01

    Data are presented on a new evaporator/sublimator concept using a hollow fiber membrane unit with a high permeability to liquid water. The aim of the program was to obtain a more reliable, lightweight and simpler Extra Vehicular Life Support System (EVLSS) cooling concept than is currently being used.

  14. Influences of the Structure of Lipids on Thermal Stability of Lipid Membranes

    NASA Astrophysics Data System (ADS)

    Hai, Nan-Nan; Zhou, Xin; Li, Ming

    2015-08-01

    The binding free energy (BFE) of lipid to lipid bilayer is a critical factor to determine the thermal or mechanical stability of the bilayer. Although the molecular structure of lipids has significant impacts on BFE of the lipid, there lacks a systematic study on this issue. In this paper we use coarse-grained molecular dynamics simulation to investigate this problem for several typical phospholipids. We find that both the tail length and tail unsaturation can significantly affect the BFE of lipids but in opposite way, namely, BFE decreases linearly with increasing length, but increases linearly with addition of unsaturated bonds. Inspired by the specific structure of cholesterol which is a crucial component of biomembrane, we also find that introduction of carbo-ring-like structures to the lipid tail or to the bilayer may greatly enhance the stability of the bilayer. Our simulation also shows that temperature can influence the bilayer stability and this effect can be significant when the bilayer undergoes phase transition. These results may be helpful to the design of liposome or other self-assembled lipid systems. Support by the National Natural Science Foundation of China under Grant Nos. 91027046 and 11105218.

  15. Distribution of membrane lipids of planktonic Crenarchaeota in the Arabian Sea.

    PubMed

    Sinninghe Damsté, Jaap S; Rijpstra, W Irene C; Hopmans, Ellen C; Prahl, Fredrick G; Wakeham, Stuart G; Schouten, Stefan

    2002-06-01

    Intact core tetraether membrane lipids of marine planktonic Crenarchaeota were quantified in water column-suspended particulate matter obtained from four depth intervals ( approximately 70, 500, 1,000 and 1,500 m) at seven stations in the northwestern Arabian Sea to investigate the distribution of the organisms at various depths. Maximum concentrations generally occurred at 500 m, near the top of the oxygen minimum zone, and the concentrations at this depth were, in most cases, slightly higher than those in surface waters. In contrast, lipids derived from eukaryotes (cholesterol) and from eukaryotes and bacteria (fatty acids) were at their highest concentrations in surface waters. This indicates that these crenarchaeotes are not restricted to the photic zone of the ocean, which is consistent with the results of recent molecular biological studies. Since the Arabian Sea has a strong oxygen minimum zone between 100 and 1,000 m, with minimum oxygen levels of <1 microM, the abundance of crenarchaeotal membrane lipids at 500 m suggests that planktonic Crenarchaeota are probably facultative anaerobes. The cell numbers we calculated from the concentrations of membrane lipids are similar to those reported for the Central Pacific Ocean, supporting the recent estimation of M. B. Karner, E. F. DeLong, and D. M. Karl ( Nature 409:507-510, 2001) that the world's oceans contain ca. 10(28) cells of planktonic Crenarchaeota.

  16. A Hybrid Model for Erythrocyte Membrane: A Single Unit of Protein Network Coupled with Lipid Bilayer

    PubMed Central

    Zhu, Qiang; Vera, Carlos; Asaro, Robert J.; Sche, Paul; Sung, L. Amy

    2007-01-01

    To investigate the nanomechanics of the erythrocyte membrane we developed a hybrid model that couples the actin-spectrin network to the lipid bilayer. This model features a Fourier space Brownian dynamics model of the bilayer, a Brownian dynamics model of the actin protofilament, and a modified wormlike-chain model of the spectrin (including a cable-dynamics model to predict the oscillation in tension). This model enables us to predict the nanomechanics of single or multiple units of the protein network, the lipid bilayer, and the effect of their interactions. The present work is focused on the attitude of the actin protofilament at the equilibrium states coupled with the elevations of the lipid bilayer through their primary linkage at the suspension complex in deformations. Two different actin-spectrin junctions are considered at the junctional complex. With a point-attachment junction, large pitch angles and bifurcation of yaw angles are predicted. Thermal fluctuations at bifurcation may lead to mode-switching, which may affect the network and the physiological performance of the membrane. In contrast, with a wrap-around junction, pitch angles remain small, and the occurrence of bifurcation is greatly reduced. These simulations suggest the importance of three-dimensional molecular junctions and the lipid bilayer/protein network coupling on cell membrane mechanics. PMID:17449663

  17. Cryoprotection of Lipid Membranes for High-Resolution Solid-State NMR Studies of Membrane Peptides and Proteins at Low Temperature

    PubMed Central

    Lee, Myungwoon; Hong, Mei

    2014-01-01

    Solid-state NMR spectra of membrane proteins often show significant line broadening at cryogenic temperatures. Here we investigate the effects of several cryoprotectants to preserve the spectral resolution of lipid membranes and membrane peptides at temperatures down to ~200 K. Trehalose, glycerol, dimethylsulfoxide (DMSO), dimethylformamide (DMF), and polyethylene glycol (PEG), were chosen. These compounds are commonly used in protein crystallography and cryobiology. 13C and 1H MAS spectra of several types of lipid membranes show that DMSO provides the best resolution enhancement over unprotected membranes and also best retards ice formation at low temperature. DMF and PEG-400 show slightly weaker cryoprotection, while glycerol and trehalose neither prevent membrane line broadening nor prevent ice formation under the conditions of our study. Neutral saturated-chain phospholipids are the most amenable to cryoprotection, whereas negatively charged and unsaturated lipids attenuate cryoprotection. 13C-1H dipolar couplings and 31P chemical shift anisotropies indicate that high spectral resolution at low temperature is correlated with stronger immobilization of the lipids at high temperature, indicating that line narrowing results from reduction of the conformational space sampled by the lipid molecules at high temperature. DMSO selectively narrowed the linewidths of the most disordered residues in the influenza M2 transmembrane peptide, while residues that exhibit narrow linewidths in the unprotected membrane are less impacted. A relatively rigid β-hairpin antimicrobial peptide, PG-1, showed a linewidth increase of ~0.5 ppm over a ~70 K temperature drop both with and without cryoprotection. Finally, a short-chain saturated lipid, DLPE, exhibits excellent linewidths, suggesting that it may be a good medium for membrane protein structure determination. The three best cryoprotectants found in this work – DMSO, PEG, and DMF - should be useful for low

  18. Cryoprotection of lipid membranes for high-resolution solid-state NMR studies of membrane peptides and proteins at low temperature.

    PubMed

    Lee, Myungwoon; Hong, Mei

    2014-08-01

    Solid-state NMR spectra of membrane proteins often show significant line broadening at cryogenic temperatures. Here we investigate the effects of several cryoprotectants to preserve the spectral resolution of lipid membranes and membrane peptides at temperatures down to ~200 K. Trehalose, glycerol, dimethylsulfoxide (DMSO), dimethylformamide (DMF), and polyethylene glycol (PEG), were chosen. These compounds are commonly used in protein crystallography and cryobiology. 13C and 1H magic-angle-spinning spectra of several types of lipid membranes show that DMSO provides the best resolution enhancement over unprotected membranes and also best retards ice formation at low temperature. DMF and PEG-400 show slightly weaker cryoprotection, while glycerol and trehalose neither prevent membrane line broadening nor prevent ice formation under the conditions of our study. Neutral saturated-chain phospholipids are the most amenable to cryoprotection, whereas negatively charged and unsaturated lipids attenuate cryoprotection. 13C-1H dipolar couplings and 31P chemical shift anisotropies indicate that high spectral resolution at low temperature is correlated with stronger immobilization of the lipids at high temperature, indicating that line narrowing results from reduction of the conformational space sampled by the lipid molecules at high temperature. DMSO selectively narrowed the linewidths of the most disordered residues in the influenza M2 transmembrane peptide, while residues that exhibit narrow linewidths in the unprotected membrane are less impacted. A relatively rigid β-hairpin antimicrobial peptide, PG-1, showed a linewidth increase of ~0.5 ppm over a ~70 K temperature drop both with and without cryoprotection. Finally, a short-chain saturated lipid, DLPE, exhibits excellent linewidths, suggesting that it may be a good medium for membrane protein structure determination. The three best cryoprotectants found in this work-DMSO, PEG, and DMF-should be useful for low

  19. Effect of Pulsed Electric Field on Membrane Lipids and Oxidative Injury of Salmonella typhimurium

    PubMed Central

    Yun, Ou; Zeng, Xin-An; Brennan, Charles S.; Han, Zhong

    2016-01-01

    Salmonella typhimurium cells were subjected to pulsed electric field (PEF) treatment at 25 kV/cm for 0–4 ms to investigate the effect of PEF on the cytoplasmic membrane lipids and oxidative injury of cells. Results indicated that PEF treatment induced a decrease of membrane fluidity of Salmonella typhimurium (S. typhimuriumi), possibly due to the alterations of fatty acid biosynthesis-associated gene expressions (down-regulation of cfa and fabA gene expressions and the up-regulation of fabD gene expression), which, in turn, modified the composition of membrane lipid (decrease in the content ratio of unsaturated fatty acids to saturated fatty acids). In addition, oxidative injury induced by PEF treatment was associated with an increase in the content of malondialdehyde. The up-regulation of cytochrome bo oxidase gene expressions (cyoA, cyoB, and cyoC) indicated that membrane damage was induced by PEF treatment, which was related to the repairing mechanism of alleviating the oxidative injury caused by PEF treatment. Based on these results, we achieved better understanding of microbial injury induced by PEF, suggesting that micro-organisms tend to decrease membrane fluidity in response to PEF treatment and, thus, a greater membrane fluidity might improve the efficiency of PEF treatment to inactivate micro-organisms. PMID:27556460

  20. Research on the Changes to the Lipid/Polymer Membrane Used in the Acidic Bitterness Sensor Caused by Preconditioning

    PubMed Central

    Harada, Yuhei; Noda, Junpei; Yatabe, Rui; Ikezaki, Hidekazu; Toko, Kiyoshi

    2016-01-01

    A taste sensor that uses lipid/polymer membranes can evaluate aftertastes felt by humans using Change in membrane Potential caused by Adsorption (CPA) measurements. The sensor membrane for evaluating bitterness, which is caused by acidic bitter substances such as iso-alpha acid contained in beer, needs an immersion process in monosodium glutamate (MSG) solution, called “MSG preconditioning”. However, what happens to the lipid/polymer membrane during MSG preconditioning is not clear. Therefore, we carried out three experiments to investigate the changes in the lipid/polymer membrane caused by the MSG preconditioning, i.e., measurements of the taste sensor, measurements of the amount of the bitterness substance adsorbed onto the membrane and measurements of the contact angle of the membrane surface. The CPA values increased as the preconditioning process progressed, and became stable after 3 d of preconditioning. The response potentials to the reference solution showed the same tendency of the CPA value change during the preconditioning period. The contact angle of the lipid/polymer membrane surface decreased after 7 d of MSG preconditioning; in short, the surface of the lipid/polymer membrane became hydrophilic during MSG preconditioning. The amount of adsorbed iso-alpha acid was increased until 5 d preconditioning, and then it decreased. In this study, we revealed that the CPA values increased with the progress of MSG preconditioning in spite of the decrease of the amount of iso-alpha acid adsorbed onto the lipid/polymer membrane, and it was indicated that the CPA values increase because the sensor sensitivity was improved by the MSG preconditioning. PMID:26891299

  1. Mechanism of interaction of monovalent ions with phosphatidylcholine lipid membranes.

    PubMed

    Vácha, Robert; Jurkiewicz, Piotr; Petrov, Michal; Berkowitz, Max L; Böckmann, Rainer A; Barucha-Kraszewska, Justyna; Hof, Martin; Jungwirth, Pavel

    2010-07-29

    Interactions of different anions with phospholipid membranes in aqueous salt solutions were investigated by molecular dynamics simulations and fluorescence solvent relaxation measurements. Both approaches indicate that the anion-membrane interaction increases with the size and softness of the anion. Calculations show that iodide exhibits a genuine affinity for the membrane, which is due to its pairing with the choline group and its propensity for the nonpolar region of the acyl chains, the latter being enhanced in polarizable calculations showing that the iodide number density profile is expanded toward the glycerol level. Solvent relaxation measurements using Laurdan confirm the influence of large soft ions on the membrane organization at the glycerol level. In contrast, chloride exhibits a peak at the membrane surface only in the presence of a surface-attracted cation, such as sodium but not potassium, suggesting that this behavior is merely a counterion effect.

  2. Reticulated lipid probe fluorescence reveals MDCK cell apical membrane topography.

    PubMed

    Colarusso, Pina; Spring, Kenneth R

    2002-02-01

    High spatial resolution confocal microscopy of young MDCK cells stained with the lipophilic probe 1,1'-dihexadecyl-3,3,3',3'- tetramethylindocarbocyanine perchlorate (DiIC(16)) revealed a reticulated fluorescence pattern on the apical membrane. DiIC(16) was delivered as crystals to live cells to minimize possible solvent perturbations of the membrane lipids. The ratio of the integrated fluorescence intensities in the bright versus dim regions was 1.6 +/- 0.1 (n = 13). Deconvolved images of the cells were consistent with exclusive plasma membrane staining. Multi-spectral and fluorescence anisotropy microscopy did not reveal differences between bright and dim regions. Bright regions coincided with microvilli and microridges observed by differential interference contrast microscopy and were stable for several minutes. Fluorescence recovery after photobleaching yielded similar diffusion coefficients (pooled D = 1.5 +/- 0.6 x 10(-9) cm(2)/s, n = 40) for both bright and dim regions. Line fluorescence recovery after photobleaching showed that the reticulated pattern was maintained as the fluorescence recovered in the bleached areas. Cytochalasin D did not affect the staining pattern, but the pattern was eliminated by cholesterol depletion with methyl-beta-cyclodextrin. We conclude that the reticulated fluorescence pattern was caused by increased optical path lengths through the microvilli and microridges compared with the flat areas on the apical membrane.

  3. Reticulated lipid probe fluorescence reveals MDCK cell apical membrane topography.

    PubMed Central

    Colarusso, Pina; Spring, Kenneth R

    2002-01-01

    High spatial resolution confocal microscopy of young MDCK cells stained with the lipophilic probe 1,1'-dihexadecyl-3,3,3',3'- tetramethylindocarbocyanine perchlorate (DiIC(16)) revealed a reticulated fluorescence pattern on the apical membrane. DiIC(16) was delivered as crystals to live cells to minimize possible solvent perturbations of the membrane lipids. The ratio of the integrated fluorescence intensities in the bright versus dim regions was 1.6 +/- 0.1 (n = 13). Deconvolved images of the cells were consistent with exclusive plasma membrane staining. Multi-spectral and fluorescence anisotropy microscopy did not reveal differences between bright and dim regions. Bright regions coincided with microvilli and microridges observed by differential interference contrast microscopy and were stable for several minutes. Fluorescence recovery after photobleaching yielded similar diffusion coefficients (pooled D = 1.5 +/- 0.6 x 10(-9) cm(2)/s, n = 40) for both bright and dim regions. Line fluorescence recovery after photobleaching showed that the reticulated pattern was maintained as the fluorescence recovered in the bleached areas. Cytochalasin D did not affect the staining pattern, but the pattern was eliminated by cholesterol depletion with methyl-beta-cyclodextrin. We conclude that the reticulated fluorescence pattern was caused by increased optical path lengths through the microvilli and microridges compared with the flat areas on the apical membrane. PMID:11806917

  4. Detergent interaction with tethered bilayer lipid membranes for protein reconstitution

    NASA Astrophysics Data System (ADS)

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

    2009-03-01

    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.

  5. Lipids that determine detergent resistance of MDCK cell membrane fractions.

    PubMed

    Manni, Marco M; Cano, Ainara; Alonso, Cristina; Goñi, Félix M

    2015-10-01

    A comparative lipidomic study has been performed of whole Madin-Darby canine kidney epithelial cells and of the detergent-resistant membrane fraction (DRM) obtained after treating the cells with the non-ionic detergent Triton X-100. The DRM were isolated following a standard procedure that is extensively used in cell biology studies. Significant differences were found in the lipid composition of the whole cells and of DRM. The latter were enriched in all the analyzed sphingolipid classes: sphingomyelins, ceramides and hexosylceramides. Diacylglycerols were also preferentially found in DRM. The detergent-resistant fraction was also enriched in saturated over unsaturated fatty acyl chains, and in sn-1 acyl chains containing 16 carbon atoms, over the longer and shorter ones. The glycerophospholipid species phosphatidylethanolamines and phosphatidylinositols, that were mainly unsaturated, did not show a preference for DRM. Phosphatidylcholines were an intermediate case: the saturated, but not the unsaturated species were found preferentially in DRM. The question remains on whether these DRM, recovered from detergent-membrane mixtures by floatation over a sucrose gradient, really correspond to membrane domains existing in the cell membrane prior to detergent treatment.

  6. Effects of seaweed sterols fucosterol and desmosterol on lipid membranes.

    PubMed

    Mouritsen, Ole G; Bagatolli, Luis A; Duelund, Lars; Garvik, Olav; Ipsen, John H; Simonsen, Adam Cohen

    2017-03-30

    Higher sterols are universally present in large amounts (20-30%) in the plasma membranes of all eukaryotes whereas they are universally absent in prokaryotes. It is remarkable that each kingdom of the eukaryotes has chosen, during the course of evolution, its preferred sterol: cholesterol in animals, ergosterol in fungi and yeast, phytosterols in higher plants, and e.g., fucosterol and desmosterol in algae. The question arises as to which specific properties do sterols impart to membranes and to which extent do these properties differ among the different sterols. Using a range of biophysical techniques, including calorimetry, fluorescence microscopy, vesicle-fluctuation analysis, and atomic force microscopy, we have found that fucosterol and desmosterol, found in red and brown macroalgae (seaweeds), similar to cholesterol support liquid-ordered membrane phases and induce coexistence between liquid-ordered and liquid-disordered domains in lipid bilayers. Fucosterol and desmosterol induce acyl-chain order in liquid membranes, but less effectively than cholesterol and ergosterol in the order: cholesterol>ergosterol>desmosterol>fucosterol, possibly reflecting the different molecular structure of the sterols at the hydrocarbon tail.

  7. Probing the importance of lipid diversity in cell membranes via molecular simulation.

    PubMed

    Khakbaz, Pouyan; Klauda, Jeffery B

    2015-11-01

    Lipid membranes in prokaryotes and eukaryotes have a wide array of lipids that are necessary for proper membrane structure and function. In this paper, an introduction to lipid diversity in biology and a mini-review on how molecular simulations have been used to model biological membranes (primarily limited to one to three lipid types in most simulation-based models) is provided, which motivates the use of all-atom molecular dynamics (MD) simulations to study the effect of lipid diversity on properties of realistic membrane models of prokaryotes and eukaryotes. As an example, cytoplasmic membrane models of Escherichia coli were developed at different stages of the colony growth cycle (early-log, mid-log, stationary and overnight). The main difference between lipid compositions at each stage was the concentration of a cyclopropane-containing moiety on the sn-2 lipid acyl chain (cyC17:0). Triplicate MD simulations for each stage were run for 300 ns to study the influence of lipid diversity on the surface area per lipid, area compressibility modulus, deuterium order parameters, and electron density profiles. The overnight stage (also known as the death stage) had the highest average surface area per lipid, highest rigidity, and lowest bilayer thickness compare to other stages of E. coli cytoplasmic membrane. Although bilayer thickness did depend on the growth stage, the changes between these were small suggesting that the hydrophobic core of transmembrane proteins fit well with the membrane in all growth stages. Although it is still common practise in MD simulations of membrane proteins to use simple one- or two-component membranes, it can be important to use diverse lipid model membranes when membrane protein structure and function are influenced by changes in lipid membrane composition.

  8. Proving lipid rafts exist: membrane domains in the prokaryote Borrelia burgdorferi have the same properties as eukaryotic lipid rafts.

    PubMed

    LaRocca, Timothy J; Pathak, Priyadarshini; Chiantia, Salvatore; Toledo, Alvaro; Silvius, John R; Benach, Jorge L; London, Erwin

    2013-01-01

    Lipid rafts in eukaryotic cells are sphingolipid and cholesterol-rich, ordered membrane regions that have been postulated to play roles in many membrane functions, including infection. We previously demonstrated the existence of cholesterol-lipid-rich domains in membranes of the prokaryote, B. burgdorferi, the causative agent of Lyme disease [LaRocca et al. (2010) Cell Host & Microbe 8, 331-342]. Here, we show that these prokaryote membrane domains have the hallmarks of eukaryotic lipid rafts, despite lacking sphingolipids. Substitution experiments replacing cholesterol lipids with a set of sterols, ranging from strongly raft-promoting to raft-inhibiting when mixed with eukaryotic sphingolipids, showed that sterols that can support ordered domain formation are both necessary and sufficient for formation of B. burgdorferi membrane domains that can be detected by transmission electron microscopy or in living organisms by Förster resonance energy transfer (FRET). Raft-supporting sterols were also necessary and sufficient for formation of high amounts of detergent resistant membranes from B. burgdorferi. Furthermore, having saturated acyl chains was required for a biotinylated lipid to associate with the cholesterol-lipid-rich domains in B. burgdorferi, another characteristic identical to that of eukaryotic lipid rafts. Sterols supporting ordered domain formation were also necessary and sufficient to maintain B. burgdorferi membrane integrity, and thus critical to the life of the organism. These findings provide compelling evidence for the existence of lipid rafts and show that the same principles of lipid raft formation apply to prokaryotes and eukaryotes despite marked differences in their lipid compositions.

  9. Structural basis for the transcriptional regulation of membrane lipid homeostasis

    SciTech Connect

    Miller, Darcie J.; Zhang, Yong-Mei; Subramanian, Chitra; Rock, Charles O.; White, Stephen W.

    2010-11-09

    DesT is a transcriptional repressor that regulates the genes that control the unsaturated:saturated fatty acid ratio available for membrane lipid synthesis. DesT bound to unsaturated acyl-CoA has a high affinity for its cognate palindromic DNA-binding site, whereas DesT bound to saturated acyl-CoA does not bind this site. Structural analyses of the DesT-oleoyl-CoA-DNA and DesT-palmitoyl-CoA complexes reveal that acyl chain shape directly influences the packing of hydrophobic core residues within the DesT ligand-binding domain. These changes are propagated to the paired DNA-binding domains via conformational changes to modulate DNA binding. These structural interpretations are supported by the in vitro and in vivo characterization of site-directed mutants. The regulation of DesT by the unsaturated:saturated ratio of acyl chains rather than the concentration of a single ligand is a paradigm for understanding transcriptional regulation of membrane lipid homeostasis.

  10. Interaction of aldehydes derived from lipid peroxidation and membrane proteins

    PubMed Central

    Pizzimenti, Stefania; Ciamporcero, Eric; Daga, Martina; Pettazzoni, Piergiorgio; Arcaro, Alessia; Cetrangolo, Gianpaolo; Minelli, Rosalba; Dianzani, Chiara; Lepore, Alessio; Gentile, Fabrizio; Barrera, Giuseppina

    2013-01-01

    A great variety of compounds are formed during lipid peroxidation of polyunsaturated fatty acids of membrane phospholipids. Among them, bioactive aldehydes, such as 4-hydroxyalkenals, malondialdehyde (MDA) and acrolein, have received particular attention since they have been considered as toxic messengers that can propagate and amplify oxidative injury. In the 4-hydroxyalkenal class, 4-hydroxy-2-nonenal (HNE) is the most intensively studied aldehyde, in relation not only to its toxic function, but also to its physiological role. Indeed, HNE can be found at low concentrations in human tissues and plasma and participates in the control of biological processes, such as signal transduction, cell proliferation, and differentiation. Moreover, at low doses, HNE exerts an anti-cancer effect, by inhibiting cell proliferation, angiogenesis, cell adhesion and by inducing differentiation and/or apoptosis in various tumor cell lines. It is very likely that a substantial fraction of the effects observed in cellular responses, induced by HNE and related aldehydes, be mediated by their interaction with proteins, resulting in the formation of covalent adducts or in the modulation of their expression and/or activity. In this review we focus on membrane proteins affected by lipid peroxidation-derived aldehydes, under physiological and pathological conditions. PMID:24027536

  11. Tight binding of NAP-22 with acidic membrane lipids.

    PubMed

    Maekawa, Shohei; Kobayashi, Yuumi; Morita, Mitsuhiro; Suzaki, Toshinobu

    2015-07-23

    Recovery of various signal transduction molecules in the detergent-resistant membrane microdomain (DRM) fraction suggests the importance of this region in cellular functions. Insolubility of the outer leaflet of DRM to the non-ionic detergent is ascribed to the tight association of cholesterol and sphingolipid. Since, poor localization of sphingolipid is observed in the inner leaflet, the physicochemical background of the insolubility of the inner leaflet is hence still an enigma. NAP-22 (also called BASP1 or CAP-23) is a neuron-enriched calmodulin-binding protein and one of the major proteins in the DRM of the neuronal cell membrane. A previous study showed the presence of several lipids in a NAP-22 fraction after the process of extraction and column chromatography. In this study, the effect of lipid extraction on NAP-22 was studied through native-gel electrophoresis, ultracentrifugation, and electron microscopic observation. The mobility of NAP-22 in native-PAGE was shifted from low to high after delipidation. Delipidated NAP-22 bound phosphatidylserine (PS), phosphatidylinosotol, and ganglioside. Some part of the mixture of PS and NAP-22 was recovered in the insoluble fraction after Triton X-100 treatment and the addition of cholesterol enhanced the amount of NAP-22 in the insoluble fraction.

  12. Vascular endothelial cell membranes differentiate between stretch and shear stress through transitions in their lipid phases.

    PubMed

    Yamamoto, Kimiko; Ando, Joji

    2015-10-01

    Vascular endothelial cells (ECs) respond to the hemodynamic forces stretch and shear stress by altering their morphology, functions, and gene expression. However, how they sense and differentiate between these two forces has remained unknown. Here we report that the plasma membrane itself differentiates between stretch and shear stress by undergoing transitions in its lipid phases. Uniaxial stretching and hypotonic swelling increased the lipid order of human pulmonary artery EC plasma membranes, thereby causing a transition from the liquid-disordered phase to the liquid-ordered phase in some areas, along with a decrease in membrane fluidity. In contrast, shear stress decreased the membrane lipid order and increased membrane fluidity. A similar increase in lipid order occurred when the artificial lipid bilayer membranes of giant unilamellar vesicles were stretched by hypotonic swelling, indicating that this is a physical phenomenon. The cholesterol content of EC plasma membranes significantly increased in response to stretch but clearly decreased in response to shear stress. Blocking these changes in the membrane lipid order by depleting membrane cholesterol with methyl-β-cyclodextrin or by adding cholesterol resulted in a marked inhibition of the EC response specific to stretch and shear stress, i.e., phosphorylation of PDGF receptors and phosphorylation of VEGF receptors, respectively. These findings indicate that EC plasma membranes differently respond to stretch and shear stress by changing their lipid order, fluidity, and cholesterol content in opposite directions and that these changes in membrane physical properties are involved in the mechanotransduction that activates membrane receptors specific to each force.

  13. Analysis of Membrane Lipids of Airborne Micro-Organisms

    NASA Technical Reports Server (NTRS)

    MacNaughton, Sarah

    2006-01-01

    A method of characterization of airborne micro-organisms in a given location involves (1) large-volume filtration of air onto glass-fiber filters; (2) accelerated extraction of membrane lipids of the collected micro-organisms by use of pressurized hot liquid; and (3) identification and quantitation of the lipids by use of gas chromatography and mass spectrometry. This method is suitable for use in both outdoor and indoor environments; for example, it can be used to measure airborne microbial contamination in buildings ("sick-building syndrome"). The classical approach to analysis of airborne micro-organisms is based on the growth of cultureable micro-organisms and does not provide an account of viable but noncultureable micro-organisms, which typically amount to more than 90 percent of the micro-organisms present. In contrast, the present method provides an account of all micro-organisms, including cultureable, noncultureable, aerobic, and anaerobic ones. The analysis of lipids according to this method makes it possible to estimate the number of viable airborne micro-organisms present in the sampled air and to obtain a quantitative profile of the general types of micro-organisms present along with some information about their physiological statuses.

  14. The role of sulfatide lipid domains in the membrane pore-forming activity of cobra cardiotoxin.

    PubMed

    Wu, Po-Long; Chiu, Chang-Ru; Huang, Wei-Ning; Wu, Wen-Guey

    2012-05-01

    Cobra CTX A3, the major cardiotoxin (CTX) from Naja atra, is a cytotoxic, basic β-sheet polypeptide that is known to induce a transient membrane leakage of cardiomyocytes through a sulfatide-dependent CTX membrane pore formation and internalization mechanism. The molecular specificity of CTX A3-sulfatide interaction at atomic levels has also been shown by both nuclear magnetic resonance (NMR) and X-ray diffraction techniques to reveal a role of CTX-induced sulfatide conformational changes for CTX A3 binding and dimer formation. In this study, we investigate the role of sulfatide lipid domains in CTX pore formation by various biophysical methods, including fluorescence imaging and atomic force microscopy, and suggest an important role of liquid-disordered (ld) and solid-ordered (so) phase boundary in lipid domains to facilitate the process. Fluorescence spectroscopic studies on the kinetics of membrane leakage and CTX oligomerization further reveal that, although most CTXs can oligomerize on membranes, only a small fraction of CTXs oligomerizations form leakage pores. We therefore suggest that CTX binding at the boundary between the so and so/ld phase coexistence sulfatide lipid domains could form effective pores to significantly enhance the CTX-induced membrane leakage of sulfatide-containing phosphatidylcholine vesicles. The model is consistent with our earlier observations that CTX may penetrate and lyse the bilayers into small aggregates at a lipid/protein molar ratio of about 20 in the ripple P(β)' phase of phosphatidylcholine bilayers and suggest a novel mechanism for the synergistic action of cobra secretary phospholipase A2 and CTXs.

  15. Renaturing Membrane Proteins in the Lipid Cubic Phase, a Nanoporous Membrane Mimetic

    PubMed Central

    Li, Dianfan; Caffrey, Martin

    2014-01-01

    Membrane proteins play vital roles in the life of the cell and are important therapeutic targets. Producing them in large quantities, pure and fully functional is a major challenge. Many promising projects end when intractable aggregates or precipitates form. Here we show how such unfolded aggregates can be solubilized and the solution mixed with lipid to spontaneously self-assemble a bicontinuous cubic mesophase into the bilayer of which the protein, in a confined, chaperonin-like environment, reconstitutes with 100% efficiency. The test protein, diacylglycerol kinase, reconstituted in the bilayer of the mesophase, was then crystallized in situ by the in meso or lipid cubic phase method providing an X-ray structure to a resolution of 2.55 Å. This highly efficient, inexpensive, simple and rapid approach should find application wherever properly folded, membrane reconstituted and functional proteins are required where the starting material is a denatured aggregate. PMID:25055873

  16. Remodeling of membrane lipids in iron-starved Chlamydomonas.

    PubMed

    Urzica, Eugen I; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M Dudley; Casero, David; Gallaher, Sean D; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S

    2013-10-18

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24-48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii.

  17. Remodeling of Membrane Lipids in Iron-starved Chlamydomonas*

    PubMed Central

    Urzica, Eugen I.; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M. Dudley; Casero, David; Gallaher, Sean D.; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S.

    2013-01-01

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24–48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii. PMID:23983122

  18. Pulmonary lipid phosphate phosphohydrolase in plasma membrane signalling platforms.

    PubMed Central

    Nanjundan, M; Possmayer, F

    2001-01-01

    Lipid phosphate phosphohydrolase (LPP) has recently been proposed to have roles in signal transduction, acting sequentially to phospholipase D (PLD) and in attenuating the effects of phospholipid growth factors on cellular proliferation. In this study, LPP activity is reported to be enriched in lipid-rich signalling platforms isolated from rat lung tissue, isolated rat type II cells and type II cell-mouse lung epithelial cell lines (MLE12 and MLE15). Lung and cell line caveolin-enriched domains (CEDs), prepared on the basis of their detergent-insolubility in Triton X-100, contain caveolin-1 and protein kinase C isoforms. The LPP3 isoform was predominantly localized to rat lung CEDs. These lipid-rich domains, including those from isolated rat type II cells, were enriched both in phosphatidylcholine plus sphingomyelin (PC+SM) and cholesterol. Saponin treatment of MLE15 cells shifted the LPP activity, cholesterol, PC+SM and caveolin-1 from lipid microdomains to detergent-soluble fractions. Elevated LPP activity and LPP1/1a protein are present in caveolae from MLE15 cells prepared using the cationic-colloidal-silica method. In contrast, total plasma membranes had a higher abundance of LPP1/1a protein with low LPP activity. Phorbol ester treatment caused a 3.8-fold increase in LPP specific activity in MLE12 CEDs. Thus the activated form of LPP1/1a may be recruited into caveolae/rafts. Transdifferentiation of type II cells into a type I-like cell demonstrated enrichment in caveolin-1 levels and LPP activity. These results indicate that LPP is localized in caveolae and/or rafts in lung tissue, isolated type II cells and type II cell lines and is consistent with a role for LPP in both caveolae/raft signalling and caveolar dynamics. PMID:11535125

  19. Phase segregation of polymerizable lipids to construct filters for separating lipid-membrane-embedded species

    PubMed Central

    Hu, Shu-Kai; Chen, Ya-Ming; Chao, Ling

    2014-01-01

    Supported lipid bilayer (SLB) platforms have been developed to transport and separate membrane-embedded species in the species' native bilayer environment. In this study, we used the phase segregation phenomenon of lipid mixtures containing a polymerizable diacetylene phospholipid, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DiynePC), and a nonpolymerizable phospholipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), to create filter barrier structures in SLBs. Upon exposing the phase segregated samples to UV light, the DiynePC-rich domains could become crosslinked and remain fixed on the surface of the support, while the DOPC-rich regions, where no crosslinking could happen, could be removed later by detergent washing, and thus became the void regions in the filter. During the filter fabrication process, we used the laminar flow configuration in a microfluidic channel to control the spatial locations of the feed region and filter region in the SLB. The flow in a microfluidic channel was also used to apply a strong hydrodynamic shear stress to the SLB to transport the membrane-embedded species from the feed region to the filter region. We varied the DiynePC/DOPC molar ratio from 60/40 to 80/20 to adjust the cutoff size of the filter barriers and used two model membrane-embedded species of different sizes to examine the filtering capability. One of the model species, Texas Red 1,2-dihexa-decanoyl-sn-glycero-3-phosphoethanolamine triethylammonium salt (Texas Red DHPE), had a single-lipid size, and the other species, cholera toxin subunit B-GM1 complex, had a multilipid size. When the DiynePC/DOPC molar ratio was 60/40, both species had high penetration ratios in the filter region. However, when the ratio was increased to 70/30, only the Texas Red DHPE, which was the smaller of the two model species, could penetrate the filter to a considerable extent. When the ratio was increased to 80/20, neither of the model species could penetrate the filter

  20. Membrane-mimetic films of asymmetric phosphatidylcholine lipid bolaamphiphiles.

    PubMed

    Sun, Xue-Long; Biswas, Nilanjana; Kai, Toshitsugu; Dai, Zhifei; Dluhy, Richard A; Chaikof, Elliot L

    2006-01-31

    Membrane-spanning phospholipid bolaamphiphiles either alone or as a constituent of a multicomponent lipid membrane may prove to be facile building blocks for generating robust bioactive membrane-mimetic assemblies. We have previously reported the synthesis of asymmetric dialkyl phospholipid bolaamphiphiles that contain ester linked phosphatidylcholine and amine functionalities at opposite chain ends. In this report, we describe the synthesis of phospholipid bolaamphiphiles that are conjugated to biotin via the terminal amine with or without a poly(ethylene oxide) spacer arm of varying chain length. The behavior of biotinylated bolaamphiphiles as a self-assembled monolayer at an air-water interface was characterized by epi-fluorescence microscopy and revealed that domain structure and pi-A isotherms were substantially influenced by linker type and size. Substrate bound assemblies were produced by Langmuir-Blodgett deposition onto planar substrates coated with an avidin derivatized polyelectrolyte multilayer. Significantly, external reflectance infrared spectroscopy confirmed the fabrication of bolaamphiphile thin films that display extended stability in vitro.

  1. Membrane lipid unsaturation as physiological adaptation to animal longevity

    PubMed Central

    Naudí, Alba; Jové, Mariona; Ayala, Victòria; Portero-Otín, Manuel; Barja, Gustavo; Pamplona, Reinald

    2013-01-01

    The appearance of oxygen in the terrestrial atmosphere represented an important selective pressure for ancestral living organisms and contributed toward setting up the pace of evolutionary changes in structural and functional systems. The evolution of using oxygen for efficient energy production served as a driving force for the evolution of complex organisms. The redox reactions associated with its use were, however, responsible for the production of reactive species (derived from oxygen and lipids) with damaging effects due to oxidative chemical modifications of essential cellular components. Consequently, aerobic life required the emergence and selection of antioxidant defense systems. As a result, a high diversity in molecular and structural antioxidant defenses evolved. In the following paragraphs, we analyze the adaptation of biological membranes as a dynamic structural defense against reactive species evolved by animals. In particular, our goal is to describe the physiological mechanisms underlying the structural adaptation of cellular membranes to oxidative stress and to explain the meaning of this adaptive mechanism, and to review the state of the art about the link between membrane composition and longevity of animal species. PMID:24381560

  2. The Interaction of Melittin with Dimyristoyl Phosphatidylcholine-Dimyristoyl Phosphatidylserine Lipid Bilayer Membranes

    SciTech Connect

    Rai, Durgesh K.; Qian, Shuo; Heller, William T.

    2016-08-13

    We report that membrane-active peptides (MAPs), which interact directly with the lipid bilayer of a cell and include toxins and host defense peptides, display lipid composition-dependent activity. Phosphatidylserine (PS) lipids are anionic lipids that are found throughout the cellular membranes of most eukaryotic organisms where they serve as both a functional component and as a precursor to phosphatidylethanolamine lipids. The inner leaflet of the plasma membrane contains more PS than the outer one, and the asymmetry is actively maintained. Here, the impact of the MAP melittin on the structure of lipid bilayer vesicles made of a mixture of phosphatidylcholine and phosphatidylserine was studied. Small-angle neutron scattering of the MAP associated with selectively deuterium-labeled lipid bilayer vesicles revealed how the thickness and lipid composition of phosphatidylserine-containing vesicles change in response to melittin. The peptide thickens the lipid bilayer for concentrations up to P/L = 1/500, but membrane thinning results when P/L = 1/200. The thickness transition is accompanied by a large change in the distribution of DMPS between the leaflets of the bilayer. The change in composition is driven by electrostatic interactions, while the change in bilayer thickness is driven by changes in the interaction of the peptide with the headgroup region of the lipid bilayer. Lastly, the results provide new information about lipid-specific interactions that take place in mixed composition lipid bilayer membranes.

  3. The Interaction of Melittin with Dimyristoyl Phosphatidylcholine-Dimyristoyl Phosphatidylserine Lipid Bilayer Membranes

    DOE PAGES

    Rai, Durgesh K.; Qian, Shuo; Heller, William T.

    2016-08-13

    We report that membrane-active peptides (MAPs), which interact directly with the lipid bilayer of a cell and include toxins and host defense peptides, display lipid composition-dependent activity. Phosphatidylserine (PS) lipids are anionic lipids that are found throughout the cellular membranes of most eukaryotic organisms where they serve as both a functional component and as a precursor to phosphatidylethanolamine lipids. The inner leaflet of the plasma membrane contains more PS than the outer one, and the asymmetry is actively maintained. Here, the impact of the MAP melittin on the structure of lipid bilayer vesicles made of a mixture of phosphatidylcholine andmore » phosphatidylserine was studied. Small-angle neutron scattering of the MAP associated with selectively deuterium-labeled lipid bilayer vesicles revealed how the thickness and lipid composition of phosphatidylserine-containing vesicles change in response to melittin. The peptide thickens the lipid bilayer for concentrations up to P/L = 1/500, but membrane thinning results when P/L = 1/200. The thickness transition is accompanied by a large change in the distribution of DMPS between the leaflets of the bilayer. The change in composition is driven by electrostatic interactions, while the change in bilayer thickness is driven by changes in the interaction of the peptide with the headgroup region of the lipid bilayer. Lastly, the results provide new information about lipid-specific interactions that take place in mixed composition lipid bilayer membranes.« less

  4. Lipid composition affects the rate of photosensitized dissipation of cross-membrane diffusion potential on liposomes

    PubMed Central

    Ytzhak, Shany; Wuskell, Joseph P.; Loew, Leslie M.; Ehrenberg, Benjamin

    2010-01-01

    Hydrophobic or amphiphilic tetrapyrrole sensitizers are taken up by cells and are usually located in cellular lipid membranes. Singlet oxygen is photogenerated by the sensitizer and it diffuses in the membrane and causes oxidative damage to membrane components. This damage can occur to membrane lipids and to membrane-localized proteins. Depolarization of the Nernst electric potential on cells’ membranes has been observed in cellular photosensitization, but it was not established whether lipid oxidation is a relevant factor leading to abolishing the resting potential of cells’ membranes and to their death. In this work we studied the effect of liposomes’ lipid composition on the kinetics of hematoporphyrin-photosensitized dissipation of K+-diffusion electric potential that was generated across the membranes. We employed an electrochromic voltage-sensitive spectroscopic probe that possesses a high fluorescence signal response to the potential. We found a correlation between the structure and unsaturation of lipids and the leakage of the membrane, following photosensitization. As the extent of non-conjugated unsaturation of the lipids is increased from 1 to 6 double bonds, the kinetics of depolarization become faster. We also found that the kinetics of depolarization is affected by the percentage of the unsaturated lipids in the liposome: as the fraction of the unsaturated lipids increases the leakage trough the membrane is enhanced. When liposomes are composed of a lipid mixture similar to that of natural membranes and photosensitization is being carried out under usual photodynamic therapy (PDT) conditions, photodamage to the lipids is not likely to cause enhanced permeability of ions through the membrane, which would have been a mechanism that leads to cell death. PMID:20536150

  5. Anomalous and normal diffusion of proteins and lipids in crowded lipid membranes.

    PubMed

    Javanainen, Matti; Hammaren, Henrik; Monticelli, Luca; Jeon, Jae-Hyung; Miettinen, Markus S; Martinez-Seara, Hector; Metzler, Ralf; Vattulainen, Ilpo

    2013-01-01

    Lateral diffusion plays a crucial role in numerous processes that take place in cell membranes, yet it is quite poorly understood in native membranes characterized by, e.g., domain formation and large concentration of proteins. In this article, we use atomistic and coarse-grained simulations to consider how packing of membranes and crowding with proteins affect the lateral dynamics of lipids and membrane proteins. We find that both packing and protein crowding have a profound effect on lateral diffusion, slowing it down. Anomalous diffusion is observed to be an inherent property in both protein-free and protein-rich membranes, and the time scales of anomalous diffusion and the exponent associated with anomalous diffusion are found to strongly depend on packing and crowding. Crowding with proteins also has a striking effect on the decay rate of dynamical correlations associated with lateral single-particle motion, as the transition from anomalous to normal diffusion is found to take place at macroscopic time scales: while in protein-poor conditions normal diffusion is typically observed in hundreds of nanoseconds, in protein-rich conditions the onset of normal diffusion is tens of microseconds, and in the most crowded systems as large as milliseconds. The computational challenge which results from these time scales is not easy to deal with, not even in coarse-grained simulations. We also briefly discuss the physical limits of protein motion. Our results suggest that protein concentration is anything but constant in the plane of cell membranes. Instead, it is strongly dependent on proteins' preference for aggregation.

  6. Lipid raft detecting in membranes of live erythrocytes.

    PubMed

    Mikhalyov, Ilya; Samsonov, Andrey

    2011-07-01

    The fluorescent probe N-(BODIPY(®)-FL-propionyl)-neuraminosyl-GM(1) (BODIPY-GM(1)) was used to detect lipid rafts in living red blood cells (RBCs) membranes. The probe was detected with fluorescence video microscopy and was found to be uniformly distributed along plasma membrane at room temperature (23°C). At 4°C some probe clearly phase-separated to yield detectable bright spots that were smaller than spatial resolution. As measured by spectrofluorometry, in addition to a major fluorescence peak caused by emissions from monomers, the probe exhibited a red-shifted peak that is characteristic of a BODIPY fluorophore at high local concentrations, indicating that some probe had clustered. Red-shifted fluorescence was the greatest at 4°C, intermediate at 23°C, and the smallest at 37°C. Treating the RBCs with methyl-β-cyclodextrin to remove cholesterol eliminated the red-shifted peak. This strongly indicates that the presence of cholesterol was essential for phase separation of the probe. Fluorometry experiments indicate that rafts exist at 23°C and at 37°C, even though the membrane appears to be uniform at the resolution of microscope. The distinct GM(1) patches distributed over entire membrane of the erythrocytes were observed at both 23°C and at 37°C in RBCs stained with Alexa FL 647 cholera toxin subunit B conjugate (CTB-A647 ). Based on both fluorometry and fluorescence microscopy, some rafts clearly exist at 37°C.

  7. Unique Lipid Chemistry of Synaptic Vesicle and Synaptosome Membrane Revealed Using Mass Spectrometry.

    PubMed

    Lewis, Kenneth T; Maddipati, Krishna R; Naik, Akshata R; Jena, Bhanu P

    2017-03-02

    Synaptic vesicles measuring 30-50 nm in diameter containing neurotransmitters either completely collapse at the presynaptic membrane or dock and transiently fuse at the base of specialized 15 nm cup-shaped lipoprotein structures called porosomes at the presynaptic membrane of synaptosomes to release neurotransmitters. Recent study reports the unique composition of major lipids associated with neuronal porosomes. Given that lipids greatly influence the association and functions of membrane proteins, differences in lipid composition of synaptic vesicle and the synaptosome membrane was hypothesized. To test this hypothesis, the lipidome of isolated synaptosome, synaptosome membrane, and synaptic vesicle preparation were determined by using mass spectrometry in the current study. Results from the study demonstrate the enriched presence of triacyl glycerols and sphingomyelins in synaptic vesicles, as opposed to the enriched presence of phospholipids in the synaptosome membrane fraction, reflecting on the tight regulation of nerve cells in compartmentalization of membrane lipids at the nerve terminal.

  8. More Than a Pore: The Interplay of Pore-Forming Proteins and Lipid Membranes.

    PubMed

    Ros, Uris; García-Sáez, Ana J

    2015-06-01

    Pore-forming proteins (PFPs) punch holes in their target cell membrane to alter their permeability. Permeabilization of lipid membranes by PFPs has received special attention to study the basic molecular mechanisms of protein insertion into membranes and the development of biotechnological tools. PFPs act through a general multi-step mechanism that involves (i) membrane partitioning, (ii) insertion into the hydrophobic core of the bilayer, (iii) oligomerization, and (iv) pore formation. Interestingly, PFPs and membranes show a dynamic interplay. As PFPs are usually produced as soluble proteins, they require a large conformational change for membrane insertion. Moreover, membrane structure is modified upon PFPs insertion. In this context, the toroidal pore model has been proposed to describe a pore architecture in which not only protein molecules but also lipids are directly involved in the structure. Here, we discuss how PFPs and lipids cooperate and remodel each other to achieve pore formation, and explore new evidences of protein-lipid pore structures.

  9. Effects of phloretin on lipid organization in the erythrocyte membrane as measured by EPR

    NASA Astrophysics Data System (ADS)

    Abumrad, Nada A.; Perkins, Ray C.; Dalton, Larry R.; Park, Charles R.; Park, Jane H.

    Phloretin is a lipophilic compound which has been widely studied as a broad spectrum effector of metabolite transport in red blood cells (RBC). Phloretin effects on the organization of lipids in the RBC membrane are investigated using the spin-labeled fatty acids, 5 and 16-nitroxyl stearate (5-NS and 16-NS, respectively). Phloretin at different concentrations produced biphasic effects on the lineshape of the EPR response from 16-NS-labeled RBC. The dependence of these changes on the flat cell orientation with respect to the magnetic field suggested that phloretin promoted lipid order at low concentrations (5 to 40 μ M) and disorder at high concentrations (40 to 250 μ M). The biphasic effects of phloretin occurred at concentrations which parallel its dual actions on metabolite transfer. Phloretin generally inhibits transport (protein-mediated) and stimulates diffusion (lipid-mediated) processes. The spectroscopic effects were best characterized through second-harmonic, in-phase detection. The possible contribution of other factors to the spectroscopic changes is discussed. When RBC were spin labeled with 5-NS, higher concentrations of the probe were required for adequate detection and only monophasic effects of phoretin were observed. The results suggest that membrane lipids are important in phloretin effects on transport and diffusion processes.

  10. Experimental Observations of Dynamic Critical Phenomena in a Lipid Membrane

    NASA Astrophysics Data System (ADS)

    Honerkamp-Smith, Aurelia R.; Machta, Benjamin B.; Keller, Sarah L.

    2012-06-01

    Near a critical point, the time scale of thermally induced fluctuations diverges in a manner determined by the dynamic universality class. Experiments have verified predicted three-dimensional dynamic critical exponents in many systems, but similar experiments in two dimensions have been lacking for the case of conserved order parameter. Here we analyze the time-dependent correlation functions of a quasi-two-dimensional lipid bilayer in water to show that its critical dynamics agree with a recently predicted universality class. In particular, the effective dynamic exponent zeff crosses over from ˜2 to ˜3 as the correlation length of fluctuations exceeds a hydrodynamic length set by the membrane and bulk viscosities.

  11. Fluorescence Lifetime Imaging of Membrane Lipid Order with a Ratiometric Fluorescent Probe

    PubMed Central

    Kilin, Vasyl; Glushonkov, Oleksandr; Herdly, Lucas; Klymchenko, Andrey; Richert, Ludovic; Mely, Yves

    2015-01-01

    To monitor the lateral segregation of lipids into liquid-ordered (Lo) and -disordered (Ld) phases in lipid membranes, environment-sensitive dyes that partition in both phases but stain them differently have been developed. Of particular interest is the dual-color F2N12S probe, which can discriminate the two phases through the ratio of its two emission bands. These bands are associated with the normal (N∗) and tautomer (T∗) excited-state species that result from an excited-state intramolecular proton transfer. In this work, we investigated the potency of the time-resolved fluorescence parameters of F2N12S to discriminate lipid phases in model and cell membranes. Both the long and mean lifetime values of the T∗ form of F2N12S were found to differ by twofold between Ld and Lo phases as a result of the restriction in the relative motions of the two aromatic moieties of F2N12S imposed by the highly packed Lo phase. This differed from the changes in the ratio of the two emission bands between the two phases, which mainly resulted from the decreased hydration of the N∗ form in the Lo phase. Importantly, the strong difference in lifetimes between the two phases was preserved when cholesterol was added to the Ld phase. The two phases could be imaged with high contrast by fluorescence lifetime imaging microscopy (FLIM) on giant unilamellar vesicles. FLIM images of F2N12S-labeled live HeLa cells confirmed that the plasma membrane was mainly in the Lo-like phase. Furthermore, the two phases were found to be homogeneously distributed all over the plasma membrane, indicating that they are highly mixed at the spatiotemporal resolution of the FLIM setup. Finally, FLIM could also be used to sensitively monitor the change in lipid phase upon cholesterol depletion and apoptosis. PMID:25992730

  12. Fluorescence lifetime imaging of membrane lipid order with a ratiometric fluorescent probe.

    PubMed

    Kilin, Vasyl; Glushonkov, Oleksandr; Herdly, Lucas; Klymchenko, Andrey; Richert, Ludovic; Mely, Yves

    2015-05-19

    To monitor the lateral segregation of lipids into liquid-ordered (Lo) and -disordered (Ld) phases in lipid membranes, environment-sensitive dyes that partition in both phases but stain them differently have been developed. Of particular interest is the dual-color F2N12S probe, which can discriminate the two phases through the ratio of its two emission bands. These bands are associated with the normal (N(∗)) and tautomer (T(∗)) excited-state species that result from an excited-state intramolecular proton transfer. In this work, we investigated the potency of the time-resolved fluorescence parameters of F2N12S to discriminate lipid phases in model and cell membranes. Both the long and mean lifetime values of the T(∗) form of F2N12S were found to differ by twofold between Ld and Lo phases as a result of the restriction in the relative motions of the two aromatic moieties of F2N12S imposed by the highly packed Lo phase. This differed from the changes in the ratio of the two emission bands between the two phases, which mainly resulted from the decreased hydration of the N(∗) form in the Lo phase. Importantly, the strong difference in lifetimes between the two phases was preserved when cholesterol was added to the Ld phase. The two phases could be imaged with high contrast by fluorescence lifetime imaging microscopy (FLIM) on giant unilamellar vesicles. FLIM images of F2N12S-labeled live HeLa cells confirmed that the plasma membrane was mainly in the Lo-like phase. Furthermore, the two phases were found to be homogeneously distributed all over the plasma membrane, indicating that they are highly mixed at the spatiotemporal resolution of the FLIM setup. Finally, FLIM could also be used to sensitively monitor the change in lipid phase upon cholesterol depletion and apoptosis.

  13. Studying lipid organization in biological membranes using liposomes and EPR spin labeling

    PubMed Central

    Subczynski, Witold K.; Raguz, Marija; Widomska, Justyna

    2015-01-01

    Summary Electron paramagnetic resonance (EPR) spin-labeling methods provide a unique opportunity to determine the lateral organization of lipid bilayer membranes by discrimination of coexisting membrane domains or coexisting membrane phases. In some cases, the coexisting membrane domains can be characterized by profiles of alkyl chain order, fluidity, hydrophobicity, and oxygen diffusion-concentration product in situ, without the need for their physical separation. This chapter briefly explains how the EPR spin-labeling methods can be used to obtain the above mentioned profiles across the lipid bilayer membranes (liposomes) derived from the lipid extract of certain biological membranes. These procedures will be illustrated by EPR measurements performed for multilamellar liposomes made of the lipid extracts from cortical and nuclear fractions of the fiber cell plasma membranes of a cow eye lens. To elucidate better the major factors that determine membrane properties, the results for eye lens lipid membranes will be compared with those obtained for simple model membranes resembling basic lipid composition of biological membranes. PMID:20013402

  14. Atomic-level description of protein-lipid interactions using an accelerated membrane model.

    PubMed

    Baylon, Javier L; Vermaas, Josh V; Muller, Melanie P; Arcario, Mark J; Pogorelov, Taras V; Tajkhorshid, Emad

    2016-07-01

    Peripheral membrane proteins are structurally diverse proteins that are involved in fundamental cellular processes. Their activity of these proteins is frequently modulated through their interaction with cellular membranes, and as a result techniques to study the interfacial interaction between peripheral proteins and the membrane are in high demand. Due to the fluid nature of the membrane and the reversibility of protein-membrane interactions, the experimental study of these systems remains a challenging task. Molecular dynamics simulations offer a suitable approach to study protein-lipid interactions; however, the slow dynamics of the lipids often prevents sufficient sampling of specific membrane-protein interactions in atomistic simulations. To increase lipid dynamics while preserving the atomistic detail of protein-lipid interactions, in the highly mobile membrane-mimetic (HMMM) model the membrane core is replaced by an organic solvent, while short-tailed lipids provide a nearly complete representation of natural lipids at the organic solvent/water interface. Here, we present a brief introduction and a summary of recent applications of the HMMM to study different membrane proteins, complementing the experimental characterization of the presented systems, and we offer a perspective of future applications of the HMMM to study other classes of membrane proteins. This article is part of a Special Issue entitled: Membrane proteins edited by J.C. Gumbart and Sergei Noskov.

  15. Exploiting Lipid Permutation Symmetry to Compute Membrane Remodeling Free Energies

    NASA Astrophysics Data System (ADS)

    Bubnis, Greg; Risselada, Herre Jelger; Grubmüller, Helmut

    2016-10-01

    A complete physical description of membrane remodeling processes, such as fusion or fission, requires knowledge of the underlying free energy landscapes, particularly in barrier regions involving collective shape changes, topological transitions, and high curvature, where Canham-Helfrich (CH) continuum descriptions may fail. To calculate these free energies using atomistic simulations, one must address not only the sampling problem due to high free energy barriers, but also an orthogonal sampling problem of combinatorial complexity stemming from the permutation symmetry of identical lipids. Here, we solve the combinatorial problem with a permutation reduction scheme to map a structural ensemble into a compact, nondegenerate subregion of configuration space, thereby permitting straightforward free energy calculations via umbrella sampling. We applied this approach, using a coarse-grained lipid model, to test the CH description of bending and found sharp increases in the bending modulus for curvature radii below 10 nm. These deviations suggest that an anharmonic bending term may be required for CH models to give quantitative energetics of highly curved states.

  16. Control of plasma membrane lipid homeostasis by the extended synaptotagmins

    PubMed Central

    Saheki, Yasunori; Bian, Xin; Schauder, Curtis M.; Sawaki, Yujin; Surma, Michal A.; Klose, Christian; Pincet, Frederic; Reinisch, Karin M.; De Camilli, Pietro

    2016-01-01

    Acute metabolic changes of plasma membrane (PM) lipids, such as those mediating signaling reactions, are rapidly compensated by homeostatic responses whose molecular basis is poorly understood. Here we show that the Extended-Synaptotagmins (E-Syts), ER proteins which function as PI(4,5)P2 and Ca2+-regulated tethers to the PM, participate in these responses. E-Syts transfer glycerolipids between bilayers in vitro and such transfer requires Ca2+ and their SMP domain, a lipid-harboring module. Genome edited cells lacking E-Syts do not exhibit abnormalities in the major glycerolipids at rest, but display enhanced and sustained accumulation of PM diacylglycerol (DAG) upon PI(4,5)P2 hydrolysis by PLC activation, which can be rescued by expression of E-Syt1, but not by mutant E-Syt1 lacking the SMP domain. The formation of E-Syts-dependent ER-PM tethers in response to stimuli that cleave PI(4,5)P2 and elevate Ca2+ may help reverse accumulation of DAG in the PM by transferring it to the ER for metabolic recycling. PMID:27065097

  17. On ripples and rafts: Curvature induced nanoscale structures in lipid membranes

    NASA Astrophysics Data System (ADS)

    Schmid, Friederike; Dolezel, Stefan; Lenz, Olaf; Meinhardt, Sebastian

    2014-03-01

    We develop an elastic theory that predicts the spontaneous formation of nanoscale structures in lipid bilayers which locally phase separate between two phases with different spontaneous monolayer curvature. The theory rationalizes in a unified manner the observation of a variety of nanoscale structures in lipid membranes: Rippled states in one-component membranes, lipid rafts in multicomponent membranes. Furthermore, we report on recent observations of rippled states and rafts in simulations of a simple coarse-grained model for lipid bilayers, which are compatible with experimental observations and with our elastic model.

  18. On Physical Properties of Tetraether Lipid Membranes: Effects of Cyclopentane Rings

    PubMed Central

    Chong, Parkson Lee-Gau; Ayesa, Umme; Prakash Daswani, Varsha; Hur, Ellah Chay

    2012-01-01

    This paper reviews the recent findings related to the physical properties of tetraether lipid membranes, with special attention to the effects of the number, position, and configuration of cyclopentane rings on membrane properties. We discuss the findings obtained from liposomes and monolayers, composed of naturally occurring archaeal tetraether lipids and synthetic tetraethers as well as the results from computer simulations. It appears that the number, position, and stereochemistry of cyclopentane rings in the dibiphytanyl chains of tetraether lipids have significant influence on packing tightness, lipid conformation, membrane thickness and organization, and headgroup hydration/orientation. PMID:23028246

  19. Voltage- and Tension-Dependent Lipid Mobility in the Outer Hair Cell Plasma Membrane

    NASA Astrophysics Data System (ADS)

    Oghalai, John S.; Zhao, Hong-Bo; Kutz, J. Walter; Brownell, William E.

    2000-01-01

    The mechanism responsible for electromotility of outer hair cells in the ear is unknown but is thought to reside within the plasma membrane. Lipid lateral diffusion in the outer hair cell plasma membrane is a sigmoidal function of transmembrane potential and bathing media osmolality. Cell depolarization or hyposmotic challenge shorten the cell and reduce membrane fluidity by half. Changing the membrane tension with amphipathic drugs results in similar reductions. These dynamic changes in membrane fluidity represent the modulation of membrane tension by lipid-protein interactions. The voltage dependence may be associated with the force-generating motors that contribute to the exquisite sensitivity of mammalian hearing.

  20. Hydration dependent studies of highly aligned multilayer lipid membranes by neutron scattering

    NASA Astrophysics Data System (ADS)

    Trapp, Marcus; Gutberlet, Thomas; Juranyi, Fanni; Unruh, Tobias; Demé, Bruno; Tehei, Moeava; Peters, Judith

    2010-10-01

    We investigated molecular motions on a picosecond timescale of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) model membranes as a function of hydration by using elastic and quasielastic neutron scattering. Two different hydrations corresponding to approximately nine and twelve water molecules per lipid were studied, the latter being the fully hydrated state. In our study, we focused on head group motions by using chain deuterated lipids. Information on in-plane and out-of-plane motions could be extracted by using solid supported DMPC multilayers. Our studies confirm and complete former investigations by König et al. [J. Phys. II (France) 2, 1589 (1992)] and Rheinstädter et al. [Phys. Rev. Lett. 101, 248106 (2008)] who described the dynamics of lipid membranes, but did not explore the influence of hydration on the head group dynamics as presented here. From the elastic data, a clear shift of the main phase transition from the Pβ ripple phase to the Lα liquid phase was observed. Decreasing water content moves the transition temperature to higher temperatures. The quasielastic data permit a closer investigation of the different types of head group motion of the two samples. Two different models are needed to fit the elastic incoherent structure factor and corresponding radii were calculated. The presented data show the strong influence hydration has on the head group mobility of DMPC.

  1. A novel squarylium dye for monitoring oxidative processes in lipid membranes.

    PubMed

    Trusova, Valeriya M; Gorbenko, Galyna P; Deligeorgiev, Todor; Gadjev, Nikolai; Vasilev, Aleksey

    2009-11-01

    A novel squaraine probe SQ-1 has been found to be appropriate for monitoring the peroxidation processes in membrane systems. Formation of free radicals was triggered by methemoglobin (metHb) or cytochrome c (cyt c) binding to the model lipid membranes composed of zwitterionic lipid phosphatidylcholine (PC) and anionic lipid cardiolipin (CL). Protein association with the lipid vesicles was followed by drastic quenching of SQ-1 fluorescence. The observed spectral changes were suppressed in the presence of free radical scavengers, butylated hydroxytoluene (BHT) and thiourea (TM) suggesting that SQ-1 decolorization can be attributed to its reactions with lipid radicals.

  2. Activation of integrin α5 mediated by flow requires its translocation to membrane lipid rafts in vascular endothelial cells.

    PubMed

    Sun, Xiaoli; Fu, Yi; Gu, Mingxia; Zhang, Lu; Li, Dan; Li, Hongliang; Chien, Shu; Shyy, John Y-J; Zhu, Yi

    2016-01-19

    Local flow patterns determine the uneven distribution of atherosclerotic lesions. Membrane lipid rafts and integrins are crucial for shear stress-regulated endothelial function. In this study, we investigate the role of lipid rafts and integrin α5 in regulating the inflammatory response in endothelial cells (ECs) under atheroprone versus atheroprotective flow. Lipid raft proteins were isolated from ECs exposed to oscillatory shear stress (OS) or pulsatile shear stress, and then analyzed by quantitative proteomics. Among 396 proteins redistributed in lipid rafts, integrin α5 was the most significantly elevated in lipid rafts under OS. In addition, OS increased the level of activated integrin α5 in lipid rafts through the regulation of membrane cholesterol and fluidity. Disruption of F-actin-based cytoskeleton and knockdown of caveolin-1 prevented the OS-induced integrin α5 translocation and activation. In vivo, integrin α5 activation and EC dysfunction were observed in the atheroprone areas of low-density lipoprotein receptor-deficient (Ldlr(-/-)) mice, and knockdown of integrin α5 markedly attenuated EC dysfunction in partially ligated carotid arteries. Consistent with these findings, mice with haploinsufficency of integrin α5 exhibited a reduction of atherosclerotic lesions in the regions under atheroprone flow. The present study has revealed an integrin- and membrane lipid raft-dependent mechanotransduction mechanism by which atheroprone flow causes endothelial dysfunction.

  3. The effect of charged lipids on bacteriorhodopsin membrane reconstitution and its photochemical activities

    SciTech Connect

    Wang Zhen; Bai Jing; Xu Yuhong

    2008-07-11

    Bacteriorhodopsin (BR) was reconstituted into artificial lipid membrane containing various charged lipid compositions. The proton pumping activity of BR under flash and continuous illumination, proton permeability across membrane, as well as the decay kinetics of the photocycle intermediate M{sub 412} were studied. The results showed that lipid charges would significantly affect the orientation of BR inserted into lipid membranes. In liposomes containing anionic lipids, BRs were more likely to take natural orientation as in living cells. In neutral or positively charged liposomes, most BRs were reversely assembled, assuming an inside out orientation. Moreover, the lipids charges also affect BR's M intermediate kinetics, especially the slow component in M intermediate decay. The half-life M{sub 412s} increased significantly in BRs in liposomes containing cationic lipids, while decreased in those in anionic liposomes.

  4. Method of fabricating lipid bilayer membranes on solid supports

    NASA Technical Reports Server (NTRS)

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

    2012-01-01

    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.

  5. Mechanics of water pore formation in lipid membrane under electric field

    NASA Astrophysics Data System (ADS)

    Bu, Bing; Li, Dechang; Diao, Jiajie; Ji, Baohua

    2017-02-01

    Transmembrane water pores are crucial for substance transport through cell membranes via membrane fusion, such as in neural communication. However, the molecular mechanism of water pore formation is not clear. In this study, we apply all-atom molecular dynamics and bias-exchange metadynamics simulations to study the process of water pore formation under an electric field. We show that water molecules can enter a membrane under an electric field and form a water pore of a few nanometers in diameter. These water molecules disturb the interactions between lipid head groups and the ordered arrangement of lipids. Following the movement of water molecules, the lipid head groups are rotated and driven into the hydrophobic region of the membrane. The reorientated lipid head groups inside the membrane form a hydrophilic surface of the water pore. This study reveals the atomic details of how an electric field influences the movement of water molecules and lipid head groups, resulting in water pore formation.

  6. Methods for lipid nanostructure investigation at neutron and synchrotron sources

    NASA Astrophysics Data System (ADS)

    Kiselev, M. A.

    2011-03-01

    A lipid membrane is a main component of biological membranes. Contemporary bionanotechnologies use phospholipids and ceramides as basic components of drugs and cosmetic preparations. Phospholipids-based nanoparticles are used as drug carriers. Effective development of bionanotechnologies in Russia calls for creation of physical methods to diagnose the particle nanostructure which would be promising for application in pharmacology. Radiation with wavelengths of 1-10 Å is an adequate instrument for detecting the nanostructure of lipid bi- and monolayers. The review deals with methods that apply neutron scattering and synchrotron radiation for studying nanostructures of lipid membranes, phospholipid nanoparticles, and phospholipid monolayers on a water surface by techniques of diffraction, small-angle scattering, and reflectometry. The importance of the mutually complementary application of neutron and synchrotron radiation for solving urgent problems of membrane biophysics, microbiology, dermapharmacology, and bionanotechnologies is demonstrated by particular examples of studies of phospholipid membranes and ceramide-based membranes. The efficiency of development and application of new methods for solving urgent problems of biophysics is shown. The review is written on the basis of results obtained over the period of 1999-2010 at the Joint Institute for Nuclear Research (JINR) Laboratory of Neutron Physics in collaboration with the Pharmaceutical Departments of universities of France (Paris-Sud, Chatenay Malabry) and Germany (Martin Luther University, Halle). The experiments were performed at various European and Russian neutron and synchrotron sources.

  7. In vivo cluster formation of nisin and lipid II is correlated with membrane depolarization.

    PubMed

    Tol, Menno B; Morales Angeles, Danae; Scheffers, Dirk-Jan

    2015-01-01

    Nisin and related lantibiotics kill bacteria by pore formation or by sequestering lipid II. Some lantibiotics sequester lipid II into clusters, which were suggested to kill cells through delocalized peptidoglycan synthesis. Here, we show that cluster formation is always concomitant with (i) membrane pore formation and (ii) membrane depolarization. Nisin variants that cluster lipid II kill L-form bacteria with similar efficiency, suggesting that delocalization of peptidoglycan synthesis is not the primary killing mechanism of these lantibiotics.

  8. Analysis of the Torus Surrounding Planar Lipid Bilayer Membranes

    PubMed Central

    White, Stephen H.

    1972-01-01

    The characteristics and behavior of the torus (annulus) surrounding planar lipid bilayer membranes formed across a cylindrical aperture are analyzed using equations for the shape and volume of the annulus derived by the methods of variational calculus. The analysis leads to the following results: (a) Design criteria for the aperture can be established. (b) The transition region between thin film and thick annulus can be defined quantitatively and its effect on the measurement of specific capacitance determined. (c) At fixed annulus volume the diameter of the thin membrane is a function of the thin film-annulus contact angle. This suggests a new method for examining changes in free energy of the thin film, and explains why the area of thin film increases reversibly when potentials are present across the film. (d) In the absence of buoyant forces, the equations for the shape and volume of the annulus consist of incomplete elliptic integrals of the first and second kinds; however, the shape of the annulus in the transition region can be described with good accuracy by an approximate equation of greater simplicity. PMID:5019479

  9. Analysis of the torus surrounding planar lipid bilayer membranes.

    PubMed

    White, S H

    1972-04-01

    The characteristics and behavior of the torus (annulus) surrounding planar lipid bilayer membranes formed across a cylindrical aperture are analyzed using equations for the shape and volume of the annulus derived by the methods of variational calculus. The analysis leads to the following results: (a) Design criteria for the aperture can be established. (b) The transition region between thin film and thick annulus can be defined quantitatively and its effect on the measurement of specific capacitance determined. (c) At fixed annulus volume the diameter of the thin membrane is a function of the thin film-annulus contact angle. This suggests a new method for examining changes in free energy of the thin film, and explains why the area of thin film increases reversibly when potentials are present across the film. (d) In the absence of buoyant forces, the equations for the shape and volume of the annulus consist of incomplete elliptic integrals of the first and second kinds; however, the shape of the annulus in the transition region can be described with good accuracy by an approximate equation of greater simplicity.

  10. Ionic protein-lipid interaction at the plasma membrane: what can the charge do?

    PubMed

    Li, Lunyi; Shi, Xiaoshan; Guo, Xingdong; Li, Hua; Xu, Chenqi

    2014-03-01

    Phospholipids are the major components of cell membranes, but they have functional roles beyond forming lipid bilayers. In particular, acidic phospholipids form microdomains in the plasma membrane and can ionically interact with proteins via polybasic sequences, which can have functional consequences for the protein. The list of proteins regulated by ionic protein-lipid interaction has been quickly expanding, and now includes membrane proteins, cytoplasmic soluble proteins, and viral proteins. Here we review how acidic phospholipids in the plasma membrane regulate protein structure and function via ionic interactions, and how Ca(2+) regulates ionic protein-lipid interactions via direct and indirect mechanisms.

  11. Structural Basis for Ca2+-mediated Interaction of the Perforin C2 Domain with Lipid Membranes*

    PubMed Central

    Yagi, Hiromasa; Conroy, Paul J.; Leung, Eleanor W. W.; Law, Ruby H. P.; Trapani, Joseph A.; Voskoboinik, Ilia; Whisstock, James C.; Norton, Raymond S.

    2015-01-01

    Natural killer cells and cytotoxic T-lymphocytes deploy perforin and granzymes to kill infected host cells. Perforin, secreted by immune cells, binds target membranes to form pores that deliver pro-apoptotic granzymes into the target cell. A crucial first step in this process is interaction of its C2 domain with target cell membranes, which is a calcium-dependent event. Some aspects of this process are understood, but many molecular details remain unclear. To address this, we investigated the mechanism of Ca2+ and lipid binding to the C2 domain by NMR spectroscopy and x-ray crystallography. Calcium titrations, together with dodecylphosphocholine micelle experiments, confirmed that multiple Ca2+ ions bind within the calcium-binding regions, activating perforin with respect to membrane binding. We have also determined the affinities of several of these binding sites and have shown that this interaction causes a significant structural rearrangement in CBR1. Thus, it is proposed that Ca2+ binding at the weakest affinity site triggers changes in the C2 domain that facilitate its interaction with lipid membranes. PMID:26306037

  12. Chronic cigarette smoking alters erythrocyte membrane lipid composition and properties in male human volunteers.

    PubMed

    Padmavathi, Pannuru; Reddy, Vaddi Damodara; Kavitha, Godugu; Paramahamsa, Maturu; Varadacharyulu, Nallanchakravarthula

    2010-11-01

    Cigarette smoking is a major lifestyle factor influencing the health of human beings. The present study investigates smoking induced alterations on the erythrocyte membrane lipid composition, fluidity and the role of nitric oxide. Thirty experimental and control subjects (age 35+/-8) were selected for the study. Experimental subjects smoke 12+/-2 cigarettes per day for 7-10 years. In smokers elevated nitrite/nitrate levels in plasma and red cell lysates were observed. Smokers showed increased hemolysis, erythrocyte membrane lipid peroxidation, protein carbonyls, C/P ratio (cholesterol and phospholipid ratio), anisotropic (gamma) value with decreased Na(+)/K(+)-ATPase activity and sulfhydryl groups. Alterations in smokers erythrocyte membrane individual phospholipids were also evident from the study. Red cell lysate nitric oxide positively correlated with C/P ratio (r=0.565) and fluorescent anisotropic (gamma) value (r=0.386) in smokers. Smoking induced generation of reactive oxygen/nitrogen species might have altered erythrocyte membrane physico-chemical properties.

  13. Membrane lipids and sodium pumps of cattle and crocodiles: an experimental test of the membrane pacemaker theory of metabolism.

    PubMed

    Wu, B J; Hulbert, A J; Storlien, L H; Else, P L

    2004-09-01

    The influence of membrane lipid composition on the molecular activity of a major membrane protein (the sodium pump) was examined as a test of the membrane pacemaker theory of metabolism. Microsomal membranes from the kidneys of cattle (Bos taurus) and crocodiles (Crocodylus porosus) were found to possess similar sodium pump concentrations, but cattle membranes showed a four- to fivefold higher enzyme (Na(+)-K(+)-ATPase) activity when measured at 37 degrees C. The molecular activity of the sodium pumps (ATP/min) from both species was fully recoverable when delipidated pumps were reconstituted with membrane from the original source (same species). The results of experiments involving species membrane crossovers showed cattle sodium pump molecular activity to progressively decrease from 3,245 to 1,953 (P < 0.005) to 1,031 (P < 0.003) ATP/min when subjected to two cycles of delipidation and reconstitution with crocodile membrane as a lipid source. In contrast, the molecular activity of crocodile sodium pumps progressively increased from 729 to 908 (P < 0.01) to 1,476 (P = 0.01) ATP/min when subjected to two cycles of delipidation and reconstitution with cattle membrane as a lipid source. The lipid composition of the two membrane preparations showed similar levels of saturated ( approximately 31-34%) and monounsaturated ( approximately 23-25%) fatty acids. Cattle membrane had fourfold more n-3 polyunsaturated fatty acids (11.2 vs. 2.9%) but had a reduced n-6 polyunsaturate content (29 vs. 43%). The results support the membrane pacemaker theory of metabolism and suggest membrane lipids and their polyunsaturates play a significant role in determining the molecular activity of the sodium pump.

  14. Conversion of membrane lipid acyl groups to triacylglycerol and formation of lipid bodies upon nitrogen starvation in biofuel green algae Chlorella UTEX29.

    PubMed

    Goncalves, Elton C; Johnson, Jodie V; Rathinasabapathi, Bala

    2013-11-01

    Algal lipids are ideal biofuel sources. Our objective was to determine the contributors to triacylglycerol (TAG) accumulation and lipid body formation in Chlorella UTEX29 under nitrogen (N) deprivation. A fivefold increase in intracellular lipids following N starvation for 24 h confirmed the oleaginous characteristics of UTEX29. Ultrastructural studies revealed increased number of lipid bodies and decreased starch granules in N-starved cells compared to N-replete cells. Lipid bodies were observed as early as 3 h after N removal and plastids collapsed after 48 h of stress. Moreover, the identification of intracellular pyrenoids and differences in the expected nutritional requirements for Chlorella protothecoides (as UTEX29 is currently classified) led us to conduct a phylogenetic study using 18S and actin cDNA sequences. This indicated UTEX29 to be more phylogenetically related to Chlorella vulgaris. To investigate the fate of different lipids after N starvation, radiolabeling using ¹⁴C-acetate was used. A significant decrease in ¹⁴C-galactolipids and phospholipids matched the increase in ¹⁴C-TAG starting at 3 h of N starvation, consistent with acyl groups from structural lipids as sources for TAG under N starvation. These results have important implications for the identification of key steps controlling oil accumulation in N-starved biofuel algae and demonstrate membrane recycling during lipid body formation.

  15. Effects of cholesterol concentration on the interaction of cytarabine with lipid membranes: a molecular dynamics simulation study.

    PubMed

    Karami, Leila; Jalili, Seifollah

    2015-01-01

    Liposomal cytarabine, DepoCyt, is a chemotherapy agent which is used in cancer treatment. This form of cytarabine has more efficacy and fewer side effects relative to the other forms. Since DepoCyt contains the cytarabine encapsulated within phosphatidylcholine and the sterol molecules, we modeled dioleoylphosphatidylcholine (DOPC)/cholesterol bilayer membrane as a carrier for cytarabine to study drug-bilayer interactions. For this purpose, we performed a series of united-atom molecular dynamics (MD) simulations for 25 ns to investigate the interactions between cytarabine and cholesterol-containing DOPC lipid bilayers. Only the uncharged form of cytarabine molecule was investigated. In this study, different levels of the cholesterol content (0, 20, and 40%) were used. MD simulations allowed us to determine dynamical and structural properties of the bilayer membrane and to estimate the preferred location and orientation of the cytarabine molecule inside the bilayer membrane. Properties such as membrane thickness, area per lipid, diffusion coefficient, mass density, bilayer packing, order parameters, and intermolecular interactions were examined. The results show that by increasing the cholesterol concentration in the lipid bilayers, the bilayer thickness increases and area per lipid decreases. Moreover, in accordance with the experiments, our calculations show that cholesterol molecules have ordering effect on the hydrocarbon acyl chains. Furthermore, the cytarabine molecule preferentially occupies the polar region of the lipid head groups to form specific interactions (hydrogen bonds). Our results fully support the experimental data. Our finding about drug-bilayer interaction is crucial for the liposomal drug design.

  16. Monte Carlo Simulation of Protein-Induced Lipid Demixing in a Membrane with Interactions Derived from Experiment

    PubMed Central

    Almeida, Paulo F.; Best, Alexis; Hinderliter, Anne

    2011-01-01

    Lipid domain formation induced by annexin was investigated in mixtures of phosphatidylcholine (PC), phosphatidylserine (PS), and cholesterol (Chol), which were selected to mimic the inner leaflet of a eukaryotic plasma membrane. Annexins are ubiquitous and abundant cytoplasmic, peripheral proteins, which bind to membranes containing PS in the presence of calcium ions (Ca2+), but whose function is unknown. Prompted by indications of interplay between the presence of cholesterol in PS/PC mixtures and the binding of annexins, we used Monte Carlo simulations to investigate protein and lipid domain formation in these mixtures. The set of interaction parameters between lipids and proteins was assigned by matching experimental observables to corresponding variables in the calculations. In the case of monounsaturated phospholipids, the PS-PC and PC-Chol interactions are weakly repulsive. The interaction between protein and PS was determined based on experiments of annexin binding to PC/PS mixtures in the presence of Ca2+. Based on the proposal that PS and cholesterol form a complex in model membranes, a favorable PS-Chol interaction was postulated. Finally, protein-protein favorable interactions were also included, which are consistent with observations of large, two-dimensional, regular arrays of annexins on membranes. Those net interactions between pairs of lipids, proteins and lipids, and between proteins are all small, of the order of the average kinetic energy. We found that annexin a5 can induce formation of large PS domains, coincident with protein domains, but only if cholesterol is present. PMID:22004747

  17. Lipids in the assembly of membrane proteins and organization of protein supercomplexes: implications for lipid-linked disorders.

    PubMed

    Bogdanov, Mikhail; Mileykovskaya, Eugenia; Dowhan, William

    2008-01-01

    Lipids play important roles in cellular dysfunction leading to disease. Although a major role for phospholipids is in defining the membrane permeability barrier, phospholipids play a central role in a diverse range of cellular processes and therefore are important factors in cellular dysfunction and disease. This review is focused on the role of phospholipids in normal assembly and organization of the membrane proteins, multimeric protein complexes, and higher order supercomplexes. Since lipids have no catalytic activity, it is difficult to determine their function at the molecular level. Lipid function has generally been defined by affects on protein function or cellular processes. Molecular details derived from genetic, biochemical, and structural approaches are presented for involvement of phosphatidylethanolamine and cardiolipin in protein organization. Experimental evidence is presented that changes in phosphatidylethanolamine levels results in misfolding and topological misorientation of membrane proteins leading to dysfunctional proteins. Examples are presented for diseases in which proper protein folding or topological organization is not attained due to either demonstrated or proposed involvement of a lipid. Similar changes in cardiolipin levels affects the structure and function of individual components of the mitochondrial electron transport chain and their organization into supercomplexes resulting in reduced mitochondrial oxidative phosphorylation efficiency and apoptosis. Diseases in which mitochondrial dysfunction has been linked to reduced cardiolipin levels are described. Therefore, understanding the principles governing lipid-dependent assembly and organization of membrane proteins and protein complexes will be useful in developing novel therapeutic approaches for disorders in which lipids play an important role.

  18. MOLECULAR GENETIC AND BIOCHEMICAL APPROACHES FOR DEFINING LIPID-DEPENDENT MEMBRANE PROTEIN FOLDING

    PubMed Central

    Dowhan, William; Bogdanov, Mikhail

    2011-01-01

    We provide an overview of lipid-dependent polytopic membrane protein folding and topogenesis. Lipid dependence of this process was determined by employing Escherichia coli cells in which specific lipids can be eliminated, substituted, tightly titrated or controlled temporally during membrane protein synthesis and assembly. The secondary transport protein lactose permease (LacY) was used to establish general principles underlying the molecular basis of lipid-dependent effects on protein domain folding, protein transmembrane domain (TM) orientation, and function. These principles were then extended to several other secondary transport proteins of E. coli. The methods used to follow proper conformational organization of protein domains and the topological organization of protein TMs in whole cells and membranes are described. The proper folding of an extramembrane domain of LacY that is crucial for energy dependent uphill transport function depends on specific lipids acting as non-protein molecular chaperones. Correct TM topogenesis is dependent on charge interactions between the cytoplasmic surface of membrane proteins and a proper balance of the membrane surface net charge defined by the lipid head groups. Short-range interactions between the nascent protein chain and the translocon are necessary but not sufficient for establishment of final topology. After release from the translocon short-range interactions between lipid head groups and the nascent protein chain, partitioning of protein hydrophobic domains into the membrane bilayer, and long–range interactions within the protein thermodynamically drive final membrane protein organization. Given the diversity of membrane lipid compositions throughout nature, it is tempting to speculate that during the course of evolution the physical and chemical properties of proteins and lipids have co-evolved in the context of the lipid environment of membrane systems in which both are mutually depend on each other for

  19. One-dimensional potential of mean force underestimates activation barrier for transport across flexible lipid membranes

    NASA Astrophysics Data System (ADS)

    Kopelevich, Dmitry I.

    2013-10-01

    Transport of a fullerene-like nanoparticle across a lipid bilayer is investigated by coarse-grained molecular dynamics (MD) simulations. Potentials of mean force (PMF) acting on the nanoparticle in a flexible bilayer suspended in water and a bilayer restrained to a flat surface are computed by constrained MD simulations. The rate of the nanoparticle transport into the bilayer interior is predicted using one-dimensional Langevin models based on these PMFs. The predictions are compared with the transport rates obtained from a series of direct (unconstrained) MD simulations of the solute transport into the flexible bilayer. It is observed that the PMF acting on the solute in the flexible membrane underestimates the transport rate by more than an order of magnitude while the PMF acting on the solute in the restrained membrane yields an accurate estimate of the activation energy for transport into the flexible membrane. This paradox is explained by a coexistence of metastable membrane configurations for a range of the solute positions inside and near the flexible membrane. This leads to a significant reduction of the contribution of the transition state to the mean force acting on the solute. Restraining the membrane shape ensures that there is only one stable membrane configuration corresponding to each solute position and thus the transition state is adequately represented in the PMF. This mechanism is quite general and thus this phenomenon is expected to occur in a wide range of interfacial systems. A simple model for the free energy landscape of the coupled solute-membrane system is proposed and validated. This model explicitly accounts for effects of the membrane deformations on the solute transport and yields an accurate prediction of the activation energy for the solute transport.

  20. One-dimensional potential of mean force underestimates activation barrier for transport across flexible lipid membranes.

    PubMed

    Kopelevich, Dmitry I

    2013-10-07

    Transport of a fullerene-like nanoparticle across a lipid bilayer is investigated by coarse-grained molecular dynamics (MD) simulations. Potentials of mean force (PMF) acting on the nanoparticle in a flexible bilayer suspended in water and a bilayer restrained to a flat surface are computed by constrained MD simulations. The rate of the nanoparticle transport into the bilayer interior is predicted using one-dimensional Langevin models based on these PMFs. The predictions are compared with the transport rates obtained from a series of direct (unconstrained) MD simulations of the solute transport into the flexible bilayer. It is observed that the PMF acting on the solute in the flexible membrane underestimates the transport rate by more than an order of magnitude while the PMF acting on the solute in the restrained membrane yields an accurate estimate of the activation energy for transport into the flexible membrane. This paradox is explained by a coexistence of metastable membrane configurations for a range of the solute positions inside and near the flexible membrane. This leads to a significant reduction of the contribution of the transition state to the mean force acting on the solute. Restraining the membrane shape ensures that there is only one stable membrane configuration corresponding to each solute position and thus the transition state is adequately represented in the PMF. This mechanism is quite general and thus this phenomenon is expected to occur in a wide range of interfacial systems. A simple model for the free energy landscape of the coupled solute-membrane system is proposed and validated. This model explicitly accounts for effects of the membrane deformations on the solute transport and yields an accurate prediction of the activation energy for the solute transport.

  1. Interactions between HIV-1 Neutralizing Antibodies and Model Lipid Membranes imaged with AFM

    NASA Astrophysics Data System (ADS)

    Zauscher, Stefan; Hardy, Gregory; Alam, Munir; Shapter, Joseph

    2012-02-01

    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.

  2. Membrane interaction of antimicrobial peptides using E. coli lipid extract as model bacterial cell membranes and SFG spectroscopy.

    PubMed

    Soblosky, Lauren; Ramamoorthy, Ayyalusamy; Chen, Zhan

    2015-04-01

    Supported lipid bilayers are used as a convenient model cell membrane system to study biologically important molecule-lipid interactions in situ. However, the lipid bilayer models are often simple and the acquired results with these models may not provide all pertinent information related to a real cell membrane. In this work, we use sum frequency generation (SFG) vibrational spectroscopy to study molecular-level interactions between the antimicrobial peptides (AMPs) MSI-594, ovispirin-1 G18, magainin 2 and a simple 1,2-dipalmitoyl-d62-sn-glycero-3-phosphoglycerol (dDPPG)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) bilayer. We compared such interactions to those between the AMPs and a more complex dDPPG/Escherichia coli (E. coli) polar lipid extract bilayer. We show that to fully understand more complex aspects of peptide-bilayer interaction, such as interaction kinetics, a heterogeneous lipid composition is required, such as the E. coli polar lipid extract. The discrepancy in peptide-bilayer interaction is likely due in part to the difference in bilayer charge between the two systems since highly negative charged lipids can promote more favorable electrostatic interactions between the peptide and lipid bilayer. Results presented in this paper indicate that more complex model bilayers are needed to accurately analyze peptide-cell membrane interactions and demonstrates the importance of using an appropriate lipid composition to study AMP interaction properties.

  3. A model for surface diffusion of trans-membrane proteins on lipid bilayers

    NASA Astrophysics Data System (ADS)

    Agrawal, Ashutosh; Steigmann, David J.

    2011-06-01

    The equilibrium theory of lipid membranes is modified to include the effects of a continuous distribution of trans-membrane proteins. These influence membrane shape and evolve in accordance with a diffusive balance law. The model is purely mechanical in the absence of the proteins. Conditions ensuring energy dissipation in the presence of diffusion are given and an example constitutive function is used to simulate the coupled inertia-less interplay between membrane shape and protein distribution. The work extends an earlier continuum theory of equilibrium configurations of composite lipid-protein membranes to accommodate surface diffusion.

  4. Analysis and quantification of plant membrane lipids by thin-layer chromatography and gas chromatography.

    PubMed

    Wewer, Vera; Dörmann, Peter; Hölzl, Georg

    2013-01-01

    Galactolipids represent the predominant membrane lipid class in plants. In general, galactolipids are restricted to plastids, but during phosphate deficiency, they also accumulate in extraplastidial membranes. Two groups of plants can be distinguished based on the presence of a specific fatty acid, hexadecatrienoic acid (16:3), in chloroplast lipids. Plants that contain galactolipids with 16:3 acids are designated "16:3-plants"; the other group of plants which lack 16:3 contain mostly 18:3 in their galactolipids ("18:3-plants"). The methods in this chapter describe the extraction of membrane lipids from whole leaves, or from subcellular fractions, and their analysis via thin-layer chromatography (TLC) with different staining methods. Furthermore, a protocol for membrane lipid quantification is presented starting with the separation via TLC, transmethylation of the isolated lipids to fatty acid methyl esters, and their quantitative analysis via gas chromatography (GC).

  5. Interaction of anionic phenylene ethynylene polymers with lipids: from membrane embedding to liposome fusion.

    PubMed

    Karam, Pierre; Hariri, Amani A; Calver, Christina F; Zhao, Xiaoyong; Schanze, Kirk S; Cosa, Gonzalo

    2014-09-09

    Here we report spectroscopic studies on the interaction of negatively charged, amphiphilic polyphenylene ethynylene (PPE) polymers with liposomes prepared either from negative, positive or zwitterionic lipids. Emission spectra of PPEs of 7 and 49 average repeat units bearing carboxylate terminated side chains showed that the polymer embeds within positively charged lipids where it exists as free chains. No interaction was observed between PPEs and negatively charged lipids. Here the polymer remained aggregated giving rise to broad emission spectra characteristic of the aggregate species. In zwitterionic lipids, we observed that the majority of the polymer remained aggregated yet a small fraction readily embedded within the membrane. Titration experiments revealed that saturation of zwitterionic lipids with polymer typically occurred at a polymer repeat unit to lipid mole ratio close to 0.05. No further membrane embedding was observed above that point. For liposomes prepared from positively charged lipids, saturation was observed at a PPE repeat unit to lipid mole ratio of ∼0.1 and liposome precipitation was observed above this point. FRET studies showed that precipitation was preceded by lipid mixing and liposome fusion induced by the PPEs. This behavior was prominent for the longer polymer and negligible for the shorter polymer at a repeat unit to lipid mole ratio of 0.05. We postulate that fusion is the consequence of membrane destabilization whereby the longer polymer gives rise to more extensive membrane deformation than the shorter polymer.

  6. Accumulation of raft lipids in T-cell plasma membrane domains engaged in TCR signalling

    PubMed Central

    Zech, Tobias; Ejsing, Christer S; Gaus, Katharina; de Wet, Ben; Shevchenko, Andrej; Simons, Kai; Harder, Thomas

    2009-01-01

    Activating stimuli for T lymphocytes are transmitted through plasma membrane domains that form at T-cell antigen receptor (TCR) signalling foci. Here, we determined the molecular lipid composition of immunoisolated TCR activation domains. We observed that they accumulate cholesterol, sphingomyelin and saturated phosphatidylcholine species as compared with control plasma membrane fragments. This provides, for the first time, direct evidence that TCR activation domains comprise a distinct molecular lipid composition reminiscent of liquid-ordered raft phases in model membranes. Interestingly, TCR activation domains were also enriched in plasmenyl phosphatidylethanolamine and phosphatidylserine. Modulating the T-cell lipidome with polyunsaturated fatty acids impaired the plasma membrane condensation at TCR signalling foci and resulted in a perturbed molecular lipid composition. These results correlate the accumulation of specific molecular lipid species with the specific plasma membrane condensation at sites of TCR activation and with early TCR activation responses. PMID:19177148

  7. Communication: Activation energy of tension-induced pore formation in lipid membranes.

    PubMed

    Karal, Mohammad Abu Sayem; Yamazaki, Masahito

    2015-08-28

    Tension plays a vital role in pore formation in biomembranes, but the mechanism of pore formation remains unclear. We investigated the temperature dependence of the rate constant of constant tension (σ)-induced pore formation in giant unilamellar vesicles of lipid membranes using an experimental method we developed. By analyzing this result, we determined the activation energy (Ua) of tension-induced pore formation as a function of tension. A constant (U0) that does not depend on tension was found to contribute significantly to Ua. Analysis of the activation energy clearly indicated that the dependence of Ua on σ in the classical theory is correct, but that the classical theory of pore formation is not entirely correct due to the presence of U0. We can reasonably consider that U0 is a nucleation free energy to form a hydrophilic pre-pore from a hydrophobic pre-pore or a region with lower lateral lipid density. After obtaining U0, the evolution of a pre-pore follows a classical theory. Our data provide valuable information that help explain the mechanism of tension-induced pore formation in biomembranes and lipid membranes.

  8. Binding of Hemagglutinin and Influenza Virus to a Peptide-Conjugated Lipid Membrane

    PubMed Central

    Matsubara, Teruhiko; Shibata, Rabi; Sato, Toshinori

    2016-01-01

    Hemagglutinin (HA) plays an important role in the first step of influenza virus (IFV) infection because it initiates the binding of the virus to the sialylgalactose linkages of the receptors on the host cells. We herein demonstrate that a HA-binding peptide immobilized on a solid support is available to bind to HA and IFV. We previously obtained a HA-binding pentapeptide (Ala-Arg-Leu-Pro-Arg), which was identified by phage-display selection against HAs from random peptide libraries. This peptide binds to the receptor-binding site of HA by mimicking sialic acid. A peptide-conjugated lipid (pep-PE) was chemically synthesized from the peptide and a saturated phospholipid. A lipid bilayer composed of pep-PE and an unsaturated phospholipid (DOPC) was immobilized on a mica plate; and the interaction between HA and the pep-PE/DOPC membrane was investigated using atomic force microscopy. The binding of IFV to the pep-PE/DOPC membrane was detected by an enzyme-linked immunosorbent assay and real-time reverse transcription PCR. Our results indicate that peptide-conjugated lipids are a useful molecular device for the detection of HA and IFV. PMID:27092124

  9. Membrane lipid profiles of coral responded to zinc oxide nanoparticle-induced perturbations on the cellular membrane.

    PubMed

    Tang, Chuan-Ho; Lin, Ching-Yu; Lee, Shu-Hui; Wang, Wei-Hsien

    2017-03-31

    Zinc oxide nanoparticles (nZnOs) released from popular sunscreens used during marine recreation apparently endanger corals; however, the known biological effects are very limited. Membrane lipids constitute the basic structural element to create cell a dynamic structure according to the circumstance. Nano-specific effects have been shown to mechanically perturb the physical state of the lipid membrane, and the cells accommodating the actions of nZnOs can be involved in the alteration of the membrane lipid composition. To gain insight into the effects of nanoparticles on coral, glycerophosphocholine (GPC) profiling of the coral Seriatopora caliendrum exposed to nZnOs was performed in this study. Increasing lyso-GPCs, docosapentaenoic acid-possessing GPCs and docosahexaenoic acid-possessing GPCs and decreasing arachidonic acid-possessing GPCs were the predominant changes responded to nZnO exposure in the coral. A backfilling of polyunsaturated plasmanylcholines was observed in the coral exposed to nZnO levels over a threshold. These changes can be logically interpreted as an accommodation to nZnOs-induced mechanical disturbances in the cellular membrane based on the biophysical properties of the lipids. Moreover, the coral demonstrated a difference in the changes in lipid profiles between intra-colonial functionally differentiated polyps, indicating an initial membrane composition-dependent response. Based on the physicochemical properties and physiological functions of these changed lipids, some chronic biological effects can be incubated once the coral receives long-term exposure to nZnOs.

  10. Biophysics of cell membrane lipids in cancer drug resistance: Implications for drug transport and drug delivery with nanoparticles.

    PubMed

    Peetla, Chiranjeevi; Vijayaraghavalu, Sivakumar; Labhasetwar, Vinod

    2013-11-01

    In this review, we focus on the biophysics of cell membrane lipids, particularly when cancers develop acquired drug resistance, and how biophysical changes in resistant cell membrane influence drug transport and nanoparticle-mediated drug delivery. Recent advances in membrane lipid research show the varied roles of lipids in regulating membrane P-glycoprotein function, membrane trafficking, apoptotic pathways, drug transport, and endocytic functions, particularly endocytosis, the primary mechanism of cellular uptake of nanoparticle-based drug delivery systems. Since acquired drug resistance alters lipid biosynthesis, understanding the role of lipids in cell membrane biophysics and its effect on drug transport is critical for developing effective therapeutic and drug delivery approaches to overcome drug resistance. Here we discuss novel strategies for (a) modulating the biophysical properties of membrane lipids of resistant cells to facilitate drug transport and regain endocytic function and (b) developing effective nanoparticles based on their biophysical interactions with membrane lipids to enhance drug delivery and overcome drug resistance.

  11. Alteration of interleaflet coupling due to compounds displaying rapid translocation in lipid membranes

    PubMed Central

    Reigada, Ramon

    2016-01-01

    The spatial coincidence of lipid domains at both layers of the cell membrane is expected to play an important role in many cellular functions. Competition between the surface interleaflet tension and a line hydrophobic mismatch penalty are conjectured to determine the transversal behavior of laterally heterogeneous lipid membranes. Here, by a combination of molecular dynamics simulations, a continuum field theory and kinetic equations, I demonstrate that the presence of small, rapidly translocating molecules residing in the lipid bilayer may alter its transversal behavior by favoring the spatial coincidence of similar lipid phases. PMID:27596355

  12. Lipid Replacement Therapy: a natural medicine approach to replacing damaged lipids in cellular membranes and organelles and restoring function.

    PubMed

    Nicolson, Garth L; Ash, Michael E

    2014-06-01

    Lipid Replacement Therapy, the use of functional oral supplements containing cell membrane phospholipids and antioxidants, has been used to replace damaged, usually oxidized, membrane glycerophospholipids that accumulate during aging and in various clinical conditions in order to restore cellular function. This approach differs from other dietary and intravenous phospholipid interventions in the composition of phospholipids and their defense against oxidation during storage, ingestion, digestion and uptake as well as the use of protective molecules that noncovalently complex with phospholipid micelles and prevent their enzymatic and bile disruption. Once the phospholipids have been taken in by transport processes, they are protected by several natural mechanisms involving lipid receptors, transport and carrier molecules and circulating cells and lipoproteins until their delivery to tissues and cells where they can again be transferred to intracellular membranes by specific and nonspecific transport systems. Once delivered to membrane sites, they naturally replace and stimulate removal of damaged membrane lipids. Various chronic clinical conditions are characterized by membrane damage, mainly oxidative but also enzymatic, resulting in loss of cellular function. This is readily apparent in mitochondrial inner membranes where oxidative damage to phospholipids like cardiolipin and other molecules results in loss of trans-membrane potential, electron transport function and generation of high-energy molecules. Recent clinical trials have shown the benefits of Lipid Replacement Therapy in restoring mitochondrial function and reducing fatigue in aged subjects and patients with a variety of clinical diagnoses that are characterized by loss of mitochondrial function and include fatigue as a major symptom. This Article is Part of a Special Issue Entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy.

  13. Membrane Permeabilization Induced by Sphingosine: Effect of Negatively Charged Lipids

    PubMed Central

    Jiménez-Rojo, Noemi; Sot, Jesús; Viguera, Ana R.; Collado, M. Isabel; Torrecillas, Alejandro; Gómez-Fernández, J.C.; Goñi, Félix M.; Alonso, Alicia

    2014-01-01

    Sphingosine [(2S, 3R, 4E)-2-amino-4-octadecen-1, 3-diol] is the most common sphingoid long chain base in sphingolipids. It is the precursor of important cell signaling molecules, such as ceramides. In the last decade it has been shown to act itself as a potent metabolic signaling molecule, by activating a number of protein kinases. Moreover, sphingosine has been found to permeabilize phospholipid bilayers, giving rise to vesicle leakage. The present contribution intends to analyze the mechanism by which this bioactive lipid induces vesicle contents release, and the effect of negatively charged bilayers in the release process. Fluorescence lifetime measurements and confocal fluorescence microscopy have been applied to observe the mechanism of sphingosine efflux from large and giant unilamellar vesicles; a graded-release efflux has been detected. Additionally, stopped-flow measurements have shown that the rate of vesicle permeabilization increases with sphingosine concentration. Because at the physiological pH sphingosine has a net positive charge, its interaction with negatively charged phospholipids (e.g., bilayers containing phosphatidic acid together with sphingomyelins, phosphatidylethanolamine, and cholesterol) gives rise to a release of vesicular contents, faster than with electrically neutral bilayers. Furthermore, phosphorous 31-NMR and x-ray data show the capacity of sphingosine to facilitate the formation of nonbilayer (cubic phase) intermediates in negatively charged membranes. The data might explain the pathogenesis of Niemann-Pick type C1 disease. PMID:24940775

  14. LM cell growth and membrane lipid adaptation to sterol structure.

    PubMed

    Rujanavech, C; Silbert, D F

    1986-06-05

    Using a sterol auxotroph of the LM cell mouse fibroblast, we demonstrate that relatively few cholesterol analogues can substitute for cholesterol as a growth factor. The auxotroph grows normally on desmosterol and trans-22-dehydrocholesterol and at reduced rates on dihydrocholesterol, campesterol, and 22,23-dihydrobrassicasterol. It does not grow with beta-sitosterol, stigmasterol, ergosterol, or cis-22-dehydrocholesterol when the sterol is present as sole supplement but does grow at normal rates when the analogue is supplied with suboptimal amounts of cholesterol. Two contrasting types of membrane lipid changes are observed in cells grown on cholesterol analogues. In cells grown with dihydrocholesterol, a marked increase in desaturation and elongation of fatty acids is noted. Conversely, when cells are grown with cis-22-dehydrocholesterol, desaturation and elongation of fatty acids are severely curtailed. Cells grown on alkyl sterols respond like cells grown on cis-22-dehydrocholesterol but in a less pronounced fashion. The effects of sterol substitution in mammalian cells versus in lower eukaryotes are compared, and an explanation for the secondary changes in fatty acid composition in terms of phospholipid phase behavior is suggested.

  15. Femtosecond crystallography of membrane proteins in the lipidic cubic phase

    PubMed Central

    Liu, Wei; Wacker, Daniel; Wang, Chong; Abola, Enrique; Cherezov, Vadim

    2014-01-01

    Despite recent technological advances in heterologous expression, stabilization and crystallization of membrane proteins (MPs), their structural studies remain difficult and require new transformative approaches. During the past two years, crystallization in lipidic cubic phase (LCP) has started gaining a widespread acceptance, owing to the spectacular success in high-resolution structure determination of G protein-coupled receptors (GPCRs) and to the introduction of commercial instrumentation, tools and protocols. The recent appearance of X-ray free-electron lasers (XFELs) has enabled structure determination from substantially smaller crystals than previously possible with minimal effects of radiation damage, offering new exciting opportunities in structural biology. The unique properties of LCP material have been exploited to develop special protocols and devices that have established a new method of serial femtosecond crystallography of MPs in LCP (LCP-SFX). In this method, microcrystals are generated in LCP and streamed continuously inside the same media across the intersection with a pulsed XFEL beam at a flow rate that can be adjusted to minimize sample consumption. Pioneering studies that yielded the first room temperature GPCR structures, using a few hundred micrograms of purified protein, validate the LCP-SFX approach and make it attractive for structure determination of difficult-to-crystallize MPs and their complexes with interacting partners. Together with the potential of femtosecond data acquisition to interrogate unstable intermediate functional states of MPs, LCP-SFX holds promise to advance our understanding of this biomedically important class of proteins. PMID:24914147

  16. Chemical properties of lipids strongly affect the kinetics of the membrane-induced aggregation of α-synuclein

    PubMed Central

    Brown, James W. P.; Ouberai, Myriam M.; Flagmeier, Patrick; Vendruscolo, Michele; Buell, Alexander K.; Sparr, Emma; Dobson, Christopher M.

    2016-01-01

    Intracellular α-synuclein deposits, known as Lewy bodies, have been linked to a range of neurodegenerative disorders, including Parkinson’s disease. α-Synuclein binds to synthetic and biological lipids, and this interaction has been shown to play a crucial role for both α-synuclein’s native function, including synaptic plasticity, and the initiation of its aggregation. Here, we describe the interplay between the lipid properties and the lipid binding and aggregation propensity of α-synuclein. In particular, we have observed that the binding of α-synuclein to model membranes is much stronger when the latter is in the fluid rather than the gel phase, and that this binding induces a segregation of the lipids into protein-poor and protein-rich populations. In addition, α-synuclein was found to aggregate at detectable rates only when interacting with membranes composed of the most soluble lipids investigated here. Overall, our results show that the chemical properties of lipids determine whether or not the lipids can trigger the aggregation of α-synuclein, thus affecting the balance between functional and aberrant behavior of the protein. PMID:27298346

  17. Lipid and protein oxidation in hepatic homogenates and cell membranes exposed to bile acids.

    PubMed

    Fuentes-Broto, Lorena; Martínez-Ballarín, Enrique; Miana-Mena, Javier; Berzosa, Cesar; Piedrafita, Eduardo; Cebrián, Igor; Reiter, Russel J; García, Joaquín J

    2009-01-01

    Cholestasis occurs in a variety of hepatic diseases and causes damage due to accumulation of bile acids in the liver. The aim was to investigate the effect of several bile acids, i.e. chenodeoxycholic, taurochenodeoxycholic, deoxycholic, taurodeoxycholic, ursodeoxycholic, lithocholic and taurolithocholic (TLC), in inducing oxidative damage. Hepatic tissue of male Sprague-Dawley rats was incubated with or without 1 mM of each bile acid, with or without 0.1 mM FeCl(3) and 0.1 mM ascorbic acid for the purpose of generating free radicals. Several bile acids increased lipid and protein oxidation, with TLC being the most pro-oxidative (657% and 175% in homogenates and 350% and 311% in membranes, respectively). TLC also enhanced iron-induced oxidative stress to lipids (21% in homogenates and 29% in membranes) and to proteins (74% in membranes). This enhancement was dose- and time-dependent and was reduced by melatonin. These results suggest that bile acids differentially mediate hepatic oxidative stress and may be involved in the physiopathology of cholestasis.

  18. Probing the Interaction between Nanoparticles and Lipid Membranes by Quartz Crystal Microbalance with Dissipation Monitoring

    PubMed Central

    Yousefi, Nariman; Tufenkji, Nathalie

    2016-01-01

    There is increasing interest in using quartz crystal microbalance with dissipation monitoring (QCM-D) to investigate the interaction of nanoparticles (NPs) with model surfaces. The high sensitivity, ease of use and the ability to monitor interactions in real-time has made it a popular technique for colloid chemists, biologists, bioengineers, and biophysicists. QCM-D has been recently used to probe the interaction of NPs with supported lipid bilayers (SLBs) as model cell membranes. The interaction of NPs with SLBs is highly influenced by the quality of the lipid bilayers. Unlike many surface sensitive techniques, by using QCM-D, the quality of SLBs can be assessed in real-time, hence QCM-D studies on SLB-NP interactions are less prone to the artifacts arising from bilayers that are not well formed. The ease of use and commercial availability of a wide range of sensor surfaces also have made QCM-D a versatile tool for studying NP interactions with lipid bilayers. In this review, we summarize the state-of-the-art on QCM-D based techniques for probing the interactions of NPs with lipid bilayers. PMID:27995125

  19. Probing the interaction between nanoparticles and lipid membranes by quartz crystal microbalance with dissipation monitoring

    NASA Astrophysics Data System (ADS)

    Yousefi, Nariman; Tufenkji, Nathalie

    2016-12-01

    There is increasing interest in using quartz crystal microbalance with dissipation monitoring (QCM-D) to investigate the interaction of nanoparticles (NPs) with model surfaces. The high sensitivity, ease of use and the ability to monitor interactions in real-time has made it a popular technique for colloid chemists, biologists, bioengineers and biophysicists. QCM-D has been recently used to probe the interaction of NPs with supported lipid bilayers (SLBs) as model cell membranes. The interaction of NPs with SLBs is highly influenced by the quality of the lipid bilayers. Unlike many surface sensitive techniques, using QCM-D, the quality of SLBs can be assessed in real-time, hence QCM-D studies on SLB-NP interactions are less prone to the artefacts arising from bilayers that are not well formed. The ease of use and commercial availability of a wide range of sensor surfaces also have made QCM-D a versatile tool for studying NP interactions with lipid bilayers. In this review, we summarize the state-of-the-art on QCM-D based techniques for probing the interactions of NPs with lipid bilayers.

  20. Characterization of M-laurdan, a versatile probe to explore order in lipid membranes

    PubMed Central

    Mazeres, Serge; Joly, Etienne; Lopez, Andre; Tardin, Catherine

    2014-01-01

    Microdomains corresponding to localized partition of lipids between ordered and less ordered environments are the subject of intensive investigations, because of their putative participation in modulating cellular responses. One popular approach in the field consists in labelling membranes with solvatochromic fluorescent probes such as laurdan and C-laurdan. In this report, we describe a high-yield procedure for the synthesis of laurdan, C-laurdan and two new fluorophores, called MoC-laurdan and M-laurdan, as well as their extensive photophysical characterization. We find that the latter probe, M-laurdan, is particularly suited to discriminate lipid phases independently of the chemical nature of the lipids, as measured by both fluorescence Generalized Polarization (GP) and anisotropy in large unilamellar vesicles made of various lipid compositions. In addition, staining of live cells with M-laurdan shows a good stability over time without any apparent toxicity, as well as a wider distribution in the various cell compartments than the other probes. PMID:25485094

  1. Lipid demixing and protein-protein interactions in the adsorption of charged proteins on mixed membranes.

    PubMed Central

    May, S; Harries, D; Ben-Shaul, A

    2000-01-01

    The adsorption free energy of charged proteins on mixed membranes, containing varying amounts of (oppositely) charged lipids, is calculated based on a mean-field free energy expression that accounts explicitly for the ability of the lipids to demix locally, and for lateral interactions between the adsorbed proteins. Minimization of this free energy functional yields the familiar nonlinear Poisson-Boltzmann equation and the boundary condition at the membrane surface that allows for lipid charge rearrangement. These two self-consistent equations are solved simultaneously. The proteins are modeled as uniformly charged spheres and the (bare) membrane as an ideal two-dimensional binary mixture of charged and neutral lipids. Substantial variations in the lipid charge density profiles are found when highly charged proteins adsorb on weakly charged membranes; the lipids, at a certain demixing entropy penalty, adjust their concentration in the vicinity of the adsorbed protein to achieve optimal charge matching. Lateral repulsive interactions between the adsorbed proteins affect the lipid modulation profile and, at high densities, result in substantial lowering of the binding energy. Adsorption isotherms demonstrating the importance of lipid mobility and protein-protein interactions are calculated using an adsorption equation with a coverage-dependent binding constant. Typically, at bulk-surface equilibrium (i.e., when the membrane surface is "saturated" by adsorbed proteins), the membrane charges are "overcompensated" by the protein charges, because only about half of the protein charges (those on the hemispheres facing the membrane) are involved in charge neutralization. Finally, it is argued that the formation of lipid-protein domains may be enhanced by electrostatic adsorption of proteins, but its origin (e.g., elastic deformations associated with lipid demixing) is not purely electrostatic. PMID:11023883

  2. Arabidopsis chloroplast lipid transport protein TGD2 disrupts membranes and is part of a large complex.

    PubMed

    Roston, Rebecca; Gao, Jinpeng; Xu, Changcheng; Benning, Christoph

    2011-06-01

    In most plants the assembly of the photosynthetic thylakoid membrane requires lipid precursors synthesized at the endoplasmic reticulum (ER). Thus, the transport of lipids from the ER to the chloroplast is essential for biogenesis of the thylakoids. TGD2 is one of four proteins in Arabidopsis required for lipid import into the chloroplast, and was found to bind phosphatidic acid in vitro. However, the significance of phosphatidic acid binding for the function of TGD2 in vivo and TGD2 interaction with membranes remained unclear. Developing three functional assays probing how TGD2 affects lipid bilayers in vitro, we show that it perturbs membranes to the point of fusion, causes liposome leakage and redistributes lipids in the bilayer. By identifying and characterizing five new mutant alleles, we demonstrate that these functions are impaired in specific mutants with lipid phenotypes in vivo. At the structural level, we show that TGD2 is part of a protein complex larger than 500 kDa, the formation of which is disrupted in two mutant alleles, indicative of the biological relevance of this TGD2-containing complex. Based on the data presented, we propose that TGD2, as part of a larger complex, forms a lipid transport conduit between the inner and outer chloroplast envelope membranes, with its N terminus anchored in the inner membrane and its C terminus binding phosphatidic acid in the outer membrane.

  3. Interaction of LL-37 with Model Membrane Systems of Different Complexity: Influence of the Lipid Matrix

    PubMed Central

    Sevcsik, E.; Pabst, G.; Richter, W.; Danner, S.; Amenitsch, H.; Lohner, K.

    2008-01-01

    As the main difference between bacterial and mammalian cell membranes is their net charge, the focal point of consideration in many model membrane experiments with antimicrobial peptides is lipid headgroup charge. We studied the interaction of the human multifunctional peptide LL-37 with single phospholipid monolayers, bilayers, and bilayers composed of binary mixtures of the four phospholipid species predominantly used in model membrane experiments (phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylserine). We found that 1), the effects on single lipid monolayers are not comparable to those on the corresponding bilayers; 2), there are four different effects of LL-37 on bilayers of the four lipids; 3), the preference of LL-37 for the specific lipids is roughly inversely related to chain packing density; and 4), in the binary lipid mixtures, one lipid—and not necessarily the charged one—generally governs the mode of lipid/peptide interaction. Thus, our results show that lipid net charge is not the decisive factor determining the membrane-perturbing mechanism of LL-37, but only one of several parameters, among them packing density, the ability to form intermolecular H-bonds, and lipid molecular shape, which emphasizes how profoundly the choice of the model system can influence the outcome of a study of lipid/peptide interaction. PMID:18326643

  4. Dynamical Clustering and a Mechanism for Raft-like Structures in a Model Lipid Membrane

    PubMed Central

    Starr, Francis W.; Hartmann, Benedikt; Douglas, Jack F.

    2014-01-01

    We use molecular dynamics simulations to examine the dynamical heterogeneity of a model single-component lipid membrane using a coarse-grained representation of lipid molecules. This model qualitatively reproduces the known phase transitions between disordered, ordered, and gel membrane phases, and the phase transitions are accompanied by significant changes in the nature of the lipid dynamics. In particular, lipid diffusion in the liquid-ordered phase is hindered by the transient trapping of molecules by their neighbors, similar to the dynamics of a liquid approaching its glass transition. This transient molecular caging gives rise to two distinct mobility groups within a single-component membrane: lipids that are transiently trapped, and lipids with displacements on the scale of the intermolecular spacing. Most significantly, lipids within these distinct mobility states spatially segregate, creating transient “islands” of enhanced mobility having a size and time scale compatible with lipid “rafts,” dynamical structures thought to be important for cell membrane function. Although the dynamic lipid clusters that we observe do not themselves correspond to rafts (which are more complex, multicomponent structures), we hypothesize that such rafts may develop from the same universal mechanism, explaining why raft-like regions should arise, regardless of lipid structural or compositional details. These clusters are strikingly similar to the dynamical clusters found in glass-forming fluids, and distinct from phase-separation clusters. Further examination shows that mobile lipid clusters can be dissected into smaller clusters of cooperatively rearranging molecules. The geometry of these clusters can be understood in the context of branched equilibrium polymers, related to the statistics percolation theory. We discuss how these dynamical structures relate to a range observations on the dynamics of lipid membranes. PMID:24695573

  5. Steric Pressure among Membrane-Bound Polymers Opposes Lipid Phase Separation.

    PubMed

    Imam, Zachary I; Kenyon, Laura E; Carrillo, Adelita; Espinoza, Isai; Nagib, Fatema; Stachowiak, Jeanne C

    2016-04-19

    Lipid rafts are thought to be key organizers of membrane-protein complexes in cells. Many proteins that interact with rafts have bulky polymeric components such as intrinsically disordered protein domains and polysaccharide chains. Therefore, understanding the interaction between membrane domains and membrane-bound polymers provides insights into the roles rafts play in cells. Multiple studies have demonstrated that high concentrations of membrane-bound polymeric domains create significant lateral steric pressure at membrane surfaces. Furthermore, our recent work has shown that lateral steric pressure at membrane surfaces opposes the assembly of membrane domains. Building on these findings, here we report that membrane-bound polymers are potent suppressors of membrane phase separation, which can destabilize lipid domains with substantially greater efficiency than globular domains such as membrane-bound proteins. Specifically, we created giant vesicles with a ternary lipid composition, which separated into coexisting liquid ordered and disordered phases. Lipids with saturated tails and poly(ethylene glycol) (PEG) chains conjugated to their head groups were included at increasing molar concentrations. When these lipids were sparse on the membrane surface they partitioned to the liquid ordered phase. However, as they became more concentrated, the fraction of GUVs that were phase-separated decreased dramatically, ultimately yielding a population of homogeneous membrane vesicles. Experiments and physical modeling using compositions of increasing PEG molecular weight and lipid miscibility phase transition temperature demonstrate that longer polymers are the most efficient suppressors of membrane phase separation when the energetic barrier to lipid mixing is low. In contrast, as the miscibility transition temperature increases, longer polymers are more readily driven out of domains by the increased steric pressure. Therefore, the concentration of shorter polymers required

  6. Recent progress on lipid lateral heterogeneity in plasma membranes: From rafts to submicrometric domains.

    PubMed

    Carquin, Mélanie; D'Auria, Ludovic; Pollet, Hélène; Bongarzone, Ernesto R; Tyteca, Donatienne

    2016-04-01

    The concept of transient nanometric domains known as lipid rafts has brought interest to reassess the validity of the Singer-Nicolson model of a fluid bilayer for cell membranes. However, this new view is still insufficient to explain the cellular control of surface lipid diversity or membrane deformability. During the past decades, the hypothesis that some lipids form large (submicrometric/mesoscale vs nanometric rafts) and stable (>min vs s) membrane domains has emerged, largely based on indirect methods. Morphological evidence for stable submicrometric lipid domains, well-accepted for artificial and highly specialized biological membranes, was further reported for a variety of living cells from prokaryot es to yeast and mammalian cells. However, results remained questioned based on limitations of available fluorescent tools, use of poor lipid fixatives, and imaging artifacts due to non-resolved membrane projections. In this review, we will discuss recent evidence generated using powerful and innovative approaches such as lipid-specific toxin fragments that support the existence of submicrometric domains. We will integrate documented mechanisms involved in the formation and maintenance of these domains, and provide a perspective on their relevance on membrane deformability and regulation of membrane protein distribution.

  7. Autonomous transmembrane segment S4 of the voltage sensor domain partitions into the lipid membrane.

    PubMed

    Tiriveedhi, Venkataswarup; Miller, Melissa; Butko, Peter; Li, Min

    2012-07-01

    The S4 transmembrane segment in voltage-gated ion channels, a highly basic alpha helix, responds to changes in membrane potential and induces channel opening. Earlier work by others indicates that the S4 segment interacts with lipids in plasma membrane, but its mechanism is unclear. Working with synthetic tryptophan-labeled S4 peptides, we characterized binding of autonomous S4 to lipid membranes. The binding free energy (5.2 +/- 0.2 kcal/mol) of the peptide-lipid interaction was estimated from the apparent dissociation constants, determined from the changes in anisotropy of tryptophan fluorescence induced by addition of lipid vesicles with 30 mol% phosphatidylglycerol. The results are in good agreement with the prediction based on the Wimley-White hydrophobicity scale for interfacial (IF) binding of an alpha-helical peptide to the lipid bilayer (6.98 kcal/mol). High salt inhibited the interaction, thus indicating that the peptide/membrane interaction has both electrostatic and non-electrostatic components. Furthermore, the synthetic S4 corresponding to the Shaker potassium channel was found to spontaneously penetrate into the negatively charged lipid membrane to a depth of about 9 A. Our results revealed important biophysical parameters that influence the interaction of S4 with the membrane: they include fluidity, surface charge, and surface pressure of the membrane, and the at helicity and regular spacing of basic amino-acid residues in the S4 sequence.

  8. Recent progress on lipid lateral heterogeneity in plasma membranes: from rafts to submicrometric domains

    PubMed Central

    Carquin, Mélanie; D'Auria, Ludovic; Pollet, Hélène; Bongarzone, Ernesto R.; Tyteca, Donatienne

    2016-01-01

    The concept of transient nanometric domains known as lipid rafts has brought interest to reassess the validity of the Singer-Nicholson model of a fluid bilayer for cell membranes. However, this new view is still insufficient to explain the cellular control of surface lipid diversity or membrane deformability. During the past decade, the hypothesis that some lipids form large (submicrometric/mesoscale vs nanometric rafts) and stable (> min vs sec) membrane domains has emerged, largely based on indirect methods. Morphological evidence for stable submicrometric lipid domains, well-accepted for artificial and highly specialized biological membranes, was further reported for a variety of living cells from prokaryotes to yeast and mammalian cells. However, results remained questioned based on limitations of available fluorescent tools, use of poor lipid fixatives, and imaging artifacts due to non-resolved membrane projections. In this review, we will discuss recent evidence generated using powerful and innovative approaches such as lipid-specific toxin fragments that support the existence of submicrometric domains. We will integrate documented mechanisms involved in the formation and maintenance of these domains, and provide a perspective on their relevance on membrane deformability and regulation of membrane protein distribution. PMID:26738447

  9. Autonomous Transmembrane Segment S4 of the Voltage Sensor Domain Partitions into the Lipid Membrane

    PubMed Central

    Tiriveedhi, Venkataswarup; Miller, Melissa; Butko, Peter; Li, Min

    2012-01-01

    The S4 transmembrane segment in voltage-gated ion channels, a highly basic α helix, responds to changes in membrane potential and induces channel opening. Earlier work by others indicates that the S4 segment interacts with lipids in plasma membrane, but its mechanism is unclear. Working with synthetic tryptophan-labeled S4 peptides, we characterized binding of autonomous S4 to lipid membranes. The binding free energy (5.2 ± 0.2 kcal/mol) of the peptide-lipid interaction was estimated from the apparent dissociation constants, determined from the changes in anisotropy of tryptophan fluorescence induced by addition of lipid vesicles with 30 mol% phosphatidylglycerol. The results are in good agreement with the prediction based on the Wimley-White hydrophobicity scale for interfacial (IF) binding of an alpha-helical peptide to the lipid bilayer (6.98 kcal/mol). High salt inhibited the interaction, thus indicating that the peptide/membrane interaction has both electrostatic and non-electrostatic components. Furthermore, the synthetic S4 corresponding to the Shaker potassium channel was found to spontaneously penetrate into the negatively charged lipid membrane to a depth of about 9 Å. Our results revealed important biophysical parameters that influence the interaction of S4 with the membrane: they include fluidity, surface charge, and surface pressure of the membrane, and the α helicity and regular spacing of basic amino-acid residues in the S4 sequence. PMID:22465069

  10. Spherical Nanoparticle Supported Lipid Bilayers for the Structural Study of Membrane Geometry-Sensitive Molecules

    PubMed Central

    Kim, Edward Y.; Briley, Nicole E.; Tyndall, Erin R.; Xu, Jie; Li, Conggang; Ramamurthi, Kumaran S.; Flanagan, John M.; Tian, Fang

    2015-01-01

    Many essential cellular processes including endocytosis and vesicle trafficking require alteration of membrane geometry. These changes are usually mediated by proteins that can sense and/or induce membrane curvature. Using spherical nanoparticle supported lipid bilayers (SSLBs), we characterize how SpoVM, a bacterial development factor, interacts with differently curved membranes by magic angle spinning solid-state NMR. Our results demonstrate that SSLBs are an effective system for structural and topological studies of membrane geometry-sensitive molecules. PMID:26488086

  11. Responses of lipid membranes of taste sensor to astringent and pungent substances.

    PubMed

    Iiyama, S; Toko, K; Matsuno, T; Yamafuji, K

    1994-02-01

    Astringent substances and pungent substances were studied using a multichannel taste sensor with lipid membranes. The electric-potential pattern constructed of eight outputs from the membranes has information of taste quality and intensity. Pungent substances, such as capsaicin, piperine and allyl isothiocyanate, had no effect on the membrane potentials of the lipid membranes. On the other hand, astringent substances such as tannic acid, catechin, gallic acid and chlorogenic acid changed the potentials remarkably. A principal component analysis of the patterns in electric potential changes caused by the taste substances revealed the astringency is located between bitterness and sourness.

  12. Study of sweet taste evaluation using taste sensor with lipid/polymer membranes.

    PubMed

    Habara, Masaaki; Ikezaki, Hidekazu; Toko, Kiyoshi

    2004-07-15

    The higher sensitivity for sweeteners can be achieved by newly developed lipid/polymer membranes. The membrane is composed of lipids such as phosphoric acid di-n-hexadecyl ester and tetradodecylammoniumbromid, and a plasticizer, dioctyl phenylphosphonate. As a result of changing electric charge of the membrane surface, the newly developed membrane shows 5-10 times higher sensitivity for sucrose than the conventional ones. We also applied the sensor to other sugars such as sugar alcohol which is used as alternative sweetness or food additives. The experimental results of other sweeteners relatively correspond to human sensory evaluation, though the sensitivity for some sugars need to be improved.

  13. Differential effect of plant lipids on membrane organization: specificities of phytosphingolipids and phytosterols.

    PubMed

    Grosjean, Kevin; Mongrand, Sébastien; Beney, Laurent; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

    2015-02-27

    The high diversity of the plant lipid mixture raises the question of their respective involvement in the definition of membrane organization. This is particularly the case for plant plasma membrane, which is enriched in specific lipids, such as free and conjugated forms of phytosterols and typical phytosphingolipids, such as glycosylinositolphosphoceramides. This question was here addressed extensively by characterizing the order level of membrane from vesicles prepared using various plant lipid mixtures and labeled with an environment-sensitive probe. Fluorescence spectroscopy experiments showed that among major phytosterols, campesterol exhibits a stronger ability than β-sitosterol and stigmasterol to order model membranes. Multispectral confocal microscopy, allowing spatial analysis of membrane organization, demonstrated accordingly the strong ability of campesterol to promote ordered domain formation and to organize their spatial distribution at the membrane surface. Conjugated sterol forms, alone and in synergy with free sterols, exhibit a striking ability to order membrane. Plant sphingolipids, particularly glycosylinositolphosphoceramides, enhanced the sterol-induced ordering effect, emphasizing the formation and increasing the size of sterol-dependent ordered domains. Altogether, our results support a differential involvement of free and conjugated phytosterols in the formation of ordered domains and suggest that the diversity of plant lipids, allowing various local combinations of lipid species, could be a major contributor to membrane organization in particular through the formation of sphingolipid-sterol interacting domains.

  14. Specific Membrane Lipid Composition Is Important for Plasmodesmata Function in Arabidopsis

    PubMed Central

    Grison, Magali S.; Brocard, Lysiane; Fouillen, Laetitia; Nicolas, William; Wewer, Vera; Dörmann, Peter; Nacir, Houda; Benitez-Alfonso, Yoselin; Claverol, Stéphane; Germain, Véronique; Boutté, Yohann; Mongrand, Sébastien; Bayer, Emmanuelle M.

    2015-01-01

    Plasmodesmata (PD) are nano-sized membrane-lined channels controlling intercellular communication in plants. Although progress has been made in identifying PD proteins, the role played by major membrane constituents, such as the lipids, in defining specialized membrane domains in PD remains unknown. Through a rigorous isolation of “native” PD membrane fractions and comparative mass spectrometry-based analysis, we demonstrate that lipids are laterally segregated along the plasma membrane (PM) at the PD cell-to-cell junction in Arabidopsis thaliana. Remarkably, our results show that PD membranes display enrichment in sterols and sphingolipids with very long chain saturated fatty acids when compared with the bulk of the PM. Intriguingly, this lipid profile is reminiscent of detergent-insoluble membrane microdomains, although our approach is valuably detergent-free. Modulation of the overall sterol composition of young dividing cells reversibly impaired the PD localization of the glycosylphosphatidylinositol-anchored proteins Plasmodesmata Callose Binding 1 and the β-1,3-glucanase PdBG2 and altered callose-mediated PD permeability. Altogether, this study not only provides a comprehensive analysis of the lipid constituents of PD but also identifies a role for sterols in modulating cell-to-cell connectivity, possibly by establishing and maintaining the positional specificity of callose-modifying glycosylphosphatidylinositol proteins at PD. Our work emphasizes the importance of lipids in defining PD membranes. PMID:25818623

  15. Specific membrane lipid composition is important for plasmodesmata function in Arabidopsis.

    PubMed

    Grison, Magali S; Brocard, Lysiane; Fouillen, Laetitia; Nicolas, William; Wewer, Vera; Dörmann, Peter; Nacir, Houda; Benitez-Alfonso, Yoselin; Claverol, Stéphane; Germain, Véronique; Boutté, Yohann; Mongrand, Sébastien; Bayer, Emmanuelle M

    2015-04-01

    Plasmodesmata (PD) are nano-sized membrane-lined channels controlling intercellular communication in plants. Although progress has been made in identifying PD proteins, the role played by major membrane constituents, such as the lipids, in defining specialized membrane domains in PD remains unknown. Through a rigorous isolation of "native" PD membrane fractions and comparative mass spectrometry-based analysis, we demonstrate that lipids are laterally segregated along the plasma membrane (PM) at the PD cell-to-cell junction in Arabidopsis thaliana. Remarkably, our results show that PD membranes display enrichment in sterols and sphingolipids with very long chain saturated fatty acids when compared with the bulk of the PM. Intriguingly, this lipid profile is reminiscent of detergent-insoluble membrane microdomains, although our approach is valuably detergent-free. Modulation of the overall sterol composition of young dividing cells reversibly impaired the PD localization of the glycosylphosphatidylinositol-anchored proteins Plasmodesmata Callose Binding 1 and the β-1,3-glucanase PdBG2 and altered callose-mediated PD permeability. Altogether, this study not only provides a comprehensive analysis of the lipid constituents of PD but also identifies a role for sterols in modulating cell-to-cell connectivity, possibly by establishing and maintaining the positional specificity of callose-modifying glycosylphosphatidylinositol proteins at PD. Our work emphasizes the importance of lipids in defining PD membranes.

  16. Membrane lipids are key modulators of the endocannabinoid-hydrolase FAAH.

    PubMed

    Dainese, Enrico; De Fabritiis, Gianni; Sabatucci, Annalaura; Oddi, Sergio; Angelucci, Clotilde Beatrice; Di Pancrazio, Chiara; Giorgino, Toni; Stanley, Nathaniel; Del Carlo, Michele; Cravatt, Benjamin F; Maccarrone, Mauro

    2014-02-01

    Lipid composition is expected to play an important role in modulating membrane enzyme activity, in particular if the substrates are themselves lipid molecules. A paradigmatic case is FAAH (fatty acid amide hydrolase), an enzyme critical in terminating endocannabinoid signalling and an important therapeutic target. In the present study, using a combined experimental and computational approach, we show that membrane lipids modulate the structure, subcellular localization and activity of FAAH. We report that the FAAH dimer is stabilized by the lipid bilayer and shows a higher membrane-binding affinity and enzymatic activity within membranes containing both cholesterol and the natural FAAH substrate AEA (anandamide). Additionally, co-localization of cholesterol, AEA and FAAH in mouse neuroblastoma cells suggests a mechanism through which cholesterol increases the substrate accessibility of FAAH.

  17. Effect of lipid structural modifications on their intermolecular hydrogen bonding interactions and membrane functions.

    PubMed

    Boggs, J M

    1986-01-01

    The large number of different membrane lipids with various structural modifications and properties and the characteristic lipid composition of different types of membranes suggest that different lipids have specific functions in the membrane. Many of the varying properties of lipids with different polar head groups and in different ionization states can be attributed to the presence of interactive or repulsive forces between the head groups in the bilayer. The interactive forces are hydrogen bonds between hydrogen bond donating groups such as --P--OH,--OH, and--NH3+ and hydrogen bond accepting groups such as --P--O- and --COO-. These interactions increase the lipid phase transition temperature and can account for the tendency of certain lipids to go into the hexagonal phase and the dependence of this tendency on the pH and ionization state of the lipid. The presence or absence of these interactions can also affect the penetration of hydrophobic substances into the bilayer, including hydrophobic residues of membrane proteins. Evidence for this suggestion has been gathered from studies of the myelin basic protein, a water-soluble protein with a number of hydrophobic residues. In this way the lipid composition can affect the conformation and activity of membrane proteins. Since hydrogen-bonding interactions depend on the ionization state of the lipid, they can be altered by changes in the environment which affect the pK of the ionizable groups. The formation of the hexagonal phase or inverted micelles, the conformation and activity of membrane proteins, and other functions mediated by lipids could thus be regulated in this way.

  18. The selective interactions of cationic tetra-p-guanidinoethylcalix[4]arene with lipid membranes: theoretical and experimental model studies.

    PubMed

    Korchowiec, Beata; Gorczyca, Marcelina; Rogalska, Ewa; Regnouf-de-Vains, Jean-Bernard; Mourer, Maxime; Korchowiec, Jacek

    2016-01-07

    Behavior of cationic tetra-p-guanidinoethylcalix[4]arene (CX1) and its building block, p-guanidinoethylphenol (mCX1) in model monolayer lipid membranes was investigated using all atom molecular dynamics simulations and surface pressure measurements. Members of two classes of lipids were taken into account: zwitterionic 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and anionic 1,2-dimyristoyl-sn-glycero-3-phospho-l-serine sodium salt (DMPS) as models of eukaryotic and bacterial cell membranes, respectively. It was demonstrated that CX1 and mCX1 accumulate near the negatively charged DMPS monolayers. The adsorption to neutral monolayers was negligible. In contrast to mCX1, CX1 penetrated into the hydrophobic part of the monolayer. The latter effect, which is possible due to a flip-flop inversion of the CX1 orientation in the lipid layer compared to the aqueous phase, may be responsible for its antibacterial activity.

  19. Characterization of Membrane Protein-Lipid Interactions by Mass Spectrometry Ion Mobility Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Liu, Yang; Cong, Xiao; Liu, Wen; Laganowsky, Arthur

    2016-12-01

    Lipids in the biological membrane can modulate the structure and function of integral and peripheral membrane proteins. Distinguishing individual lipids that bind selectively to membrane protein complexes from an ensemble of lipid-bound species remains a daunting task. Recently, ion mobility mass spectrometry (IM-MS) has proven to be invaluable for interrogating the interactions between protein and individual lipids, where the complex undergoes collision induced unfolding followed by quantification of the unfolding pathway to assess the effect of these interactions. However, gas-phase unfolding experiments for membrane proteins are typically performed on the entire ensemble (apo and lipid bound species), raising uncertainty to the contribution of individual lipids and the species that are ejected in the unfolding process. Here, we describe the application of mass spectrometry ion mobility mass spectrometry (MS-IM-MS) for isolating ions corresponding to lipid-bound states of a model integral membrane protein, ammonia channel (AmtB) from Escherichia coli. Free of ensemble effects, MS-IM-MS reveals that bound lipids are ejected as neutral species; however, no correlation was found between the lipid-induced stabilization of complex and their equilibrium binding constants. In comparison to data obtained by IM-MS, there are surprisingly limited differences in stability measurements from IM-MS and MS-IM-MS. The approach described here to isolate ions of membrane protein complexes will be useful for other MS methods, such as surface induced dissociation or collision induced dissociation to determine the stoichiometry of hetero-oligomeric membrane protein complexes.

  20. Insulin Receptor Processing and Lipid Composition of Erythrocyte Membrane in Patients with Hyperlipidemia.

    PubMed

    Masella, R.; Cantafora, A.; Maffì, D.; Volpe, R.; Ginnetti, M.G.; Ricci, G.; Mace, N.L.; Buxton, G.M.; Peterson, S.W.

    1995-08-01

    The aim of this study was to determine whether the common forms of dyslipidemia could affect either the lipid composition or insulin receptor processing (down-regulation) of erythrocytes. The study included 22 patients with type IIa hypercholesterolemia, 15 patients with type IV hypertriglyceridemia and 12 patients with type IIb hyperlipidemia. Ten normolipidemic subjects were used as controls. Their erythrocyte membranes were analyzed for lipid composition and insulin receptor down-regulation. The results show that all the hyperlipidemias investigated were characterized by significant increases in the cholesterol to phospholipid molar ratio (0.56 +/- 0.08 in controls and 1.11 +/- 0.13, 1.09 +/- 0.14, 1.04 +/- 0.15, p < 0.001, in types IIa, IIb and IV, respectively). Surface insulin receptors of type IIa and IIb patients did not appear to down-regulate when compared to normal subjects, but rather up-regulated (+65.2% in controls, -1.0% and -8.7%, p < 0.001, in type IIa and IIb patients, respectively). Patients with type IV hypertriglyceridemia showed a residual capacity for insulin receptor internalization (10.7% down-regulation). Membranes of all the patients contained a higher proportion of phosphatidylethanolamine; the molar ratio of sphingomyelin to phosphatidylcholine was significantly higher in types IIb than in controls (1.22 +/- 0.11 and 1.12 +/- 0.10, p < 0.05, respectively); all the patients showed a lower content of polyunsaturated fatty acids in the major glycerophospholipid classes. However, type IV hypertriglyceridemics showed less variations, especially in the phosphatidylserine fraction. These results indicate that the alterations in lipoprotein pattern may affect both the lipid membrane equilibria and the processing ability of surface insulin receptors. Copyright 1995 S. Karger AG, Basel

  1. Immunocytochemical localization of acyl-lipid desaturases in cyanobacterial cells: evidence that both thylakoid membranes and cytoplasmic membranes are sites of lipid desaturation.

    PubMed Central

    Mustardy, L; Los, D A; Gombos, Z; Murata, N

    1996-01-01

    There are four acyl-lipid desaturases in the cyanobacterium Synechocystis sp. PCC 6803. Each of these desaturases introduces a double bond at a specific position, such as the Delta6, Delta9, Delta12, or omicron3 position, in C18 fatty acids. The localization of the desaturases in cyanobacterial cells was examined immunocytochemically with antibodies raised against synthetic oligopeptides that corresponded to the carboxyl-terminal regions of the desaturases. All four desaturases appeared to be located in the regions of both the cytoplasmic and the thylakoid membranes. These findings suggest that fatty acid desaturation of membrane lipids takes place in the thylakoid membranes as well as in the cytoplasmic membranes. Images Fig. 1 Fig. 2 Fig. 3 PMID:11607709

  2. Altered membrane lipid composition and functional parameters of circulating cells in cockles (Cerastoderma edule) affected by disseminated neoplasia.

    PubMed

    Le Grand, Fabienne; Soudant, Philippe; Marty, Yanic; Le Goïc, Nelly; Kraffe, Edouard

    2013-01-01

    Membrane lipid composition and morpho-functional parameters were investigated in circulating cells of the edible cockle (Cerastoderma edule) affected by disseminated neoplasia (neoplastic cells) and compared to those from healthy cockles (hemocytes). Membrane sterol levels, phospholipid (PL) class and subclass proportions and their respective fatty acid (FA) compositions were determined. Morpho-functional parameters were evaluated through total hemocyte count (THC), mortality rate, phagocytosis ability and reactive oxygen species (ROS) production. Both morpho-functional parameters and lipid composition were profoundly affected in neoplastic cells. These dedifferentiated cells displayed higher THC (5×), mortality rate (3×) and ROS production with addition of carbonyl cyanide m-chloro phenylhydrazone (1.7×) but lower phagocytosis ability (½×), than unaffected hemocytes. Total PL amounts were higher in neoplastic cells than in hemocytes (12.3 and 5.1 nmol×10(-6) cells, respectively). However, sterols and a particular subclass of PL (plasmalogens; 1-alkenyl-2-acyl PL) were present in similar amounts in both cell type membranes. This led to a two times lower proportion of these membrane lipid constituents in neoplastic cells when compared to hemocytes (20.5% vs. 42.1% of sterols in total membrane lipids and 21.7% vs. 44.2% of plasmalogens among total PL, respectively). Proportions of non-methylene interrupted FA- and 20:1n-11-plasmalogen molecular species were the most impacted in neoplastic cells when compared to hemocytes (⅓× and ¼×, respectively). These changes in response to this leukemia-like disease in bivalves highlight the specific imbalance of plasmalogens and sterols in neoplastic cells, in comparison to the greater stability of other membrane lipid components.

  3. Semiconductor particles in bilayer lipid membranes. Formation, characterization, and photoelectrochemistry

    SciTech Connect

    Zhao, X.K.; Baral, S.B.; Rolandi, R.; Fendler, J.H.

    1988-02-17

    Bilayer lipid membranes (BLMs) have been formed from bovine brain phosphatidylserine (PS), glyceryl monooleate (GMO), and a ploymerizable surfactant, (n-C/sub 15/H/sub 31/CO/sub 2/(CH/sub 2/))/sub 2/N/sup +/(CH/sub 3/)CH/sub 2/C/sub 6/H/sub 4/CH==CH/sub 2/Cl/sup -/(STYRS). These BLMs were then used to provide matrices for the in situ generation of microcrystalline CdS, CuS, Cu/sub 2/S, PbS, ZnS, HgS, and In/sub 2/S/sub 3/. Semiconductors were formed by injecting appropriate metal ion precursors and H/sub 2/S into the bathing solutions on opposite sides of the BLM. Their presence was established by voltage-dependent capacitance measurements, absorption spectroscopy, and optical microscopy. Subsequent to the injection of H/sub 2/S, the first observable change was the appearance of fairly uniform white dots on the black film. These dots rapidly moved around and grew in size, forming islands that then merged with themselves and with a second generation of dots, which ultimately led to a continuous film that continued to grow in thickness. Film formation and growth were monitored by simultaneous optical thickness and capacitance measurements. These data were treated in terms of an equivalent R-C circuit and allowed for the assessment of the semiconductor penetration depth into the BLM. This value for a GMO-BLM-supported In/sub 2/S/sub 3/ film was determined to be 24 A. Bandgap excitation, by nanosecond-pulsed or continuous illumination of the BLM-supported semiconductor film, led to observable photoelectric effects. Visible light (lambda > 350 nm) excitation into STYRS-BLM-supported CdS led to polymerization of the styrene moiety of STYRS. BLM-supported semiconductors remained stable for days.

  4. Different effects of Alzheimer's peptide Aβ(1-40) oligomers and fibrils on supported lipid membranes.

    PubMed

    Canale, Claudio; Seghezza, Silvia; Vilasi, Silvia; Carrotta, Rita; Bulone, Donatella; Diaspro, Alberto; San Biagio, Pier Luigi; Dante, Silvia

    2013-12-01

    Beta-amyloid (1-40) is one of the two most abundant species of amyloid-beta peptides present as fibrils in the extracellular senile plaques in the brain of Alzheimer's patients. Recently, the molecular aggregates constituting the early stage of fibril formation, i.e., oligomers and protofibrils, have been investigated as the main responsible for amyloid-beta cytotoxic effect. The molecular mechanism leading to neurodegeneration is still under debate, and it is common opinion that it may reside in the interaction between amyloid species and the neural membrane. In this investigation Atomic Force Microscopy and spectroscopy have been used to understand how structural (and mechanical) properties of POPC/POPS lipid bilayers, simulating the phospholipid composition and negative net charge of neuritic cell membranes, are influenced by the interaction with Aβ(1-40), in different stages of the peptide aggregation. Substantial differences in the damage caused to the lipid bilayers have been observed, confirming the toxic effect exerted especially by Aβ(1-40) prefibrillar oligomers.

  5. Modulation of folding and assembly of the membrane protein bacteriorhodopsin by intermolecular forces within the lipid bilayer.

    PubMed

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

    1999-07-20

    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.

  6. The effect of copper ions on the lipid composition of subcellular membranes in Hydrilla verticillata.

    PubMed

    Rozentsvet, Olga A; Nesterov, Viktor N; Sinyutina, Natalia F

    2012-09-01

    The paper studies changes in the content and composition of lipids in the membranes of chloroplasts, mitochondria and microsomes of the aquatic plant Hydrilla verticillata exposed to copper ions (100 μM; 1, 3, 6 and 24 h). The rate of copper accumulation and the coefficient of its extraction by the plant were also determined. The presence of copper in the incubation medium and its accumulation in the plant tissues decreased the content of photosynthetic pigments, stimulated lipid peroxidation and enhanced membrane permeability. The gradual accumulation of copper in the plant tissues was accompanied by specific changes in the composition of lipids: the content of sulfolipids (SQDG) in chloroplasts declined; the content of monogalactosyl diacylglycerols (MGDG), digalactosyl diacylglycerols (DGDG) and phosphatidyl glycerols (PG) in chloroplasts and mitochondria grew after an hour of copper exposure; and the content of all the lipids except phosphatidic acids (PA) decreased after 3 h of exposure. The decline in the content of phosphatidyl cholines (PC) was first observed in the membranes of microsomes (after an hour of exposure) and later in the membranes of chloroplasts and mitochondria (after 3-6 h of exposure). The experiments with incorporation of [2-(14)C]sodium acetate into fatty acids of polar lipids showed that in parallel with lipid destruction, there took place an intensive and specific renewal of the lipid pool of subcellular membrane fractions.

  7. Cholesterol expels ibuprofen from the hydrophobic membrane core and stabilizes lamellar phases in lipid membranes containing ibuprofen.

    PubMed

    Alsop, Richard J; Armstrong, Clare L; Maqbool, Amna; Toppozini, Laura; Dies, Hannah; Rheinstädter, Maikel C

    2015-06-28

    There is increasing evidence that common drugs, such as aspirin and ibuprofen, interact with lipid membranes. Ibuprofen is one of the most common over the counter drugs in the world, and is used for relief of pain and fever. It interacts with the cyclooxygenase pathway leading to inhibition of prostaglandin synthesis. From X-ray diffraction of highly oriented model membranes containing between 0 and 20 mol% ibuprofen, 20 mol% cholesterol, and dimyristoylphosphatidylcholine (DMPC), we present evidence for a non-specific interaction between ibuprofen and cholesterol in lipid bilayers. At a low ibuprofen concentrations of 2 mol%, three different populations of ibuprofen molecules were found: two in the lipid head group region and one in the hydrophobic membrane core. At higher ibuprofen concentrations of 10 and 20 mol%, the lamellar bilayer structure is disrupted and a lamellar to cubic phase transition was observed. In the presence of 20 mol% cholesterol, ibuprofen (at 5 mol%) was found to be expelled from the membrane core and reside solely in the head group region of the bilayers. 20 mol% cholesterol was found to stabilize lamellar membrane structure and the formation of a cubic phase at 10 and 20 mol% ibuprofen was suppressed. The results demonstrate that ibuprofen interacts with lipid membranes and that the interaction is strongly dependent on the presence of cholesterol.

  8. Determining Membrane Protein-Lipid Binding Thermodynamics Using Native Mass Spectrometry.

    PubMed

    Cong, Xiao; Liu, Yang; Liu, Wen; Liang, Xiaowen; Russell, David H; Laganowsky, Arthur

    2016-04-06

    Membrane proteins are embedded in the biological membrane where the chemically diverse lipid environment can modulate their structure and function. However, the thermodynamics governing the molecular recognition and interaction of lipids with membrane proteins is poorly understood. Here, we report a method using native mass spectrometry (MS), to determine thermodynamics of individual ligand binding events to proteins. Unlike conventional methods, native MS can resolve individual ligand binding events and, coupled with an apparatus to control the temperature, determine binding thermodynamic parameters, such as for protein-lipid interactions. We validated our approach using three soluble protein-ligand systems (maltose binding protein, lysozyme, and nitrogen regulatory protein) and obtained similar results to those using isothermal titration calorimetry and surface plasmon resonance. We also determined for the first time the thermodynamics of individual lipid binding to the ammonia channel (AmtB), an integral membrane protein from Escherichia coli. Remarkably, we observed distinct thermodynamic signatures for the binding of different lipids and entropy-enthalpy compensation for binding lipids of variable chain length. Additionally, using a mutant form of AmtB that abolishes a specific phosphatidylglycerol (PG) binding site, we observed distinct changes in the thermodynamic signatures for binding PG, implying these signatures can identify key residues involved in specific lipid binding and potentially differentiate between specific lipid binding sites.

  9. The interaction of mefloquine hydrochloride with cell membrane models at the air-water interface is modulated by the monolayer lipid composition.

    PubMed

    Goto, Thiago Eichi; Caseli, Luciano

    2014-10-01

    The antiparasitic properties of antiparasitic drugs are believed to be associated with their interactions with the protozoan membrane, encouraging research on the identification of membrane sites capable of drug binding. In this study, we investigated the interaction of mefloquine hydrochloride, known to be effective against malaria, with cell membrane models represented by Langmuir monolayers of selected lipids. It is shown that even small amounts of the drug affect the surface pressure-area isotherms as well as surface vibrational spectra of some lipid monolayers, which points to a significant interaction. The effects on the latter depend on the electrical charge of the monolayer-forming molecules, with the drug activity being particularly distinctive for negatively charged lipids. Therefore, the lipid composition of the monolayer modulates the interaction with the lipophilic drug, which may have important implications in understanding how the drug acts on specific sites of the protozoan membrane.

  10. Removal of lipopolysaccharide from protein solution using nanostructured porous supports bearing lipid membranes

    NASA Astrophysics Data System (ADS)

    Wakita, Masa-aki

    2013-11-01

    Polymeric lipid membranes of N-octadecylchitosan, which consists of 70 mol% of 2-(octadecylamino)-2-deoxy- d-glucopyranose, 17 mol% of 2-amino-2-deoxy- d-glucopyranose, and 13 mol% of 2-acetamido-2-deoxy- d-glucopyranose, were covalently immobilized to carboxylated porous supports composed of chitosan and used for the adsorption of pyrogenic lipopolysaccharide. When human serum albumin solution, including 5 mg mL-1 of albumin and 5.6 ng mL-1 of lipopolysaccharide, was passed through a column packed with the resulting porous supports bearing lipid membranes assembled in nanoscale, lipopolysaccharide was removed to as low as a detection limit of 0.020 ng mL-1 with a quantitative recovery of protein. On the other hand, in the case of directly N-octadecylated porous supports having cationic and hydrophobic ligands which are not assembled as lipid membranes, lipopolysaccharide could not be removed to the detection limit and protein recovery was lower than the porous supports bearing lipid membranes. The difference above as well as difference from conventional adsorbents suggested that the selectivity was attributable to an interaction between the cationic lipid membranes of N-octadecylchitosan and lipopolysaccharide as well as protein. The porous supports bearing lipid membranes were stable in 0.5 M NaOH and 0.1 M HCl at ambient temperature. Considering the confirmed excellent selectivity and chemical stability, their practical use as separation media in the pharmaceutical manufacturing can be expected.

  11. Quantitative electron microscopy for the nanoscale analysis of membrane lipid distribution.

    PubMed

    Fujita, Akikazu; Cheng, Jinglei; Fujimoto, Toyoshi

    2010-04-01

    An important goal of membrane biology is to define the local heterogeneity of membrane lipid composition. Here we describe a quantitative electron microscopic method that enables the localization of specific membrane lipids at the nanometer scale. The method involves freezing cells rapidly to halt the molecular motion, physically stabilizing membrane molecules in the freeze-fracture replica by the deposition of evaporated platinum and carbon layers and labeling with specific probes for electron microscopic observation. Lipids in both the outer and inner membrane leaflets can thus be labeled, and their distributions can be analyzed quantitatively by statistical methods. A major advantage of this method is that it does not require the expression of artificial probes. Therefore, this method can be applied to any cell in vitro or in vivo, and the whole procedure can be completed in 1-2 d.

  12. Red cell membrane lipid changes at 3,500 m and on return to sea level.

    PubMed

    González, Gustavo; Celedón, Gloria; Escobar, Marcela; Sotomayor, Carlos; Ferrer, Verónica; Benítez, Dixan; Behn, Claus

    2005-01-01

    Previous studies have shown that acute hypobaric hypoxia, obtained in a hypobaric chamber, and subsequent reoxygenation, give rise to modifications of the erythrocyte membrane lipid dynamics, resulting in an increased lateral diffusivity of the membrane lipids, and this was interpreted as the result of a modified lipid-protein interaction. The aim of the present study was to determine the effect of the reoxygenation condition in individuals after 3 days at an altitude of 3,500 m above sea level. Reoxygenation was a consequence of returning to sea level. Resting blood samples from both conditions were obtained, and erythrocytes were separated and immediately lysed for membrane isolation. We measured the bilayer polarity in membranes with Laurdan, a fluorescent probe. We also measured malondialdehyde in membrane lipids, an indicator of oxidative damage. We found a 12% (p = 0.016, n = 7) increase in the polarity of the membrane bilayer surface, and an increase of 70% (p = 0.005, n = 7) in the formation of malondialdehyde in the membrane after the reoxygenation condition. The membrane bilayer polarity increase is due to an oxidative modification of the phospholipid backbone after reoxygenation. People working and/or recreating at moderate altitude (3,500 m) may be at risk of erythrocyte membrane oxidative damage upon returning to sea level, and therefore a better understanding of the processes occurring upon reoxygenation may lead to proposed strategies to minimize this effect.

  13. Mapping of Membrane Lipid Order in Root Apex Zones of Arabidopsis thaliana

    PubMed Central

    Zhao, Xiaoyu; Zhang, Xiran; Qu, Yanli; Li, Ruili; Baluška, František; Wan, Yinglang

    2015-01-01

    In this study, we used the fluorescence probe, Di-4-ANEPPDHQ, to map the distribution of membrane lipid order in the apical region of Arabidopsis roots. The generalized polarization (GP) value of Di-4-ANEPPDHQ-stained roots indicated the highest lipid order in the root transition zone (RTZ). The cortical cells have higher lipid order than the epidermal cells in same regions, while the developing root hairs show very prominent cell polarity with high lipid order in apical region. Moreover, the endosomes had lower lipid order than that of the plasma membrane (PM). Brefeldin A (BFA) treatment decreased the lipid order in both the plasma and endosomal membranes of epidermal cells in the RTZ. The lipid order of BFA-induced compartments became higher than that of the PM after BFA treatment in epidermal cells. Meanwhile, the polarly growing tips of root hairs did not show the same behavior. The lipid order of the PM remained unchanged, with higher values than that of the endosomes. This suggests that the lipid ordering in the PM was affected by recycling of endosomal vesicles in epidermal cells of the root apex transition zone but not in the root hairs of Arabidopsis. PMID:26734047

  14. [Effect of microwaves on bilayer lipid membranes: role of a membrane-forming hole in the Teflon film].

    PubMed

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

    2009-01-01

    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.

  15. Structure formation of lipid membranes: Membrane self-assembly and vesicle opening-up to octopus-like micelles

    NASA Astrophysics Data System (ADS)

    Noguchi, Hiroshi

    2013-02-01

    We briefly review our recent studies on self-assembly and vesicle rupture of lipid membranes using coarse-grained molecular simulations. For single component membranes, lipid molecules self-assemble from random gas states to vesicles via disk-shaped clusters. Clusters aggregate into larger clusters, and subsequently the large disks close into vesicles. The size of vesicles are determined by kinetics than by thermodynamics. When a vesicle composed of lipid and detergent types of molecules is ruptured, a disk-shaped micelle called bicelle can be formed. When both surfactants have negligibly low critical micelle concentration, it is found that bicelles connected with worm-like micelles are also formed depending on the surfactant ratio and spontaneous curvature of the membrane monolayer.

  16. Lipid and fatty acid composition of Gluconobacter oxydans before and after intracytoplasmic membrane formation.

    PubMed Central

    Heefner, D L; Claus, G W

    1978-01-01

    Gluconobacter oxydans differentiates by forming quantities of intracytoplasmic membranes at the end of exponential growth, and this formation occurs concurrently with a 60% increase in cellular lipid. The present study was initiated to determine whether this newly synthesized lipid differed from that extracted before intracytoplasmic membrane synthesis. Undifferentiated exponential-phase cells were found to contain 30% phosphatidylcholine, 27.1% caridolipin, 25% phosphatidylethanolamine, 12.5% phosphatidylglycerol, 0.4% phosphatidic acid, 0.2% phosphatidylserine, and four additional unidentified lipids totaling less than 5%. The only change detected after formation of intracytoplasmic membranes was a slight decrease in phosphatidylethanolamine and a corresponding increase in phosphatidylcholine. An examination of lipid hydrolysates revealed 11 different fatty acids in the lipids from each cell type. Hexadecanoic acid and monounsaturated octadecenoic accounted for more than 75% of the total fatty acids for both cell types. Proportional changes were noted in all fatty acids except octadecenoate. Anteiso-pentadecanoate comprised less than 1% of the fatty acids from undifferentiated cells but more than 13% of the total fatty acids from cells containing intracytoplasmic membranes. These results suggest that anteiso-pentadecanoate formation closely parallels the formation of intracytoplasmic membranes. Increased concentrations of this fatty acid may contribute to the fluidity necessary for plasma membrane convolution during intracytoplasmic membrane development. PMID:649571

  17. Membrane lipids in Agrobacterium tumefaciens: biosynthetic pathways and importance for pathogenesis

    PubMed Central

    Aktas, Meriyem; Danne, Linna; Möller, Philip; Narberhaus, Franz

    2014-01-01

    Many cellular processes critically depend on the membrane composition. In this review, we focus on the biosynthesis and physiological roles of membrane lipids in the plant pathogen Agrobacterium tumefaciens. The major components of A. tumefaciens membranes are the phospholipids (PLs), phosphatidylethanolamine (PE), phosphatidylglycerol, phosphatidylcholine (PC) and cardiolipin, and ornithine lipids (OLs). Under phosphate-limited conditions, the membrane composition shifts to phosphate-free lipids like glycolipids, OLs and a betaine lipid. Remarkably, PC and OLs have opposing effects on virulence of A. tumefaciens. OL-lacking A. tumefaciens mutants form tumors on the host plant earlier than the wild type suggesting a reduced host defense response in the absence of OLs. In contrast, A. tumefaciens is compromised in tumor formation in the absence of PC. In general, PC is a rare component of bacterial membranes but amount to ~22% of all PLs in A. tumefaciens. PC biosynthesis occurs via two pathways. The phospholipid N-methyltransferase PmtA methylates PE via the intermediates monomethyl-PE and dimethyl-PE to PC. In the second pathway, the membrane-integral enzyme PC synthase (Pcs) condenses choline with CDP-diacylglycerol to PC. Apart from the virulence defect, PC-deficient A. tumefaciens pmtA and pcs double mutants show reduced motility, enhanced biofilm formation and increased sensitivity towards detergent and thermal stress. In summary, there is cumulative evidence that the membrane lipid composition of A. tumefaciens is critical for agrobacterial physiology and tumor formation. PMID:24723930

  18. An ultrasensitive tool exploiting hydration dynamics to decipher weak lipid membrane-polymer interactions.

    PubMed

    Cheng, Chi-Yuan; Wang, Jia-Yu; Kausik, Ravinath; Lee, Ka Yee C; Han, Songi

    2012-02-01

    We introduce a newly developed tool, (1)H Overhauser Dynamic Nuclear Polarization (ODNP), to sensitively explore weak macromolecular interactions by site-specifically probing the modulation of the translational dynamics of hydration water at the interaction interface, in the full presence of bulk water. Here, ODNP is employed on an illustrative example of a membrane-active triblock copolymer, poloxamer 188 (P188), which is known to restore the integrity of structurally compromised cell membranes. We observe a distinct change in the translational dynamics of the hydration layer interacting with the lipid membrane surface and the bilayer-interior as P188 is added to a solution of lipid vesicles, but no measurable changes in the dynamics or structure of the lipid membranes. This study shows that hydration water is an integral constituent of a lipid membrane system, and demonstrates for the first time that the modulation of its translational diffusivity can sensitively report on weak polymer-membrane interactions, as well as mediate essential lipid membrane functions. ODNP holds much promise as a unique tool to unravel molecular interactions at interfaces even in the presence of bulk water under ambient conditions.

  19. Changes in plasma membrane lipids, aquaporins and proton pump of broccoli roots, as an adaptation mechanism to salinity.

    PubMed

    López-Pérez, Luis; Martínez-Ballesta, María Del Carmen; Maurel, Christophe; Carvajal, Micaela

    2009-03-01

    Salinity stress is known to modify the plasma membrane lipid and protein composition of plant cells. In this work, we determined the effects of salt stress on the lipid composition of broccoli root plasma membrane vesicles and investigated how these changes could affect water transport via aquaporins. Brassica oleracea L. var. Italica plants treated with different levels of NaCl (0, 40 or 80mM) showed significant differences in sterol and fatty acid levels. Salinity increased linoleic (18:2) and linolenic (18:3) acids and stigmasterol, but decreased palmitoleic (16:1) and oleic (18:1) acids and sitosterol. Also, the unsaturation index increased with salinity. Salinity increased the expression of aquaporins of the PIP1 and PIP2 subfamilies and the activity of the plasma membrane H(+)-ATPase. However, there was no effect of NaCl on water permeability (P(f)) values of root plasma membrane vesicles, as determined by stopped-flow light scattering. The counteracting changes in lipid composition and aquaporin expression observed in NaCl-treated plants could allow to maintain the membrane permeability to water and a higher H(+)-ATPase activity, thereby helping to reduce partially the Na(+) concentration in the cytoplasm of the cell while maintaining water uptake via cell-to-cell pathways. We propose that the modification of lipid composition could affect membrane stability and the abundance or activity of plasma membrane proteins such as aquaporins or H(+)-ATPase. This would provide a mechanism for controlling water permeability and for acclimation to salinity stress.

  20. Rigid lipid membranes and nanometer clefts: motifs for the creation of molecular landscapes.

    PubMed

    Li, Guangtao; Fudickar, Werner; Skupin, Marc; Klyszcz, Andreas; Draeger, Christian; Lauer, Matthias; Fuhrhop, Jürgen-Hinrich

    2002-06-03

    Amphiphilic lipids associate in water spontaneously to form micelles, vesicles, monolayers, or biological membranes. These aggregates are soft and their shape can be changed easily. They behave like complex fluids because they are merely held together by weak, nondirected forces. The most important characteristic of these monolayers is their ability to dissolve hydrophobic molecules in the form of freely movable monomers. The fluid molecular layers are not suitable to anchor the components of chain reactions. However, if the alkyl chains are replaced by rigid skeletons or if the head groups are connected through intermolecular interactions, the aggregates become rigid and their fluid solvent character is lost. The construction of chiral surfaces by synkinesis (synthesis of noncovalent compounds) and of enzyme-type surface clefts of defined size can now be carried out by using rigid lipid membranes. Monolayers and nanometer pores on solid substrates attain sharp edges, and upright nanometer columns on smooth surfaces no longer dissipate. Five examples illustrate the advantages of using rigid molecular assemblies: 1) Cationic domains of rigid edge amphiphiles in fluid membranes act as manipulable ion channels. 2) Spherical micelles, micellar helical fibers, and vesicular tubes can be dried and stored as stable material. Molecular landscapes form on smooth surfaces. 3) alpha,omega-Diamide bolaamphiphiles form rigid nanometer-thick walls on smooth surfaces and these barriers cannot be penetrated by amines. Around steroids and porphyrins, they form rigid nanometer clefts whose walls and water-filled centers can be functionalized. 4) The structure of rigid oligophenylene- and quinone monolayers on electrodes can be changed drastically and reversibly by changing the potential. 5) 10(10) Porphyrin cones on a 1-cm2 gold electrode can be controlled individually by AFM- and STM-tips and investigated by electrochemical, photochemical, and mechanical means. In summary, rigid

  1. Lipopolysaccharide-Induced Dynamic Lipid Membrane Reorganization: Tubules, Perforations, and Stacks

    PubMed Central

    Adams, Peter G.; Lamoureux, Loreen; Swingle, Kirstie L.; Mukundan, Harshini; Montaño, Gabriel A.

    2014-01-01

    Lipopolysaccharide (LPS) is a unique lipoglycan, with two major physiological roles: 1), as a major structural component of the outer membrane of Gram-negative bacteria and 2), as a highly potent mammalian toxin when released from cells into solution (endotoxin). LPS is an amphiphile that spontaneously inserts into the outer leaflet of lipid bilayers to bury its hydrophobic lipidic domain, leaving the hydrophilic polysaccharide chain exposed to the exterior polar solvent. Divalent cations have long been known to neutralize and stabilize LPS in the outer membrane, whereas LPS in the presence of monovalent cations forms highly mobile negatively-charged aggregates. Yet, much of our understanding of LPS and its interactions with the cell membrane does not take into account its amphiphilic biochemistry and charge polarization. Herein, we report fluorescence microscopy and atomic force microscopy analysis of the interaction between LPS and fluid-phase supported lipid bilayer assemblies (sLBAs), as model membranes. Depending on cation availability, LPS induces three remarkably different effects on simple sLBAs. Net-negative LPS-Na+ leads to the formation of 100-μm-long flexible lipid tubules from surface-associated lipid vesicles and the destabilization of the sLBA resulting in micron-size hole formation. Neutral LPS-Ca2+ gives rise to 100-μm-wide single- or multilamellar planar sheets of lipid and LPS formed from surface-associated lipid vesicles. Our findings have important implications about the physical interactions between LPS and lipids and demonstrate that sLBAs can be useful platforms to study the interactions of amphiphilic virulence factors with cell membranes. Additionally, our study supports the general phenomenon that lipids with highly charged or bulky headgroups can promote highly curved membrane architectures due to electrostatic and/or steric repulsions. PMID:24896118

  2. Punching Holes in Membranes: How Oligomeric Pore-Forming Proteins and Lipids Cooperate to Form Aqueous Channels in Membranes

    NASA Astrophysics Data System (ADS)

    Fradin, Cécile; Satsoura, Dmitri; Andrews, David W.

    Many important biological processes are carried out by a small number of proteins working together as a team to accomplish a specific task. Cooperation between the different proteins is often accomplished through the formation of a supramolecular complex, comprised of either identical or different subunits. Although the formation of protein assemblies is a favored mechanism throughout the cell, it becomes especially important in lipid membranes, as evidenced by the numerous cellular events that are either triggered by or result in the formation of protein complexes in membranes. However, due to the difficulties associated with the study of membrane proteins, the formation of oligomers in lipid membranes is perhaps one of the least understood cellular processes. In this chapter we focus our attention on a subset of membrane complexes — namely, those formed by proteins that are able to pass from a water-soluble to a transmembrane form in order to create a water-filled channel through the lipid membrane. These pore-forming proteins (PFPs) are found in many organisms throughout different kingdoms of life, from bacteria to human. They are often involved in cell death mechanisms through their capacity to break membrane permeability barriers, which can lead to dissipation of the membrane potential as well as introduction or leakage of enzymatic proteins. In fact, a large subset of the PFPs are toxins, and referred to in the literature as pore-forming toxins (PFTs). The association of several monomers into an oligomer is almost always an important aspect of the modus operandi of these proteins. Oligomerization can be useful in several ways: it results in structures large enough to delineate nanometer-size water-filled channels in lipid bilayers, it ensures the presence of large hydrophobic surfaces that can support insertion in the membrane, and it permits cooperative formation and insertion mechanisms.

  3. Membrane Binding of HIV-1 Matrix Protein: Dependence on Bilayer Composition and Protein Lipidation

    PubMed Central

    Barros, Marilia; Nanda, Hirsh

    2016-01-01

    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

  4. Receptor-mediated membrane adhesion of lipid-polymer hybrid (LPH) nanoparticles studied by dissipative particle dynamics simulations.

    PubMed

    Li, Zhenlong; Gorfe, Alemayehu A

    2015-01-14

    Lipid-polymer hybrid (LPH) nanoparticles represent a novel class of targeted drug delivery platforms that combine the advantages of liposomes and biodegrad