Biomembrane Function Research Articles

‘New dimensions for manufacturing’ — A UK strategy for nanotechnology

This chapter discusses the nanotechnology and nano engineered membranes. Self-assembly of molecules or large groups of molecules is a key feature in nanotechnology. Nanotechnology deals with structures having at least 1D that is sufficiently small, of the order of about one to several hundred nanometers. It is concerned with materials and systems whose structures and components exhibit novel and improved physical, chemical, and biological properties. It enables the exploitation of novel phenomena and processes because of their nanoscale size. The chapter presents some examples of nano and micro engineered membrane technology and its use for nano engineered devices. A technique to pattern surfaces is to evaporate a target material through a thin membrane (shadow mask) with well-defined perforations. Photonic crystals are artificially three-dimensional structures fabricated in an optical material (crystal or amorphous) with unit cells whose dimensions are comparable to the optical wavelength. Nano and microsieves can be used as stencils for patterning, as photonic crystals, but they can also be used as a proper solid support for thin functional membranes. Nanotubes are the hollow cylindrical structures. The chapter discusses the nano printing and etching with phase separated membranes. Research goal in biomembrane science is to study the function of natural biomembranes and to engineer new types of biomembrane materials on the nanometer scale.

Peptide Aggregation in Lipid Bilayers — Is the Peptide Really in the Striated Domains?

In order to elucidate the structure and function of biomembranes, it is useful to investigate interactions between lipids and proteins, which are the main building blocks of all biological membranes. Supported lipid/peptide bilayers, which can be prepared, using established protocols, are a widely used model system to perform these investigations. Atomic Force Microscopy (AFM) offers a unique possibility to investigate these supported bilayers under physiological conditions (in water environment). We (H. Rinia et al., Biochemistry, 2000, 39, 5852-5858) previously found that synthetic transmembrane peptides self organize in highly characteristic striated domains in supported gel state bilayer. The precise morphology of the domains depends on the type of peptide and lipid. In the present study we attempt to localize the peptide in these domains by AFM using Au particles (app. 10 nm.) and thiol containing peptides. These peptides were also found to form striated domains. We also showed by Dynamic Light Scattering experiments that thiol group of a peptide, incorporated in a bilayer is available for reaction with gold. After addition of the gold particles over the supported bilayer we observe with contact mode AFM the presence of gold particles predominantly over domains and cracks, connecting them.

生体膜と人工膜の接点 精妙な生体膜のはたらきとナノバイオマテリアルの開発

生体膜と人工膜の接点 精妙な生体膜のはたらきとナノバイオマテリアルの開発

日本膜学会第２４年会特別講演 膜物性による生体膜機能の制御 測るから見るへ

The cell, which is a fundamental unit of the life, has been constructed of biomembranes. The inside of the cell in which the complicated chemical reaction progresses has been divided three-dimensionally by the biomembranes which are mainly composed of proteins and lipids. The function appears on membrane protein by the interaction with the lipid and is affected by the physical property of membrane lipid. In the cell, various biological phenomena occur in the regions divided by the biomembranes. Image analysis system under the microscope makes it possible to obtain spatial and time-related information of the biological phenomena in order to clarify the biological system of cell. We have developed the equipment that images the physical property of the biomembrane at video rate under the microscope using fluorescence dye Laurdan. This equipment was applied to the analysis of the fluidity change of the cell membrane of PC12 cell in the apoptosis process. And, it was applied to Giant liposome prepared for bilayer mixture of DMPC and DMPE, and the phase separation phenomenon was observed at the video rate.

Comparison of the effects of clozapine, chlorpromazine, and haloperidol on membrane lateral heterogeneity

The interactions of three neuroleptic drugs, clozapine (CLZ), chlorpromazine (CPZ), and haloperidol (HPD) with phospholipids were compared using DSC and Langmuir balance. Main emphasis was on the drug-induced effects on the lateral organization of lipid mixtures of the saturated zwitterionic 1,2-dipalmitoyl- sn-glycero-3-phosphatidylcholine (DPPC) and the unsaturated acidic phosphatidylserine, brainPS. In multilamellar vesicles (MLV) phase separation was observed by DSC at X PS⩾0.05. All three drugs bound to these MLVs, abolishing the pretransition at X drug⩾0.03. The main transition temperature ( T m) decreased almost linearly with increasing contents of the drugs, CLZ having the smallest effect. In distinction from the other two drugs, CLZ abolished the phase separation evident in the endotherms for DPPC/brainPS ( X PS=0.05) MLVs. Compression isotherms of DPPC/brainPS/drug ( X PS= X drug=0.05) monolayers revealed the neuroleptics to increase the average area/molecule, CLZ being the most effective. Penetration into brainPS monolayers showed strong interactions between the three drugs and this acidic phospholipid (in decreasing order CPZ>HPD>CLZ). Hydrophobic interactions demonstrated using neutral eggPC monolayers decreased in a different order, CLZ>CPZ>HPD. Fluorescence microscopy revealed domain morphology of DPPC/brainPS monolayers to be modulated by these drugs, increasing the gel-fluid domain boundary length in the phase coexistence region. To conclude, our data support the view that membrane-partitioning drugs could exert part of their effects by changing the lateral organization and thus also the functions of biomembranes.

Formation of Tethered Supported Bilayers by Vesicle Fusion onto Lipopolymer Monolayers Promoted by Osmotic Stress.

Lipid bilayer systems have been used extensively to study the structure and function of biomembranes including molecular recognition, permeation, adhesion, and fusion.1-5 Further interest arises from their potential use as biosensors.6,7 While classical black lipid bilayer membranes are highly suitable as such model systems, they suffer from their limited long-term stability.1 Additionally, to apply the broad range of surface sensitive experimental techniques developed in recent years for the characterization of ultrathin films, it is often necessary to immobilize lipid bilayers on solid supports. However, lateral fluidity of the lipid membrane is governed by the physical and chemical properties of the solid substrate. Thus, bilayer membranes prepared by the Langmuir-Blodgett (LB) technique or by vesicle fusion directly onto solid substrates are often polycrystalline or amorphous, and lateral lipid mobility within the membrane is restricted. While on hydrophilic oxidized silicon or glass a water layer (~1 nm thick) is formed which allows for free diffusion of phospholipid molecules (but not membrane-spanning proteins) within the bilayer,8-10 on most other metals and metal oxides, lipid mobility is strongly suppressed.11,12 To preserve the membrane's natural properties, one promising approach is to rest it on a water-swellable (hydrophilic) polymer or polyelectrolyte cushion which can act as a deformable and mobile substrate6,13,14 like the cytoskeletal support in living cells. Such softly supported lipid bilayers can, in principle, exist in an entirely fluid state, nearly undisturbed by the presence of the supporting polymer gel which provides an aqueous compartment between the solid substrate and the membrane. In our recent work, we have studied DMPC bilayers on a highly branched cationic polymer (polyethylenimine, PEI). Important structural information about these polymer-supported bilayer systems formed by various preparation methods was provided by neutron scattering experiments.15,16 Furthermore, measuring the intermembrane interaction forces using the surface forces apparatus (SFA) technique has shown the effect of fluidity on membrane functionality.17

Zinc Status in Human Immunodeficiency Virus Infection

There is substantial evidence to support an important role for zinc in immune processes. Adequate zinc status is essential for T-cell division, maturation and differentiation; lymphocyte response to mitogens; programmed cell death of lymphoid and myeloid origins; gene transcription; and biomembrane function. Lymphocytes are one of the types of cells activated by zinc. Zinc is the structural component of a wide variety of proteins, neuropeptides, hormone receptors and polynucleotides. Among the best known zinc-dependent hormones/enzymes are Cu, Zn superoxide dismutase, an enzyme component of the antioxidant defense system, and thymulin, which is essential for the formation of T-lymphocytes. In animals and humans, zinc deficiency results in rapid and marked atrophy of the thymus, impaired cell-mediated cutaneous sensitivity and lymphopenia. Primary and secondary antibody responses are reduced in zinc deficiency, particularly for those antigens that require T-cell help, such as those in heterologous red blood cells. In addition, antibody response and the generation of splenic cytotoxic T cells after immunization are reduced. Zinc also inhibits the production of tumor necrosis factor, which is implicated in the pathophysiology of cachexia and wasting in acquired immune deficiency syndrome.

Interactions of Hydration Water and Biological Membranes Studied by Neutron Scattering

To elucidate general features of structural and dynamical properties of hydration water and the influence of hydration water on the dynamical behavior of biomembranes, purple membranes from halobacteria and disk membranes from bovine retinae have been studied by neutron scattering techniques. Hydrated films of oriented multilamellar membrane stacks were used to measure lamellar diffraction patterns and quasielastic incoherent neutron scattering as a function of hydration level, of temperature, and of the protein/lipid ratio. These measurements revealed a strong interaction of a “first hydration layer” with the membrane surface and a reduced self-diffusion of aqueous solvent parallel to the membrane surface (the self-diffusion coefficient is about 5 times smaller as compared to excess water). The picosecond internal molecular motions of the protein/lipid complex are strongly affected by the amount of solvent interacting with the lipids and the membrane proteins. In particular, the lipids and their ability to attract solvent molecules play an important role for “hydration-induced flexibility” of biomembranes. On the basis of these measurements, the impact of the hydration process on the function of biomembranes is discussed for the light-driven proton pump bacteriorhodopsin in purple membranes.

Thermodynamic Analysis of Chain-Melting Transition Temperatures for Monounsaturated Phospholipid Membranes: Dependence on cis-Monoenoic Double Bond Position

Unsaturated phospholipid is the membrane component that is essential to the dynamic environment needed for biomembrane function. The dependence of the chain-melting transition temperature, T t, of phospholipid bilayer membranes on the position, n u, of the cis double bond in the glycerophospholipid sn-2 chain can be described by an expression of the form T t = T t c(1 + h′ c| n u − n c|)/(1 + s′ c| n u − n c|), where n c is the chain position of the double bond corresponding to the minimum transition temperature, T t c, for constant diacyl lipid chain lengths. This implies that the incremental transition enthalpy (and entropy) contributed by the sn-2 chain is greater for whichever of the chain segments, above or below the double-bond position, is the longer. The critical position, n c, of the double bond is offset from the center of the sn-2 chain by an approximately constant amount, δn c ≈ 1.5 C-atom units. The dependence of the parameters T t c, h′ c, and s′ c on sn-1 and sn-2 chain lengths can be interpreted consistently when allowance is made for the chain packing mismatch between the sn-1 and sn-2 chains. The length of the sn-2 chain is reduced by ∼0.8 C-atom units by the cis double bond, in addition to a shortening by ∼1.3 C-atom units by the bent configuration at the C-2 position. Based on this analysis, a general thermodynamic expression is proposed for the dependence of the chain-melting transition temperature on the position of the cis double bond and on the sn-1 and sn-2 chain lengths. The above treatment is restricted mostly to double-bond positions close to the center of the sn-2 chain. For double bonds positioned closer to the carboxyl or terminal methyl ends of the sn-2 chain, the effects on transition enthalpy can be considerably larger. They may be interpreted by the same formalism, but with different characteristic parameters, h′ c and s′ c, such that the shorter of the chain segments makes a considerably smaller contribution to the calorimetric properties of the chain-melting transition.

薬用外でのリポソームの利用 機能性高分子を表層に有するリポソームの特性解析

Liposomes are frequently used as model systems to clarify diverse functions of biomembranes. We have been introducing various functional polymer chains on the surface of liposomes by using a lipophilic radical initiator. In this review, our recent studies concerning specific molecular recognition on liposome surfaces were introduced.

Toxicology of mono-, di-, and triethanolamine.

The chemistry, biochemistry, toxicity, and industrial use of monoethanolamine (MEA), diethanolamine (DEA), and triethanolamine (TEA) are reviewed. The dual function groups, amino and hydroxyl, make them useful in cutting fluids and as intermediates in the production of surfactants, soaps, salts, corrosion control inhibitors, and in pharmaceutical and miscellaneous applications. In 1995, the annual U.S. production capacity for ethanolamines was 447,727 metric tons. The principal route of exposure is through skin, with some exposure occurring by inhalation of vapor and aerosols. MEA, DEA, and TEA in water penetrate rat skin at the rate of 2.9 x 10(-3), 4.36 x 10(-3) and 18 x 10(-3) cm/hr, respectively. MEA, DEA, and TEA are water-soluble ammonia derivatives, with pHs of 9-11 in water and pHa values of 9.3, 8.8, and 7.7, respectively. They are irritating to the skin, eyes, and respiratory tract, with MEA being the worst irritant, followed by DEA and TEA. The acute oral LD50s are 2.74 g/kg for MEA, 1.82 g/kg for DEA, and 2.34 g/kg for TEA (of bw), with most deaths occurring within 4 d of administration. MEA is present in nature as a nitrogenous base in phospholipids. These lipids, composed of glycerol, two fatty acid esters, phosphoric acid, and MEA, are the building blocks of biomembranes in animals. MEA is methylated to form choline, another important nitrogenous base in phospholipids and an essential vitamin. The rat dietary choline requirement is 10 mg kg-1 d-1; 30-d oral administration of MEA (160-2670 mg kg-1 d-1) to rats produced "altered" liver and kidney weights in animals ingesting 640 mg kg-1 d-1 or greater. Death occurred at dosages of 1280 mg kg-1 d-1. No treatment-related effects were noted in dogs administered as much as 22 mg kg-1 d-1 for 2 yr. DEA is not metabolized or readily eliminated from the liver or kidneys. At high tissue concentrations, DEA substitutes for MEA in phospholipids and is methylated to form phospholipids composed of N-methyl and N, N-dimethyl DEA. Dietary intake of DEA by rats for 13 wk at levels greater than 90 mg kg-1 d-1 resulted in degenerative changes in renal tubular epithelial cells and fatty degeneration of the liver. Similar effects were noted in drinking water studies. The findings are believed to be due to alterations in the structure and function of biomembranes brought about by the incorporation of DEA and methylated DEA in headgroups. TEA is not metabolized in the liver or incorporated into phospholipids. TEA, however, is readily eliminated in urine. Repeated oral administration to rats (7 d/wk, 24 wk) at dose levels up to and including 1600 mg kg-1 d-1 produced histopathological changes restricted to kidney and liver. Lesions in the liver consisted of cloudy swelling and occasional fatty changes, while cloudy swelling of the convoluted tubules and loop of Henle were observed in kidneys. Chronic administration (2 yr) of TEA in drinking water (0, 1%, or 2% w/v; 525 and 1100 mg kg-1 d-1 in males and 910 and 1970 mg kg-1 d-1 in females) depressed body and kidney weights in F-344 rats. Histopathological findings consisted of an "acceleration of so-called chronic nephropathy" commonly found in the kidneys of aging F-344 rats. In B6C3F1 mice, chronic administration of TEA in drinking water (0, 1%, or 2%) produced no significant change in terminal body weights between treated and control animals or gross pathological changes. TEA was not considered to be carcinogenic. Systemic effects in rats chronically administered TEA dermally (0, 32, 64, or 125 mg kg-1 d-1 in males; 0, 63, 125, or 250 mg kg-1 d-1 in females) 5 d/wk for 2 yr were primarily limited to hyperplasia of renal tubular epithelium and small microscopic adenomas. In a companion mouse dermal study, the most significant change was associated with nonneoplastic changes in livers of male mice consistent with chronic bacterial hepatitis.

Interaction of Zn2+Ions with Phospholipid Multilayers

Abstract Phospholipids are large natural amphipathic molecules which, in contact with water, assemble into higher molecular aggregates. The most relevant one is the bilayer for its relation to the structure, properties and functions of cell membranes. Given their structural complexity phospholipid bilayers are usually used as models for studying how biologically relevant chemicals affect the structure and functions of biomembranes. Zn is a trace element essential for the functional integrity of cell membranes. It is also a common contaminant of sea waters. Therefore, it was thought of interest to study the interaction of Zn2- ions with phospholipid bilayers in order to understand the way they interact with cell membranes. With this aim Zn2+, in a wide range of concentrations, was made to interact with multibilayers of the phospholipids dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE). This study was performed by X-ray diffraction. Besides, the interaction of Zn2+ with D...

アルコールによるリポソーム膜の指組み構造相の形成に関する研究

It was now well established that phosphatidylcholines or phosphatidylglycerols can form an interdigitated structural phase (Iphase), in which acyl chains of the lipids fully interpenetrate the hydrocarbon chains of the opposing monolayer in the presence of short chain alcohols and some other amphiphiles. It has been suggested that the interdigitated structure as well as a hexagonal II structure plays an important role in regulating many functions of biological membranes. We have demonstrated the participation ofIphase formation in ethanol-induced liposomal aggregation and fusion. We have also focused on control of permeability as one of the biomembrane functions, and the effects of ethanol on the permeability of large unilamellar vesicles composed of various phospho-lipids were studied by monitoring the leakage of the fluorescent dye, calcein, entrapped in the inner aqueous phase of the vesicles. In the cases of mixed membranes composed of DPPC and dilauroyl phosphatidylethanolamine (DLPE), as well as DPPC liposomal membranes, large permeabilities were observed in the wide range of ethanol concentrations, where the normal bilayer andIphase coexist and the membrane is in a phase-separated state. These results suggested that ethanol can disturb the normal control of biological membrane permeability.

Increased permeability of phase-separated liposomal membranes with mixtures of ethanol-induced interdigitated and non-interdigitated structures

It has been suggested by many workers using model membranes that the interdigitated structure formation, in which the acyl chains fully interpenetrate the hydrocarbon chains of the opposing monolayer, plays an important role in regulating many functions of biomembranes. In the present study the control of permeability was focused on as one of the biomembrane functions, and the effects of ethanol on the permeability of large unilamellar vesicles made by the extrusion technique (LUVET) (average diameter: about 250 nm), composed of dipalmitoyl or egg yolk phosphatidylcholines, were studied by monitoring the leakage of fluorescent dye, calcein, entrapped in the inner aqueous phase of the LUVET. The permeability was estimated from the apparent rate constant of calcein leakage at 25°C. Large permeabilities were observed in the region of 0.6 M to 1.3 M ethanol, with a concentration dependence. In this range of ethanol concentrations the normal bilayer and interdigitated structures coexist and the membrane is in a phase-separated state. The large permeability is due to the instability of the boundary regions, the interdigitated membrane being characterized by a thinner structure and more rigid hydrocarbon regions in the layer than its non-interdigitated counter part. These results suggest the possibility of biomembrane-permeability regulation by interdigitated membrane formation.

Effects of low-dose X-ray irradiation on biomembrane in brain cortex of aged rats

We previously found that low-dose X-ray irradiation or radon (weak α-ray) inhalation increases SOD activities and reduces lipid peroxide levels in various organs of 7-week-old rats or rabbit. In this study, we examined how the changes of SOD activity, lipid peroxide level, and membrane fluidity of the cerebral cortex in aged male Wistar rats (65 and 91 weeks old) were affected by low-dose X-ray irradiation (100 cGy or under) compared with those in 7-week-old rats, to elucidate the mechanism of aging inhibition. The following results were obtained: Although radiation sensitivity was observed to decreases with age, low-dose irradiation changed the Mn-SOD activity, lipid peroxide level, and membrane protein fluidity parameter of the cerebral cortex in the aged rats to be closer to those in the youth. These finding suggest that the increased SOD activity induced by low-dose irradiation enhances biomembrane functions, and that the decrease of lipid peroxide level enhances the membrane protein fluidity.

Divalent cation and lipid-protein interactions of biomembranes.

Divalent cations play an important role in the functions of biomembranes. This review deals with three topics: (1) Mg(2+)-mediated change in physical state of phospholipid induces conformation and activity change of reconstituted mitochondrial H(+)-ATPase, (2) a proper transmembrane Ca2+ gradient is essential for the higher enzymatic activity of adenylate cyclase, and (3) role of transmembrane Ca2+ gradient in the modulation of reconstituted sarcoplasmic reticulum Ca(2+)-ATPase activity.

Regulation of an electron transfer reaction of a flavin incorporated into synthetic lipid films on electrodes based on phase transition of molecular bilayers

From aspects of applications and utilization of biomembrane functions, the development of bilayer-modified electrodes with biomembrane-mimetic structures and properties should be highly attractive because they provide a new methodology for novel surface modification and functionalization. However, electrical and electrochemical studies of lipid membrane films on electrodes have been very limited [l-6]. One of our current interests in this area is to regulate electrochemical reactions at electrode surfaces based on the nature of organized bilayer membrane’s. In this communication, we have fabricated electrodes modified with polyion-complexed lipid bilayer films containing flavins as redox-active compounds and explored the possibility of the regulation of the electrochemistry of the films based on membrane phase transition, which is the most fundamental characteristic of lipid bilayers. A schematic drawing for a redox compound-doped multilayered bilayer film electrode and for bilayer phase transition are shown in Fig. 1. The synthetic lipid used here is a bilayer forming amphiphile, 2C1sN+ which is polyion-complexed with an anionic polymer, (poly&yrenesulfonate), PPS-) in order to avoid its dissolution into bulk aqueous solution. As we reported previously [ l,S], water-insoluble polyion-complexed lipid films possess fundamental bilayer characteristics similar to those of synthetic and biological aqueous bilayer membranes. Flavins are important electroactive molecules in biological systems and in this note lo-N-ethylisoalloxazine (1) and lo-N-dodecylisoalloxazine (2) which were

Ion permeability through bilayer lipid membranes for biosensor development: control by chemical modification of interfacial regions between phase domains

Based on the results of studies on cystic fibrosis, which implicated hydroxystearic acid (HSA) as a contributing factor in altered biomembrane function, solvent-free bilayer lipid membranes (BLMs) and monolayer films were prepared from a lipid mixture containing (by mass) 34% phosphatidylcholine, 19% dipalmitoylphosphatidyl serine, 47% cholesterol and variable amounts of 10- and 12-HSA (0-50%). Ion currents, resulting from K+ permeation through BLMs that were supported in 0.1 mol dm-3 KCl solutions buffered to pH 7.4, were monitored with use of a d.c. circuit. The structures of monolayer films at the air-water interface of a Langmuir-Blodgett trough were studied by pressure-area correlations and by further correlation with microscopic phase separation as revealed by fluorescence microscopy. In order to elucidate the role of the hydroxyl moieties in ion permeability, the transmembrane ion current was corrected for the effect of the negative surface charge of the carboxylic acid by replacement of the HSA component with stearic acid. The ion current was found to increase with the molar proportion of the HSAs. Two models for ion conduction through BLMs were considered: 'hopping' via hydrophilic sites within the hydrophobic zone of the BLMs, introduced by the hydroxyl moiety of 10- or 12-HSA; and transport through interfacial regions between phase domains that represent areas of low steric density and low structural order within monolayers. Although the two mechanisms are not distinct, the ion permeability results indicate a change in the response of ion current to HSA concentration at 35 mol-%, suggesting a change in the relative proportion of the mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Papers presented at the International Conference on Structure and Functions of Biomembranes. Calcutta, India, November 28‐December 2, 1989. Proceedings

Papers presented at the International Conference on Structure and Functions of Biomembranes. Calcutta, India, November 28‐December 2, 1989. Proceedings

Characteristics of fatty acid composition of adipose tissue and serum lipids in reindeer calves

Abstract: Fatty acid composition of adipose tissue and serum lipids were studied in newborn and growing (1.5-month-old) reindeer. Material was collected during 1985 and 1986 at the Kaamanen research station. Lipids were extracted with methanolchloroform and fatty acid methyl esters of lipid fractions were analysed by gas liquid chromatograph. Fatty acids of adipose tissue triglycerides in newborn calves were predominantly unsaturated (51%) referring to fluid properties of brown adipose tissue lipids. The dominant fatty acid was oleic acid (18:1) with 46% followed by palmitic acid (16:0) with 30% and stearic acid (18:0) with 15%. Triglycerides (TG) contained 3% polyunsaturated fatty acids (PUFA), the majority of which was essential fatty acids (EFA). In growing reindeer adipose tissue T G were predominantly saturated (67%). The dominant fatty acid was 16:0 (34%) followed by 18:1 (29%) and 18:0 (25%). Adipose tissue phospholipids (PL) were mainly unsaturated in both age groups. The proportion of P U F A was 23%, of which majority was EFA, necessary to normal structure and function of biomembranes. A high correlation existed between fatty acid composition of adipose tissue T G and P L with serum T G in newborn calves (r = 0.982 and 0.924). Fatty acids of serum T G were saturated by 54%. The dominant fatty acids were 18:1, 16:0 and 18:0 with percentages of 38,32 and 12, respectively.