Surface Of Bilayer Lipid Membranes Research Articles

Electrochemical Studies on Interaction of Ofloxacin with ITO-PANI Supported Bilayer Lipid Membrane

Bilayer phospholipid membrane was formed on the ITO-Pani surface in NaCl bath solution at different pH. Characterization and interaction of bilayer lipid membrane (BLM) with Ofloxacin molecules in 0.1 M NaCl solution was studied at different pH employing electrochemical impedance spectroscopy, cyclic voltammetry and chronoamperometric methods. The changes in the properties of BLM due to addition of Ofloxacin depends on the pH of bath solution. At low pH the membrane surface is positively charged and the interaction of drug molecules with the BLM surface takes place through chloride bridge. At alkaline pH the membrane surface is negatively charged and the drug molecules get attached through hydrogen bonding. The results of electrochemical impedance spectroscopy, cyclic voltammetry and chronoamperometry support each other.

Electrostatic Potentials Caused by the Release of Protons from Photoactivated Compound Sodium 2-Methoxy-5-nitrophenyl Sulfate at the Surface of Bilayer Lipid Membrane.

Lateral transport and release of protons at the water-membrane interface play crucial roles in cell bioenergetics. Therefore, versatile techniques need to be developed for investigating as well as clarifying the main features of these processes at the molecular level. Here, we experimentally measured the kinetics of binding of protons released from the photoactivated compound sodium 2-methoxy-5-nitrophenyl sulfate (MNPS) at the surface of a bilayer lipid membrane (BLM). We developed a theoretical model of this process describing the damage of MNPS coupled with the release of the protons at the membrane surface, as well as the exchange of MNPS molecules and protons between the membrane and solution. We found that the total change in the boundary potential difference across the membrane, ∆ϕb, is the sum of opposing effects of adsorption of MNPS anions and release of protons at the membrane-water interface. Steady-state change in the ∆ϕb due to protons decreased with the concentration of the buffer and increased with the pH of the solution. The change in the concentration of protons evaluated from measurements of ∆ϕb was close to that in the unstirred water layer near the BLM. This result, as well as rate constants of the proton exchange between the membrane and the bulk solution, indicated that the rate-limiting step of the proton surface to bulk release is the change in the concentration of protons in the unstirred layer. This means that the protons released from MNPS remain in equilibrium between the BLM surface and an adjacent water layer.

Electrophysiological Characterization of Microbial Rhodopsin Transport Properties: Electrometric and ΔpH Measurements Using Planar Lipid Bilayer, Collodion Film, and Fluorescent Probe Approaches.

Electrophysiological approaches to the study of the activity of retinal-containing protein bacteriorhodopsin (bR) or other proteins of this family are based usually on measurements of electrical current through a planar bilayer lipid membrane (BLM) with proteoliposomes attached to the BLM surface at one side of the membrane. Here, we describe the measurements of the pumping activity of bR and channelrhodopsin 2 (ChR2) with special attention to the study of voltage dependence of the light-induced currents. Strong voltage dependence of ChR2 suggests light-triggered ion channel activity of ChR2. We also describe electrophysiological measurements with the help of collodion film instead of BLM for the measurements of fast stages of a rhodopsin photocycle as well as the estimation of the activity of proteoliposomes without a macro membrane using fluorescent probes such as oxonol VI or 9-aminoacridine.

Inhibition of Amyloid β-Induced Lipid Membrane Permeation andAmyloid β Aggregation by K162.

Alzheimer’s disease (AD) is characterized by progressive neurodegeneration associated with amyloid β (Aβ) peptide aggregation. The aggregation of Aβ monomers (AβMs) leads to the formation of Aβ oligomers (AβOs), the neurotoxic Aβ form, capable of permeating the cell membrane. Here, we investigated the effect of a fluorene-based active drug candidate, named K162, on both Aβ aggregation and AβO toxicity toward the bilayer lipid membrane (BLM). Electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), and molecular dynamics (MD) were employed to show that K162 inhibits AβOs-induced BLM permeation, thus preserving BLM integrity. In the presence of K162, only shallow defects on the BLM surface were formed. Apparently, K162 modifies Aβ aggregation by bypassing the formation of toxic AβOs, and only nontoxic AβMs, dimers (AβDs), and fibrils (AβFs) are produced. Unlike other Aβ toxicity inhibitors, K162 preserves neurologically beneficial AβMs. This unique K162 inhibition mechanism provides an alternative AD therapeutic strategy that could be explored in the future.

Correction to "Distribution and Adsorption of Ionic Species into a Liposome Membrane and Their Dependence upon the Species and Concentration of a Coexisting Counterion".

The distribution of ions into a bilayer lipid membrane (BLM) and their adsorption on the BLM are investigated by extracting a hydrophobic cation, rhodamine 6G (R6G+), into a liposome through the dialysis membrane method. R6G+ distribution mainly depends upon the concentration of the coexisting anion and its species (Cl–, Br–, BF4–, ClO4–, and picrate). On the other hand, R6G+ adsorption on the BLM surface follows the Langmuir adsorption model and is independent of the coexisting anion in the aqueous phase. We propose an extraction model of ionic species into the BLM, to explain the dependence of extraction of ionic species upon the coexisting anion. In this model, an ion is distributed with a coexisting counterion into the BLM and then forms an ion pair in the BLM. Here, the ion adsorption equilibrium on the BLM surface is independent of the species and concentration of the coexisting counterion under the same ionic strength. On the basis of this model, we estimate the distribution constant of R6G+ and an...

Consequences of the Endogenous N-Glycosylation of Human Ribonuclease 1.

Ribonuclease 1 (RNase 1) is the most prevalent human homologue of the archetypal enzyme RNase A. RNase 1 contains sequons for N-linked glycosylation at Asn34, Asn76, and Asn88 and is N-glycosylated at all three sites in vivo. The effect of N-glycosylation on the structure and function of RNase 1 is unknown. By using an engineered strain of the yeast Pichia pastoris, we installed a heptasaccharide (Man5GlcNAc2) on the side chain of Asn34, Asn76, and Asn88 to produce the authentic triglycosylated form of human RNase 1. As a glutamine residue is not a substrate for cellular oligosaccharyltransferase, we used strategic asparagine-to-glutamine substitutions to produce the three diglycosylated and three monoglycosylated forms of RNase 1. We found that the N-glycosylation of RNase 1 at any position attenuates its catalytic activity but enhances both its thermostability and its resistance to proteolysis. N-Glycosylation at Asn34 generates the most active and stable glycoforms, in accord with its sequon being highly conserved among vertebrate species. These data provide new insight on the biological role of the N-glycosylation of a human secretory enzyme.

Adsorption and photodynamic efficiency of meso-tetrakis(p-sulfonatophenyl)porphyrin on the surface of bilayer lipid membranes

The adsorption and photodynamic efficiency of 5,10,15,20-tetrakis(p-sulfonatophenyl)porphyrin (H2TPPS4) on bilayer lipid membranes (BLM) have been studied. The adsorption of H2TPPS4 on BLM leads to rising of the potential drop on the membrane/water interface which has been detected either by the intramembrane field compensation (IFC) method, or as ζ-potential of liposomes measured by the dynamic light scattering method. The dependence of this potential on the concentration of H2TPPS4 and KCl in the solution can be described in the frame of Gouy-Chapman model of diffuse double layer assuming that the molecules of H2TPPS4 adsorb on the surface of BLM as an anions with four charged groups. The potential disappeared when the pH of solution decreased from 6 to 3 allowing the conclusion that the protonated forms of H2TPPS4 can not adsorb on the BLM probably due to change of conformation or aggregation of the molecules. The photodynamic efficiency of H2TPPS4 was evaluated by measuring the rate of damage of the targets - molecules of styryl dye (di-4-ANEPPS) by singlet oxygen generated under illumination on the surface of BLM. This rate was proportional to the surface density of H2TPPS4 molecules on the membrane which was determined from the change of surface charge of the membrane due to adsorption of the H2TPPS4. These results indicate that the di-4-ANEPPS molecules are damaged by singlet oxygen generated by monomers of H2TPPS4 molecules adsorbed on the membrane. The rate of oxidation of di-4-ANEPPS molecules adsorbed on the same (cis) side of the membrane together with the H2TPPS4 molecules was either the same or higher than that when di-4-ANEPPS molecules were adsorbed on opposite (trans) side. It indicates that the quenching of singlet oxygen by the di-4-ANEPPS molecules at cis side of the membrane was negligible, in contrast to our earlier study when singlet oxygen was generated by aluminum(III) phthalocyanines with one or two peripheral sulfo groups. The difference between these phthalocyanines and H2TPPS4 was explained by their different adsorption depth in the membrane.

Distribution and Adsorption of Ionic Species into a Liposome Membrane and Their Dependence upon the Species and Concentration of a Coexisting Counterion

The distribution of ions into a bilayer lipid membrane (BLM) and their adsorption on the BLM are investigated by extracting a hydrophobic cation, rhodamine 6G (R6G+), into a liposome through the dialysis membrane method. R6G+ distribution mainly depends upon the concentration of the coexisting anion and its species (Cl-, Br-, BF4-, ClO4-, and picrate). On the other hand, R6G+ adsorption on the BLM surface follows the Langmuir adsorption model and is independent of the coexisting anion in the aqueous phase. We propose an extraction model of ionic species into the BLM, to explain the dependence of extraction of ionic species upon the coexisting anion. In this model, an ion is distributed with a coexisting counterion into the BLM and then forms an ion pair in the BLM. Here, the ion adsorption equilibrium on the BLM surface is independent of the species and concentration of the coexisting counterion under the same ionic strength. On the basis of this model, we estimate the distribution constant of R6G+ and anion (KD), the ion-pair formation constant in the BLM (Kip), and the R6G+ adsorption constant on the BLM surface (Kad). Even for an ultrathin membrane system, such as a BLM, R6G+ is distributed with a coexisting counterion and the distribution equilibrium of the ionic species at the water-BLM interface is analyzable similar to that at the water-organic solvent interface.

Voltage-sensitive styryl dyes as singlet oxygen targets on the surface of bilayer lipid membrane

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

Peculiarities of the DNA hybridization on the surface of bilayer lipid membranes

The method of electrostriction was applied to study the peculiarities of interaction of short oligonucleotides with free standing (BLM) and supported lipid membranes (sBLM) and of the duplex formation between complementary oligonucleotides on a membrane surface. The 15-mer single stranded DNA (pentadecathymidylate—(dT) 15) was modified either with cholesterol (CH(dT) 15) or by palmitoyl chain (C16(dT) 15). The interaction of CH(dT) 15 with free standing BLM or with BLM formed on an agar or gold support was accompanied by sharp increase of elasticity modulus in direction perpendicular to the membrane plane , E ⊥, and by increase of surface potential. In contrast, C16(dT) 15 did not induce substantial changes of elasticity modulus, however, the surface potential was changed in a similar manner as for CH(dT) 15. Hybridization of DNA following addition of complementary chain (dA) 15 has been accompanied by a small decrease of elasticity modulus and by a slight increase of surface potential. Both the incorporation of chemically modified oligonucleotides into the lipid bilayer as well as hybridization of DNA are not cooperative processes as has been demonstrated by analysis using Scatchard plot of corresponding values.

Permeation of dicarboxylic acids with different terminal position of two carboxylic groups through planar bilayer lipid membranes

Electrically silent hydrogen ion fluxes across a planar bilayer lipid membrane (BLM) induced by an addition of dicarboxylic (DC) acids at one side of BLM are monitored by measuring pH changes in the unstirred layers near the BLM surface via recording protonophore-dependent potentials. Two groups of DC acids are studied: (1) 2- n-alkylmalonic acids with an alkyl chain of different length which carry both carboxylic groups at one terminus of the hydrocarbon chain (α,α-DC acids); and (2) dicarboxylic acids of different linear chain length having carboxylic groups at the opposite ends of the hydrocarbon chain (α,ω-DC acids). It is shown that the pH optimum of hydrogen ion fluxes for the DC acids is shifted considerably to acidic pH values compared to monocarboxylic acids and is located near pH 5. For both types of DC acids at pH≪5, the total transport is limited by diffusion of the anionic forms of the acids across the unstirred layers, while at pH≫5 the transport is limited by diffusion of the neutral form across the membrane. The fluxes of α,α-DC acids are similar to those of α,ω-DC acids provided that the acids have the similar number of carbon atoms, the fluxes grow with the increase in the chain length of the alkyl radical.

Flow injection monitoring and analysis of mixtures of simazine, atrazine, and propazine using filter‐supported bilayer lipid membranes (BLMs)

AbstractThis work describes a technique for the rapid and sensitive electrochemical flow injection monitoring and analysis of mixtures of the triazine herbicides simazine, atrazine and propazine using stabilized systems of filter‐supported bilayer lipid membranes (BLMs) composed of egg phosphatidylcholine (egg PC) and dipalmitoylphosphatidic acid (DPPA). Injections of herbicides were made into flowing streams of a carrier electrolyte solution and a transient current signal with a duration of seconds reproducibly appeared in less than two min after exposure of the lipid membranes to the herbicides. The magnitude of this signal was linearly related to the herbicide concentration, which could be determined at sub‐micromolar level. The mechanism of signal generation was related to a rapid adsorption of the triazine followed by a slow aggregative process of the herbicide at the surface of BLMs with a consequent rapid reorganization of the membrane electrostatics. Repetitive cycles of injection of herbicides have shown no signal degradation during each cycle. The time of appearance of the transient response was different for each triazine and increased to the order of simazine, atrazine and propazine which has allowed selective detection and analysis of these triazines in mixtures.

Electrochemical transduction of interactions of atrazine with bilayer lipid membranes

AbstractThis work investigates the interactions of atrazine with bilayer lipid membranes (BLMs) that can be used for the direct electrochemical sensing of this herbicide. Egg phosphatidylcholine (PC) and dipalmitoylphosphatidic acid (DPPA) were used for the formation of solventless BLMs. The interactions of atrazine with these membranes were found to be electrochemically transduced by BLMs in the form of a transient current signal with a duration of seconds, which reproducibly appeared within 1 min after exposure of the membranes to atrazine. The sensitivity of the response was maximized by use of BLMs composed of 35% (w./w.) DPPA, and by alteration of the phase distribution within membranes by the introduction of calcium ions in bulk solution. The mechanism of signal generation was related to the adsorption of atrazine with a consequent rapid reorganization of the membrane electrostatics due to atrazine aggregation at the surface of BLMs. Hydrogen bonding between atrazine and the carbonyl group of the lipid was explored by addition of platelet‐activating factor (PAF; an ether analog of PC) in BLMs composed of PC. Differential scanning calorimetry of vesicles composed of 15% DPPA was used to study the aggregation of atrazine in membrane domains enriched in the charged lipid. The magnitude of the transient current signal was linearly related to the concentration of atrazine in bulk solution with sub‐micromolar detection limits. This electrochemical transduction of atrazine interactions with BLMs holds prospects for flow injection monitoring of triazine herbicides.

Effect of the alkyl chain length of monocarboxylic acid on the permeation through bilayer lipid membranes

Electrically silent hydrogen ion fluxes across a planar bilayer lipid membrane (BLM) induced by an addition of monocarboxylic acid at one sive of BLM were studied by measuring pH changes in the unstirred layers near the BLM surface. The pH changes were assayed by recording protonophore-vepenvent potentials as well as by direct measurements of pH shifts in the unstirred layers close to the membrane by the pH microelectrove. It was shown that the mechanism of the acid transport changed qualitatively upon the increase of the hydrophobic chain length of the acid. In the case of short-chain acids at pH < p K a, the total transport was limited by diffusion of the anionic form of the acid across the unstirred layers, while at the alkaline pH (pH > > p K a) the transport was limited by diffusion of the neutral form across the membrane. In the alkaline pH range the pH shifts induced by short-chain acids were sensitive to the presence of cholesterol in the BLM as well as to the stirring conditions in the cell. However, in the case of long chain acids (more than 8 carbonic atoms) the transport was limited by diffusion of the anionic form of the acid in the whole range of pH studied. In the latter case, pH changes in the unstirred layers did not vepend on the presence of cholesterol in the membrane, and moreover pH shifts were not vepenvent on the thickness of the unstirred layer. It was proposed that the peculiarities of the long-chain acid-induced proton transport were associated with the formation of micelles of the acid in bathing solutions.

Calcium‐dependence of laminin binding to phospholipid membranes

Laminin is the most abundant noncollagenous protein in basement membranes. Its self-assembly has been studied in solution, and it has been established that calcium ions induce partially reversible aggregation. The behavior at a lipid membrane surface is of greater biological significance, but it is difficult to study quantitatively binding kinetics at a surface. The present work uses a powerful new integrated optics technique to measure the absorption and aggregation of the laminin-nidogen at a bilayer lipid membrane surface. It is found that the binding of a single layer of laminin at the lipid membrane is independent of the presence of calcium, but that the building up of multilayer laminin membranes requires calcium, and that these may not be destroyed by a calcium-complexing agent.

Adsorption of gentamicin onto a bilayer lipid membrane

Addition of the aminoglycoside antibiotic, gentamicin (GM), to one side of a bilayer lipid membrane (BLM) results in a potential difference across the membrane. Evidence is presented that the membrane potential is caused by the adsorption of GM, bearing four positive charges, on the BLM surface. The experimental results are subjected to a quantitative analysis based on the double-layer theory and the Langmuir adsorption isotherm. The adsorption is saturated (i.e., the BLM is fully covered) at the bulk GM concentration of about 80 μmol/l. At this point, the calculated GM-induced increase in the BLM surface charge density is σ = 0.0054 C m −2, which is equivalent to one positive charge per 50 lipids or one molecule of GM per 200 lipids.

Adsorption of gentamicin onto a bilayer lipid membrane

Addition of the aminoglycoside antibiotic, gentamicin (GM), to one side of a bilayer lipid membrane (BLM) results in a potential difference across the membrane. Evidence is presented that the membrane potential is caused by the adsorption of GM, bearing four positive charges, on the BLM surface. The experimental results are subjected to a quantitative analysis based on the double-layer theory and the Langmuir adsorption isotherm. The adsorption is saturated (i.e., the BLM is fully covered) at the bulk GM concentration of about 80 μmol/1. At this point, the calculated GM-induced increase in the BLM surface charge density is σ = 0.0054 C m −2, which is equivalent to one positive charge per 50 lipids or one molecule of GM per 200 lipids.

Formation and characterization of microcrystalline semiconductor particles on bilayer lipid membranes

Microcrystalline cadmium, indium, copper, and zinc sulfides were generated in situ on the surface of bilayer lipid membranes (BLMs) prepared from bovine-brain phosphatidylserine (PS), glyceryl monooleate (GMO), and a synthetic, polymerizable surfactant (n-C/sub 15/H/sub 31/CO/sub 2/(CH/sub 2/)/sub 2/)/sub 2/N/sup +/(CH/sub 3/)CH/sub 2/C/sub 6/H/sub 4/CH double bond CH/sub 2/, Cl/sup -/ (STYRS). Semiconductor-containing BLMs remained stable for days. Semiconductor formation on the BLM surface was monitored by optical microscopy, voltage-dependent capacitance measurements, and absorption and intracavity-laser-absorption spectroscopy. Band gap excitation of GMO- BLM-incorporated CdS resulted in the development of photovoltage. Irradiation of CdS incorporated into BLMs formed from STYRS (using a 350-nm cutoff filter) led to absorption losses due to the styrene moiety in the surfactant. Apparently, CdS sensitized the photopolymerization of STRYS BLMs.