Ion permeability of bilayer membranes formed from synthetic phospholipids in the phase transition region

Ion permeability of black lipid membranes formed from synthetic phospholipids has been studied. The resistance of BLM formed from phosphatidylcholine, tiophosphatidylcholine, threealkylphosphate and threealkyltiophosphate was 10(7)--10(8) Ohm.cm2. It was shown that the membrane potential of the 10--30 mV arised in KCl gradient indicating the preference cation conductance in synthetic lipid membranes. A sharp decrease of the membrane conductance near to the phase transition temperature was discovered. The change of conductance by phase transition temperature was sensitive to chemical nature of the polar head of phospholipids used.     

Electro-osmosis at the surface of phospholipid bilayer membranes

The electro-osmotic velocity is the velocity of a fluid near an interface produced by an electric field parallel to a surface. The velocity adjacent to fixed phospholipid bilayer membranes was measured by observing the velocity of small vesicles suspended in the fluid. The charge densities of the bilayers ranged from 0 to 1 electronic charge per lipid and experiments were performed at temperatures above and below the transition temperature of the phospholipid bilayer in 1, 10 and 100 mM NaCl solutions. The Helmholtz-Smoluchowski equation correctly predicted the electro-osmotic velocity from the known value of zeta potential of the phospholipid bilayer.     

Calcium-sensitive univalent cation channel formed by lysotriphosphoinositide in bilayer lipid membranes

A calcium sensitive univalent cation channel could be formed by lysotriphosphoinositide on an artificial bilayer membrane made of oxidized cholesterol. The modified membrane was selectively permeable to univalent cations, but was only very sparingly permeable to anions or divalent cations. Selectivity sequence among group IA cations was Rb⁺ > Cs⁺ > Na⁺ > K⁺ > Li⁺. The conductance of the membrane was increased up to a value of about 10⁻² ohm⁻¹/cm² with an increase in the concentration of univalent cation, and was drastically depressed by a relatively small increase in the concentration of calcium ion or other divalent cations. The sequence of depressing efficiency among divalent cations was Zn²⁺ > Cd²⁺ > Ca²⁺ > Sr²⁺ > Mg²⁺.     

Influence of sterols on ion transport through lipid bilayer membranes

Charge-pulse relaxation experiments with the negatively charged lipophilic ions, dipicrylamine and tetraphenylborate, (as well as with the positively charged carrier system Rb⁺-valinomycin) have been carried out in order to study the influence of sterols on the ion transport through the lipid bilayer membrane. The mol fraction of the sterols (cholesterol, epicholesterol, ergosterol, stigmasterol, dihydrocholsterol, epicoprostanol and cholesterololeate) as referred to total lipid was varied in a wide range (mol fractions 0–0.8).The monoolein/sterol or dioleoylphosphatidylcholine/sterol mixtures were dissolved in $\text{n}$-hexadecane in order to minimize effects of the sterol on the membrane thickness.Cholesterol had a strong influence on the transport of the lipophilic ions. Its incorporation into monoolein membranes increased the rate constant ${}_{\mathrm{<mtext>i</mtext>}}$ of translocation up to 8-fold, but incorporation into phosphatidylcholine membranes had virtually no influence on $\text{k}{}_{\mathrm{<mtext>i</mtext>}}$. The other sterols with one hydroxy group and cholesterololeate had no influence on the rate constant or the partition coefficient β. The results are discussed on the basis of a possible change of dipole potential of the membrane caused by cholesterol and its derivatives.In the case of valinomycin-mediated Rb⁺ transport only cholesterol had a strong influence on transport properties. The rate constants of association ($\text{k}{}_{\mathrm{<mtext>R</mtext>}}$) as well as the rate constants of translocation of the complex ($\text{k}{}_{\mathrm{<mtext>MS</mtext>}}$) and of the free carrier ($\text{k}{}_{\mathrm{<mtext>S</mtext>}}$) were reduced by incorporation of cholesterol up to eight-fold. The decrease of $\text{k}{}_{\mathrm{<mtext>S</mtext>}}$ and $\text{k}{}_{\mathrm{<mtext>MS</mtext>}}$ are possibly caused by a decrease of membrane fluidity, whereas the decrease of $\text{k}{}_{\mathrm{<mtext>R</mtext>}}$ may be due to an increase of surface potential. The different action of cholesterol on the two transport systems is discussed under the assumption that the adsorption plane of the lipophilic ion is located more towards the aqueous side and that of the ion-carrier complexes more towards the hydrocarbon side of the dipole layer.     

Properties of the large ion-permeable pores formed from protein F of Pseudomonas aeruginosa in lipid bilayer membranes

The incorporation of porin protein F from the outer membrane of Pseudomonas aeruginosa into artificial lipid bilayers results in an increase of the membrane conductance by many orders of magnitude. The membrane conductance is caused by the formation of large ion-permeable channels with a single-channel conductance in the order of 5 nS for 1 M alkali chlorides. The conductance has an ohmic current vs. voltage relationship. Further information on the structure of the pore formed by protein F was obtained by determining the single-channel conductance for various species differing in charge and size, and from zero-current potential measurements. The channel was found to be permeable for large organic ions (Tris+, N(C2H5)4+, Hepes-) and a channel diameter of 2.2 nm could be estimated from the conductance data (pore length of 7.5 nm). At neutral pH the pore is about two times more permeable for cations than for anions, possibly caused by negative charges in the pore. The consistent observation of large water filled pores formed by porin protein F in model membrane systems is discussed in the light of the known low permeability of the Ps. aeruginosa outer membrane towards antibiotics. It is suggested that this results from a relatively low proportion of open functional porin protein F pores in vivo.     

Calcium-sensitive univalent cation channel formed by lysotriphosphoinositide in bilayer lipid membranes.

A calcium sensitive univalent cation channel could be formed by lysotriphosphoinositide on an artificial bilayer membrane made of oxidized cholesterol. The modified membrane was selectively permeable to univalent cations, but was only very sparingly permeable to anions or divalent cations. Selectivity sequence among group IA cations was Rb+ greater than Cs+ greater than Na+ greater than K+ greater than Li+. The conductance of the membrane was increased up to a value of about 10-2 ohm-1/cm2 with an increase in the concentration of univalent cation, and was drastically depressed by a relatively small increase in the concentration of calcium ion or other divalent cations. The sequence of depressing efficiency among divalent cations was Zn+ greater than Cd2+ greater than Ca2+ greater than Sr2+ greater than Mg2+.     

Influence of membrane structure on ion transport through lipid bilayer membranes

Summary Charge-pulse relaxation studies with the positively charged PV-K⁺ complex (cyclo-(d-Val-l-Pro-l-Val-d-Pro)₃) and the negatively charged lipophilic ion dipicrylamine (DPA^–) have been performed in order to study the influence of structural properties on ion transport through lipid bilayer membranes. First, the thickness of monoolein membranes was varied over a wide range using differentn-alkanes and slovent-free membranes. The thickness (d) of the hydrocarbon core of these membranes varied between 4.9 and 2.5 nm. For both transport systems the partition coefficient was found to be rather insensitive to variations ind. The same was valid for the translocation rate constantk _MS of PV-K⁺, whereas a strong increase of the translocation rate constantk _i of DPA-with decreasingd was observed. In a further set of experimental conditions the structure of the lipids, such as number and position of the double bonds in the hydrocarbon chain and its chain length as well as the nature of the polar head group, was varied. The translocation constantk _MS of PV-K⁺ transport was found to be much more sensitive to these variations thank _i of DPA-.Much larger variations ink _i andk _MS were observed in membranes made from lipids with ether instead of ester linkages between glycerol backbone and hydrocarbon chain. The results are in qualitative agreement with the surface potentials of monolayers made from corresponding lipids. Increasing amounts of cholesterol in membranes of dioleoylphosphatidylcholine caused a strong decrease ofk _MS (PV-K⁺), whereask _i was found to be rather insensitive to this variation.In monoolein membranes cholesterol causes a decrease ofk _MS up to sixfold and a increase ofk _i up to eightfold. The partition coefficient of DPA^– was insensitive to cholesterol, whereas of PV-K⁺ was found to decrease about eightfold in these membranes. The influence of cholesterol onk _MS is discussed on the basis of viscosity changes in the membrane and the change ink _i of DPA^– and of PV-K⁺ on the basis of a possible change of the dipole potential of the membranes. The other sterols, epicholesterol and ergosterol cause no change in the kinetics of the two probes.The different influence of membrane properties like thickness, viscosity, and dipole potential on the two transport systems is discussed under the assumption that the adsorption planes of the two probes have different positions in a membrane. Possibly because of a larger hydrophobic interaction, the adsorption plane of PV-K⁺ is located more towards the hydrocarbon side and that of DPA^– more towards the aqueous side of the dipole layer.     

The adsorption of adrenocorticotropin-(1-24)-tetracosapeptide to lecithin bilayer membranes formed from liposomes

The interaction of the peptide hormone adrenocorticotropin (ACTH_1-24) with solvent-free planar lipid bilayers has been studied by use of the capacitance minimization method. The membranes were formed from artificial vesicles according to the method described by Schindler. In contrast to analogous studies with hexane-containing membranes, experiments with these vesicle-derived bilayers were completely reproducible and gave no indication that ACTH_1-24 spans such hexane-free bilayers.     

Gating properties of channels formed by colicin Ia in planar lipid bilayer membranes

Summary Colicin Ia forms voltage-dependent channels when incorporated into planar lipid bilayers. A membrane containing many Colicin Ia channels shows a conductance which is turned on when high positive voltages (>+10 mV) are applied to thecis side (side to which the protein is added). The ionic current flowing through the membrane in response to a voltage step shows at first an exponential and then a linear rise with time. The relationship between the steady-state conductance, achieved immediately after the exponential portion, and voltage is S-shaped and is adequately fit by a Boltzmann distribution. The time constant () of the exponential is also dependent on voltage, and the relation between these two parameters is asymmetric aroundV _o (voltage at which half of the channels are open). In both cases the steepness of the voltage dependence, a consequence of the number of effective gating particles (n) present in the channel, is greatly influenced by the pH of the bathing solutions. Thus, increasing the pH leads to a reduction inn, while acidic pH's have the opposite effects. This result is obtained either by changing the pH on both sides of the membrane or on only one side, be itcis orrans. On the other hand, changing pH on only one side by addition of an impermeant buffer fails to induce any change inn. At the single-channel level, pH had an effect both on the unitary conductance, doubling it in going from pH 4.5 to 8.2, as well as on the fraction of time the channels stay open,F _(v). For a given voltage,F _(v) is clearly diminished by increasing the pH. This titration of the voltage sensitivity leads to the conclusion that gating in the Colicin Ia molecule is accomplished by charged amino-acid residues present in the protein molecule. Our results also support the notion that these charged groups are inside the aqueous portion of the channel.     

Chaotropic anions and the surface potential of bilayer membranes.

The chaotropic anions perchlorate and thiocyanate adsorb to artificial phospholipid membranes. The negative electrostatic potential they produce at the surface of the membranes was measured by two independent techniques. The conductance produced by neutral carriers of cations and anions was measured to estimate changes in the surface potentials of planar black lipid films and the electrophoretic mobility of phospholipid vesicles was used to monitor changes in the zeta potentials of spherical bilayer membranes. Qualitatively similar results were obtained with the two techniques. The results, moreover, agreed with the change in surface potential produced by these anions at an air water interface, as measured directly with an ionizing electrode (Randles, J. E. B. (1957) Discuss. Faraday Soc. 24, 194-199). The results obtained with artificial bilayers may explain the observation (Wieth, J. O. (1970) J. Physiol. 207, 581-609) that thiocyanate increases the sodium or potassium and decreases the sulfate permeability of erythrocyte membranes.     

Intermonolayer friction and surface shear viscosity of lipid bilayer membranes

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.     

Influence of cholesterol on electroporation of bilayer lipid membranes: chronopotentiometric studies

This paper presents the results of constant-current (chronopotentiometric) measurements of the egg yolk phosphatidylcholine (PC) bilayer membrane without and with cholesterol. The experiments were performed on planar bilayer lipid membrane (BLM) formed by the Mueller-Rudin method. It is demonstrated that the constant-intensity current flow through bilayer membranes generated fluctuating pores in their structure. The presence of cholesterol in the membrane caused an increase in the value of the breakdown potential. It is postulated that greater stability of the bilayer with cholesterol can result from an increased critical pore radius (at which the bilayer would undergo irreversible rupture). This confirms that cholesterol has a stabilizing effect on BLM. Besides, our results suggest that addition of cholesterol causes shift in the distribution of pore conductance towards a smaller value. It is suggested that this can be connected with the phenomenon of domain formation in the membranes containing high concentration of cholesterol. Moreover, it is shown that chronopotentiometry with programmable current intensity is a promising method for observation of the membrane recovery process.     

Influence of dolichyl phosphate on permeability and stability of bilayer lipid membranes

The ionic permeability coefficients, ionic transference numbers, activation energy of ion transport and breakdown voltage of bilayer lipid membranes made from dioleoylphosphatidylcholine or its mixtures with dolichyl 12-phosphate have been studied. The electrical measurements showed that dolichyl phosphate in phospholipid bilayers decreases membrane permeability, changes membrane ionic selectivity and increases membrane stability. These results are discussed in light of the aggregation behavior and the intramolecular clustering of a dolichyl phosphate molecule in phospholipid membranes. From our data we suggest that the hydrophilic part of dolichyl phosphate molecules regulates their behavior in membranes.     

Influence of cholesterol on electroporation of bilayer lipid membranes: chronopotentiometric studies

This paper presents the results of constant-current (chronopotentiometric) measurements of the egg yolk phosphatidylcholine (PC) bilayer membrane without and with cholesterol. The experiments were performed on planar bilayer lipid membrane (BLM) formed by the Mueller-Rudin method. It is demonstrated that the constant-intensity current flow through bilayer membranes generated fluctuating pores in their structure. The presence of cholesterol in the membrane caused an increase in the value of the breakdown potential. It is postulated that greater stability of the bilayer with cholesterol can result from an increased critical pore radius (at which the bilayer would undergo irreversible rupture). This confirms that cholesterol has a stabilizing effect on BLM. Besides, our results suggest that addition of cholesterol causes shift in the distribution of pore conductance towards a smaller value. It is suggested that this can be connected with the phenomenon of domain formation in the membranes containing high concentration of cholesterol. Moreover, it is shown that chronopotentiometry with programmable current intensity is a promising method for observation of the membrane recovery process.     

Interfacial tension of bilayer lipid membrane formed from phosphatidylethanolamine

The dependence of the interfacial tension of a lipid membrane on the pH of the aqueous solution has been studied. Interfacial tension measurements of phosphatidylethanolamine (PE) were carried out. A theoretical equation is derived to describe this dependence in the whole pH range. A maximum corresponding to the isoelectric point appears both in the theoretical equation and in the experimental data.     

Planar bilayer membranes from pure lipids.

Planar lipid bilayer membranes are formed from mixtures of pure lipids in the absence of non-biological solvents. The solventless bilayers are characterized by a large specific capacitance (586-957 nF/cm2) comparable to that of cell membranes but considerably greater than that of conventional lipid/decane bilayers. Hydrocarbon solvents, such as n-alkanes or squalene, thicken the bilayer. Membrane dielectric thickness is used as an indicator of bilayer lipid composition. For membranes made from pure monoglyceride/triglyceride mixtures the thickness of the solventless lipid bilayer is independent of both the chain length (11-22 carbons) and mol fraction (0.1-0.9) of triglyceride in the bulk mixture. In contrast, the thickness of the bilayer (2.0-3.3 nm) depends strongly upon the length (16-24 carbons) of the monoglyceride component. Molecular volume considerations lead to the conclusion that the bulk lipid mixture disproportionates to yield bilayer membranes composed of nearly pure monoglyceride. The dielectric thickness of the monoglyceride bilayer is consistent with the notion that the lipid fatty acyl chains are fluid.     

Physical properties and surface interactions of bilayer membranes containing N-methylated phosphatidylethanolamines

The structure and physical properties of aqueous dispersions of 1,2-diacyl-sn-glycero-3-phosphoethanolamines (PE's) and their N-methylated analogues have been studied by scanning calorimetry, 31P nuclear magnetic resonance, and freeze-fracture electron microscopy. While successive N-methylations of a diacylphosphatidylethanolamine cause only modest decreases in its gel to liquid-crystalline phase transition temperature, the introduction of even a single N-methyl group sharply increases the temperature at which the lipid forms a hexagonal II phase. However, 31P nuclear magnetic resonance and electron microscopy show that unlike pure PE species, N-methylated PE's can form a variety of irregular nonlamellar structures at temperatures well below that at which a well-defined hexagonal II phase is formed. The rate of calcium-induced leakage of encapsulated carboxyfluorescein from large unilamellar vesicles composed of dioleoyl- or dielaidoylphosphatidylserine and the corresponding PE is strongly reduced when PE is replaced by N-methylated derivatives. The rate of calcium-induced intermixing of lipids of PE/phosphatidylserine (PS) vesicles steadily decreases as the PE component is successively replaced by its mono-, di-, and tri-N-methylated (phosphatidylcholine) derivatives. By correlating calorimetrically obtained phase diagrams with measurements of vesicle lipid intermixing, we conclude that dielaidoyl-N-methylphosphatidylethanolamine, like PE, can support direct interactions between the surfaces of PS/N-methyl-PE vesicles without lateral separation of a PS(Ca2+)-rich phase, while dielaidoyl-N,N-dimethyl-PE (and phosphatidylcholine) cannot.(ABSTRACT TRUNCATED AT 250 WORDS)     

Properties of the large ion-permeable pores formed from protein F of Pseudomonas aeruginosa in lipid bilayer membranes

The incorporation of porin protein F from the outer membrane of Pseudomonas aeruginosa into artificial lipid bilayers results in an increase of the membrane conductance by many orders of magnitude. The membrane conductance is caused by the formation of large ion-permeable channels with a single-channel conductance in the order of 5 nS for 1 M alkali chlorides. The conductance has an ohmic current vs. voltage relationship. Further information on the structure of the pore formed by protein F was obtained by determining the single-channel conductance for various species differing in charge and size, and from zero-current potential measurements. The channel was found to be permeable for large organic ions (Tris⁺, N(C₂H₅)₄⁺, Hepes^?) and a channel diameter of 2.2 nm could be estimated from the conductance data (pore length of 7.5 nm). At neutral pH the pore is about two times more permeable for cations than for anions, possibly caused by negative charges in the pore. The consistent observation of large water filled pores formed by porin protein F in model membrane systems is discussed in the light of the known low permeability of the Ps. aeruginosa outer membrane towards antibiotics. It is suggested that this results from a relatively low proportion of open functional porin protein F pores in vivo.     

Calcium-induced changes in bilayer membranes from oxidized cholesterol

Elastic properties of oxidized cholesterol bilayers in n-octane and membrane solvent free were studied by measuring Young modulus E perpendicular in the direction perpendicular to the membrane plane as a function of concentration of calcium ions. Interaction between calcium ions and solvent free bilayers resulted in a significant increases of Young modulus E perpendicular in the concentration range 20-40 mmol/l Ca2+. It is suggested that the hardening of the membrane is caused by some structural changes in the hydrophobic region of the membrane.     

Membrane fusion promoters and inhibitors have contrasting effects on lipid bilayer structure and undulations

It has been established that the fusion of both biological membranes and phospholipid bilayers can be modulated by altering their lipid composition (Chernomordik et al., 1995 .J. Membr. Biol. 146:3). In particular, when added exogenously between apposing membranes, monomyristoylphosphatidylcholine (MMPC) inhibits membrane fusion, whereas glycerol monoleate (GMO), oleic acid (OA), and arachidonic acid (AA) promote fusion. This present study uses x-ray diffraction to investigate the effects of MMPC, GMO, OA, and AA on the bending and stability of lipid bilayers when bilayers are forced together with applied osmotic pressure. The addition of 10 and 30 mol% MMPC to egg phosphatidylcholine (EPC) bilayers maintains the bilayer structure, even when the interbilayer fluid spacing is reduced to approximately 3 A, and increases the repulsive pressure between bilayers so that the fluid spacing in excess water increases by 5 and 15 A, respectively. Thus MMPC increases the undulation pressure, implying that the addition of MMPC promotes out-of-plane bending and decreases the adhesion energy between bilayers. In contrast, the addition of GMO has minor effects on the undulation pressure; 10 and 50 mol% GMO increase the fluid spacing of EPC in excess water by 0 and 2 A, respectively. However, x-ray diffraction indicates that, at small interbilayer separations, GMO, OA, or AA converts the bilayer to a structure containing hexagonally packed scattering units approximately 50 A in diameter. Thus GMO, OA, or AA destabilizes bilayer structure as apposing bilayers are brought into contact, which could contribute to their role in promoting membrane fusion.