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Lipid and protein molecules anisotropically oriented at a hydrocarbon-aqueous interface configure a dynamic array of self-organized molecular dipoles. Electrostatic fields applied to lipid monolayers have been shown to induce in-plane migration of domains or phase separation in a homogeneous system. In this work, we have investigated the effect of externally applied electrostatic fields on different lipid monolayers exhibiting surface immiscibility. In the monolayers studied, lipids in the condensed state segregate in discontinuous round-shaped domains, with the lipid in the liquid-expanded state forming the continuous phase. The use of fluorescent probes with selective phase partitioning allows analyzing by epifluorescence microscopy the migrations of the domains under the influence of inhomogeneous electric fields applied to the surface. Our observations indicate that a positive potential applied to an electrode placed over the monolayer promotes a repulsion of the domains until a steady state is reached, indicating the presence of a force opposed to the externally applied electric force. The experimental results were modeled by considering that the opposing force is generated by the dipole-dipole repulsion between the domains.

The catalytic activity of a glycosylphosphatidylinositol (GPI)-anchored alkaline phosphatase has been studied in Langmuir phospholipid monolayers at different surface pressures. The enzyme substrate, p-nitrophenyl phosphate, was injected into the subphase of mixed enzyme/lipid Langmuir monolayers. Its hydrolysis product was followed by monitoring the absorbance at 410 nm in situ in the monolayer subphase of the Langmuir trough. Several surface pressures, corresponding to different molecular surface densities, were attained by lateral compression of the monolayers. The morphology of the monolayers, observed by fluorescence microscopy, showed three different types of domains owing to the heterogeneous partition of the enzyme within the mixed enzyme/lipid film. The catalytic activity was modulated by the enzyme surface density, and it increased until a pressure of 18 mN/m was reached, but it decreased significantly when the equilibrium in-plane elasticity (surface compressional modulus) increased more noticeably, resulting in alterations in the interface morphology. A model for the modulation of the enzyme orientation and catalytic activity by lipid/enzyme surface morphology and enzyme surface packing at the air/liquid interface is proposed. The results might have an important impact on the comprehension of the enzymatic activity regulation of GPI-anchored proteins in biomembranes.

In this paper we report the dynamic response of the lateral topography and reflectance of monolayers of glycosphingolipids under changes of surface pressure (π) and composition. Epifluorescence and Brewster angle microscopy (BAM) observations have been carried out on pure monolayers formed with ceramide (Cer) and a series of glycosphingolipids (galactocerebroside, GM1 ganglioside, and asialo-GM1), and on mixed films formed with Cer and GM1. The curves of reflectance versus π for films of pure components are very dependent on the type and length of the oligosaccharide chain in the polar headgroup and are in agreement with the phase state of the π-area isotherms and with the overall molecular reorientation under compression shown by the variation of the perpendicular resultant dipole moments. In turn, the surface appearance of films formed with mixtures of GM1 and Cer is homogeneous as assessed by BAM, but epifluorescence microscopy reveals the coexistence of expanded and condensed phase domains that are under the limit of resolution of our BAM. The addition of Cer, which has a small headgroup, to GM1, which has the largest headgroup among the lipids studied, produced an average decrease of the dependence of reflectance on π (dI/dπ). Also, we analyzed the changes of reflectance and of dI/dπ with respect to the proportion of fluorescent and nonfluorescent phases. On the basis of these data it appears that the incorporation of GM1 in the Cer-enriched phase induces a drastic increase of reflectance without marked changes of dI/dπ with respect to phases of pure Cer. Cer incorporated into GM1-enriched phases of the mixed monolayers does not affect markedly the optical properties (reflectance and dI/dπ values) compared to films of pure GM1.

In the present paper the packing and dipolar properties of dimirystoylphosphatidylcholine and sphingolipid monolayers were studied at the air−water interface in correlation with the charge-transfer process of the K3Fe(CN)6/ K4Fe(CN)6 redox couple voltammetricaly analyzed on glassy carbon electrodes coated with the transferred monolayers. Lipid molecules having in common the lipophilic chain or the polar group were chosen for the analysis. Different degrees of electrochemical irreversibility in the voltammetric profile were observed depending on the lipid nature. The reversibility of the process was recovered after further addition of calcium chloride to the solution under study, indicating an increase in the monolayer permeability and consequently an increase in the rate of the charge-transfer process. The necessary amount of calcium to produce this effect depended also on the nature of the monolayer, which was evidence of some specific interaction between Ca²⁺ and each lipid. Surface pressure−area and surface potential−area compression isotherms at the air−water interface under different subphase composition were performed to get a better insight of this behavior. Interactions between the lipids and the ions were also observed, and a correlation between these results and the electrochemical response was found.

The transcription factor c-Fos forms stable Gibbs and Langmuir monolayers at the air-buffer interface. Its marked surface activity is enhanced by penetration into phospholipid films above the protein's own maximum adsorption surface pressure to a lipid-free interface. The protein-phospholipid stabilizing interactions at the interface depend on the lipid polar head group and the increases of lateral surface pressure generated are comparable to those of membrane-active proteins. The surface activity of c-Fos is strong enough to thermodynamically drive and retain c-Fos at the membrane interface where it may exert direct or indirect effects.

The effects of galactosylceramide (cerebroside) and sulfogalactosylceramide (sulfatide) from bovine brain on the lamellar-to-inverted hexagonal phase transition of dielaidoylphosphatidylethanolamine are examined using differential scanning calorimetry. When mixed with dielaidoylphosphatidylethanolamine, cerebroside increases the transition temperature (ca. 0.2 °C/mol % added cerebroside) and increases the transition ΔH. Sulfatide increases the transition temperature by ca. 0.4 °C/mol % added sulfatide and decreases ΔH. Both lipids are seen to hinder the formation of the nonlamellar phase, although sulfatide is more effective in this respect. When incorporated into vesicles formed by phosphatidylcholine/phosphatidylethanolamine/cholesterol (2:1:1 mole ratio), which are a good substrate for phospholipase C and undergo fusion as a consequence of the enzyme activity (Nieva et al. Biochemistry 1989, 28, 7364), cerebroside at all concentrations and sulfatide at >5 mol % inhibit enzyme activity and vesicle fusion. Cerebroside inhibition of fusion is due not only to a reduced enzyme activity but also to the impaired formation of nonlamellar phases. Sulfatide at low concentrations (e.g., 1 mol %) enhances phospholipase C activity and vesicle fusion, probably because its net negative charge causes hyperpolarization of the interface, which is known to activate phospholipase C. Under these conditions, its enzyme-activating effect predominates over its bilayer-stabilizing properties. Thus, sulfatide at low concentrations is an exception to the rule that amphiphiles hindering the lamellar−hexagonal transition inhibit both phospholipase C activity and membrane fusion.

The sphingomyelinase (Sphmase) activity degrading sphingomyelin (Sphm) monolayers shows a slow-reaction latency period before exhibiting constant rate catalysis. These two kinetic regions are regulated independently by the lateral surface pressure and by lipids that are biomodulators of cell function such as ceramide, glycosphingolipids, fatty acids, and lysophospholipids. Knowledge of the interfacial adsorption of Sphmase, precatalytic activation, initiation of effective catalysis, and the corresponding kinetic parameters is necessary for studying the level at which different lipids modulate the activity. We dissected some kinetic steps and determined the rate constants for degradation of Sphm, under controlled intermolecular organization, by Sphmase. Six models, adapted to two dimensions, were used to elucidate possible mechanisms for the interfacial activation of Sphmase during the lag time. The models consider enzyme binding to the substrate monolayer and a subsequent, essentially irreversible interfacial activation; this is supported experimentally by monolayer transfer experiments. Some mechanisms involve enzyme-substrate binding and associated states of the enzyme in the bulk subphase or at the interface, prior to complete activation. The activity of Sphmase is consistent with kinetics involving enzyme partitioning into the interface followed by substrate association, and by a process that proceeds with bimolecular kinetic dependence on the interfacial Sphmase concentration, and a subsequent slow step of activation. A possible equilibrium between the apparent monomolecular and bimolecular activated states of the interfacial enzyme, coupled to a slow activation, constitute rate-limiting steps that can explain the existence of lag time and the achievement of a maximum constant rate of degradation of Sphm monolayers by Sphmase.

Phospholipase A2 activity against mixed monolayers of dilauroylphosphatidic acid or dilauroylphosphatidylcholine with glycosphingolipids can be reversibly modulated by external constant electrostatic fields. The changes of enzymatic activity are correlated to the depolarization or hyperpolarization of the film caused by specific dipolar properties of glycosphingolipids. Hyperpolarizing fields enhance the enzymatic activity against pure dilauroylphosphatidic acid while depolarizing fields induce a decrease of activity. Compared to the pure substrate, the interface of mixed films containing neutral glycosphingolipids or gangliosides is already partially depolarized and the magnitude of activation induced by an external hyperpolarizing field is decreased; conversely, depolarizing fields cause an increased inhibition of activity. Differing from gangliosides, sulfatides bring about a hyperpolarization of the mixed lipid monolayer and external hyperpolarizing or depolarizing fields cause enhanced activation and reduced inhibition, respectively. The effects of glycosphingolipids depend on their relative proportion in the monolayer. Results were similar with dilauroylphosphosphatidylcholine but the field effects were less than half of those found with dilauroylphosphatidic acid. Our work shows that the activity of phospholipase A2 in addition to responding reversibly to external electrostatic fields, is directly modulated by the polarity and magnitude of the lipid polar head group dipole moments.

In mixtures with dipalmitoylphosphatidylcholine, ceramide induces broadening of the calorimetric main phase transition that could be deconvoluted into at least three components: the first represents isothermal melting of a phosphatidylcholine-enriched phase; the second and third represent phases with increasing proportions of ceramide melting at progressively higher temperatures. The partial phase diagram (up to 40 mole % ceramide) indicates complete or partial gel-phase immiscibility, and complete gel- and liquid-phase miscibility depending on the ceramide content. Cluster distribution function analysis of each individual transition reveals decreased cooperativity and domain size with increased amounts of ceramide. Compared to individual lipids, mixed monolayers with dipalmitoylphosphatidylcholine show unchanged mean molecular areas or slight expansions at 24 degrees C with dipole potentials exhibiting hyperpolarization; by contrast, already at 27 degrees C the mean molecular areas become condensed and dipole potentials show little changes or are slightly depolarized. This suggests that favorable ceramide;-phosphatidylcholine dipolar matching in the liquid state can be one of the local determinants for close molecular interactions while unfavorable matching may explain lateral domain segregation of ceramide-enriched gel phases. The changes are detected at relatively low proportions of Cer (1;-12 mole %) which are comparable to variations of Cer levels in membranes of cultured cells undergoing functional responses mediated by the sphingomyelin signaling pathway.

The presence of glycerol, free from surface-active impurities, modifies the molecular area, surface potential/molecule and thermodynamic parameters of compression of monolayers of galactosylceramide, sulphatide and gangliosides GM1, GD1a and GT1b. This may be due to changes of the composition and structural properties of the glycosphingolipid solvation shell with an influence on the intermolecular organization.

On t.l.c. plates 125I-cholera toxin binds to a disialoganglioside tentatively identified as GDlb with about 10 times less capacity than to ganglioside GM1. Binding of labeled toxin to both gangliosides was abolished in presence of excess amounts of unlabeled B subunit. Ganglioside extracts from human or pig intestinal mucosa showed toxin binding to gangliosides GM1 and GD1b. In ganglioside-containing lipid monolayers the penetration of the toxin was independent of the ganglioside binding capacity.

1. The interactions of four proteins (albumin, myelin basic protein, melittin and glycophorin) with eight neutral or acidic glycosphingolipids, including sulphatides and gangliosides, five zwitterionic or anionic phospholipids and some of their mixtures, were studied in lipid monolayers at the air/145 mM-NaCl interface. 2. In lipid-free interfaces, the surface pressure and surface potential reached by either soluble or integral membrane proteins did not reveal marked differences. 3. All the proteins studied showed interactions with each of the lipids but the maximal interactions were found for basic proteins with acidic glycosphingolipids. 4. Surface-potential measurements indicated that different dipolar organizations at the interface can be adopted by lipid-protein interactions showing the same value for surface free energy. 5. The individual surface properties of either the lipid of protein component are modified as a consequence of the lipid-protein interaction. 6. In mixed-lipid monolayers, the composition of the interface may affect the lipid-protein interactions in a non-proportional manner with respect to the relative amount of the individual lipid components.

1. The surface behaviour of six different gangliosides and eight chemically related glycosphingolipids was investigated in monolayers at the air-water interface. 2. Mono-, di-, tri and tetra-hexosylceramides had force-area isotherms showing similar limiting molecular areas on 145 mM-NaCl, pH 5.6. The increasing number of negatively charged sialosyl residues in mono-, di- and tri-sialogangliosides induced a progressive increase in the liquid-expanded character of the films and in the limiting area occupied per molecule, owing to electrostatic repulsions. When the ganglioside monolayers were spread on subphases at pH 1.2, the limiting area per molecule was similar to that found for neutral glycosphingolipids. 3. The monolayer collapse pressure at pH 5.6 increased with the number of uncharged carbohydrate units up to when the polar head group contained 3-4 residues. For gangliosides the collapse pressures were lower and decreased from mono- to tri-sialogangliosides. Ganglioside monolayers on subphases at pH 1.2 showed increases in their collapse pressure. 4. The glycosphingolipid monolayers studied had various surface in their collapse pressure. 4. The glycosphingolipid monolayers studied had various surface potentials according to the complexity of the polar head group of the lipid. Attempts to calculate the dipolar contributions to the surface potential from each carbohydrate residue suggest that the second and third sialosyl residues in di- and tri-sialogangliosides contributed with a vertical dipole moment opposite to that of the first sialosyl residue.

1. The penetration of alpha-tocopherol and seven of its derivatives, and five compounds in the ubiquinone series, having differing chain lengths, into monolayers at the air/water interface of 11 different synthetic phospholipids and cholesterol was investigated; the properties of mixed monolayers of the tocopherols and of ubiquinones with phospholipids were also studied. 2. Penetration of alpha-tocopherol into diarachidonylglycerylphosphorycholine was approximately constant for molar ratios of tocopherol/phospholipid ranging from 0.4:1.0 to 2.0:1.0. 3. Tocopherols with shorter or longer side chains than alpha-tocopherol had a lesser ability to penetrate monolayers of phospholipid molecules with 16 or more carbon atoms in their acyl chains. 4. All the tocopherols penetrated more readily as unsaturation in the phospholipids was increased, and their penetration into mixed monolayers of phospholipids was greatly facilitated by the presence of relatively small quantities of unsaturated phospholipid molecules. 5. There was relatively little interaction between the tocopherols and cholesterol, or between the ubiquinones and phospholipids. 6. The possible significance of the observed interactions between alpha-tocopherol and polyunsaturated phospholipids is discussed in relation to the biochemical actions of alpha-tocopherol in vivo. 7. It is suggested that fluidity of the lipid bilayer in membranes containing polyunsaturated phospholipids may allow alpha-tocopherol to interact in a dynamic manner with a number of phospholipid molecules.

Poly(ethylene glycol), glycerol and dimethyl sulphoxide markedly decrease the surface potentials of monolayers of phosphatidylcholine and phosphatidylethanolamine. This finding is discussed in relation to the properties of hen erythrocytes undergoing fusion induced by poly(ethylene glycol).

1. The behaviour of mixed monolayers of 14 different lipids with preparations of erythrocyte lipids, purified natural and synthetic phospholipids, cholesterol and galactosylceramide was investigated. 2. The mean areas occupied per molecule in mixed films containing lipids that are fusogenic for hen erythrocytes were compared with those for corresponding films containing lipids that are inactive as fusogens. 3. Fusogenic lipids were found to exhibit interactions, which were not shown by non-fusogenic lipids, in mixed monolayers with several species of phospholipid, particularly those containing a choline head group. 4. Heterogeneity in the hydrophobic chains of phosphatidylcholine, their degree of unsaturation and the presence of cholesterol had little effect on the interaction of phosphatidylcholine with fusogenic lipids. 5. Fusogenic lipids showed little specific interaction with natural or synthetic preparations of phosphatidylethanolamine. 6. The possible significance of these observations in relation to the action of fusogenic lipids on biological membranes is discussed in the light of the asymmetrical distribution of phospholipids in erythrocyte membranes.

We studied the effect of gangliosides GD1a and GM1 on the lamellar-to-hexagonal II phase transition of mixtures of dioleoylphosphatidylethanolamine/dioleoylphosphatidyl choline, 3:1, and of transphosphatidylated phosphatidylethanolamine with dioleoylglycerol by high-sensitivity differential scanning calorimetry, 31P-NMR, and pyrene fluorescence of a phosphatidylcholine probe. Gangliosides had a dual effect. Below 1 mol % ganglioside the hexagonal II phase transition was affected but still occurred at lower temperature than in the absence of gangliosides. The presence of between 1 and 2 mol % gangliosides increased the temperature for formation of the hexagonal II phase and progressively decreased its cooperativity. Above 3 mol % gangliosides totally inhibited the formation of both the temperature-induced and composition-induced hexagonal phase, probably by opposing the geometric distortions necessary for the inverted micellar structures.

The thermotropic behavior of the natural glycosphingolipids galactosylceramide, asialo-Gal beta 1-3GalNAc beta 1-4Gal(3-2 alpha NeuAc)beta 1-4Glc beta 1-Cer (GM1), sulfatide, GM1, NeuAc alpha 2-3Gal beta 1-3GalNAc beta 1-4Gal(3-2 alpha NeuAc)beta 1-4Glc beta 1-1Cer (GD1a), and NeuAc alpha 2-3Gal beta 1-3GalNAc beta 1-4Gal(3-2 alpha NeuAc8-2 alpha NeuAc)beta 1-4Glc beta 1-1 Cer (GT1b), and their mixtures with dipalmitoylphosphatidylcholine (DPPC) in the presence of myelin basic protein (MBP) was studied by high sensitivity differential scanning calorimetry. The transition temperature of DPPC, galactosylceramide, and asialo-GM1 is affected little by MBP while their transition enthalpy is decreased in proportion to the amount of protein in the mixture. The thermotropic behavior of anionic glycosphingolipids is considerably perturbed by MBP. The transition temperature of gangliosides increases in the presence of MBP, whereas that of sulfatide decreases. The enthalpy of the transition of anionic glycosphingolipids increases markedly in the presence of MBP. The excess heat capacity function of these systems can be resolved into two independent phase transitions. Phase separation of enriched lipid/protein domains occurs in a magnitude that depends on the amount of MBP; the rest of the lipid phase exhibits some altered thermodynamic properties. In mixtures of glycosphingolipids with DPPC, phase separation is also present but no phase transition with the characteristic of pure DPPC is found. MBP is changing the properties of the lipid mixture as a whole and does not interact exclusively with the glycosphingolipids. The proportion of MBP required to produce the maximal changes is greater the greater the complexity of the glycosphingolipids polar head group. Relatively small variations of the amount of MBP induce large shifts in the proportion of the different phases present.

The effect of neutral (galactocerebroside and asialo-ganglioside GM1) or anionic (sulphatide and gangliosides GM1, GD1a and GT1b) glycosphingolipids on the activity of phospholipase A2 from pig pancreas was studied in mixed monolayers of dilauroyl phosphatidylcholine with the glycosphingolipids in different molar fractions at various constant surface pressures. The activity of the enzyme depends on the proportion and type of glycosphingolipid in the interface. Sulphatide activates the enzyme at all proportions, whereas galactocerebroside shows inhibition or activation depending on its proportion in the film. Asialo-ganglioside GM1 and gangliosides GM1, GD1a and GT1b can strongly inhibit the enzyme at relatively low molar fractions in the film in the following order: asialo-ganglioside GM1 less than ganglioside GM1 less than ganglioside GT1b less than ganglioside GD1a. The changes of activity are not due to a direct action of the lipids on the active centre or interfacial recognition region of the enzyme.

The penetration of melittin and myelin basic protein into glycosphingolipid monolayers depends on the lipid polar head group, the protein concentration available and the initial surface pressure. The lipid-protein interaction leads to modification of the surface properties of both the glycosphingolipid and the proteins.

1. The interactions among five different gangliosides and three chemically related glycosphingolipids and their behaviour in mixed monolayers with six different phospholipids were investigated at the air/145 mM-NaCl interface at pH 5.6. 2. The mixed monolayers of any of the different gangliosides showed an immiscible behaviour at high surface pressures, with absence of interactions among them revealed by an ideal behaviour for mean molecular area and surface potential per molecule. 3. This behaviour was probably the consequence of steric hindrance and electrostatic repulsions between their polar head groups. 4. Di- and tri-sialogangliosides could be differentiated from neutral sphingolipids and monosialogangliosides on the basis of their interactions with phospholipids, which were correlated to the perpendicular electric field at the interface contributed by the carbohydrate residues. 5. The presence of the phosphocholine polar head group in phosphatidylcholine was important to establish interactions with di- and tri-sialogangliosides revealed by negative deviations from the ideal behaviour for mean molecular areas and mean surface potential per molecule. 6. The possible significance of these observations is discussed in relation to the participation of gangliosides in the organization of membranes and to their capability of inducing membrane fusion.

1. The surface potentials of mixed monolayers of synthetic phospholipids with lipids that are fusogenic for hen erythrocytes were investigated. 2. At pH 5.6 and 10, but not at pH2, mixed monolayers of the fusogenic lipid, glycerol mono-oleate, with phosphatidylcholine exhibited negative deviations from the ideality rule in surface potential per molecule which were accompanied by negative deviations in mean molecular area. 3. Interactions of this type were not seen with chemically related but non-fusogenic lipids, nor were they found in mixed monolayers of any of the lipids with phosphatidylethanolamine. 4. Experiments with dihexadecyl phosphate and hexadecyltrimethyl-ammonium indicated that the complete head group of phosphatidylcholine is required for its observed behaviour with fusogenic lipids. 5. Bivalent cations (Ca2+, UO2(2+) or Zn2+) in the subphase at pH 5.6 significantly modified the behaviour of mixed monolayers of fusogenic lipids with phospholipids; there was a parallel perturbing effect of fusogenic lipids on interactions between monolayers of phospholipids and bivalent cations. 6. Possible molecular interactions of fusogenic lipids with membrane phospholipids, and the role of Ca2+, are discussed which may be relevant to cell fusion in erythrocytes induced by low-melting lipids in the presence of Ca2+.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Eleven diaminoalkanes such as (I) are coupled with the salicylate (II) to produce the title compounds (III).

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.