Interaction between PAF and human platelet membranes: a fluorescence study

The interaction between PAF and human platelet membranes was investigated by measuring the steadystate fluorescence anisotropy and fluorescence decay of 1 (4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) incorporated in platelet plasma membranes. PAF induced a time-limited and significant increase of the lipid order in the exterior part of the membrane and a decrease in membrane heterogeneity. These changes were blocked in the presence of the PAF antagonists, L-659,989 and 1-O-hexadecyl-2-acetyl-sn-glycero-3-phospho(N,N,N-trimethyl)hexanolamine.H(2)O. These results indicate that the observed changes in the physico-chemical properties of the membrane are attributed to the PAF-receptor interaction and signal transduction.     

Interaction of benzocaine with model membranes

We measured the absorption properties, water solubility and partition coefficients (P) between n-octanol, egg phosphatidylcholine (EPC) liposomes and erythrocyte ghosts/water for benzocaine (BZC), an ester-type always uncharged local anesthetic. The interaction of BZC with EPC liposomes was followed using Electron Paramagnetic Resonance, with spin labels at different positions in the acyl chain (5, 7, 12, 16-doxylstearic acid methyl ester). Changes in lipid organization upon BZC addition allowed the determination of P values, without phase separation. The effect of BZC in decreasing membrane organization (maximum of 11.6% at approx. 0.8:1 BZC:EPC) was compared to those caused by the local anesthetics tetracaine and lidocaine. Hemolytic tests revealed a biphasic (protective/inductive) concentration-dependent hemolytic effect for BZC upon rat erythrocytes, with an effective BZC:lipid molar ratio in the membrane for protection (RePROT), onset of hemolysis (ReSAT) and 100% membrane solubilization (ReSOL) of 1.0:1, 1.1:1 and 1.3:1, respectively. The results presented here reinforce the importance of considering hydrophobic interactions in the interpretation of the effects of anesthetics on membranes.     

Interaction of mitochondrial creatine kinase with model membranes. A monolayer study

The interaction of mitochondrial creatine kinase (Mi-CK; EC 2.7.3.2) with phospholipid monolayers and spread mitochondrial membranes at the air/water interface has been investigated. It appeared that Mi-CK penetrated into these monolayers as evidenced by an increase in surface pressure upon incorporation of Mi-CK. The increase in surface pressure was dependent on (1) the amount and (2) the oligomeric form of Mi-CK in the subphase, as well as on (3) the initial surface pressure and (4) the phospholipid composition of the monolayer. In this experimental system Mi-CK was able to interact equally well with both inner and outer mitochondrial membranes.     

Interaction between model membranes and a new class of surfactants with antioxidant function.

The effect of two series of amphiphilic quaternary ammonium salts on some properties of phospholipid membranes was studied. The compounds of one series, N-benzyl-N,N-dimethyl-N-alkyl ammonium bromides, exert a destructive effect on membranes and are treated as reference compounds. The compounds of the other series, N-(3,5-di-t-butyl-4-hydroxy)benzyl-N,N-dimethyl-N-alkyl ammonium bromides, are derivatives of the former ones, exhibit antioxidant properties, and do only relatively slight damage to the membranes. The aim of the work was to explain the difference in molecular interaction with membranes between the two kinds of hydrophobic compounds. Thermodynamic methods, a new mixing technique, and monolayer and quantum calculation methods were used. It has been shown that the antioxidant molecules are less hydrophobic than those of the reference compounds and disturb the membrane organization to a lesser extent. On the basis of monolayer data, we suggest that the studied antioxidant behaves like a substitutional impurity, whereas the reference behaves like an interstitial one.     

Interaction of the pitavastatin with model membranes

Pitavastatin is a statin drug that, by competitively inhibiting 3-hydroxy-3-methylglutaryl-coenzyme A reductase, can lower serum cholesterol levels of low-density lipoprotein (LDL) accompanied by side effects due to pleiotropic effects leading to statin intolerance. These effects can be explained by the lipophilicity of statins, which creates membrane affinity and causes statin localization in cellular membranes. In the current report, the interaction of pitavastatin with POPC model membranes and its influence on the membrane structure were investigated using ¹H, ²H and ³¹P solid-state NMR spectroscopy. Our experiments show the average localization of pitavastatin at the lipid/water interface of the membrane, which is biased towards the hydrocarbon core in comparison to other statin molecules. The membrane binding of pitavastatin also introduced an isotropic component into the ³¹P NMR powder spectra, suggesting that some of the lamellar POPC molecules are converted into highly curved structures.     

Interaction between amyloid beta-protein aggregates and membranes.

The conversion of soluble, nontoxic amyloid beta-protein (Abeta) to aggregated, toxic Abeta rich in beta-sheet structures is considered to be the key step in the development of Alzheimer's disease. Therefore, extensive studies have been carried out on the mechanisms involved in Abeta aggregation and the characterization of Abeta aggregates formed in aqueous solutions mimicking biological fluids. On the other hand, several investigators pointed out that membranes play an important role in Abeta aggregation. However, it remains unclear whether Abeta aggregates formed in solution and membranes are identical and whether the former can bind to membranes. In this study, using a dye-labeled Abeta-(1-40) as well as native Abeta-(1-40), the properties of Abeta aggregates formed in buffer and raft-like membranes composed of monosialoganglioside GM1/cholesterol/sphingomyelin were compared. Fourier transform infrared spectroscopic measurements suggested that Abeta aggregates formed in buffer and in membranes have different beta-sheet structures. Fluorescence experiments revealed that Abeta aggregated in buffer did not show any affinity for membranes.     

Interaction of diphtheria toxin with model membranes

Low pH is believed to trigger membrane penetration by diphtheria toxin in vivo. The effect of pH upon the binding of the toxin to unilamellar model membrane vesicles was determined by using a fluorescence quenching assay. A series of studies were undertaken to determine the effect of lipid composition upon the binding of lipids to the toxin. The binding of toxin to various small unilamellar vesicles of zwitterionic or anionic lipids was similar in extent and was accompanied by deep penetration of the toxin into the fatty acyl chains, in agreement with previous studies. However, the transition pH, which is the pH at and below which toxin binding becomes significant, depended upon the fraction of anionic lipids, being highest with model membranes composed totally of anionic lipids (pH 5.8) and lowest with membranes composed of zwitterionic lipids (pH 5.2). Except for vesicle charge, the transition pH was independent of the nature of the lipid polar groups used. High ionic strength, which had no effect on the transition pH with zwitterionic vesicles, was found to shift the transition pH with totally anionic vesicles to pH 5.2. This suggests that both direct protein-lipid electrostatic interactions and the ionic double layer, which gives rise to a low local pH around anionic vesicles, contribute to the shift in the transition pH. The effect of lipid composition upon the kinetics and strength of binding was also examined. At low pH, binding was rapid and tight. Binding to vesicles containing 20 wt % anionic phosphatidylglycerol was faster and tighter than binding to vesicles of zwitterionic phosphatidylcholine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interaction of dystrophin fragments with model membranes

The interaction with membrane lipids of recombinant fragments of human dystrophin, corresponding to a single structural repeating unit of the rod domain, was examined. Surface plasmon resonance, constant-pressure isotherms in a Langmuir surface film balance, and interfacial rheology were used to observe binding of the polypeptides and its effects on the properties of the lipid film. Modification of the monolayer properties was found to depend on the presence of phosphatidylserine in the lipid mixture and on the native tertiary fold of the polypeptide; thus a fragment with the minimum chain length required for folding (117 residues) or longer caused a contraction of the surface area at constant pressure, whereas fragments of 116 residues or less had no effect. The full extent of contraction was reached at a surface concentration of lipid corresponding to an average area of about 42 A2 per lipid molecule. A dystrophin fragment with the native, folded conformation induced a large increase in surface shear viscosity of the lipid film, whereas an unfolded fragment had no effect. Within a wide range of applied shear, the shear viscosity remained Newtonian. Binding of liposomes to immobilized dystrophin fragments could be observed by surface plasmon resonance and was again related to the conformational state of the polypeptide and the presence of phosphatidylserine in the liposomes. Our results render it likely that intact dystrophin interacts directly and strongly with the sarcolemmal lipid bilayer and grossly modifies its material properties.     

Interaction between ubiquinone and ATPase in mitochondrial membranes

The extraction of ubiquinone from mitochondrial membranes produces alterations of ATPase activity including a reversible loss of oligomycin sensitivity which is restored by long-chain Q-homologs. Short-chain ubiquinones like Q₃ produce a loss of oligomycin and dicyclohexylcarbodiimide (DCCD) sensitivity in submitochondrial particles. The effect shows uncompetitive or noncompetitive kinetics with respect to oligomycin or DCCD respectively. Long-chain ubiquinones have a competitive effect with Q₃, thus restoring oligomycin sensitivity; they behave, however, in about the same way as Q₃ in lowering the DCCD sensitivity in submitochondrial particles. On the basis of these observations we suggest that ubiquinone may be a physiological modulator of ATPase activity in the mitochondrial membrane.Abbreviations used: BHM, beef heart mitochondria; DCCD, dicyclohexylcarbodiimide; ETP, electron transfer particles (submitochondrial particles); Q, ubiquinone.     

Interaction between Ca2+ and dipalmitoylphosphatidylcholine membranes. II. Fluorescence anisotropy study

The effect of Ca2+ on the molecular mobility in dipalmitoylphosphatidylcholine membranes was studied by steady-state and time-resolved measurements of fluorescence anisotropy. The fluorescence anisotropy decay of 1,6-diphenyl-1,3,5-hexatriene in the hydrocarbon region indicated that the free volume of molecular rotation became more restricted when the Ca2+ concentration was increased. The decrease of the molecular mobility was observed from 1 mM Ca2+, at which the number of bound Ca2+ is much less than that of the total lipid molecules. A distinct difference between Ca2+ and Mg2+ effects suggested that the change in various membrane properties was induced by the binding of these ions. From these results we propose a long-range attractive interaction between bound Ca2+ and the polar head groups of distant phosphatidylcholine molecules.     

Interaction between inclusions embedded in membranes.

We calculate the membrane-induced interaction between inclusions, in terms of the membrane stretching and bending moduli and the spontaneous curvature. We find that the membrane-induced interaction between inclusions varies nonmonotonically as a function of the inclusion spacing. The location of the energy minimum depends on the spontaneous curvature and the membrane perturbation decay length, where the latter is set by the membrane moduli. The membrane perturbation energy increases with the inclusion radius. The Ornstein-Zernike theory, with the Percus-Yevick closure, is used to calculate the radial distribution function of inclusions. We find that when the spontaneous curvature is zero, the interaction between inclusions due to the membrane deformation is qualitatively similar to the hard-core interaction. However, in the case of finite spontaneous curvature, the effective interaction is dramatically modified.     

Interaction of polynucleotides with natural and model membranes.

Polynucleotides adsorb on natural and model phospholipid membranes in the presence of Mg2+-cations. Adsorption of nucleic acids on membranes results in a considerable change of their secondary structure. The presence of model phosphatidylcholine membranes greatly stimulates the rate of the synthesis of RNA by E. coli RNA-polymerase on DNA template.     

Interaction of Hsp90 with phospholipid model membranes

Heat shock protein 90 (Hsp90) is an essential molecular chaperone with versatile functions in cell homeostatic control under both normal and stress conditions. Hsp90 has been found to be expressed on the cell surface, but the mechanism of Hsp90 association to the membrane remains obscure. In this study, the direct interaction of Hsp90 and phospholipid vesicles was characterized, and the role of Hsp90 on membrane physical state was explored. Using surface plasmon resonance (SPR), we observed a strong interaction between Hsp90 and different compositions of lipid. Hsp90 had a preference to bind with more unsaturated phospholipid species and the affinity was higher with negatively charged lipids than zwitterionic lipids. Increasing the mole fraction of cholesterol in the phospholipid led to a decrease of binding affinity to Hsp90. Circular dichroism (CD) spectroscopy of Hsp90 in PC membranes showed more α-helix structure than in aqueous buffer. The differential scanning calorimeter (DSC) and fluorescence polarization results showed Hsp90 could affect the transition temperature and fluidity of the bilayer. We postulate from these results that the association between Hsp90 and membranes may involve both electrostatic and hydrophobic force, and constitute a possible mechanism that modulates membrane lipid order during thermal fluctuations.     

Interaction of cationic antimicrobial peptides with model membranes

A series of natural and synthetic cationic antimicrobial peptides from various structural classes, including alpha-helical, beta-sheet, extended, and cyclic, were examined for their ability to interact with model membranes, assessing penetration of phospholipid monolayers and induction of lipid flip-flop, membrane leakiness, and peptide translocation across the bilayer of large unilamellar liposomes, at a range of peptide/lipid ratios. All peptides were able to penetrate into monolayers made with negatively charged phospholipids, but only two interacted weakly with neutral lipids. Peptide-mediated lipid flip-flop generally occurred at peptide concentrations that were 3- to 5-fold lower than those causing leakage of calcein across the membrane, regardless of peptide structure. With the exception of two alpha-helical peptides V681(n) and V25(p,) the extent of peptide-induced calcein release from large unilamellar liposomes was generally low at peptide/lipid molar ratios below 1:50. Peptide translocation across bilayers was found to be higher for the beta-sheet peptide polyphemusin, intermediate for alpha-helical peptides, and low for extended peptides. Overall, whereas all studied cationic antimicrobial peptides interacted with membranes, they were quite heterogeneous in their impact on these membranes.     

Interaction of psychotropic preparations with model lipid membranes

Interaction of psychotropic drugs with model phosphatidylcholine membranes was investigated by fluorescent probes. The data obtained indicate different affinity of the drugs for phosphatidylcholine. The tranquilizers were not bound to the model membranes. The antidepressants were localized in the lipid polar groups area whereas the neuroleptics in the lipid polar groups area and deeper regions of the lipid bilayer.     

Interaction of enkephalin peptides with anionic model membranes

According to the model for passive transport across the membranes, the total flow of permeant molecules is related to the product of the water-membrane partition coefficient and the diffusion coefficient, and to the water-membrane interfacial barrier. The effect of membrane surface charge on the permeability and interaction of analgesic peptide ligands with model membranes was investigated. A mixture of zwitterionic phospholipids with cholesterol was used as a model membrane. The lipid membrane charge density was controlled by the addition of anionic 1-palmitoyl-2-oleoylphosphatidylserine. Two classes of highly potent analgesic peptides were studied, c[D-Pen²,D-Pen⁵]enkephalin (DPDPE) and biphalin, a dimeric analog of enkephalin. The effect of increased surface charge on the permeability of the zwitterionic DPDPE is a relatively modest decrease, that appears to be due to a diminished partition coefficient. On the other hand the binding of the dicationic biphalin ligands to membranes increases proportionally with increased negative surface charge. This effect translates into a significant reduction of biphalin permeability by reducing the diffusion of the peptide across the bilayer. These experiments show the importance of electrostatic effects on the peptide-membrane interactions and suggest that the negative charge naturally present in cell membranes may hamper the membrane transport of some peptide drugs, especially cationic ones, unless there are cationic transporters present.     

Interaction between lanthanum and lipid moieties of biological membranes

The binding of colloidal lanthanum to isolated biological membranes and lipid vesicles was studied by optical and X-ray spectroscopy, and by electron microscopy. Excellent correlation was observed using these techniques. The increase in turbidity and electron density appeared to be directly proportional to the extent of negative charge in the system. X-ray microanalysis confirmed the presence of lanthanum on the lipid vesicles. Among the negatively charged phospholipids, phosphatidylinositol showed the greatest affinity for lanthanum.     

The study on the interaction between phytosterols and phospholipids in model membranes

Sterols are one of the major components of cellular membranes. Although in mammalian membranes cholesterol is a predominant sterol, in the human organism plant sterols (phytosterols) can also be found. Phytosterols, especially if present in concentrations higher than normal (phytosterolemia), may strongly affect membrane properties. In this work, we studied phytosterol-phospholipid interactions in mixed Langmuir monolayers serving as model membranes. Investigated were two phytosterols, beta-sitosterol and stigmasterol and a variety of phospholipids, both phosphatidylethanolamines and phosphatidylcholines. The phospholipids had different polar heads, different length and saturation of their hydrocarbon chains. The interactions between molecules in mixed sterol/phospholipid films were characterized with the mean area per molecule (A(12)) and the excess free energy of mixing (DeltaG(Exc)). The effect of the sterols on the molecular organization of the phospholipid monolayers was analyzed based on the compression modulus values. It was found that the incorporation of the phytosterols into the phospholipid monolayers increased their condensation. The plant sterols revealed higher affinity towards phosphatidylcholines as compared to phosphatidylethanolamines. The phytosterols interacted more strongly with phospholipids possessing longer and saturated chains. Moreover, both the length and the saturation of the phosphatidylcholines influenced the stoichiometry of the most stable complexes. Our results, compared with those presented previously for cholesterol/phospholipid monolayers, allowed us to draw a conclusion that the structure of sterol (cholesterol, beta-sitosterol, stigmasterol) does not affect the stoichiometry of the most stable complexes formed with particular phospholipids, but influences their stability. Namely, the strongest interactions were found for cholesterol/phospholipids mixtures, while the weakest for mixed systems containing stigmasterol.