Diazeniumdiolate reactivity in model membrane systems.

The effect of small unilamellar phospholipid vesicles on the acid-catalyzed dissociation of nitric oxide from diazeniumdiolate ions, R(1)R(2)N[N(O)NO](-), [1: R(1)=H(2)N(CH(2))(3)-, R(2)=H(2)N(CH(2))(3)NH(CH(2))(4)-; 2: R(1)=R(2)=H(2)N(CH(2))(3)-; 3: R(1)=n-butyl-, R(2)=n-butyl-NH2+(CH(2))(6)-; 4: R(1)=R(2)=nPr-] has been examined at pH 7.4 and 37 degrees C. NO release was catalyzed by anionic liposomes (DPPG, DOPG, DMPS, POPS and DOPA) and by mixed phosphatidylglycerol/phosphatidylcholine (DPPG/DPPC and DOPG/DPPC) covesicles, while cationic liposomes derived from 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and the zwitterionic liposome DMPC did not significantly affect the dissociation rates of the substrates examined. Enhancement of the dissociation rate constant in DPPG liposome media (0.010M phosphate buffer, pH 7.4, 37 degrees C) at 10mM phosphoglycerol levels, ranged from 37 for 1 to 1.2 for the anionic diazeniumdiolate 4, while DOPA effected the greatest rate enhancement, achieving 49-fold rate increases with 1 under similar conditions. The observed catalysis decreases with increase in the bulk concentration of electrolytes in the reaction media. Quantitative analysis of catalytic effects has been obtained through the application of pseudo-phase kinetic models and equilibrium binding constants at different liposome interfaces are compared. The stoichiometry of nitric oxide release from 1 and 2 in DPPG/DPPC liposome media has been obtained through oxyhemoglobin assay. DPPG=1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], DOPG=1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], DMPS=1,2-dimyristoyl-sn-glycero-3-[phospho-l-serine], POPS=1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-l-serine], DOPA=1,2-dioleoyl-sn-glycero-3-phosphate; DPPC=1,2-dipalmitoyl-sn-glycero-3-phosphocholine, DMPC=1,2-dimyristoyl-sn-glycero-3-phosphocholine, DOTAP=1,2-dioleoyl-3-trimethylammonium-propane.     

Lipid membrane templates the ordering and induces the fibrillogenesis of Alzheimer's disease amyloid-beta peptide

The lipid membrane has been shown to mediate the fibrillogenesis and toxicity of Alzheimer's disease (AD) amyloid-beta (Abeta) peptide. Electrostatic interactions between Abeta40 and the phospholipid headgroup have been found to control the association and insertion of monomeric Abeta into lipid monolayers, where Abeta exhibited enhanced interactions with charged lipids compared with zwitterionic lipids. To elucidate the molecular-scale structural details of Abeta-membrane association, we have used complementary X-ray and neutron scattering techniques (grazing-incidence X-ray diffraction, X-ray reflectivity, and neutron reflectivity) in this study to investigate in situ the association of Abeta with lipid monolayers composed of either the anionic lipid 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), the zwitterionic lipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), or the cationic lipid 1,2-dipalmitoyl 3-trimethylammonium propane (DPTAP) at the air-buffer interface. We found that the anionic lipid DPPG uniquely induced crystalline ordering of Abeta at the membrane surface that closely mimicked the beta-sheet structure in fibrils, revealing an intriguing templated ordering effect of DPPG on Abeta. Furthermore, incubating Abeta with lipid vesicles containing the anionic lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) induced the formation of amyloid fibrils, confirming that the templated ordering of Abeta at the membrane surface seeded fibril formation. This study provides a detailed molecular-scale characterization of the early structural fluctuation and assembly events that may trigger the misfolding and aggregation of Abeta in vivo. Our results implicate that the adsorption of Abeta to anionic lipids, which could become exposed to the outer membrane leaflet by cell injury, may serve as an in vivo mechanism of templated-aggregation and drive the pathogenesis of AD.     

Interaction of the pore-forming protein equinatoxin II with model lipid membranes: A calorimetric and spectroscopic study.

The interactions of equinatoxin II (EqTxII) with zwitterionic (DPPC) and anionic (DPPG) phospholipids and an equimolar mixture of the two phospholipids (DPPC/DPPG) have been investigated by differential scanning calorimetry (DSC), CD-spectropolarimetry, intrinsic emission fluorescence spectroscopy, and ultrasonic velocimetry. EqTxII binds to small unilamellar vesicles formed from negatively charged DPPG lipids, causing a marked reduction in the cooperativity and enthalpy of their gel/liquid-crystalline phase transition. This transition is completely abolished at a lipid-to-protein ratio, L/P, of 10. For the mixed DPPC/DPPG vesicles, a 2-fold greater lipid-to-protein ratio (L/P = 20) is required to abolish the phase transition, which corresponds to the same negative charge (-10) of lipid molecules per EqTxII molecule. The disappearance of the phase transition of the lipids apparently corresponds to the precipitation of the lipid-protein complex, as suggested by our sound velocity measurements. Based on the far-UV CD spectra, EqTxII undergoes two structural transitions in the presence of negatively charged vesicles (DPPG). The first transition coincides with the gel/liquid-crystalline phase transition of the lipids, which suggests that the liquid-crystalline form of negatively charged lipids triggers structural changes in EqTxII. The second transition involves the formation of alpha-helical structure. Based on these observations, we propose that, in addition to electrostatic interactions, hydrophobic interactions play an important role in EqTxII-membrane association.     

Effects of hydrostatic pressure on the molecular structure and endothermic phase transitions of phosphatidylcholine bilayers: a Raman scattering study

The temperature dependences of the Raman spectra of aqueous dispersions of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) were monitored at different but constant pressures between 1 and 1210 bar. The changes observed in these Raman spectra are discussed in terms of the effects of high pressure on the phase state and molecular structure of lipid bilayers. It is demonstrated that the temperature of the endothermic gel to liquid-crystal phase transition, as well as the temperature of the pretransition, increases linearly with increasing hydrostatic pressure. The dTm/dP values obtained from a wide range of pressures are 20.8 degrees C X kbar-1 for DPPC and 20.1 degrees C X kbar-1 for DMPC. The dTp/dP value for DPPC is 16.2 degrees C X kbar-1. It is also shown that the volume change that occurs at the gel to liquid-crystal transition is not constant; i.e., d delta Vm/dP decreases by 6.2% (DPPC) or 6.3% (DMPC) per kilobar pressure. The volume change at the pretransition is also pressure dependent; the d delta Vp/dP value of DPPC decreases by 4.7% per kilobar pressure.     

Surface-enhanced resonance Raman scattering of porphyrins on gold nanoparticles attached to silanized glass plates

Surface-enhanced resonance Raman scattering (SERRS) spectra of cationic 5,10,15,20-tetrakis(1-methyl-4-pyridyl) porphyrin (TMPyP) and anionic 5,10,15,20-tetrakis(4-sulfonatophenyl) porphyrin (TSPP) were measured from gold surfaces prepared by attaching citrate-reduced colloidal nanoparticles to glass slides silanized by 3-aminopropyltrimethoxysilane. SERRS spectra of both porphyrins obtained in a large concentration range (1 x 10(-4) to 1 x 10(-7)M) of primary solution do not show any sign of porphyrin metalation or perturbation of its native structure. Optimal adsorption time (15-20 min) and covering concentration limit (lower than 1 x 10(-5)M) of porphyrins have been estimated from the concentration and soaking time dependences of SERRS spectra.     

Thermotropic behavior and electronmicroscopic structures of mixtures of gangliosides and dipalmitoylphosphatidylcholine

The calorimetric properties and morphological structures of dispersed mixtures of 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and highly purified human brain gangliosides, GM2, GM1, GD1a, GD1b, and GT1b, were studied using a high-sensitivity differential scanning calorimeter and an electron-microscope, as a function of the ganglioside molar fraction. No thermal phase transitions of pure gangliosides in aqueous dispersions could be detected. In the mixtures of DPPC and gangliosides, the gel to liquid crystalline phase transition occurred at a higher temperature than in pure DPPC dispersions and progressed over a wide temperature range. As increasing amounts of the pure ganglioside species were added to DPPC, the temperature for the main transition gradually increased. The phase transition progressed differently among different gangliosides/DPPC mixtures. The enthalpy values were found to decrease linearly as the number of sialic acid residues increased. Electron-microscopically the ganglioside/DPPC mixtures formed multilamellar structures at lower concentrations of the gangliosides, and the structures changed to cylindrical and spherical micelles as the ganglioside concentration was increased. The polysialoganglioside/DPPC mixtures showed the micellar form even at lower ganglioside concentrations, contrary to the case of the monosialoganglioside/DPPC mixtures. The morphological changes of gangliosides/DPPC mixtures corresponded with changes in the calorimetric properties. These results show that individual gangliosides have different physicochemical effects on model membranes, possibly because of the interaction of their negatively charged head groups.     

Investigating the effects of L- to D-amino acid substitution and deamidation on the activity and membrane interactions of antimicrobial peptide anoplin

Isolated from the venom sac of solitary spider wasp, Anoplius samariensis, anoplin is the smallest linear α-helical antimicrobial peptide found naturally with broad spectrum activity against both Gram-positive and Gram-negative bacteria, and little hemolytic activity toward human erythrocytes. Deamidation was found to decrease the peptide's antibacterial properties. In the present work, interactions of amidated (Ano-NH2) and deamidated (Ano-OH) forms of anoplin as well as Ano-NH2 composed of all D-amino acids (D-Ano-NH2) with model cell membranes were investigated by means of Langmuir Blodgett (LB) technique, atomic force microscopy (AFM), X-ray photoemission electron microscopy (X-PEEM) and carboxyfluorescein leakage assay in order to gain a better understanding of the effect of these peptide modifications on membrane binding and lytic properties. According to LB, all three peptides form stable monolayers at the air/water interface with Ano-NH2 occupying a slightly greater area per molecule than Ano-OH. All three forms of the peptide interact preferentially with anionic 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), rather than zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid monolayer. Peptides form nanoscale clusters in zwitterionic but not in anionic monolayers. Finally, membrane lytic activity of all derivatives was found to depend strongly on membrane composition and lipid/peptide ratio. The results suggest that amidated forms of peptides are likely to possess higher membrane binding affinity due to the increased charge.     

Electric field increases the phase transition temperature in the bilayer membrane of phosphatidic acid

The effect of the electric field on the phase transition temperature (Tc) of acidic 1,2-dipalmitoyl-sn-glycero-3-phosphate (DPPA) and 1,2-dipalmitoyl-sn-glycero-3-thionphosphate (thion-DPPA) and zwitterion, i.e. 1,2-dipalmitoyl-rac-3-phosphocholine and 1,2-distearoyl-rac-glycero-3-phosphocholine (DPPC and DSPC), lipids has been investigated. The phase transition was detected using the jump-like increase effect in the conductance of the planar bilayer membrane. A voltage increase to 150 mV has been shown to increase the phase transition temperature in a bilayer lipid membrane (BLM) of phosphatidic acids (DPPA and thion-DPPA) by 8-12 degrees C while the transition temperature in the bilayer of zwitterion lipids (DPPC and DSPC) increases insignificantly. The increasing of Tt in BLM of acidic lipids is attributed to the voltage-induced changes in the molecule packing density.     

Effect of hydrophobic structure on the catalysis of nitric oxide release from zwitterionic diazeniumdiolates in surfactant and liposome media.

The effect of phospholipid liposomes and surfactant micelles on the rate of nitric oxide release from zwitterionic diazeniumdiolates, R1R2N[N(O)NO]-, with significant hydrophobic structure, has been explored. The acid-catalyzed dissociation of NO has been examined in phosphate-buffered solutions of sodium dodecylsulfate (SDS) micelles and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-[phospho-(1-glycerol)] sodium salt (DPPG) phospholipid liposomes. The reaction behavior of dibenzylamine-, monobenzylamine-, and dibutylamine-derived substrates [1]: R1 = C6H5CH2, R2 = C6H5CH2 NH2+(CH2)2, 2: R1 = C6H5CH2, R2 = NH3+(CH2)2, and 3: R1 = n-butyl, R2 = n-butyl-NH2+(CH2)6] has been compared with that of SPER/NO, 4: R1 = H2N(CH2)3, R2 = H2N(CH2) 3NH2+(CH2)4]. Catalysis of NO release is observed in both micellar and liposome media. Hydrophobic interactions contribute to micellar binding for 1-3 and appear to be the main factor facilitating catalysis by charge neutral DPPC liposomes. Binding constants for the association of 1 and 3 with SDS micelles were 3-fold larger than those previously obtained with comparable zwitterionic substrates lacking their hydrophobic structure. Anionic DPPG liposomes were much more effective in catalyzing NO release than either DPPC liposomes or SDS micelles. DPPG liposomes (at 10 mM total lipid) induced a 30-fold increase in the NO dissociation rate of SPER/NO compared to 12- and 14-fold increases in that of 1 and 3.     

The cyclic antimicrobial peptide RTD-1 induces stabilized lipid-peptide domains more efficiently than its open-chain analogue

The effects of a mammalian cyclic antimicrobial peptide, rhesus theta defensin 1 (RTD-1) and its open chain analogue (oRTD-1), on the phase behaviour and structure of model membrane systems (dipalmitoyl phosphatidylcholine, DPPC and dipalmitoyl phosphatidylglycerol, DPPG) were studied. The increased selectivity of RTD-1 for anionic DPPG over zwitterionic DPPC was shown by differential scanning calorimetry. RTD-1, at a molar peptide-lipid ratio of 1:100, induced considerable changes in the phase behaviour of DPPG, but not of DPPC. The main transition temperature, Tm, was unchanged, but additional phase transitions appeared above Tm. oRTD-1 induced similar effects. However, the effects were not observable below a peptide:lipid molar ratio of 1:50, which correlates with the weaker biological activity of oRTD-1. Small- and wide-angle X-ray scattering revealed for DPPG the appearance of additional structural features induced by RTD-1 above Tm, which were interpreted as correlated lamellar structures, with increased order of the fatty acyl side chains of the lipid. It is proposed that after initial electrostatic interaction of the cationic rim of the peptide with the anionic DPPG headgroups, leading to stabilized lipid-peptide clusters, the hydrophobic face of the peptide assists in its interaction with the fatty acyl side chains eventually leading to membrane disruption.     

The surfactant peptide KL4 in lipid monolayers: phase behavior, topography, and chemical distribution

Studies of different fragments and mutants of SP-B suggest that the function related structural and compositional characteristics in SP-B are its positive charges with intermittent hydrophobic domains. KL4 ([lysine-(leucine)4]4-lysine) is a synthetic peptide based on SP-B structure and is the major constituent of Surfaxin, a potential therapeutic agent for respiratory distress syndrome in premature infants. There is, however, no clear understanding about the possible lipid-KL4 interactions behind its function, which is an inevitable knowledge to design improved therapeutic agents. To examine the phase behavior, topography, and lipid specificity of KL4/lipid systems, we aimed to study different surfactant model systems containing KL4, neutral dipalmitoylphosphatidylcholine (DPPC) and/or negatively charged dipalmitoylphosphatidylglycerol (DPPG) in the presence of Ca2+ ions. Surface pressure-area isotherms, fluorescence microscopic images, scanning force microscopy as well as time-of-flight secondary ion mass spectrometry suggest (i) that KL4 is not miscible with DPPC and therefore forms peptide aggregates in DPPC/KL4 mixtures; (ii) that KL4 specifically interacts with DPPG via electrostatic interactions and induces percolation of DPPG-rich phases; (iii) that existing DPPG-Ca2+ interactions are too strong to be overcome by KL4, the reason why the peptide remains excluded from condensed DPPG domains and passively colocalizes with DPPC in a demixed fluid phase; and (iv) that the presence of negatively charged lipid is necessary for the formation of bilayer protrusions. These results indicate that the capability of the peptide to induce the formation of a defined surface-confined reservoir depends on the lipid environment, especially on the presence of anionic lipids.     

Interaction of the cholesterol reducing agent simvastatin with zwitterionic DPPC and charged DPPG phospholipid membranes

Simvastatin is a lipid-lowering drug in the pharmaceutical group statins. Interaction of a drug with lipids may define its role in the system and be critical for its pharmacological activity. We examined the interactions of simvastatin with zwitterionic dipalmitoyl phosphatidylcholine (DPPC) and anionic dipalmitoyl phosphatidylglycerol (DPPG) multilamellar vesicles (MLVs) as a function of temperature at different simvastatin concentrations using Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). The FTIR results indicate that the effect of simvastatin on membrane structure and dynamics depends on the type of membrane lipids. In anionic DPPG MLVs, high simvastatin concentrations (12, 18, 24 mol%) change the position of the CH₂ antisymmetric stretching mode to lower wavenumber values, implying an ordering effect. However, in zwitterionic DPPC MLVs, high concentrations of simvastatin disorder systems both in the gel and liquid crystalline phases. Moreover, in DPPG and DPPC MLVs, simvastatin has opposite dual effects on membrane dynamics. The bandwidth of the CH₂ antisymmetric stretching modes increases in DPPG MLVs, implying an increase in the dynamics, whereas it decreases in DPPC MLVs. Simvastatin caused broadening of the phase transition peaks and formation of shoulders on the phase transition peaks in DSC curves, indicating multi-domain formations in the phospholipid membranes. Because physical features of membranes such as lipid order and fluidity may be changed with the bioactivity of drugs, opposing effects of simvastatin on the order and dynamics of neutral and charged phospholipids may be critical to deduce the action mechanism of the drug and estimate drug-membrane interactions.     

Effects of divalent cations on the ultrasonic absorption coefficient of negatively charged liposomes (LUV) near their phase transition temperature

The ultrasonic absorption coefficient per wavelength (alpha lambda), as a function of temperature and frequency, was determined for large unilamellar vesicles (LUV) in the vicinity of their phospholipid phase transition temperature, using a double crystal acoustic interferometer. (The vesicles were composed of a 4:1 (w/w) mixture of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG). It has been found that alpha lambda reaches a maximum (alpha lambda)max at the phase transition temperature (tm) of the phospholipids in the bilayer, at an ultrasonic relaxation frequency of 2.1 MHz. Divalent cations (Ca2+ and Mg2+), added to LUV suspensions, shifted (alpha lambda)max to higher temperatures, dependent upon the concentration of divalent cation. It was also found that the shape of the alpha lambda versus t curve was significantly changed, representing changes in the Van't Hoff enthalpy of the transition, and therefore, the cooperative unit of the transition. This suggests that divalent cations interact individually with the negatively charged phospholipid headgroups of DPPG and with DPPC headgroups, thus decreasing the cooperative unit of the transition. The observed upward shift in tm suggests an interaction that increases the activation energy and, therefore, the temperature of the phase transition. However, alpha lambda as a function of frequency did not change with the addition of divalent cations and, thus, the relaxation time of the event responsible for the absorption of ultrasound is not changed by the addition of divalent cations.     

Antimicrobial peptide-lipid binding interactions and binding selectivity

Surface pressure measurements, external reflection-Fourier transform infrared spectroscopy, and neutron reflectivity have been used to investigate the lipid-binding behavior of three antimicrobial peptides: melittin, magainin II, and cecropin P1. As expected, all three cationic peptides were shown to interact more strongly with the anionic lipid, 1,2 dihexadecanoyl-sn-glycerol-3-(phosphor-rac-(1-glycerol)) (DPPG), compared to the zwitterionic lipid, 1,2 dihexadecanoyl-sn-glycerol-3-phosphocholine (DPPC). All three peptides have been shown to penetrate DPPC lipid layers by surface pressure, and this was confirmed for the melittin-DPPC interaction by neutron reflectivity measurements. Adsorption of peptide was, however, minimal, with a maximum of 0.4 mg m(-2) seen for melittin adsorption compared to 2.1 mg m(-2) for adsorption to DPPG (from 0.7 microM solution). The mode of binding to DPPG was shown to depend on the distribution of basic residues within the peptide alpha-helix, although in all cases adsorption below the lipid layer was shown to dominate over insertion within the layer. Melittin adsorption to DPPG altered the lipid layer structure observed through changes in the external reflection-Fourier transform infrared lipid spectra and neutron reflectivity. This lipid disruption was not observed for magainin or cecropin. In addition, melittin binding to both lipids was shown to be 50% greater than for either magainin or cecropin. Adsorption to the bare air-water interface was also investigated and surface activity followed the trend melittin>magainin>cecropin. External reflection-Fourier transform infrared amide spectra revealed that melittin adopted a helical structure only in the presence of lipid, whereas magainin and cecropin adopted helical structure also at an air-water interface. This behavior has been related to the different charge distributions on the peptide amino acid sequences.     

Interaction of antimycobacterial and anti-pneumocystis drugs with phospholipid membranes

Liposomes can be used as carriers of drugs in the treatment of viral, bacterial and protozoal infections. The potential for liposome-mediated therapy of Mycobacterium avium-intracellulare complex infections, one of the most common opportunistic infections in AIDS, is currently under study. Here, we have investigated the effect of the lipid-soluble antimycobacterial drugs ansamycin, clofazimine and CGP7040 on the thermotropic behavior of liposomes composed of dipalmitoylphosphatidylcholine (DPPC) or dipalmitoylphosphatidylglycerol (DPPG) using differential scanning calorimetry (DSC). In the presence of ansamycin (rifabutine), the peak gel-liquid crystalline phase transition temperature (Tm) of DPPG was reduced, as was the sub-transition temperature (Ts), whereas the Tm of DPPC was reduced only slightly. The temperature of the pre-transition (Tp) of DPPC was lowered, while the pre-transition of DPPG was abolished. Ansamycin also caused the broadening of the transition endotherm of both lipids. Equilibration of the drug/lipid complex for 1 or 5 days produced different thermotropic behavior. In the presence of clofazimine, the cooperativity of the phase transition of DPPG decreased. Above 10 mol% clofazimine formed two complexes with DPPG, as indicated by two distinguishable peaks in DSC thermograms. The Tm of both peaks were lowered as the mole fraction increased. Clofazimine had minimal interaction with DPPC. In contrast, CGP7040 interacted more effectively with DPPC than with DPPG, causing a reduction of the size of the cooperative unit of DPPC even at 2 mol%. The main transition of DPPC split into 3 peaks at 5 mol% drug. The pre-transition was abolished at all drug concentrations and the sub-transition disappeared at 10 mol% CGP7040. These studies suggest that maximal encapsulation of clofazimine in liposomes would require a highly negatively charged membrane, while that of CGP7040 would necessitate a zwitterionic membrane. We have also investigated the interaction of the water-soluble antibiotic pentamidine, which has been used against Pneumocystis carinii, the most lethal of AIDS-related opportunistic pathogens. Aerosol administration of this drug leads to long-term sequestration of the drug in the lungs. The DPPG/pentamidine complex exhibited a pre-transition at 3.5 degrees C, an endothermic peak at 42 degrees C, and an exothermic peak at 44.5 degrees C, followed by another endothermic peak at 55 degrees C. The exotherm depended on the history of the sample, requiring pre-incubation for several minutes below the 42 degrees C transition. These observations suggest that upon melting of the DPPG chains at 42 degrees C, the DPPG crystallizes as a DPPG/pentamidine complex that melts at 55 degrees C.     

The effects of radioprotectant and potential antioxidant agent amifostine on the structure and dynamics of DPPC and DPPG liposomes

Agents capable of scavenging ROS have attracted attention recently because of their potential use as antioxidative agents. Amifostine, a ROS scavenger, has the potential to be used as an antioxidant in therapeutic applications. In this study, the effect of amifostine on neutral zwitterionic dipalmitoylphosphatidylcholine (DPPC) and anionic dipalmitoylphosphatidylglycerol (DPPG) model membranes' structure and dynamics is aimed to be examined by Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). Our results revealed that amifostine at concentrations used (1–24 mol%) does not induce any important alteration in the shape of phase transition curve and phase transition temperature in the DPPC and DPPG membranes. High concentrations of amifostine slightly increased the acyl chain flexibility of DPPC membranes in the liquid crystalline phase and DPPG membranes in the gel phase. A lessening in the dynamics of DPPC liposomes was observed for all concentrations of amifostine in both phases but slight dual effect was observed only in the gel phase as a decrease in dynamics at low concentrations and an increase at higher concentrations of amifostine in DPPG liposomes. Additionally, strong hydrogen bonding was observed for both CO and PO₂⁻ groups in case of DPPC and for PO₂⁻ groups in case of DPPG. Dehydration around the CO regions occurred in case of DPPG. Accordingly, amifostine is proposed to be interacting strongly with zwitterionic and negatively charged membrane head groups and glycerol backbone in all concentrations and because of this interaction it causes some changes in lipid order and dynamics especially at high concentrations.     

The characteristics of the radiation-initiated peroxidation of the phosphatidylcholine making up liposomes containing phospholipids which are susceptible to free-radical fragmentation

The regularities of accumulation of conjugated dienes and thiobarbituric acid (TBA)-reactive substances under gamma-irradiation of liposomes from rat liver phosphatidylcholine (PC) and its mixtures with the resistant to lipid peroxidation saturated phospholipids and bovine brain sphingomyelin (SM) were studied. It was established that the incorporation of negatively charged dipalmitoylphosphatidylglycerol (DPPG) and dipalmitoylphosphatidylethanol (DPPET) into lipid bilayer resulted in the increase of primary and secondary products of LPO, whereas neutral dipalmitoylphosphatidylcholine (DPPC) and SM involving in the phospholipid mixtures inhibited the peroxidation of PC. For anionic phospholipids, DPPG had more profound activating action on LPO, amongst the neutral phospholipids SM was more potent inhibitor of the reaction. Unlike DPPET and DPPC, DPPG and SM were subjected to free radical fragmentation on gamma-radiation. It is suggested that the intermediates and products of free radical fragmentation may modulate the progress of LPO.     

Comparative study of the gel phases of ether- and ester-linked phosphatidylcholines

Calorimetric, X-ray diffraction, and 31P nuclear magnetic resonance (NMR) studies of aqueous dispersions of 1,2-dihexadecyl-sn-glycero-3-phosphocholine (DHPC) gel phases at low temperatures (-60 to 22 degrees C) show thermal, structural, and dynamic differences when compared to aqueous dispersions of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) gel phases at corresponding temperatures. Differential scanning calorimetry of DHPC dispersions demonstrates a reversible, low-enthalpy "subtransition" at 4 degrees C in contrast to the conditionally reversible, high-enthalpy subtransition observed at 17 degrees C for annealed DPPC bilayers. X-ray diffraction studies indicate that DHPC dispersions form a lamellar gel phase with dav congruent to 46 A both above and below the "subtransition". It is suggested that the reduced dav observed for DHPC (46 A as compared to 64 A in DPPC) is due to an interdigitated lamellar gel phase which exists at all temperatures below the pretransition at 35 degrees C. 31P NMR spectra of DHPC gel-phase bilayers show an axially symmetric chemical shift anisotropy powder pattern which remains sharp down to -20 degrees C, suggesting the presence of fast axial diffusion. In contrast, 31P spectra of DPPC bilayers indicate this type of motion is frozen out at approximately 0 degrees C.     

The phase behavior of mixed aqueous dispersions of dipalmitoyl derivatives of phosphatidylcholine and diacylglycerol.

The phases and transition sequences for aqueous dispersions of mixtures of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycerol (1,2-DPG) have been studied by differential scanning calorimetry, dynamic x-ray diffraction, freeze-fracture electron microscopy, 31P-nuclear magnetic resonance spectroscopy, and Fourier-transform infrared spectroscopy. The results have been used to construct a dynamic phase diagram of the binary mixture as a function of temperature over the range 20 degrees-90 degrees C. It is concluded that DPPC and 1,2-DPG form two complexes in the gel phase, the first one with a DPPC/1,2-DPG molar ratio of 55:45 and the second one at a molar ratio of approximately 1:2, defining three different regions in the phase diagram. Two eutectic points are postulated to occur: one at a very low 1,2-DPG concentration and the other at a 1,2-DPG concentration slightly higher than 66 mol%. At temperatures higher than the transition temperature, lamellar phases were predominant at low 1,2-DPG concentrations, but nonlamellar phases were found to be predominant at high proportions of 1,2-DPG. A very important aspect of these DPPC/1,2-DPG mixtures was that, in the gel phase, they showed a ripple structure, as seen by freeze-fracture electron microscopy and consistent with the high lamellar repeat spacings seen by x-ray diffraction. Ripple phase characteristics were also found in the fluid lamellar phases occurring at concentrations up to 35.6 mol% of 1,2-DPG. Evidence was obtained by Fourier transform infrared spectroscopy of the dehydration of the lipid-water interface induced by the presence of 1,2-DPG. The biological significance of the presence of diacylglycerol in membrane lipid domains is discussed.     

Interaction of poly(l-lysines) with negatively charged membranes: an FT-IR and DSC study

The influence of the binding of poly(l-lysine) (PLL) to negatively charged membranes containing phosphatidylglycerols (PG) was studied by DSC and FT-IR spectroscopy. We found a general increase in the main transition temperature as well as increase in hydrophobic order of the membrane upon PLL binding. Furthermore we observed stronger binding of hydration water to the lipid head groups after PLL binding. The secondary structure of the PLL after binding was studied by FT-IR spectroscopy. We found that PLL binds in an α-helical conformation to negatively charged DPPG membranes or membranes with DPPG-rich domains. Moreover we proved that PLL binding induces domain formation in the gel state of mixed DPPC/DPPG or DMPC/DPPG membranes as well as lipid remixing in the liquid–crystalline state. We studied these effects as a function of PLL chain length and found a significant dependence of the secondary structure, phase transition temperature and domain formation capacity on PLL chain length and also a correlation between the peptide secondary structure and the phase transition temperature of the membrane. We present a system in which the membrane phase transition triggers a highly cooperative secondary structure transition of the membrane-bound peptide from α-helix to random coil. Dedicated to Prof. K. Arnold on the occasion of his 65th birthday.