Thermotropic phase behaviour of alpha-dipalmitoylphosphatidylcholine multibilayers is influenced to various extents by carotenoids containing different structural features--evidence from differential scanning calorimetry

Carotenoids are the effective modulators of physical properties of model and natural membranes. To demonstrate the relationship between the structure of carotenoids and their effect on the molecular dynamics of membranes, we have investigated the influence of five structurally different carotenoids: beta-carotene, lycopene, lutein, violaxanthin, zeaxanthin and additionally carotane--a fully saturated derivative of beta-carotene, on thermotropic phase behaviour of dipalmitoylphosphatidylcholine (DPPC) multilamellar vesicles by means of differential scanning calorimetry (DSC). The results obtained indicate that the carotenoids used modulated the thermotropic properties of multibilayers to various extents, broadening the pretransition and the main phase transition peaks and shifting them to lower temperatures. Pronounced decrease of pretransition enthalpy (DeltaH(p)) proves that carotenoids very strongly alter the membrane properties in its gel phase. Comparison of the influence of several carotenoids shows that a rigid, polyisoprenoid chain plays a basic role in altering the thermotropic properties of such membranes and the presence of rings without oxygen-containing groups has a minor significance for the observed interactions. Carotenoids containing epoxy and/or hydroxy groups attached to their rings modify the thermotropic phase behaviour of DPPC multilamellar vesicles stronger than carotenes--a result of their orientation in the DPPC bilayer.     

Molecular characteristics of astaxanthin and beta-carotene in the phospholipid monolayer and their distributions in the phospholipid bilayer.

The molecular characteristics of the monolayers of astaxanthin with polar group on the beta-ionone ring in the molecule and beta-carotene without polar group and their interactions in mixed carotenoid-phospholipid monolayers and the effects of carotenoids on the phase behavior of the phospholipid bilayers were examined by the monolayer technique and differential scanning calorimetry (DSC). We found from the monolayer study that beta-carotene had an amphiphilic nature. The molecular assembly of astaxanthin in the monolayer at the hydrophobic/hydrophilic interface was more stable than that of beta-carotene. Dimyristoylphosphatidylcholine (DMPC) in the monolayer was miscible with astaxanthin in the range of 0-0.4 mol fractions of astaxanthin, but not fully miscible with beta-carotene even at low concentrations below 0.1 mol fraction of beta-carotene. Surface potential and compression/expansion cycles of beta-carotene monolayer indicated the formation of molecular aggregates by itself. DSC study showed that when small amount of astaxanthin was added, the transition temperature of dipalmitoylphosphatidylcholine (DPPC) was markedly shifted to lower temperatures and that the transition peak was asymmetrically broadened, indicative of a significant depression in cooperativity of the gel to liquid-crystalline transition. The asymmetric DSC endothermic bands of DPPC incorporating small amounts of astaxanthin were well fit by deconvolution into two to three domains containing different concentrations of astaxanthin. On the contrary, the incorporation of beta-carotene resulted in a small depression of the main transition temperature with a slight broadening of the transition peak, suggesting a small miscibility of beta-carotene with the phospholipid bilayer or a formation of aggregates of beta-carotene in the membranes. These results suggest that there would be a high localized concentration in the phase separated membrane for astaxanthin or beta-carotene to function effectively as scavenger.     

Thermotropic phase behaviour of α-dipalmitoylphosphatidylcholine multibilayers is influenced to various extents by carotenoids containing different structural features- evidence from differential scanning calorimetry

Carotenoids are the effective modulators of physical properties of model and natural membranes. To demonstrate the relationship between the structure of carotenoids and their effect on the molecular dynamics of membranes, we have investigated the influence of five structurally different carotenoids: β-carotene, lycopene, lutein, violaxanthin, zeaxanthin and additionally carotane- a fully saturated derivative of β-carotene, on thermotropic phase behaviour of dipalmitoylphosphatidylcholine (DPPC) multilamellar vesicles by means of differential scanning calorimetry (DSC). The results obtained indicate that the carotenoids used modulated the thermotropic properties of multibilayers to various extents, broadening the pretransition and the main phase transition peaks and shifting them to lower temperatures. Pronounced decrease of pretransition enthalpy (ΔH_p) proves that carotenoids very strongly alter the membrane properties in its gel phase. Comparison of the influence of several carotenoids shows that a rigid, polyisoprenoid chain plays a basic role in altering the thermotropic properties of such membranes and the presence of rings without oxygen-containing groups has a minor significance for the observed interactions. Carotenoids containing epoxy and/or hydroxy groups attached to their rings modify the thermotropic phase behaviour of DPPC multilamellar vesicles stronger than carotenes- a result of their orientation in the DPPC bilayer.     

Interactions of the AT1 antagonist valsartan with dipalmitoyl-phosphatidylcholine bilayers

Valsartan is a marketed drug with high affinity to the type 1 angiotensin (AT1) receptor. It has been reported that AT1 antagonists may reach the receptor site by diffusion through the plasma membrane. For this reason we have applied a combination of differential scanning calorimetry (DSC), Raman spectroscopy and small and wide angle X-ray scattering (SAXS and WAXS) to investigate the interactions of valsartan with the model membrane of dipalmitoyl-phosphatidylcholine (DPPC). Hence, the thermal, dynamic and structural effects in bulk as well as local dynamic properties in the bilayers were studied with different valsartan concentrations ranging from 0 to 20 mol%. The DSC experimental results showed that valsartan causes a lowering and broadening of the phase transition. A splitting of the main transition is observed at high drug concentrations. In addition, valsartan causes an increase in enthalpy change of the main transition, which can be related to the induction of interdigitation of the lipid bilayers in the gel phase. Raman spectroscopy revealed distinct interactions between valsartan with the lipid interface localizing it in the polar head group region and in the upper part of the hydrophobic core. This localization of the drug molecule in the lipid bilayers supports the interdigitation view. SAXS measurements confirm a monotonous bilayer thinning in the fluid phase, associated with a steady increase of the root mean square fluctuation of the bilayers as the valsartan concentration is increased. At high drug concentrations these fluctuations are mainly governed by the electrostatic repulsion of neighboring membranes. Finally, valsartans' complex thermal and structural effects on DPPC bilayers are illustrated and discussed on a molecular level.     

Effects of non-steroid anti-inflammatory drugs in membrane bilayers

The thermal effects of non-steroidal anti-inflammatory drugs (NSAIDs) meloxicam, tenoxicam, piroxicam and lornoxicam have been studied in dipalmitoylphosphatidylcholine (DPPC) membrane bilayers using neutral and acidic environments (pH 2.5). The strength of the perturbing effect of the drugs is summarized to a lowering of the main phase transition temperature and a broadening of the phase transition temperature as well as broadening or abolishment of the pretransition of DPPC bilayers. The thermal profiles in the two environments were very similar. Among the NSAIDs studied meloxicam showed the least perturbing effect. The differential scanning calorimetry results (DSC) in combination with molecular modeling studies point out that NSAIDs are characterized by amphoteric interactions and are extended between the polar and hydrophobic segments of lipid bilayers. The effects of NSAIDs in membrane bilayers were also investigated using Raman spectroscopy. Meloxicam showed a gauche:trans profile similar to DPPC bilayers while the other NSAIDs increased significantly the gauche:trans ratio. In conclusion, both techniques show that in spite of the close structural similarity of the NSAIDs studied, meloxicam appears to have the lowest membrane perturbing effects probably attributed to its highest lipophilicity.     

Competitive carotenoid and cholesterol incorporation into liposomes: effects on membrane phase transition, fluidity, polarity and anisotropy

Pure 1,2-dipalmitoyl-sn-glycero-3-phosphorylcholine (DPPC) or mixed DPPC:1,2-dipalmitoyl phosphatidyletanolamine (DPPE):1,2-dipalmitoyl diphosphatidylserine (DPPS) (17:5:3) liposomes were incorporated with 5 mol% dietary carotenoids (beta-carotene, lutein and zeaxanthin) or with cholesterol (16 and 48 mol%) in the absence or presence of 15 mol% carotenoids, respectively. The carotenoid incorporation yields ranged from 0.42 in pure to 0.72 in mixed phospholipid liposomes. They decreased significantly, from 3 to 14%, in the corresponding cholesterol-doped liposomes, respectively. Highest incorporation yields were achieved by zeaxanthin and lutein in phospholipid liposomes while in cholesterol-containing liposomes, lutein was highest incorporated. The effects on membrane structure and dynamics were determined by differential scanning calorimetry, steady-state fluorescence and anisotropy measurements. Polar carotenoids and cholesterol cause similar, dose-dependent effects: ordering and rigidification revealed by broadening of the transition peak, and increase of anisotropy. Membrane hydrophobicity is determined by cholesterol content and carotenoid polarity. In cholesterol-doped liposomes, beta-carotene is less incorporated than in cholesterol-free liposomes. Our observations suggest effects of carotenoids, even at much lower effective concentrations than cholesterol (8 to 80-fold), on membrane structure and dynamics. Although they are minor constituents of animal membranes, carotenoids may act as modulators of membrane phase transition, fluidity, polarity and permeability, and therefore, can influence the membrane physiology and pathology.     

Effect of lipid composition on the "membrane response" induced by a fusion peptide

To understand the initial stages of membrane destabilization induced by viral proteins, the factors important for binding of fusion peptides to cell membranes must be identified. In this study, effects of lipid composition on the mode of peptides' binding to membranes are explored via molecular dynamics (MD) simulations of the peptide E5, a water-soluble analogue of influenza hemagglutinin fusion peptide, in two full-atom hydrated lipid bilayers composed of dimyristoyl- and dipalmitoylphosphatidylcholine (DMPC and DPPC, respectively). The results show that, although the peptide has a common folding motif in both systems, it possesses different modes of binding. The peptide inserts obliquely into the DMPC membrane mainly with its N-terminal alpha helix, while in DPPC, the helix lies on the lipid/water interface, almost parallel to the membrane surface. The peptide seriously affects structural and dynamical parameters of surrounding lipids. Thus, it induces local thinning of both bilayers and disordering of acyl chains of lipids in close proximity to the binding site. The "membrane response" significantly depends upon lipid composition: distortions of DMPC bilayer are more pronounced than those in DPPC. Implications of the observed effects to molecular events on initial stages of membrane destabilization induced by fusion peptides are discussed.     

Membrane topology of a 14-mer model amphipathic peptide: a solid-state NMR spectroscopy study

We have investigated the interaction between a synthetic amphipathic 14-mer peptide and model membranes by solid-state NMR. The 14-mer peptide is composed of leucines and phenylalanines modified by the addition of crown ethers and forms a helical amphipathic structure in solution and bound to lipid membranes. To shed light on its membrane topology, 31P, 2H, 15N solid-state NMR experiments have been performed on the 14-mer peptide in interaction with mechanically oriented bilayers of dilauroylphosphatidylcholine (DLPC), dimyristoylphosphatidylcholine (DMPC), and dipalmitoylphosphatidylcholine (DPPC). The 31P, 2H, and 15N NMR results indicate that the 14-mer peptide remains at the surface of the DLPC, DMPC, and DPPC bilayers stacked between glass plates and perturbs the lipid orientation relative to the magnetic field direction. Its membrane topology is similar in DLPC and DMPC bilayers, whereas the peptide seems to be more deeply inserted in DPPC bilayers, as revealed by the greater orientational and motional disorder of the DPPC lipid headgroup and acyl chains. 15N{31P} rotational echo double resonance experiments have also been used to measure the intermolecular dipole-dipole interaction between the 14-mer peptide and the phospholipid headgroup of DMPC multilamellar vesicles, and the results indicate that the 14-mer peptide is in contact with the polar region of the DMPC lipids. On the basis of these studies, the mechanism of membrane perturbation of the 14-mer peptide is associated to the induction of a positive curvature strain induced by the peptide lying on the bilayer surface and seems to be independent of the bilayer hydrophobic thickness.     

Enhancing the carotenoid content of <Emphasis Type="Italic">Brassica napus</Emphasis> seeds by downregulating lycopene epsilon cyclase

The accumulation of carotenoids in higher plants is regulated by the environment, tissue type and developmental stage. In Brassica napus leaves, beta-carotene and lutein were the main carotenoids present while petals primarily accumulated lutein and violaxanthin. Carotenoid accumulation in seeds was developmentally regulated with the highest levels detected at 35-40 days post anthesis. The carotenoid biosynthesis pathway branches after the formation of lycopene. One branch forms carotenoids with two beta rings such as beta-carotene, zeaxanthin and violaxanthin, while the other introduces both beta- and epsilon-rings in lycopene to form alpha-carotene and lutein. By reducing the expression of lycopene epsilon-cyclase (epsilon-CYC) using RNAi, we investigated altering carotenoid accumulation in seeds of B. napus. Transgenic seeds expressing this construct had increased levels of beta-carotene, zeaxanthin, violaxanthin and, unexpectedly, lutein. The higher total carotenoid content resulting from reduction of epsilon-CYC expression in seeds suggests that this gene is a rate-limiting step in the carotenoid biosynthesis pathway. epsilon-CYC activity and carotenoid production may also be related to fatty acid biosynthesis in seeds as transgenic seeds showed an overall decrease in total fatty acid content and minor changes in the proportions of various fatty acids.     

Insight into the structural role of carotenoids in the photosystem I: a quantum chemical analysis

The structural stabilization role of carotenoids in the formation of photosynthetic pigment-protein complexes is investigated theoretically. The pi-pi stacking and CH-pi interactions between beta-carotenes and their surrounding chlorophylls (and/or aromatic residues) in Photosystem I (PS1) from the cyanobacterium Synechococcus elongatus were studied by means of the supermolecular approach at the level of the second-order M?ller-Plesset perturbation method. PS1 features a core integral antenna system consisting of 22 beta-carotenes intertwined with 90 chlorophyll molecules. The binding environments of all 22 beta-carotenes were systematically analyzed. For 21 out of the 22 cases, one or more chlorophyll molecules exist within van der Waals' contacts of the beta-carotene molecule. The calculated strengths of pi-pi stacking interactions between the conjugated core of beta-carotene and the aromatic tetrapyrrole rings of chlorophyll are substantial, ranging from -3.54 kcal/mol for the perpendicular-positioned BCR4004...CHL1217 pair to -16.01 kcal/mol for the parallel-oriented BCR4007...CHL1122 pair. A strong dependence of the pi-pi stacking interaction energies on the intermolecular configurations of the two interacting pi-planes is observed. The parallel-oriented beta-carotene and chlorophyll pair is energetically much more stable than the perpendicular-positioned pair. The larger the extent of pi-pi overlapping, the stronger the interaction strength. In many cases, the beta-ring ends of beta-carotene molecules are found to interact with the tetrapyrrole rings of chlorophyll via CH-pi interactions. For the latter interactions, the calculated interaction strengths vary from -7.03 to -11.03 kcal/mol, depending on the intermolecular configuration. This work leads to the conclusion that pi-pi stacking and CH-pi interactions between beta-carotene and their surrounding chlorophylls and aromatic residues play an essential role in binding beta-carotenes in PS1 from S. elongatus. Consequently, the molecular basis of the structural stabilization function of carotenoids in formation of the photosynthetic pigment-protein complexes is established.     

Thermodynamic characterization of interactions between ornithine transcarbamylase leader peptide and phospholipid bilayer membranes

The interactions of the targeting sequence of the mitochondrial enzyme ornithine transcarbamylase with phospholipid bilayers of different molecular compositions have been studied by high-sensitivity heating and cooling differential scanning calorimetry, high-sensitivity isothermal titration calorimetry, fluorescence spectroscopy, and electron microscopy. These studies indicate that the leader peptide interacts strongly with dipalmitoylphosphatidylcholine (DPPC) bilayer membranes containing small mole percents of the anionic phospholipids dipalmitoylphosphatidylglycerol (DPPG) or brain phosphatidylserine (brain PS) but not with pure phosphatidylcholines. For the first time, the energetics of the leader peptide-membrane interaction have been measured directly by using calorimetric techniques. At 20 degrees C, the association of the peptide with the membrane is exothermic and characterized by an association constant of 2.3 X 10(6) M-1 in the case of phosphatidylglycerol-containing and 0.35 X 10(6) M-1 in the case of phosphatidylserine-containing phospholipid bilayers. In both cases, the enthalpy of association is -60 kcal/mol of peptide. Additional experiments using fluorescence techniques suggest that the peptide does not penetrate deeply into the hydrophobic core of the membrane. The addition of the leader peptide to DPPC/DPPG (5:1) or DPPC/brain PS (5:1) small sonicated vesicles results in vesicle fusion. The fusion process is dependent on peptide concentration and is maximal at the phase transition temperature of the vesicles and minimal at temperatures below the phase transition.     

Distribution and favorable binding sites of pyrroloquinoline and its analogues in a lipid bilayer studied by molecular dynamics simulations

The distribution of 1H-pyrrolo[3,2-h]quinoline (PQ), 11H-dipyrido[2,3-a]carbazole (PC) and 7-azaindole (7AI) at a water/membrane interface has been investigated by molecular dynamics (MD) simulations. The MD study focused on favorable binding sites of the azaaromatic probes across a dipalmitoylphosphatidylcholine (DPPC) bilayer. Our simulations show that PQ and PC are preferably accommodated at the hydrocarbon core of the bilayer below the glycerol moiety. In addition, it is found that the hydrophobic aromatic parts of the probes are located inside a more ordered region of DPPC, consisting of hydrophobic lipid chains. In contrast to PQ and PC, 7AI is characterized by a broad distribution between a DPPC interface and water, so that the three preferable binding sites are found across a water/membrane interface. It is found that in the sequence 7AI-PQ-PC, due to the increase of the number of aromatic rings and, hence, the hydrophobic character of the probes, the depth of the probe localization is gradually shifted deeper inside the hydrocarbon core of the bilayer. We found that the probe-lipid hydrogen-bonding contributes weakly to the favorable localizations of the azaaromatic probes inside the DPPC bilayer, so that the probe localization is mainly driven by electrostatic dipole-dipole and van der Waals interactions.     

The interfacial region of dipalmitoylphosphatidylcholine bilayers is perturbed by fusogenic amphipaths.

Several structural methods were used to probe the influence of three fusogenic and four nonfusogenic amphipaths on large, unilamellar dipalmitoylphosphatidylcholine (DPPC) vesicles. For four of these structural measurements there was a correlation observed between the ability of an amphipath to favor poly(ethylene glycol) (PEG)-induced fusion and the structural perturbation reported by each method. First, the fluorescence anisotropy of 1-[4-(trimethylamino)phenyl]-6-phenyhexa-1,3,5-triene (TMA-DPH), which probes the upper region of the bilayer, decreased in the range of PEG concentrations previously found to cause fusion of membranes containing fusogenic amphipaths. For nonfusogenic amphipaths, the anisotropy increased monotonically with PEG concentration. The properties of similar probes that locate in the hydrophobic core of the bilayer showed no correlation with fusogenicity, nor did the properties of probes purported to sense the aqueous surface of the membrane. Second, the frequency of the C=O stretch increased and then decreased dramatically as fusogenic but not nonfusogenic membranes were heated through their phase transition. Third, there was a dramatic increase in the frequency of the C-O-C ester stretch at the membrane order/disorder phase transition for membranes containing fusogenic amphipaths, twice the increase observed for nonfusogenic amphipaths. The spectral characteristics of phosphate, choline, and acyl chain motions showed no such correlation with fusogenicity. Finally, calorimetric measurements showed that low levels of fusogenic amphipaths eliminated the "pretransition" (L beta-->P beta) in DPPC membranes, whereas other amphipaths shifted but did not eliminate this transition. Taken together, these results indicate that fusogenic amphipaths perturb the interface or "backbone" region of the bilayer rather than the hydrophobic core, the headgroup, or the water interface regions of DPPC bilayers.     

HgCl2 disrupts the structure of the human erythrocyte membrane and model phospholipid bilayers

The structural effects of Hg(II) ions on the erythrocyte membrane were studied through the interactions of HgCl2 with human erythrocytes and their isolated resealed membranes. Studies were carried out by scanning electron microscopy and fluorescence spectroscopy, respectively. Hg(II) induced shape changes in erythrocytes, which took the form of echinocytes and stomatocytes. This finding means that Hg(II) locates in both the outer and inner monolayers of the erythrocyte membrane. Fluorescence spectroscopy results indicate strong interactions of Hg(II) ions with phospholipid amino groups, which also affected the packing of the lipid acyl chains at the deep hydrophobic core of the membrane. HgCl2 also interacted with bilayers of dimyristoylphosphatidylcholine and dimyristoylphosphatidylethanolamine, representative of phospholipid classes located in the outer and inner monolayers of the erythrocyte membrane, respectively. X-ray diffraction indicated that Hg(II) ions induced molecular disorder to both phospholipid bilayers, while fluorescence spectroscopy of dimyristoylphosphatidylcholine large unilamellar vesicles confirmed the interaction of Hg(II) ions with the lipid polar head groups. All these findings point to the important role of the phospholipid bilayers in the interaction of Hg(II) on cell membranes.     

Variations in the carotenoid content of Chlamydomonas reinhardii throughout the cell cycle.

Synchronous cultures of Chlamydomonas reinhardii have been examined for the total amounts of carotenoid and chlorophyll present throughout a 12 hrs light -- 4 hrs dark life cycle. Variations in the carotenoid distribution at different points within the cell cycle have been found. During the greater part of the light period all major carotenoids increased at a proportionally similar rate. However, the increases in lutein and violaxanthin preceded those in beta-carotene and neoxanthin by some 2 hrs and that in loroxanthin, and algal xanthophyll, by abour 3 hrs. A marked drop in total carotenoid accumulation, corresponding to similar temporary falling away in the accumulation of beta-carotene, lutein and violaxanthin occurred at 9 hrs. The correspondence of this with the established drop in RNA accumulation and the break-up of the nucleolus was pointed out. Considerable redistribution among the carotenoids occurred during the dark period, notably the amount of beta-carotene increased relative to the total xanthophylls. The full significance of these results can not be estimated in the absence of comparative data on related organisms.     

Lipid phase separations induced by the association of cholera toxin to phospholipid membranes containing ganglioside GM1

The interactions of cholera toxin and their isolated binding and active subunits with phospholipid bilayers containing the toxin receptor ganglioside GM1 have been studied by using high-sensitivity differential scanning calorimetry and steady-state and time-resolved fluorescence and phosphorescence spectroscopy. The results of this investigation indicate that cholera toxin associates with phospholipid bilayers containing ganglioside GM1, independent of the physical state of the membrane. In the absence of Ca2+, calorimetric scans of intact cholera toxin bound to dipalmitoylphosphatidylcholine (DPPC) large unilamellar vesicles containing ganglioside GM1 result in a broadening of the lipid phase transition peak and a slight decrease (less than 5%) in the transition enthalpy. In the presence of Ca2+ concentrations sufficient to cause ganglioside phase separation, the association of the intact toxin to the membrane results in a significant decrease of enthalpy change for the lipid transition, indicating that under these conditions the toxin molecule perturbs the hydrophobic core of the bilayer. Calorimetric scans using isolated binding subunits lacking the hydrophobic toxic subunit did not exhibit a decrease in the phospholipid transition enthalpy even in the presence of Ca2+, indicating that the binding subunits per se do not perturb the hydrophobic core of the bilayer. On the other hand, the hydrophobic A1 subunit by itself was able to reduce the phospholipid transition enthalpy when reconstituted into DPPC vesicles. These calorimetric observations were confirmed by fluorescence experiments using pyrene phospholipids.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modelling the phase equilibria in two-component membranes of phospholipids with different acyl-chain lengths

A phenomenological model is proposed to describe the membrane phase equilibria in binary mixtures of saturated phospholipids with different acyl-chain lengths. The model is formulated in terms of thermodynamic and thermomechanic properties of the pure lipid bilayers, specifically the chain-melting transition temperature and enthalpy, the hydrophobic bilayer thickness, and the lateral area compressibility modulus. The model is studied using a regular solution theory made up of a set of interaction parameters which directly identify that part of the lipid-lipid interaction which is due to hydrophobic mismatch of saturated chains of different lengths. It is then found that there is effectively a single universal interaction parameter which, in the full composition range, describes the phase equilibria in mixtures of DMPC/DPPC, DPPC/DSPC, DMPC/DSPC, and DLPC/DSPC, in excellent agreement with experimental measurements. The model is used to predict the variation with temperature and composition of the specific heat, as well as of the average membrane thickness and area in each of the phases. Given the value of the universal interaction parameter, the model is then used to predict the phase diagrams of binary mixtures of phospholipids with different polar head groups, e.g., DPPC/DPPE, DMPC/DPPE and DMPE/DSPC. By comparison with experimental results for these mixtures, it is shown that difference in acyl-chain lengths gives the major contribution to deviation from ideal mixing. Application of the model to mixtures with non-saturated lipids is also discussed.     

Investigation of temperature-induced phase transitions in DOPC and DPPC phospholipid bilayers using temperature-controlled scanning force microscopy

Under physiological conditions, multicomponent biological membranes undergo structural changes which help define how the membrane functions. An understanding of biomembrane structure-function relations can be based on knowledge of the physical and chemical properties of pure phospholipid bilayers. Here, we have investigated phase transitions in dipalmitoylphosphatidylcholine (DPPC) and dioleoylphosphatidylcholine (DOPC) bilayers. We demonstrated the existence of several phase transitions in DPPC and DOPC mica-supported bilayers by both atomic force microscopy imaging and force measurements. Supported DPPC bilayers show a broad L(beta)-L(alpha) transition. In addition to the main transition we observed structural changes both above and below main transition temperature, which include increase in bilayer coverage and changes in bilayer height. Force measurements provide valuable information on bilayer thickness and phase transitions and are in good agreement with atomic force microscopy imaging data. A De Gennes model was used to characterize the repulsive steric forces as the origin of supported bilayer elastic properties. Both electrostatic and steric forces contribute to the repulsive part of the force plot.     

Efficient radical trapping at the surface and inside the phospholipid membrane is responsible for highly potent antiperoxidative activity of the carotenoid astaxanthin

The effects of the carotenoids beta-carotene and astaxanthin on the peroxidation of liposomes induced by ADP and Fe(2+) were examined. Both compounds inhibited production of lipid peroxides, astaxanthin being about 2-fold more effective than beta-carotene. The difference in the modes of destruction of the conjugated polyene chain between beta-carotene and astaxanthin suggested that the conjugated polyene moiety and terminal ring moieties of the more potent astaxanthin trapped radicals in the membrane and both at the membrane surface and in the membrane, respectively, whereas only the conjugated polyene chain of beta-carotene was responsible for radical trapping near the membrane surface and in the interior of the membrane. The efficient antioxidant activity of astaxanthin is suggested to be due to the unique structure of the terminal ring moiety.     

Interactions of the dipeptide paralysin beta-Ala-Tyr and the aminoacid Glu with phospholipid bilayers

Existing evidence points out that the biological activity of beta-Ala-Tyr may in part related to its interactions with the cell membranes. For comparative reasons the effects of Glu were also examined using identical techniques and conditions. In order to examine their thermal and dynamic effects on membrane bilayers a combination of DSC, Raman and solid state NMR spectroscopy on DPPC/water model membranes were applied and the results were compared. DSC data showed that Glu perturbs to a greater degree the model membrane compared to beta-Ala-Tyr. Thus, alteration of the phase transition temperature and half width of the peaks, abolishment of the pretransition and influence on the enthalpy of the phase transition were more pronounced in the Glu loaded bilayers. Raman spectroscopy showed that incorporation of Glu in DPPC/water bilayers increased the order in the bilayers in contrast to the effect of the dipeptide. Several structural and dynamical properties of the DPPC multilamellar bilayers with and without the dipeptide or Glu were compared using high resolution C-13 MAS (Magic Angle Spinning) spectra and spectral simulations of inhomogeneously broadened, stationary P-31 NMR lineshapes measured under CP (Cross-polarization) conditions. These methods revealed that the aminoacid Glu binds in the close realm of the phosphate in the hydrophilic headgroup of DPPC while beta-Ala-Tyr is located more deeply inside the hydrophobic zone of the bilayer. The P-31 NMR simulations indicated restricted fast rotary motion of the phospholipids about their long axes in the organized bilayer structure. Finally, by the applied methodologies it is concluded that the two molecules under study exert dissimilar thermal and dynamic effects on lipid bilayers, the Glu improving significantly the packing of the lipids in contrast to the smaller and opposite effect of the dipeptide.