Fluid Lamellar Phase Research Articles

Effect of angiotensin II non-peptide AT1 antagonist losartan on phosphatidylethanolamine membranes

Losartan was found to affect both the thermotropic behavior and molecular mobility of dimyristoyl- and dipalmitoyl-phosphatidylcholine membranes (Theodoropoulou and Marsh, Biochim. Biophys. Acta 1461 (1999) 135–146). At low concentrations, the antagonist is located close to the interfacial region of the phosphatidylcholine bilayer while at high mole fractions it inserts deeper in the bilayers. In the present study, we investigated the interactions of losartan with phosphatidylethanolamine membranes using differential scanning calorimetry (DSC), electron spin resonance (ESR) and 31P nuclear magnetic resonance (NMR) spectroscopy. DSC showed that the antagonist affected the thermotropic transitions of dimyristoyl-, dipalmitoyl- and dielaidoyl-phosphatidylethanolamine membranes (DMPE, DPPE and DEPE, respectively). ESR spectroscopy showed that the interaction of losartan with phosphatidylethanolamine membranes is more superficial than in the case of phosphatidylcholine bilayers. Additionally, losartan increased the spin-spin broadening of 12-PESL spin labels in the gel phase of DMPE and DPPE membranes, while in the case of DEPE membranes the opposite effect was observed. 31P-NMR showed that the antagonist stabilizes the fluid lamellar phase of DEPE membranes relative to the hexagonal HII phase. Our results show that losartan affects the thermotropic behavior of phosphatidylethanolamine membranes, while the molecular mobility of the membranes is not affected greatly. Furthermore, its interactions with phosphatidylethanolamine membranes are more superficial than with phosphatidylcholine bilayers.

Estimations of lipid bilayer geometry in fluid lamellar phases

The excess water bilayer thickness, d l,0, and molecular area, A 0, of lipid amphiphiles in the fluid lamellar phases of dioleoylphosphatidylcholine (DOPC) and dipalmitoleoylphosphatidylcholine (DPolPC) have been estimated between 15 and 50°C and for dimyristoylphosphatidylcholine (DMPC) between 25 and 50°C. These determinations have been made from X-ray measurements on samples of known water composition. With respect to temperature, T, d l,0 and A 0 are well fitted to a linear equation. We find d l,0 (Å)=(35.68±0.02)−(0.0333±0.0006) T (°C) and A 0 (Å 2)=(70.97±0.05)+(0.136±0.001) T (°C) for DOPC, d l,0 (Å)=(35.2±0.1)−(0.068±0.003) T (°C) and A 0 (Å 2)=(59.7±0.2)+(0.210±0.006) T (°C) for DMPC, and d l,0 (Å)=(34.54±0.03)−(0.0531±0.0009) T (°C) and A 0 (Å 2)=(67.12±0.09)+(0.173±0.003) T (°C) for DPolPC. The accuracy of these estimates depends largely on how accurately the excess water point is determined. Ideally, reliable X-ray and compositional data will be available around the excess water and it may be found by simple inspection, but this is the exception rather than the rule, since samples close to water excess normally sequester sizeable amounts of water in defects, which lead to an underestimate of d l,0. and overestimate of A 0. In this paper, we report a methodology for identifying and removing such data points and fitting the remaining data in order to determine the excess water point.

Cardiolipin, α- d-glucopyranosyl, and l-lysylcardiolipin from Gram-positive bacteria: FAB MS, monofilm and X-ray powder diffraction studies

Cardiolipin preparations from Streptococcus B, Listeria welshimeri, Staphylococcus aureus, and a glucosyl and lysyl derivative of cardiolipin were analysed for fatty acid composition and fatty acid combinations. Three different fatty acid patterns are described and up to 17 molecular species were identified in Streptococcus B lipids by high resolution FAB MS. The physicochemical properties of these lipids were characterised in the sodium salt form by monofilm experiments and X-ray powder diffraction. All lipids formed stable monofilms. The minimal space requirement of unsubstituted cardiolipin was dictated by the fatty acid pattern. Substitution with l-lysine led to a decrease of the molecular area, substitution with d-glucopyranosyl to an increase. On self assembly at 100% relative humidity, all preparations adopted lamellar structures. They showed a high degree of order, in spite of the heterogeneous fatty acid compositions and numerous fatty acid combinations. The repeat distances in lamellar fluid phase varied between 4.99 and 5.52 nm, the bilayer thickness between 3.70 and 4.46 nm. Surprising were the low values of sorbed water per molecule of the glucosyl and lysyl derivatives which were 58 and 60%, compared with those of the respective cardiolipin. When Na + was replaced as counterion by Ba 2+, the bilayer structure was retained, but the lipids were in the lamellar gel phase and the fatty acids were tilted between 32 and 53° away from the bilayer normal. Wide angle X-ray diffraction studies and electron density profiles are also reported. Particular properties of glucosyl cardiolipin are discussed.

Influence of the physical state of the membrane on the enzymatic activity and energy of activation of protein kinase C alpha.

The activation of protein kinase C alpha was studied by using a lipid system consisting of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS) (molar ratio 4:1) and different proportions of 1-palmitoyl-2-oleoyl-sn-glycerol (POG). The phase behavior of the lipidic system was characterized by using differential scanning calorimetry and 31P NMR, and a phase diagram was elaborated. The results suggested the formation of two diacylglycerol/phospholipid complexes, one at 15 mol % of POG and the second at 30 mol % of POG. These two complexes would define the three regions of the phase diagram: in the first region (concentrations of POG lower than 15 mol %) there is gel-gel immiscibility at temperatures below that of the phase transition between C1 and pure phospholipid, and a fluid lamellar phase above of the phase transition. In the second region (between 15 and 30 mol % of POG), gel-gel immiscibility between C1 and C2 with fluid-fluid immiscibility was observed, while inverted hexagonal HII and isotropic phases were detected by 31P NMR. In the third region (concentrations of POG higher than 30 mol %), gel-gel immiscibility seemed to occur between C2 and pure POG along with fluid-fluid immiscibility, while an isotropic phase was detected by 31P NMR. When PKC alpha activity was measured, as a function of POG concentration, maximum activity was found at POG concentrations as low as 5-10 mol %; the activity slightly decreased as POG concentration was increased to 45 mol % at 32 degrees C (above Tc) whereas activity did not change with increasing concentrations of POG at 5 degrees C (below Tc). When the activity was studied as a function of temperature, at different POG concentrations, and depicted as Arrhenius plots, it was found that the activity increased with increasing temperatures, showing a discontinuity at a temperature very close to the phase transition of the system and a lower activation energy at the upper slope of the graph, indicating that the physical state of the membrane affected the interaction of PKC alpha with the membrane.

Synthesis and Phase Behavior of New Amphiphilic PEG-Based Triblock Copolymers as Gelling Agents for Lamellar Liquid Crystalline Phases

We report the first evidence of the potential of new amphiphilic ABA-triblock copolymers as gelling agents for lamellar liquid crystalline Lα phases. Recently, we described a new type of lamellar hydrogels which are not based on a polymer network swollen in water but are obtained by the addition of small amounts of a nonionic polymer surfactant to the fluid lamellar Lα phase of the dimyristoyl−phosphatidyl−choline(DMPC)/pentanol/water system. In contrast with these previously reported gelling agents which were all AB-diblock copolymers, the novel ABA-triblock copolymers consist of double chain hydrophobic moieties (A) attached to each end of a poly(ethylene glycol) chain (B). The synthesis of these new macromolecules and their gelation properties are described. The comparison of these novel lamellar hydrogels with those based on the AB-diblock copolymers provides direct evidence for the existence of cross-bridging conformations of the ABA-triblock copolymers between adjacent membranes.

Polymorphism of the 1-Palmitoyl-2-arachidonoyl-phosphatidyl-ethanolamine/Dimyristoyl-phosphatidylmethanol Mixture, a Phospholipase A2 Substrate

The polymorphism of the equimolar mixture of 1-palmitoyl-2-arachidonoyl-phosphatidylethanolamine (PAPE) and dimyristoyl-phosphatidylmethanol (DMPM) was examined by infrared and31P nuclear magnetic resonance spectroscopy to determine the suitability of this widely used but yet uncharacterized lipid mixture as a phospholipase A2 substrate. The results show that the mixture undergoes a gel-to-liquid crystalline phase transition between 12 and 35°C. The transitions of the individual lipids were also examined. We report that the temperature of the gel-to-liquid crystalline phase transition of DMPM is 40°C. At 2°C, PAPE exists in the fluid lamellar form. This lipid undergoes a lamellar-to-inverted hexagonal phase transition at 31°C. In conclusion, the equimolar PAPE/DMPM mixture forms fluid lamellar phase at the physiological temperature. However, the upper end of the gel-to-liquid crystalline phase transition of the mixture is really close to the physiological temperature and this situation is a serious source of potential artefacts.

A model membrane approach to the epidermal permeability barrier: an X-ray diffraction study.

The permeability of mammalian skin is determined in large part by lamellar lipid domains packed between cells of the upper layer of the epidermis, the stratum comeum. Although these lamellae have features in common with typical biological membranes, they differ in having a lipid population composed mainly of ceramides, cholesterol, and free fatty acids. In our initial studies of the relationship between lipid composition and phase behavior in this unusual system, we used deuterium NMR [Kitson et al. (1994) Biochemistry 33, 6707-6715] to examine aqueous dispersions of nonhydroxylated bovine brain ceramide, cholesterol, and perdeuterated palmitic acid, and found complex phase behavior as a function of temperature and pH, whereas analogous dispersions in which sphingomyelin replaced ceramide resulted in spectra consistent with a fluid lamellar phase under the same conditions. To extend these observations, we examined the same dispersions at pH 5.2 by means of X-ray diffraction. The significant findings are as follows: (1) the ceramide dispersions form complex crystalline phases between room temperature and about 40 degrees C; (2) the majority of the crystalline cholesterol is not in a separate phase; and (3) the analogous sphingomyelin dispersions form a fluid lamellar phase under the same conditions. We conclude that ceramides, even in the presence of considerable mole fractions of cholesterol, can form crystalline lamellar structures. We suggest that the existence of such structures in stratum corneum may be important in the function of the epidermal permeability barrier, and that the interaction between ceramide and cholesterol in other biological membranes may result in regions having unique physical properties.

Lipid mixing in dimyristoyl phosphatidylcholine-dimyristoyl glycerol dispersions: spin label ESR studies

The lipid transfer and mixing properties in hydrated dispersions of dimyristoyl phosphatidylcholine (DMPC)/dimyristoyl glycerol (DMG) binary mixtures have been investigated by using electron spin resonance spectroscopy of the spin-labelled lipid components. The assay system is based on the reduction in spectral broadening from spin–spin interactions that takes place by dilution of the spin-labelled lipid on transfer to a 9-fold excess of dispersions that contain no spin label. Lipid dispersions with DMPC:DMG compositions of 70:30, 40:60 and 20:80 mol/mol, for which the fluid phases have lamellar, inverted hexagonal and isotropic structures, respectively, have been studied. Essentially no transfer of spin-labelled lipid takes place for any of the lipid mixtures in the lamellar gel phase, or in dispersions of DMPC alone at all temperatures studied. The greatest degrees of transfer are found in the fluid phase of the DMPC/DMG mixtures. In general, the extent of lipid transfer is greater for the diacylglycerol component than for the phosphatidylcholine component. The extent of transfer of phosphatidylcholine is very low in the fluid lamellar phase of the 70:30 mol/mol DMPC/DMG, as compared with that of diacylglycerol. Only in the case of the 40:60 mol/mol DMPC/DMG mixture, in the inverted hexagonal phase, are the extents of transfer comparable for both phosphatidylcholine and diacylglycerol components, indicating a bulk transfer of lipid within the dispersions. The largest extent of transfer is found for diacylglyerol in the 20:80 mol/mol DMPC/DMG mixture in the isotropic phase.

Dynamic chain conformations in dimyristoyl glycerol-dimyristoyl phosphatidylcholine mixtures. 2H-NMR studies

The dynamic molecular lipid chain conformations in fully hydrated dimyristoyl phosphatidylcholine (DMPC)-dimyristoyl glycerol (DMG) mixtures have been investigated by 2H-NMR spectroscopy of the individual lipid components, the sn-2 chains of which were perdeuterated or, in the case of DMG, specifically deuterated at the C-2 position. Mixtures of compositions corresponding to the three different regions of the binary phase diagram in which the fluid phase is lamellar (DMPC:DMG 70:30 mol/mol), inverted hexagonal (DMPC:DMG 45:55 and 40:60 mol/mol), or isotropic (DMPC:DMG 20:80 mol/mol) were investigated. The gel phase in all three regions of the phase diagram has a lamellar structure, with the lipid chains rotating about the molecular long axis but executing only limited angular excursions. In the fluid lamellar phase of the 70:30 mol/mol DMPC-DMG mixture the profile of segmental chain flexibility is similar to that in single-component phospholipid bilayers and is characterized by an order parameter plateau for both lipid components. The chain order of the DMPC component is greater than in bilayers of DMPC alone and is also greater than that of the DMG component. In the inverted hexagonal phase of the 45:55 mol/mol DMPC-DMG mixture the chain flexibility profile is characterized by more widely spaced segmental order parameters off the plateau region. The intrinsic degree of chain order in the inverted hexagonal phase is less than in the lamellar phase of the 70:30 mol/mol mixture, and the difference in chain order between the DMPC and DMG components is reduced relative to that in the lamellar phase. The unique conformational features at the C-2 position of the sn-2 chain that characterize bilayers of diacyl phospholipids are found also for the diacylglycerol molecules in the fluid lamellar phase and most probably also in the inverted hexagonal phase. The DMG molecules are therefore integrated in the membrane (or nonlamellar lipid phase) in a configuration that is similar to that of the phospholipids and different from the crystal structure of diacylglycerols.

Lipid chain dynamics in diacylglycerol-phosphatidylcholine mixtures studied by slow-motional simulations of spin label ESR spectra

Simulation of the ESR spectra from spin-labelled phosphatidylcholine and from spin-labelled diacylglycerol has been used to study the lipid chain dynamics in mixtures of dimyristoyl glycerol with dimyristoyl phosphatidylcholine, as a function of temperature and composition. Successful simulation of the ESR spectra required taking into account slow motional components which was done by using the very anisotropic reorientation model. Chain rotational rates varied with temperature, but not greatly with composition and chain position, which primarily affected chain order. The latter decreased progressively with increasing diacylglycerol content from the lamellar, to inverted hexagonal, to isotropic phase and was consistently lower for the diacylglycerol component than for the phosphatidylcholine component. The order profile of the spin-labelled chains was obtained for both the phosphatidylcholine and diacylglycerol components in a 58:42 mol/mol mixture in the fluid lamellar phase.

Effects of COR6.6 and COR15am Polypeptides Encoded by COR (Cold-Regulated) Genes of Arabidopsis thaliana on Dehydration-Induced Phase Transitions of Phospholipid Membranes

Cold acclimation of Arabidopsis thaliana includes the expression of cold-regulated (COR) genes and the accumulation of COR polypeptides. The hydration characteristics of two COR polypeptides, COR6.6 and COR15am, have been determined and their effects on the dehydration-induced liquid crystalline-to-gel and lamellar-to-hexagonal II phase transitions in phospholipid mixtures have been examined. After dehydration at osmotic pressures between 8 and 150 MPa, the water content of the COR polypeptides was less than that of bovine serum albumin, with COr15am the least hydrated: bovine serum albumin > COR6.6 > COR15am. Neither COR6.6 nor COR15am altered the dehydration-induced gel lamellar --> fluid lamellar phase transition temperature of either dipalmitoylphosphatidylcholine or dioleoylphosphatidylcholine (DOPC). In multilamellar vesicles of dioleoylphosphatidylethanolamine:DOPC (1:1, mol:mol) prepared by either freeze-thaw or reverse-phase evaporation methods, neither COR6.6, COR15am, nor bovine serum albumin altered the incidence of the dehydration-induced formation of the inverted hexagonal phase as a function of osmotic pressure. However, a specific ultrastructural alteration--the formation of a striated surface morphology in the lamellar domains--was observed in mixtures of dioleoylphosphatidylethanolamine:DOPC that were dehydrated in the presence of COR15am. Nevertheless, neither COR6.6 nor COR15am appears to participate in a specific protein-phospholipid interaction that alters the dehydration-induced phase behavior of phospholipid vesicles.

Lamellar Biogels: Fluid-Membrane-Based Hydrogels Containing Polymer Lipids

A class of lamellar biological hydrogels comprised of fluid membranes of lipids and surfactants with small amounts of low molecular weight poly(ethylene glycol)-derived polymer lipids (PEG-lipids) were studied by x-ray diffraction, polarized light microscopy, and rheometry. In contrast to isotropic hydrogels of polymer networks, these membrane-based birefringent liquid crystalline biogels, labeled L-alpha,g, form the gel phase when water is added to the liquid-like lamellar L-alpha phase, which reenters a liquid-like mixed phase upon further dilution. Furthermore, gels with larger water content require less PEG-lipid to remain stable. Although concentrated (approximately 50 weight percent) mixtures of free PEG (molecular weight, 5000) and water do not gel, gelation does occur in mixtures containing as little as 0.5 weight percent PEG-lipid. A defining signature of the L-alpha,g regime as it sets in from the fluid lamellar L-alpha phase is the proliferation of layer-dislocation-type defects, which are stabilized by the segregation of PEG-lipids to the defect regions of high membrane curvature that connect the membranes.

Lipid chain dynamics and molecular location of diacylglycerol in hydrated binary mixtures with phosphatidylcholine: spin label ESR studies.

The lipid chain motions in hydrated binary mixtures of dimyristoylglycerol (DMG) with dimyristoylphosphatidylcholine (DMPC) have been studied by using ESR spectroscopy of analogues of both components that are spin-labeled at one of eight different positions along the sn-2 chain. The phase diagram of the binary mixtures divides into three separate regions along the composition axis, corresponding to the formation of isothermally melting compounds, with DMPC/DMG stoichiometries of approximately 1:1 and 1:2 mol/mol in the gel phase [Heimburg, T., Wurz, U., & Marsh, D. (1992) Biophys. J. 63, 1369-1378]. In the first region (up to 50 mol % DMG), comparison of the chain flexibility profiles, and the chain profiles of the polarity-dependent isotropic hyperfine coupling constant, of the two different spin-labeled components indicates that DMG is incorporated in the fluid lipid bilayer in a manner similar to that of the host DMPC but is situated approximately two CH2 groups deeper into the hydrophobic interior. At lower contents of DMG, the chain packing is increased by the addition of DMG, whereas at higher DMG contents the lipid chain order decreases rapidly, on reaching the inverted hexagonal phase of the second region of the phase diagram. In the second region of the phase diagram (50-67 mol % DMG), the DMG fits better into the fluid inverted hexagonal phase than into the fluid lamellar phase of the first region and is located only approximately one CH2 group deeper than the corresponding DMPC. In these first two regions of the phase diagram, the ESR spectra of both spin-labeled components display an axial anisotropy that evidences the increasing angular amplitude of motion with position down the chain that is characteristic of liquid crystalline fluid phases. In the third region of the phase diagram (above 67 mol % DMG), the fluid phase consists of isotropically tumbling DMG molecules in which the DMPC molecules are incorporated as inverted micelles as indicated by the residual anisotropic motion of the spin-labeled phosphatidylcholine analogues.

Unexpected Formation of Vesicular Aggregates in Aqueous Solutions of n-Octyl 1-Thio-.alpha.-D-talopyranoside. Marked Effects of Intramolecular Hydrogen Bonding

Carbohydrate-derived amphiphiles are the subject of intensive investigations, mainly due to their potential as biodegradable, nontoxic, and nonallergenic surface-active components of detergent formulations, pharmaceuticals, and foodstuffs. Our interest is focused on the relationship between molecular structure and physical properties of nonionic carbohydrate-derived amphiphiles. In contrast to ionic amphiphiles, the type of aggregate formed in aqueous solution changes upon chain elongation. We have established a firm relationship between the number of hydrogen-bond-forming functionalities in the headgroup and the type of aggregate that is formed in aqueous solution at a given alkyl chain length. This theory predicts that hexopyranose derivatives with a single alkyl chain up to at least n-nonyl will form micelles in solution. However, this approach cannot predict the differences that may arise from variation in carbohydrate stereochemistry. To study the influence of the stereochemistry of the headgroup, several novel alkyl 1-thioglycopyranosides have been synthesized. Relatively short (n-hexyl to n-octyl) alkyl chains were chosen to avoid solubility problems. Lyotropic behavior can be screened conveniently with the aid of the contact preparation method. We found that n-heptyl 1-thio-α-D-glucopyranoside forms normal hexagonal (HI), bicontinuous cubic (VI), and fluid lamellar (Lα) phases as a function of increasing surfactant concentration, as anticipated on the basis of elegant work by Chung and Jeffrey. Lyotropic mesophases result from a packing of the aggregates in aqueous solution into ordered structures. Therefore, the nature of the mesophase at the interface with bulk water (i.e., at maximum hydration)

The effect of incorporation of phosphonic acid dibutyl ester in POPC model membranes on the hydration force

The hydration and swelling properties of lipid model membranes formed by mixtures of 1-palmitoyl-2-oleoylglycerophosphatidyl-choline (POPC) and different molar ratios R a l of dodecyl phosphonic acid dibutyl ester (PAB 12) in water were studied at 25°C. It is shown by X-ray diffraction and 31P NMR spectroscopy that the system exists in a fluid lamellar phase at R a l = 0.25 and 0.5 over a wide range of water concentration. Intermembrane forces were determined by the osmotic stress technique. The hydration repulsion was found to decrease exponentially with increasing intermembrane water layer thickness. The values of the pre-exponential factor of the hydration forces which slightly decrease with increasing R a l were discussed according to the theory of water polarization near structured water/lipid interfaces (Cevc).

Lyotropic phase behaviour of n-octyl-1-O-?-D-glucopyranoside and its thio derivative n-octyl-1-S-?-D-glucopyranoside

We have determined the phase diagrams of n-octyl-I -O-p-D-glucopyranoside and its thio derivative n-octyl-I -S-p-D-glucopyranoside in water, using polarizing microscopy and X-ray diffraction. Comparison of the two compounds gives us an insight into the complex interactions which determine phase behaviour. Both compounds form type I (normal) lyotropic phases, with the adoption of a fluid lamellar phase at low hydrations and a cubic phase at higher hydrations. However, the hexagonal phase observed in the n-octyl- I-O-p-D-glucopyranoside/water system adjacent to the micellar wlution is suppressed in the n-octyl-I-S-p-D-glucopyranoside/water system. In addition, we have found that in both systems the cubic phase spontaneously grows into large monodomains, as revealed by faceting of entrapped air bubbles. Indexing of the monodomain diffraction patterns shows that in both cases the space group of the cubic phase is Ia3d (no. 230).

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.

Structure of vitaminylated lipids in aqueous dispersion: X-ray diffraction and phosphorus-31 NMR studies of N-biotinylphosphatidylethanolamines

The structures of the phases formed in excess buffer (at pH 7.4) by a homologous series of saturated diacylphosphatidylethanolamines in which the headgroup is N-derivatized with biotin have been investigated for chain lengths of C(12:0) to C(20:0), using both 31P nuclear magnetic resonance (NMR) spectroscopy and small-angle X-ray diffraction. In 1 M NaCl, all lipids display 31P NMR spectra characteristic of a lamellar gel phase at low temperature. In the fluid phase, the lipids of C(12:0) and C(14:0) chain lengths display isotropic 31P NMR spectra, corresponding to aggregated phases with high surface curvature, whereas those with C(18:0) and C(20:0) chain lengths display sharp axial powder patterns characteristic of a lamellar (L alpha) phase. The lipid of intermediate C(16:0) chain length displays a more complex temperature dependence of the 31P NMR spectra in the fluid phase. The spectra convert from an axial powder pattern of unusually low chemical shift anisotropy to one characteristic of a fluid lamellar (L alpha) phase with increasing temperature. The small-angle X-ray diffraction patterns of the lipids in 1 M NaCl have lamellar repeat spacings in the gel phase which increase linearly with chain length and are consistently lower than those in the fluid phase [for chain lengths of C(16:0) to C(20:0)]. In addition, the gradient in long spacing with chain length in the gel phase is approximately half that expected for a gel phase with untilted, all-trans chains, indicating that the lipid chains are interdigitated in the gel phase (L beta i).(ABSTRACT TRUNCATED AT 250 WORDS)

Dipyrenylphosphatidylcholine as a probe of bilayer pressures

We have used dipyrenylphosphatidylcholines (dipyPCs) to study the pressure in the fluid lamellar phase formed by mixtures of fully hydrated dioleoylphosphatidylcholine (DOPC) and dioleoylphosphatidylethanolamine (DOPE). As we increase the DOPE mole fraction at 25°C we observe a linear increase in the ratio of the excimer-to-monomer signal (E/M1). We argue that this observation can be understood in terms of an increase in the lateral pressure in the chain region, i.e., in the bilayer plane. This change itself is driven by the decrease in lateral pressure between headgroups as we add DOPE. We expect the lateral pressure to vary in magnitude as we probe the bilayer at different depths [1]. We have confirmed this by recording E/M1 using di[10-(pyren-1-yl)decanoyl]phosphatidylcholine (10dipyPC) and di[4-(pyren-1-yl)butanoyl]phosphatidylcholine (4dipyPC). We find that in 100% DOPC the E/M1 for 4dipyPC is 2.5 times greater than that for 10dipyPC. The above observations can all be rationalized in terms of changes in the lateral pressure profile. An inverse hexagonal liquid crystalline phase is found in the range 100-83% DOPE [2]. In this region of the phase diagram we observe a quadratic variation in E/M1, with a minimum at 95% DOPE. We hypothesize that this variation reflects the chain stretching that is necessitated by the geometrical packing constraints of the hexagonal phase [3]. Again, we find that the E/M1 for 4dipyPC is greater than that for 10dipyPC, but in this phase only by a factor of two.

Nonideal mixing of phosphatidylserine and phosphatidylcholine in the fluid lamellar phase

The mixing of phosphatidylserine (PS) and phosphatidylcholine (PC) in fluid bilayer model membranes was studied by measuring binding of aqueous Ca2+ ions. The measured [Ca2+]aq was used to derive the activity coefficient for PS, gamma PS, in the lipid mixture. For (16:0, 18:1) PS in binary mixtures with either (16:0, 18:1)PC, (14:1, 14:1)PC, or (18:1, 18:1)PC, gamma PS > 1; i.e., mixing is nonideal, with PS and PC clustered rather than randomly distributed, despite the electrostatic repulsion between PS headgroups. To understand better this mixing behavior, Monte Carlo simulations of the PS/PC distributions were performed, using Kawasaki relaxation. The excess energy was divided into an electrostatic term Uel and one adjustable term including all other nonideal energy contributions, delta Em. Uel was calculated using a discrete charge theory. Kirkwood's coupling parameter method was used to calculate the excess free energy of mixing, delta GEmix, hence In gamma PS,calc. The values of In gamma PS,calc were equalized by adjusting delta Em in order to find the simulated PS/PC distribution that corresponded to the experimental results. We were thus able to compare the smeared charge calculation of [Ca2+]surf with a calculation ("masked evaluation method") that recognized clustering of the negatively charged PS: clustering was found to have a modest effect on [Ca2+]surf, relative to the smeared charge model. Even though both PS and PC tend to cluster, the long-range nature of the electrostatic repulsion reduces the extent of PS clustering at low PS mole fraction compared to PC clustering at an equivalent low PC mole fraction.