Interfacial behavior of phospholipid monolayers revealed by mesoscopic simulation

A mesoscopic model with molecular resolution is presented for dipalmitoyl phosphatidylcholine (DPPC) and palmitoyl oleoyl phosphatidylcholine (POPC) monolayer simulations at the air-water interface using many-body dissipative particle dynamics (MDPD). The parameterization scheme is rigorously based on reproducing the physical properties of water and alkane and the interfacial property of the phospholipid monolayer by comparison with experimental results. Using much less computing cost, these MDPD simulations yield a similar surface pressure-area isotherm as well as similar pressure-related morphologies as all-atom simulations and experiments. Moreover, the compressibility modulus, order parameter of lipid tails, and thickness of the phospholipid monolayer are quantitatively in line with the all-atom simulations and experiments. This model also captures the sensitive changes in the pressure-area isotherms of mixed DPPC/POPC monolayers with altered mixing ratios, indicating that the model is promising for applications with complex natural phospholipid monolayers. These results demonstrate a significant improvement of quantitative phospholipid monolayer simulations over previous coarse-grained models.     

Interaction of poly(ethylene-glycols) with air-water interfaces and lipid monolayers: investigations on surface pressure and surface potential.

We have characterized the surface activity of different-sized poly(ethylene-glycols) (PEG; M(r) 200-100,000 Da) in the presence or absence of lipid monolayers and over a wide range of bulk PEG concentrations (10(-8)-10% w/v). Measurements of the surface potential and surface pressure demonstrate that PEGs interact with the air-water and lipid-water interfaces. Without lipid, PEG added either to the subphase or to the air-water interface forms relatively stable monolayers. Except for very low molecular weight polymers (PEGs < 1000 Da), low concentrations of PEG in the subphase (between 10(-5) and 10(-4)% w/v) increase the surface potential from zero (with respect to the potential of a pure air-water interface) to a plateau value of approximately 440 mV. At much higher polymer concentrations, > 10(-1)% (w/v), depending on the molecular weight of the PEG and corresponding to the concentration at which the polymers in solution are likely to overlap, the surface potential decreases. High concentrations of PEG in the subphase cause a similar decrease in the surface potential of densely packed lipid monolayers spread from either diphytanoyl phosphatidylcholine (DPhPC), dipalmitoyl phosphatidylcholine (DPPC), or dioleoyl phosphatidylserine (DOPS). Adding PEG as a monolayer at the air-water interface also affects the surface activity of DPhPC or DPPC monolayers. At low lipid concentration, the surface pressure and potential are determined by the polymer. For intermediate lipid concentrations, the surface pressure-area and surface potential-area isotherms show that the effects due to lipid and PEG are not always additive and that the polymer's effect is distinct for the two lipids. When PEG-lipid-mixed monolayers are compressed to surface pressures greater than the collapse pressure for a PEG monolayer, the surface pressure-area and surface potential-area isotherms approach that of the lipid alone, suggesting that for this experimental condition PEG is expelled from the interface.     

Molecular organization of antibiotic amphotericin B in dipalmitoylphosphatidylcholine monolayers induced by K(+) and Na(+) ions: the Langmuir technique study

The effect of potassium (K(+)) and sodium (Na(+)) ions on the self-association of antibiotic amphotericin B (AmB) in the lipid membrane was reported. Mixed Langmuir monolayers of AmB and dipalmitoylphosphatidylcholine (DPPC) were investigated by recording surface pressure-area isotherms spread on aqueous buffers containing physiological concentration of K(+) and Na(+) ions. The analyses of the π-A isotherms and compressional modulus curves indicate the interactions in the AmB-DPPC system. The strength of the AmB-DPPC interactions and the stability of the mixed monolayers were examined on the basis of the excess free energy of mixing values. The obtained results proved a high affinity of AmB towards lipids induced by the presence of K(+) than Na(+) ions. The most stable monolayers in the presence of K(+) and Na(+) ions were formed by AmB and DPPC with the 1:1 and 2:1 stoichiometry. The understanding of the AmB aggregation processes at the molecular level should contribute to elucidate the mechanisms of action and toxicity of this widely used drug. The presented results are potentially valuable in respect to develop more efficient and less toxic AmB formulations.     

Liquid crystalline/gel state phase separation in docosahexaenoic acid-containing bilayers and monolayers

The phase behavior of lipid mixtures containing 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (18:0, 22:6 PC) with 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) was studied with bilayers using differential scanning calorimetry (DSC), and with monolayers monitoring pressure/area isotherms and surface elasticity, and lipid domain formation followed by epifluorescence microscopy. From DSC studies it is concluded that DPPC/18:0, 22:6 PC phase separates into DPPC-rich and 18:0, 22:6 PC-rich phases. In monolayers, phase separation is indicated by changes in pressure-area isotherms implying phase separation where 18:0, 22:6 PC is 'squeezed out' of the remaining DPPC monolayer. Phase separation into lipid domains in the mixed PC monolayer is quantified by epifluorescence microscopy using the fluorescently labeled phospholipid membrane probe, 1, 2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl). These results further describe the ability of docosahexaenoic acid to participate in lipid phase separations in membranes.     

Microstructure and dynamic surface properties of surfactant protein SP-B/dipalmitoylphosphatidylcholine interfacial films spread from lipid-protein bilayers

 Suspensions of dipalmitoylphosphatidylcholine (DPPC) bilayers containing 5, 10 or 20% (w/w) surfactant protein SP-B have been reconstituted and spread at air-liquid interfaces. Compression isotherms of DPPC/SP-B monolayers spread from these preparations were qualitatively comparable to the isotherms of the corresponding DPPC/SP-B monolayers spread from solvents. SP-B was squeezed-out at higher pressures from vesicle-spread films than from solvent-spread monolayers. SP-B caused a marked decrease on the rate of relaxation of DPPC collapse phases to equilibrium pressures in all the lipid/protein films assayed. This stabilizing effect was higher in vesicle-spread than in solvent-spread monolayers. Inclusion in the films of traces of the fluorescent probe NBD-PC (1 mol%) and use of a fluorescent derivative of SP-B labeled with a rhodamine derivative, Texas Red, allowed for direct observation of protein and lipid domains at the interface by epifluorescence microscopy. Upon compression, SP-B altered the packing of phospholipids in the bilayer-spread films, observed as a SP-B-induced reduction of the area of liquid-condensed domains, in a way similar to its effect in solvent-spread monolayers. SP-B was not associated with condensed regions of the films. Fluorescence images from vesicle-spread films showed discrete fluorescent aggregates that could be consistent with the existence of lipid-protein vesicles in close association with the monolayer. Both the retention of SP-B at higher surface pressures and the greater stability of collapse phases of DPPC/SP-B films prepared by spreading from liposomes in comparison to those spread from solvents can be interpreted as a consequence of formation of complex bilayer-monolayer interacting systems. Received: 1 December 1999 / Revised version: 2 March 2000 / Accepted: 2 March 2000     

The detrimental effect of serum albumin on the re-spreading of a dipalmitoyl phosphatidylcholine Langmuir monolayer is counteracted by a fluorocarbon gas

We have recently reported that fluorocarbon gases exhibit an effective fluidizing effect on Langmuir monolayers of dipalmitoyl phosphatidylcholine (DPPC), preventing them from crystallizing up to surface pressures of ∼ 40 mN m^− 1, i.e. well above the DPPC's equilibrium surface pressure. We now report that gaseous perfluorooctyl bromide (gPFOB) promotes the re-spreading of DPPC Langmuir monolayers compressed on a bovine serum albumin (BSA)-containing sub-phase. The latter protein is known to maintain a concentration-dependent surface pressure that can exceed the re-spreading pressure of collapsed monolayers. This phenomenon was proposed to be responsible for lung surfactant inactivation. Compression/expansion isotherms and fluorescence microscopy experiments were carried out to assess the monolayers' physical state. We have found that, during expansion under gPFOB-containing air, the surface pressure of a DPPC monolayer on a BSA-containing sub-phase decreased to much lower values than when the DPPC monolayer was expanded in the presence of BSA under air (∼ 0 mN m^− 1 vs. ∼ 7.5 mN m^− 1 at 120 Å², respectively). Moreover, fluorescence images showed that, during expansion, the BSA-coupled DPPC monolayers, in contact with gPFOB, remained in the liquid-expanded state for surface pressures lower than 10 mN m^− 1, whereas they were in a liquid-condensed semi-crystalline state, even at large molecular areas (120 Å²), when expanded under air. The re-incorporation of the PFOB molecules in the DPPC monolayer during expansion thus competes with the re-incorporation of BSA, thus preventing the latter from penetrating into the DPPC monolayer. We suggest that combinations of DPPC and a fluorocarbon gas may be useful in the treatment of lung conditions resulting from a deterioration of the native lung surfactant function due to plasma proteins, such as in the acute respiratory distress syndrome.     

Lipid headgroup discrimination by antimicrobial peptide LL-37: insight into mechanism of action

Interaction of the human antimicrobial peptide LL-37 with lipid monolayers has been investigated by a range of complementary techniques including pressure-area isotherms, insertion assay, epifluorescence microscopy, and synchrotron x-ray scattering, to analyze its mechanism of action. Lipid monolayers were formed at the air-liquid interface to mimic the surface of the bacterial cell wall and the outer leaflet of erythrocyte cell membrane by using phosphatidylglycerol (DPPG), phosphatidylcholine (DPPC), and phosphatidylethanolamine (DPPE) lipids. LL-37 is found to readily insert into DPPG monolayers, disrupting their structure and thus indicating bactericidal action. In contrast, DPPC and DPPE monolayers remained virtually unaffected by LL-37, demonstrating its nonhemolytic activity and lipid discrimination. Specular x-ray reflectivity data yielded considerable differences in layer thickness and electron-density profile after addition of the peptide to DPPG monolayers, but little change was seen after peptide injection when probing monolayers composed of DPPC and DPPE. Grazing incidence x-ray diffraction demonstrated significant peptide insertion and lateral packing order disruption of the DPPG monolayer by LL-37 insertion. Epifluorescence microscopy data support these findings.     

Molecular organization of surfactin-phospholipid monolayers: Effect of phospholipid chain length and polar head

Mixed monolayers of the surface-active lipopeptide surfactin-C₁₅ and various lipids differing by their chain length (DMPC, DPPC, DSPC) and polar headgroup (DPPC, DPPE, DPPS) were investigated by atomic force microscopy (AFM) in combination with molecular modeling (Hypermatrix procedure) and surface pressure-area isotherms. In the presence of surfactin, AFM topographic images showed phase separation for each surfactin-phospholipid system except for surfactin-DMPC, which was in good agreement with compression isotherms. On the basis of domain shape and line tension theory, we conclude that the miscibility between surfactin and phospholipids is higher for shorter chain lengths (DMPC > DPPC > DSPC) and that the polar headgroup of phospholipids influences the miscibility of surfactin in the order DPPC > DPPE > DPPS. Molecular modeling data show that mixing surfactin and DPPC has a destabilizing effect on DPPC monolayer while it has a stabilizing effect towards DPPE and DPPS molecular interactions. Our results provide valuable information on the activity mechanism of surfactin and may be useful for the design of surfactin delivery systems.     

Miscibility of ternary mixtures of phospholipids and cholesterol in monolayers, and application to bilayer systems

We investigate miscibility transitions of two different ternary lipid mixtures, DOPC/DPPC/Chol and POPC/PSM/Chol. In vesicles, both of these mixtures of an unsaturated lipid, a saturated lipid, and cholesterol form micron-scale domains of immiscible liquid phases for only a limited range of compositions. In contrast, in monolayers, both of these mixtures produce two distinct regions of immiscible liquid phases that span all compositions studied, the alpha-region at low cholesterol and the beta-region at high cholesterol. In other words, we find only limited overlap in miscibility phase behavior of monolayers and bilayers for the lipids studied. For vesicles at 25 degrees C, the miscibility phase boundary spans portions of both the monolayer alpha-region and beta-region. Within the monolayer beta-region, domains persist to high pressures, yet within the alpha-region, miscibility phase transition pressures always fall below 15 mN/m, far below the bilayer equivalent pressure of 32 mN/m. Approximately equivalent phase behavior is observed for monolayers of DOPC/DPPC/Chol and for monolayers of POPC/PSM/Chol. As expected, pressure-area isotherms of our ternary lipid mixtures yield smaller molecular area and compressibility for monolayers containing more saturated acyl chains and cholesterol. All monolayer experiments were conducted under argon. We show that exposure of unsaturated lipids to air causes monolayer surface pressures to decrease rapidly and miscibility transition pressures to increase rapidly.     

Aggregation of puroindoline in phospholipid monolayers spread at the air-liquid interface

Puroindolines, cationic and cystine-rich low molecular weight lipid binding proteins from wheat seeds, display unique foaming properties and antimicrobial activity. To unravel the mechanism involved in these properties, the interaction of puroindoline-a (PIN-a) with dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) monolayers was studied by coupling Langmuir-Blodgett and imaging techniques. Compression isotherms of PIN-a/phospholipid monolayers and adsorption of PIN-a to lipid monolayers showed that the protein interacted strongly with phospholipids, especially with the anionic DPPG. The electrostatic contribution led to the formation of a highly stable lipoprotein monolayer. Confocal laser scanning microscopy and atomic force microscopy showed that PIN-a was mainly inserted in the liquid-expanded phase of the DPPC, where it formed an aggregated protein network and induced the fusion of liquid-condensed domains. For DPPG, the protein partitioned in both the liquid-expanded and liquid-condensed phases, where it was aggregated. The extent of protein aggregation was related both to the physical state of phospholipids, i.e., condensed or expanded, and to the electrostatic interactions between lipids and PIN-a. Aggregation of PIN-a at air-liquid and lipid interfaces could account for the biological and technological properties of this wheat lipid binding protein.     

Structural alterations of phospholipid film domain morphology induced by cholesterol

Structures of the monolayer films of dipalmitoylphosphatidylcholine (DPPC) mixed with different amounts of cholesterol were studied at air-water interface using surface pressure-area measurements, epifluorescence microscopy and atomic force microscopy (AFM). Pure DPPC, cholesterol or DPPC-cholesterol mixtures were dissolved in organic solvents with a small amount of fluorescently labeled phospholipid probe (NBD-PC) and spread onto the air-water interface. Surface pressure-area isotherms and epifluorescence microscopy of such films at the air-water interface suggested that DPPC undergoes a gas to fluid to condensed phase transition, while cholesterol undergoes a gas to solid-like transition. A shift of the surface pressure-area curve to lower area per molecule was observed when cholesterol was mixed with DPPC. Epifluorescence microscopy showed the formation of spiral shaped domains for mixed monolayers. Increase in cholesterol content abolished domain characteristics possibly due to fluidizing property of cholesterol. AFM measurements of monolayers, transferred onto freshly cleaved mica by Langmuir-Blodgett technique, revealed the alterations caused by cholesterol on the gel and fluid domains of such films. AFM measurements re-established similar trend in domain characteristics as evidenced in epifluorescence microscopy.     

The interaction of meso-tetraphenylporphyrin with phospholipid monolayers

In this article, we investigate the interaction of meso-tetraphenylporphyrin (TPP) with phospholipid monolayers. Pure TPP molecules form films at the air-water interface with large extension of aggregation, which is confirmed by UV-vis spectra of transferred monolayers. For mixed films of TPP with dipalmitoyl phosphatidyl choline (DPPC) or dipalmitoyl phosphatidyl glycerol (DPPG), on the other hand, aggregation is only significant at high surface pressures or high concentrations of TPP (above 0.1 molar ratio). This was observed via Brewster angle microscopy (BAM) for the Langmuir films and UV-vis spectroscopy for transferred layers onto solid substrates. TPP indeed causes the DPPC and DPPG monolayers to expand, especially at the liquid-expanded to liquid-condensed phase transition for DPPC. The effects from TPP cannot be explained using purely geometrical considerations, as the area per TPP molecule obtained from the isotherms is at least twice the expected value from the literature. Therefore, interaction between TPP and DPPC or DPPG should be cooperative, so that more phospholipid molecules are affected than just the first neighbors to a TPP molecule.     

Coarse-Grained Molecular Models of Water: A Review

Coarse-grained (CG) models have proven to be very effective tools in the study of phenomena or systems that involve large time- and length-scales. By decreasing the degrees of freedom in the system and using softer interactions than seen in atomistic models, larger timesteps can be used and much longer simulation times can be studied. CG simulations are widely used to study systems of biological importance that are beyond the reach of atomistic simulation, necessitating a computationally efficient and accurate CG model for water. In this review, we discuss the methods used for developing CG water models and the relative advantages and disadvantages of the resulting models. In general, CG water models differ with regards to how many waters each CG group or bead represents, whether analytical or tabular potentials have been used to describe the interactions, and how the model incorporates electrostatic interactions. Finally, how the models are parameterized depends on their application, so, while some are fitted to experimental properties such as surface tension and density, others are fitted to radial distribution functions extracted from atomistic simulations.     

Interpretation of H-NMR Experiments on the Orientation of the Transmembrane Helix WALP23 by Computer Simulations

Orientation, dynamics, and packing of transmembrane helical peptides are important determinants of membrane protein structure, dynamics, and function. Because it is difficult to investigate these aspects by studying real membrane proteins, model transmembrane helical peptides are widely used. NMR experiments provide information on both orientation and dynamics of peptides, but they require that motional models be interpreted. Different motional models yield different interpretations of quadrupolar splittings (QS) in terms of helix orientation and dynamics. Here, we use coarse-grained (CG) molecular dynamics (MD) simulations to investigate the behavior of a well-known model transmembrane peptide, WALP23, under different hydrophobic matching/mismatching conditions. We compare experimental ²H-NMR QS (directly measured in experiments), as well as helix tilt angle and azimuthal rotation (not directly measured), with CG MD simulation results. For QS, the agreement is significantly better than previously obtained with atomistic simulations, indicating that equilibrium sampling is more important than atomistic details for reproducing experimental QS. Calculations of helix orientation confirm that the interpretation of QS depends on the motional model used. Our simulations suggest that WALP23 can form dimers, which are more stable in an antiparallel arrangement. The origin of the preference for the antiparallel orientation lies not only in electrostatic interactions but also in better surface complementarity. In most cases, a mixture of monomers and antiparallel dimers provides better agreement with NMR data compared to the monomer and the parallel dimer. CG MD simulations allow predictions of helix orientation and dynamics and interpretation of QS data without requiring any assumption about the motional model.     

Persistence of phase coexistence in disaturated phosphatidylcholine monolayers at high surface pressures

Prior reports that the coexistence of the liquid-expanded (LE) and liquid-condensed (LC) phases in phospholipid monolayers terminates in a critical point have been compromised by experimental difficulties with Langmuir troughs at high surface pressures and temperatures. The studies reported here used the continuous interface of a captive bubble to minimize these problems during measurements of the phase behavior for monolayers containing the phosphatidylcholines with the four different possible combinations of palmitoyl and/or myristoyl acyl residues. Isothermal compression produced surface pressure-area curves for dipalmitoyl phosphatidylcholine (DPPC) that were indistinguishable from previously published data obtained with Langmuir troughs. During isobaric heating, a steep increase in molecular area corresponding to the main LC-LE phase transition persisted for all four compounds to 45 mN/m, at which collapse of the LE phase first occurred. No other discontinuities to suggest other phase transitions were apparent. Isobars for DPPC at higher pressures were complicated by collapse of the monolayer, but continued to show evidence up to 65 mN/m for at least the onset of the LC-LE transition. The persistence of the main phase transition to high surface pressures suggests that a critical point for these monolayers of disaturated phospholipids is either nonexistent or inaccessible at an air-water interface.     

The detrimental effect of serum albumin on the re-spreading of a dipalmitoyl phosphatidylcholine Langmuir monolayer is counteracted by a fluorocarbon gas

We have recently reported that fluorocarbon gases exhibit an effective fluidizing effect on Langmuir monolayers of dipalmitoyl phosphatidylcholine (DPPC), preventing them from crystallizing up to surface pressures of approximately 40 mN m(-1), i.e. well above the DPPC's equilibrium surface pressure. We now report that gaseous perfluorooctyl bromide (gPFOB) promotes the re-spreading of DPPC Langmuir monolayers compressed on a bovine serum albumin (BSA)-containing sub-phase. The latter protein is known to maintain a concentration-dependent surface pressure that can exceed the re-spreading pressure of collapsed monolayers. This phenomenon was proposed to be responsible for lung surfactant inactivation. Compression/expansion isotherms and fluorescence microscopy experiments were carried out to assess the monolayers' physical state. We have found that, during expansion under gPFOB-containing air, the surface pressure of a DPPC monolayer on a BSA-containing sub-phase decreased to much lower values than when the DPPC monolayer was expanded in the presence of BSA under air ( approximately 0 mN m(-1) vs. approximately 7.5 mN m(-1) at 120 A(2), respectively). Moreover, fluorescence images showed that, during expansion, the BSA-coupled DPPC monolayers, in contact with gPFOB, remained in the liquid-expanded state for surface pressures lower than 10 mN m(-1), whereas they were in a liquid-condensed semi-crystalline state, even at large molecular areas (120 A(2)), when expanded under air. The re-incorporation of the PFOB molecules in the DPPC monolayer during expansion thus competes with the re-incorporation of BSA, thus preventing the latter from penetrating into the DPPC monolayer. We suggest that combinations of DPPC and a fluorocarbon gas may be useful in the treatment of lung conditions resulting from a deterioration of the native lung surfactant function due to plasma proteins, such as in the acute respiratory distress syndrome.     

Interactions between two sheets of a bilayer membrane and its internal lateral pressure

We have modelled a phospholipid bilayer as two monolayer sheets which interact with each other by a coupling which depends upon the states of the lipid hydrocarbon chains in each sheet. We make use of a model (Georgallas and Pink 1982a) and its parameters, already used to study monolayer phase changes at the LC-LE transition, in order to study the lipid main transition. Although the monolayer coexistence curve can be calculated exactly, we have made use of high-temperature series expansions to calculate the critical point of the bilayer. We also present the results of computer simulations on triangular lattices for the pressure-area isotherms. We find: (i) the interaction between the sheets of a DPPC bilayer is about 1.5–2% of the maximum interaction within the plane of each sheet; (ii) the internal lateral pressure of a DPPC bilayer is about 30.5 dyne/cm; (iii) the bilayer transition enthalpy depends sensitively upon the coupling between the sheets. Should this coupling vary from sample to sample (due, possibly, to its preparation) then very different values of transition enthalpy may be measured. (iv) We present a rough rule-of-thumb for estimating the internal lateral pressure of a bilayer from a knowledge of the corresponding monolayer pressure-area isotherms.Abbreviations LC-LE liquid condensed — liquid expanded - DPPC dipalmitoylphosphatidylcholine - Q transition enthalpy Work supported in part by the Natural Sciences and Engineering Research Council of Canada     

The liquid condensed diffusional transition of dipalmitoylphosphoglycerocholine in monolayers

The fine details of the phase transition of dipalmitoylphosphoglycerocholine (DPPC) monolayers at air/NaCl solution interfaces were investigated at 21 +/- 1 degrees C by using the fluorescence after photobleaching technique employing 12-(9-anthroyloxy)stearic acid as fluorescent probe. The mode of compression of the monolayer (i.e., continuous compression or successive additions of the lipid at fixed area) together with the ionic strength of the subphase (0.1 or 1.0 M NaCl) were particularly studied. The photobleaching results show that the lateral diffusion coefficient of the probe molecules decreases drastically within the liquid-condensed phase, i.e., from the end of the liquid-expanded-liquid-condensed phase transition to the beginning of the solid-condensed phase. The molecular areas at which the phase transition occurs under the various experimental conditions, together with a parallel analysis of the hydration states and related molecular areas of the DPPC molecules in multilayers, strongly suggest that the steric hindrance associated with the hydration water of the polar head of DPPC molecules in the monolayer is responsible for the drastic decrease in diffusion coefficient in the liquid-condensed phase. Furthermore, the fluorescence characteristics of the probe molecules also show that, together with the aforementioned reorganization of the polar head, a structural reorganization of the aliphatic chains of the lipid molecules also takes place in the liquid-condensed phase. The liquid-condensed phase therefore appears as a transition region from liquid to solid phases in which the lipid molecules present a significant decrease in their lateral diffusion related to a structural reorganization of both their polar and aliphatic components.     

Mitochondrial creatine kinase interaction with heterogeneous monolayers: Effect on lipid lateral organization

Our study highlights the tight relationship between protein binding to monolayers and the phase-state of the phospholipids. Interaction of mitochondrial creatine kinase with phospholipidic membranes was analysed using a two-phase monolayer system containing anionic phospholipids under chain mismatch conditions. Monolayers were made up of mixtures of DMPC/DPPG or DPPC/DMPG containing 40% negatively charged phospholipids which is approximately the negative charge content of the mitochondrial inner membrane. Langmuir isotherms of these monolayers showed that they underwent a phase transition from a liquid expanded state to a liquid-condensed phase at about 2 mN/m and 5 mN/m respectively. Interface morphology modifications caused by injection of mtCK under these monolayers at low or high surface pressure were monitored by Brewster angle microscopy. This work provides evidence that the presence at the air/water interface of discrete domains with increased charge density, may lead to difference in partition of soluble proteins such as mtCK, interacting with the lipid monolayer. Conversely these proteins may help to organize charged phospholipid domains in a membrane.     

Condensing and fluidizing effects of ganglioside GM1 on phospholipid films

Mixed monolayers of the ganglioside G_M1 and the lipid dipalmitoylphosphatidlycholine (DPPC) at air-water and solid-air interfaces were investigated using various biophysical techniques to ascertain the location and phase behavior of the ganglioside molecules in a mixed membrane. The effects induced by G_M1 on the mean molecular area of the binary mixtures and the phase behavior of DPPC were followed for G_M1 concentrations ranging from 5 to 70 mol %. Surface pressure isotherms and fluorescence microscopy imaging of domain formation indicate that at low concentrations of G_M1 (<25 mol %), the monolayer becomes continually more condensed than DPPC upon further addition of ganglioside. At higher G_M1 concentrations (>25 mol %), the mixed monolayer becomes more expanded or fluid-like. After deposition onto a solid substrate, atomic force microscopy imaging of these lipid monolayers showed that G_M1 and DPPC pack cooperatively in the condensed phase domain to form geometrically packed complexes that are more ordered than either individual component as evidenced by a more extended total height of the complex arising from a well-packed hydrocarbon tail region. Grazing incidence x-ray diffraction on the DPPC/G_M1 binary mixture provides evidence that ordering can emerge when two otherwise fluid components are mixed together. The addition of G_M1 to DPPC gives rise to a unit cell that differs from that of a pure DPPC monolayer. To determine the region of the G_M1 molecule that interacts with the DPPC molecule and causes condensation and subsequent expansion of the monolayer, surface pressure isotherms were obtained with molecules modeling the backbone or headgroup portions of the G_M1 molecule. The observed concentration-dependent condensing and fluidizing effects are specific to the rigid, sugar headgroup portion of the G_M1 molecule.