Purified myelin lipids display a critical mixing point at low surface pressure

Lipids extracted from Purified Myelin Membranes (LPMM) were spread as monomolecular films at the air/aqueous interface. The films were visualized by Brewster Angle Microscopy (BAM) at different lateral pressures (π) and ionic environments. Coexistence of Liquid-Expanded (LE) and cholesterol-enriched (CE) rounded domains persisted up to π ≈ 5 mN/m but the monolayers became homogeneous at higher surface pressures. Before mixing, the domains distorted to non-rounded domains. We experimentally measured the line tension (λ) for the lipid monolayers at the domain borders by a shape relaxation technique using non-homogeneous electric fields. Regardless of the subphase conditions, the obtained line tensions are of the order of pN and tended to decrease as lateral pressure increased toward the mixing point. From the mean square displacement of nested trapped domains, we also calculated the dipole density difference between phases (μ). A non-linear drop was detected in this parameter as the mixing point is approached. Here we quantitively evaluated the π-dependance of both parameters with proper power laws in the vicinity of the critical mixing surface pressure, and the exponents showed to be consistent with a critical phenomenon in the two-dimensional Ising universality class. This idea of bidimensionality was found to be compatible only for simplified lipidic systems, while for whole myelin monolayers, that means including proteins, no critical mixing point was detected.Finally, the line tension values were related with the thickness differences between phases (Δt) near the critical point.     

Effect of calcium and temperature on mixed lipid-valinomycin monolayers. A comparison of glycosphingolipids (ganglioside GT1b, sulphatides) and phosphatidylcholine

The influence of calcium and temperature on pure lipid (bovine brain PC, sulphatides, ganglioside GT1b), valinomycin and mixed lipid-valinomycin monolayers at the air/water interface was studied. In mixed films, evidence was found that the two components were miscible. On the other hand, at higher surface pressures, phase separation occurs in the cases of PC and sulphatides. Measuring the area requirement and the collapse pressure the stability of both lipid and the peptide was increased in particular due to ganglioside-valinomycin interaction. The addition of 10(-5) M calcium into the subphase at 20 and 37 degrees C and surface pressures of 10 and 20 mN/m led to a condensing effect in ganglioside mixtures, with formation of aggregates as indicated also by the nearly ideal behaviour of two component monolayers.     

Rapid compression transforms interfacial monolayers of pulmonary surfactant

Films of pulmonary surfactant in the lung are metastable at surface pressures well above the equilibrium spreading pressure of 45 mN/m but commonly collapse at that pressure when compressed in vitro. The studies reported here determined the effect of compression rate on the ability of monolayers containing extracted calf surfactant at 37 degrees C to maintain very high surface pressures on the continuous interface of a captive bubble. Increasing the rate from 2 A(2)/phospholipid/min (i.e., 3% of (initial area at 40 mN/m)/min) to 23%/s produced only transient increases to 48 mN/m. Above a threshold rate of 32%/s, however, surface pressures reached > 68 mN/m. After the rapid compression, static films maintained surface pressures within +/- 1 mN/m both at these maximum values and at lower pressures following expansion at < 5%/min to > or = 45 mN/m. Experiments with dimyristoyl phosphatidylcholine at 37 degrees C produced similar results. These findings indicate that compression at rates comparable to values in the lungs can transform at least some phospholipid monolayers from a form that collapses readily at the equilibrium spreading pressure to one that is metastable for prolonged periods at higher pressures. Our results also suggest that transformation of surfactant films can occur without refinement of their composition.     

Differential effects of surfactant protein A on regional organization of phospholipid monolayers containing surfactant protein B or C

Epifluorescence microscopy combined with a surface balance was used to study monolayers of dipalmitoylphosphatidylcholine (DPPC)/egg phosphatidylglycerol (PG) (8:2, mol/mol) plus 17 wt % SP-B or SP-C spread on subphases containing SP-A in the presence or absence of 5 mM Ca(2+). Independently of the presence of Ca(2+) in the subphase, SP-A at a bulk concentration of 0.68 microg/ml adsorbed into the spread monolayers and caused an increase in the molecular areas in the films. Films of DPPC/PG formed on SP-A solutions showed a pressure-dependent coexistence of liquid-condensed (LC) and liquid-expanded (LE) phases. Apart from these surface phases, a probe-excluding phase, likely enriched in SP-A, was seen in the films between 7 mN/m < or = pi < or = 20 mN/m. In monolayers of SP-B/(DPPC/PG) spread on SP-A, regardless of the presence of calcium ions, large clusters of a probe-excluding phase, different from probe-excluding lipid LC phase, appeared and segregated from the LE phase at near-zero surface pressures and coexisted with the conventional LE and LC phases up to approximately 35 mN/m. Varying the levels of either SP-A or SP-B in films of SP-B/SP-A/(DPPC/PG) revealed that the formation of the probe-excluding clusters distinctive for the quaternary films was influenced by the two proteins. Concanavalin A in the subphase could not replace SP-A in its ability to modulate the textures of films of SP-B/(DPPC/PG). In films of SP-C/SP-A/(DPPC/PG), in the absence of calcium, regions consisting of a probe-excluding phase, likely enriched in SP-A, were detected at surface pressures between 2 mN/m and 20 mN/m in addition to the lipid LE and LC phases. Ca(2+) in the subphase appeared to disperse this phase into tiny probe-excluding particles, likely comprising Ca(2+)-aggregated SP-A. Despite their strikingly different morphologies, the films of DPPC/PG that contained combinations of SP-B/SP-A or SP-C/SP-A displayed similar distributions of LC and LE phases with LC regions occupying a maximum of 20% of the total monolayer area. Combining SP-A and SP-B reorganized the morphology of monolayers composed of DPPC and PG in a Ca(2+)-independent manner that led to the formation of a separate potentially protein-rich phase in the films.     

The role of surfactant proteins in DPPC enrichment of surface films

A pressure-driven captive bubble surfactometer was used to determine the role of surfactant proteins in refinement of the surface film. The advantage of this apparatus is that surface films can be spread at the interface of an air bubble with a different lipid/protein composition than the subphase vesicles. Using different combinations of subphase vesicles and spread surface films a clear correlation between dipalmitoylphosphatidylcholine (DPPC) content and minimum surface tension was observed. Spread phospholipid films containing 50% DPPC over a subphase containing 50% DPPC vesicles did not form stable surface films with a low minimum surface tension. Addition of surfactant protein B (SP-B) to the surface film led to a progressive decrease in minimum surface tension toward 1 mN/m upon cycling, indicating an enrichment in DPPC. Surfactant protein C (SP-C) had no such detectable refining effect on the film. Surfactant protein A (SP-A) had a positive effect on refinement when it was present in the subphase. However, this effect was only observed when SP-A was combined with SP-B and incubated with subphase vesicles before addition to the air bubble containing sample chamber. Comparison of spread films with adsorbed films indicated that refinement induced by SP-B occurs by selective removal of non-DPPC lipids upon cycling. SP-A, combined with SP-B, induces a selective adsorption of DPPC from subphase vesicles into the surface film. This is achieved by formation of large lipid structures which might resemble tubular myelin.     

Structure of SP-B/DPPC mixed films studied by neutron reflectometry

The structures of films of pulmonary surfactant protein B (SP-B) and mixtures of SP-B and dipalmitoylphosphatidylcholine (DPPC) at the air/water interface have been studied by neutron reflectometry and Langmuir film balance methods. From the film balance studies, we observe that the isotherms of pure DPPC and SP-B/DPPC mixtures very nearly overlay one another at very high pressures, suggesting that the SP-B is being excluded from the film. The use of multiple contrasts with neutron reflectometry at a range of surface pressures has enabled the mixing and squeeze out of the DPPC and SP-B mixtures to be studied. We can identify the SP-B component of the interfacial structure and its position as a function of surface pressure. The mixtures are initially a homogeneous layer at low surface pressures. At higher surface pressures, the SP-B is squeezed out of the lipid layer into the subphase, with the first signs detected at 30 mN m⁻¹. At 50 mN m⁻¹, the subphase is almost completely excluded from the DPPC layer, with the SP-B content significantly reduced. Only a small amount of DPPC appears to be associated with the squeezed out SP-B.     

Adsorption of pulmonary surfactant protein SP-A to monolayers of phospholipids containing hydrophobic surfactant protein SP-B or SP-C: potential differential role for tertiary interaction of lipids, hydrophobic proteins, and SP-A

Surface balance techniques were used to study the interactions of surfactant protein SP-A with monolayers of surfactant components preformed at the air-water interface. SP-A adsorption into the monolayers was followed by monitoring the increase in the surface pressure Deltapi after injection of SP-A beneath the films. Monolayers of dipalmitoylphosphatidylcholine (DPPC):egg phosphatidylglycerol (PG) (8:2, mol/mol) spread at initial surface pressure pi(i) = 5 mN/m did not promote the adsorption of SP-A at a subphase concentration of 0.68 microg/mL as compared to its adsorption to the monolayer-free surface. Surfactant proteins, SP-B or SP-C, when present in the films of DPPC:PG spread at pi(i) = 5 mN/m, enhanced the incorporation of SP-A in the monolayers to a similar extent; the Deltapi values being dependent on the levels of SP-B or SP-C, 3-17 wt %, in the lipid films. Calcium in the subphase did not affect the intrinsic surface activity of SP-A but reduced the Deltapi values produced by the adsorption of the protein to all the preformed films independently of their compositions and charges. The divalent ions likely modified the interaction of SP-A with the monolayers through their effects on the conformation, self-association, and charge state of SP-A. Values of Deltapi produced by adsorption of SP-A to the films of DPPC:PG with or without SP-B or SP-C were a function of the initial surface pressure of the films, pi(i). In the range of pressures 5 相似文献   

Spectrin maintains the lateral order in phosphatidylserine monolayers

We investigate the effect of the skeletal protein spectrin on the lateral order in dipalmitoyl phosphatidylserine monolayers spread on aqueous surfaces using grazing incidence X-ray diffraction. Without spectrin, the condensed lipid monolayer exhibits two-dimensional hexagonal packing, characterized by monotonic decrease in the d-spacing and increase in the degree of order with increasing surface pressure between 17 and 36 mN/m. Addition of spectrin to the aqueous subphase at high pressures preserves the monolayers structural parameters unchanged from 36 to 25 mN/m. These results demonstrate for the first time that spectrin could participate in sustaining the two-dimensional order in lipid domains through a direct interaction with phosphatidylserine species.     

Interaction of C-phycocyanin with lipid monolayers under nitrogen and in the presence of air

The surface interaction of C-phycocyanin with lipids was studied using the monolayer technique. The surface activity of the protein was found to be higher at the lipid-water interface than at the nitrogen-water interface, particularly at high surface pressures of the lipid monolayer. The maximum initial surface pressures beyond which phycocyanin could not penetrate the dipalmitoylphosphatidylcholine and monogalactosyldiglycerol monolayers were 27 and 30 mN m-1, respectively. Below these values the protein demonstrated preferential interaction with the monogalactosyldiglycerol monolayer. The surface properties of the unfolded protein at pH 2.5 at the lipid-water interface were compared with those of the protein at pH 7.0. Higher affinity of the three-dimensional structure of the protein to lipid monolayers was observed, in particular by high subphase protein concentration. When the lipid films were subjected to oxidation stress by exposure to air, the surface properties of C-phycocyanin and dipalmitoylphosphatidylcholine were not greatly affected but the surface activity of monogalactosyldiacylglycerol was reduced dramatically by autoxidation. The oxidation of monogalactosyldiacylglycerol could not be prevented by the introduction of C-phycocyanin molecules at the lipid-water interface.     

Discrepancy between phase behavior of lung surfactant phospholipids and the classical model of surfactant function

The studies reported here used fluorescence microscopy and Brewster angle microscopy to test the classical model of how pulmonary surfactant forms films that are metastable at high surface pressures in the lungs. The model predicts that the functional film is liquid-condensed (LC) and greatly enriched in dipalmitoyl phosphatidylcholine (DPPC). Both microscopic methods show that, in monolayers containing the complete set of phospholipids from calf surfactant, an expanded phase persists in coexistence with condensed domains at surface pressures approaching 70 mN/m. Constituents collapsed from the interface above 45 mN/m, but the relative area of the two phases changed little, and the LC phase never occupied more than 30% of the interface. Calculations based on these findings and on isotherms obtained on the continuous interface of a captive bubble estimated that collapse of other constituents increased the mol fraction of DPPC to no higher than 0.37. We conclude that monolayers containing the complete set of phospholipids achieve high surface pressures without forming a homogeneous LC film and with a mixed composition that falls far short of the nearly pure DPPC predicted previously. These findings contradict the classical model.     

Ca2+-induced lateral phase separation in phosphatidic acid/phosphatidylcholine monolayers as revealed by fluorescence microscopy

Phase separation in mixed monolayers of phosphatidylcholine (PC) and pyrene-labeled phosphatidic acid (PA) was observed by fluorescence microscopy on an air/water interface as a function of subphase Ca2+ concentration and lateral packing pressure of the film. Below 45 mN m-1 and in the absence of Ca2+ no indications of phase immiscibility were observed. Addition of 1 mM Ca2+ caused extensive phase separation, which was evident immediately after spreading of the film. Further increase in Ca2+ concentration up to 30 mM increased the pyrene excimer intensity of the separated phosphatidic acid enriched domains. In the presence of Ca2+ (1-30 mM) and at surface pressures below 10 mN m-1 phase separation was always evident. However, as surface pressure exceeded 10 mN m-1, mixing of PC and PA occurred. Upon decompression of the film, phase separation reappeared at surface pressures close to 10 mN m-1. The surface textures of the film before and after the compression and subsequent relaxation were different. Inclusion of 30 mol% cholesterol increased the number and decreased the size of the PA domains. In films containing 50 mol% cholesterol no phase separation could be detected at the resolution available.     

Flunitrazepam-membrane non-specific binding and unbinding: two pathways with different energy barriers

The effect of molecular packing on flunitrazepam's ability to interact with bio-membranes was studied using dipalmitoylphosphatidylcholine monomolecular layers at the air-water interface as a model membrane. Flunitrazepam penetrated from the subphase into monolayers at lateral pressures below 44.8 mN/m and induced their concentration-dependent expansion. As inferred from the values of compressibility modulus, the elasticity of the liquid-condensed phase decreased in the presence of flunitrazepam. Although this drug hardly penetrated into high-packed monolayers, it was easily incorporated in the low-packed ones at an extent sufficient to reach the partition equilibrium. Below a molecular area of 75 A(2), contrary to what would be expected, the drug surface concentration increased as a function of surface pressure, suggesting that after its penetration in disordered phases, it became energetically or physically trapped in newly-formed liquid condensed clusters. The phenomenon of flunitrazepam penetration and release would have different energy barriers depending on the membrane phase-state.     

Ubiquinones have surface-active properties suited to transport electrons and protons across membranes

Surface-active properties of ubiquinones and ubiquinols have been investigated by monomolecular-film techniques. Stable monolayers are formed at an air/water interface by the fully oxidized and reduced forms of the coenzyme; collapse pressures and hence stability of the films tend to increase with decreasing length of the isoprenoid side chain and films of the reduced coenzymes are more stable than those of their oxidized counterparts. Ubiquinone with a side chain of two isoprenoid units does not form stable monolayers at the air/water interface. Mixed monolayers of ubiquinol-10 or ubiquinone-10 with 1,2-dimyristoyl phosphatidylcholine, soya phosphatidylcholine and diphosphatidylglycerol do not exhibit ideal mixing characteristics. At surface pressures less than the collapse pressure of pure ubiquinone-10 monolayers (approx. 12mN.m(-1)) the isoprenoid chain is located substantially within the region occupied by the fatty acyl residues of the phospholipids. With increasing surface pressure the ubiquinones and their fully reduced equivalents are progressively squeezed out from between the phospholipid molecules until, at a pressure of about 35mN.m(-1), the film has surface properties consistent with that of the pure phospholipid monolayer. This suggests that the ubiquinone(ol) forms a separate phase overlying the phospholipid monolayer. The implications of this energetically poised situation, where the quinone(ol) is just able to penetrate the phospholipid film, are considered in terms of the function of ubiquinone(ol) as electron and proton carriers of energy-transducing membranes.     

Adsorption of apolipoprotein A-IV to phospholipid monolayers spread at the air/water interface. A model for its labile binding to high density lipoproteins.

The mechanisms that mediate the labile binding of apolipoprotein A-IV (apoA-IV) to high density lipoproteins (HDL) are not known. We therefore used a surface balance and surface radioactivity detector to investigate the adsorption of apoA-IV to egg phosphatidylcholine monolayers spread at the air/water interface. ApoA-IV bound rapidly and reversibly to phospholipid monolayers and generated a maximum increase in surface pressure of 19 millinewtons (mN)/m at a subphase concentration of 2 x 10(-5) g/dl. Binding decreased linearly with increasing initial surface pressure; at pressures greater than 28-29 mN/m, apoA-IV could no longer penetrate the lipid monolayer. The area occupied by the amino acid residues in apoA-IV reached an unusually low limiting molecular area of 10-12 A2/residue at surface saturation. The surface pressure of native HDL3 was calculated to be 33 mN/m, and it rapidly decreased with the action of lecithin:cholesterol acyltransferase on the particle surface. We conclude that the surface activity of apoA-IV is lower than that of any other human apolipoprotein; its binding and surface conformation are particularly sensitive to pressure; and at saturation, a significant portion of the molecule is excluded from the interface. The exclusion pressure of apoA-IV may be only slightly lower than the surface pressure of HDL; in vivo, the action of lecithin:cholesterol acyltransferase and lipid transfer proteins may cause the HDL3 surface pressure to oscillate about a narrow range that spans the exclusion pressure of apoA-IV. The resultant labile association of apoA-IV and HDL may be of central importance to its role in lipoprotein metabolism.     

Phase transitions in films of lung surfactant at the air-water interface.

Pulmonary surfactant maintains a putative surface-active film at the air-alveolar fluid interface and prevents lung collapse at low volumes. Porcine lung surfactant extracts (LSE) were studied in spread and adsorbed films at 23 +/- 1 degrees C using epifluorescence microscopy combined with surface balance techniques. By incorporating small amounts of fluorescent probe 1-palmitoyl-2-nitrobenzoxadiazole dodecanoyl phosphatidylcholine (NBD-PC) in LSE films the expanded (fluid) to condensed (gel-like) phase transition was studied under different compression rates and ionic conditions. Films spread from solvent and adsorbed from vesicles both showed condensed (probe-excluding) domains dispersed in a background of expanded (probe-including) phase, and the appearance of the films was similar at similar surface pressure. In quasistatically compressed LSE films the appearance of condensed domains occurred at a surface pressure (pi) of 13 mN/m. Such domains increased in size and amounts as pi was increased to 35 mN/m, and their amounts appeared to decrease to 4% upon further compression to 45 mN/m. Above pi of 45 mN/m the LSE films had the appearance of filamentous materials of finely divided dark and light regions, and such features persisted up to a pi near 68 mN/m. Some of the condensed domains had typical kidney bean shapes, and their distribution was similar to those seen previously in films of dipalmitoylphosphatidylcholine (DPPC), the major component of surfactant. Rapid cyclic compression and expansion of LSE films resulted in features that indicated a possible small (5%) loss of fluid components from such films or an increase in condensation efficiency over 10 cycles. Calcium (5 mM) in the subphase of LSE films altered the domain distribution, decreasing the size and increasing the number and total amount of condensed phase domains. Calcium also caused an increase in the value of pi at which the maximum amount of independent condensed phase domains were observed to 45 mN/m. It also induced formation of large amounts of novel, nearly circular domains containing probe above pi of 50 mN/m, these domains being different in appearance than any seen at lower pressures with calcium or higher pressures in the absence of calcium. Surfactant protein-A (SP-A) adsorbed from the subphase onto solvent-spread LSE films, and aggregated condensed domains in presence of calcium. This study indicates that spread or adsorbed lung surfactant films can undergo expanded to condensed, and possibly other, phase transitions at the air-water interface as lateral packing density increases. These phase transitions are affected by divalent cations and SP-A in the subphase, and possibly by loss of material from the surface upon cyclic compression and expansion.     

The interaction of cytochrome c with monolayers of phosphatidylethanolamine

1. The interaction between [(14)C]carboxymethylated cytochrome c and monolayers of egg phosphatidylethanolamine at the air/water interface has been investigated by measurements of surface radioactivity, pressure and potential. 2. On adding (14)C-labelled cytochrome c to the subphase under monolayers with a surface pressure below 24dynes/cm. there was an initial surface pressure increment as the protein penetrated, followed by an adsorption that could be detected only by a continued increase in the surface radioactivity. 3. Above film pressures of 24dynes/cm. only adsorption was observed, i.e. an increment in surface radioactivity with none in surface pressure. 4. The changes in surface parameters with penetration of cytochrome c added to the subphase were indirectly proportional to the initial pressure of the monolayer. With hydrogenated phosphatidylethanolamine the constant of proportionality was increased but penetration again ceased at 24dynes/cm. 5. On compressing a phosphatidylethanolamine film containing penetrated cytochrome c to 40dynes/cm. only a proportion of the protein was ejected on a subphase of 10mm-sodium chloride, whereas on a subphase of m-sodium chloride nearly all the protein was lost. 6. With both penetration and adsorption only a small proportion of the added cytochrome c interacted with the phospholipid films, and initially the amount bound was proportional to the added protein concentration. There was no evidence of a stoicheiometric relationship between the protein and phospholipid or the build-up of multilayers. The bonded protein was not released by removing cytochrome c from the subphase. 7. The addition of m-sodium chloride to the subphase delays the rate of protein penetration into low-pressure films, but the final surface-pressure increment is not appreciably decreased. In contrast, m-sodium chloride almost completely stops adsorption on to films at all pressures. 8. When sodium chloride is added to the subphase below cytochrome c adsorbed to monolayers at high pressures, so that the final concentration is 1m, only a proportion of the protein is desorbed and this decreases as the time of the interaction increases. This indicates that adsorption is initially electrostatic, followed by the formation of non-ionic bonds. 9. Alteration of the subphase pH under a high-pressure film leads to a steady increase in adsorption from pH3 to 8.5 followed by a rapid fall to zero adsorption at pH11. 10. The penetration into phospholipid monolayers at 10dynes/cm. shows a rate that is consistent with the relative electrostatic status of the two components of the interaction as the subphase pH is varied between 3 and 10.5. The final equilibrium penetration shows a pronounced peak in the increments of surface pressure at pH9.0 although a similar peak is not observed in the surface radioactivity. This indicates that more residues of the protein are penetrating into the film at about this pH. 11. Determinations were made of the electrophoretic mobilities of phosphatidylethanolamine particles both alone and after interaction with cytochrome c. 12. The electrophoretic mobilities of cytochrome c adsorbed on lipid particles showed an isoelectric point below that of cytochrome c. This and the observations on the monolayers suggest that, with cytochrome c, protein-protein interactions are weak compared with other proteins.     

Influence of stearic acid monolayers upon the procaine adsorption from underlying alkaline aqueous solutions

Adsorption of procaine at the air/water interface and its penetration into stearic acid monolayers from aqueous subphase of pH 8 are studied by measuring surface tension of aqueous procaine solutions and by recording surface pressure vs. mean molecular area curves for stearic acid monolayers spread onto procaine solutions of different concentrations. The amount of procaine in the interface is derived by means of Gibbs' equation. Results are compared to those obtained earlier at pH 2 and on unbuffered subphases. With increasing pH an increasing procaine adsorption and procaine penetration is observed. This phenomenon is interpreted in terms of protolytic equilibria in which participate both surfactants procaine and stearic acid.     

Nonequilibrium behavior in supported lipid membranes containing cholesterol

We investigate lateral organization of lipid domains in vesicles versus supported membranes and monolayers. The lipid mixtures used are predominantly DOPC/DPPC/Chol and DOPC/BSM/Chol, which have been previously shown to produce coexisting liquid phases in vesicles and monolayers. In a monolayer at an air-water interface, these lipids have miscibility transition pressures of approximately 12-15 mN/m, which can rise to 32 mN/m if the monolayer is exposed to air. Lipid monolayers can be transferred by Langmuir-Sch?fer deposition onto either silanized glass or existing Langmuir-Blodgett supported monolayers. Micron-scale domains are present in the transferred lipids only if they were present in the original monolayer before deposition. This result is valid for transfers at 32 mN/m and also at lower pressures. Domains transferred to glass supports differ from liquid domains in vesicles because they are static, do not align in registration across leaflets, and do not reappear after temperature is cycled. Similar static domains are found for vesicles ruptured onto glass surfaces. Although supported membranes on glass capture some aspects of vesicles in equilibrium (e.g., gel-liquid transition temperatures and diffusion rates of individual lipids), the collective behavior of lipids in large liquid domains is poorly reproduced.     

Molecular dynamics simulations of lung surfactant lipid monolayers

Pulmonary surfactant provides for a lipid rich film at the lung air-water interface, which prevents alveolar collapse at the end of expiration. The films are likely enriched in the major surfactant component dipalmitoylphosphatidylcholine (DPPC), which, due to its saturated fatty acid chains, can withstand high surface pressures up to 70 mN/m, thereby reducing surface tension in that interface to very low values (close to 1 mN/m). Despite many experimental measurements in situ, as well as in vitro for native lung surfactant films, the exact mechanism by which other fluid lipid components of surfactant, in combination with surfactant proteins, allow for such low surface tension values to be reached is not well understood. We have performed molecular dynamics simulation of films composed of DPPC alone and in mixtures with other fluid and acidic lipid components of surfactant at the high densities relevant to the low surface tension regime. 10-50 ns simulations were performed with the software GROMACS, with 40-64 lipids molecules plus water, using 5 different lipid compositions and 7 different areas per lipid. The primary focus was to learn how differences in lipid composition affect the response of the monolayer to compression, such as the development of curvature or the loss of lipids to the exterior of the monolayer. The systems studied exhibit features of two of the major schools of thought of lung surfactant mechanisms, in that although unsaturated lipids did not appear to prevent the monolayers from achieving high surface pressure, POPG did appear to be selectively squeezed out of the DPPC/POPG monolayers at high lipid densities.     

Pulmonary surfactant proteins SP-B and SP-C in spread monolayers at the air-water interface: II. Monolayers of pulmonary surfactant protein SP-C and phospholipids.

The interaction of the hydrophobic pulmonary surfactant protein SP-C with dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG) and DPPC:DPPG (7:3, mol:mol) in spread monolayers at the air-water interface has been studied. At low concentrations of SP-C (about 0.5 mol% or 3 weight%protein) the protein-lipid films collapsed at surface pressures of about 70 mN.m-1, comparable to those of the lipids alone. At initial protein concentrations higher than 0.8 mol%, or 4 weight%, the isotherms displayed kinks at surface pressures of about 50 mN.m-1 in addition to the collapse plateaux at the higher pressures. The presence of less than 6 mol%, or 27 weight%, of SP-C in the protein-lipid monolayers gave a positive deviation from ideal behavior of the mean areas in the films. Analyses of the mean areas in the protein-lipid films as functions of the monolayer composition and surface pressure showed that SP-C, associated with some phospholipid (about 8-10 lipid molecules per molecule of SP-C), was squeezed out from the monolayers at surface pressures of about 55 mN.m-1. The results suggest a potential role for SP-C to modify the composition of the monolayer at the air-water interface in the alveoli.