Comprehensive Study of the Liquid Expanded-Liquid Condensed Phase Transition in 1,2-Dimyristoyl-sn-glycero-3-phospho-l-serine Monolayers: Surface Pressure, Volta Potential, X-ray Reflectivity, and Molecular Dynamics Modeling.

An integrated approach is applied to reveal fine changes in the surface-normal structure of 1,2-dimyristoyl-sn-glycero-3-phospho-l-serine (DMPS) monolayers at the air-lipid-water interface occurring in a liquid expanded (LE)-liquid condensed (LC) transition. The combination of the Langmuir monolayer technique, X-ray reflectometry, and molecular dynamics (MD) modeling provides new insight into the molecular nature of electrostatic phenomena in different stages of lipid compression. A homemade setup with a laboratory X-ray source (λ = 1.54 Å) offers a nondestructive way to reveal the structural difference between the LE and LC phases of the lipid. The electron density profile in the direction normal to the interface is recovered from the X-ray reflectivity data with the use of both model-independent and model-based approaches. MD simulations of the DMPS monolayer are performed for several areas per lipid using the all-atom force field. Using the conventional theory of capillary waves, a comparison is made between the electron density profiles reconstructed from the X-ray data and those calculated directly from MD modeling, which demonstrates remarkable agreement between the experiment and simulations for all selected lipid densities. This confirms the validity of the simulations and allows an analysis of the contributions of the hydrophobic tails and hydrated polar groups to the electron density profile and to the dipole component of the electric field at the interface. According to the MD data, the dependence of the Volta potential on the area per lipid in the monolayer has a different molecular nature below and above the phase transition. In the LE state of the monolayer, the potential is determined mostly by the oriented water molecules in the polar region of the lipid. In the LE-LC transition, these molecules are displaced to the bulk, and their effect on the Volta potential becomes insignificant compared with the contribution of the hydrophobic tails. The hydrophobic tails are highly ordered in the state of the liquid crystal so that their dipole moments entirely determine the growth of the potential upon compression up to the monolayer collapse.

Adsorption and photodynamic efficiency of meso-tetrakis(p-sulfonatophenyl)porphyrin on the surface of bilayer lipid membranes.

The adsorption and photodynamic efficiency of 5,10,15,20-tetrakis(p-sulfonatophenyl)porphyrin (H2TPPS4) on bilayer lipid membranes (BLM) have been studied. The adsorption of H2TPPS4 on BLM leads to rising of the potential drop on the membrane/water interface which has been detected either by the intramembrane field compensation (IFC) method, or as ζ-potential of liposomes measured by the dynamic light scattering method. The dependence of this potential on the concentration of H2TPPS4 and KCl in the solution can be described in the frame of Gouy-Chapman model of diffuse double layer assuming that the molecules of H2TPPS4 adsorb on the surface of BLM as an anions with four charged groups. The potential disappeared when the pH of solution decreased from 6 to 3 allowing the conclusion that the protonated forms of H2TPPS4 can not adsorb on the BLM probably due to change of conformation or aggregation of the molecules. The photodynamic efficiency of H2TPPS4 was evaluated by measuring the rate of damage of the targets - molecules of styryl dye (di-4-ANEPPS) by singlet oxygen generated under illumination on the surface of BLM. This rate was proportional to the surface density of H2TPPS4 molecules on the membrane which was determined from the change of surface charge of the membrane due to adsorption of the H2TPPS4. These results indicate that the di-4-ANEPPS molecules are damaged by singlet oxygen generated by monomers of H2TPPS4 molecules adsorbed on the membrane. The rate of oxidation of di-4-ANEPPS molecules adsorbed on the same (cis) side of the membrane together with the H2TPPS4 molecules was either the same or higher than that when di-4-ANEPPS molecules were adsorbed on opposite (trans) side. It indicates that the quenching of singlet oxygen by the di-4-ANEPPS molecules at cis side of the membrane was negligible, in contrast to our earlier study when singlet oxygen was generated by aluminum(III) phthalocyanines with one or two peripheral sulfo groups. The difference between these phthalocyanines and H2TPPS4 was explained by their different adsorption depth in the membrane.

Lateral stress profile and fluorescent lipid probes. FRET pair of probes that introduces minimal distortions into lipid packing.

The modern theory of lipid membrane structure incorporates the concept of lateral stress profile. The latter represents the forces that act on any solute inside the membrane. We used this concept to propose two lipid probes that introduce minimal distortions into the lipid bilayer packing: the surface pressure isotherms and volt-potentials of the pure and mixed (probe-containing) lipid monolayers are equal. The probes represent a FRET pair. They are applicable in lipid transfer and vesicle fusion experiments.

Contrasting behavior of zwitterionic and cationic polymers bound to anionic liposomes.

Zwitterionic polymers were prepared by quaternizing polyvinylpyridine (DP = 1100) with bromoacids (Br(CH2)nCOOH, where n = 1, 2, 3, and 5). The resulting polymers were then added to unilamellar liposomes composed of egg lecithin or dipalmitoylphosphatidylcholine admixed with 20 mol % of cardiolipin (a phospholipid with two negative charges). These systems were compared (along with polyethylvinylpyridinium chloride, a polycation) by light scattering, electrophoretic mobility, fluorescence, and high-sensitivity differential scanning calorimetry. The external zwitterionic polymers induce no flip-flop of cardiolipin from the inner leaflet to the outer leaflet as does the polycation. Aside from this similarity, the four zwitterionic polymers all behave differently from each other toward the anionic liposomes: (a) For n = 1, there is no detectable interaction between the polymer and the liposomes. (b) For n = 2, electrostatic attraction induces polymer-liposome association (reversed by the addition of NaCl) that maintains the original negative charge on the liposome. Aggregation of the liposomes accompanies polymer adsorption. (c) For n = 3, electrostatic binding also occurs along with aggregation. However, the binding is so strong that NaCl is unable to induce polymer/liposome dissociation. (d) For n = 5, there is polymer binding and NaCl-promoted dissociation but no substantial aggregation. These differences among the closely related polymers are discussed and analyzed in molecular terms.

Grafting of polylysine with polyethylenoxide prevents demixing of O-pyromellitylgramicidin in lipid membranes.

Both natural and synthetic polycations can induce demixing of negatively charged components in artificial and possibly in natural membranes. This process can result in formation of clusters (binding of several components to a polycation chain) and/or domains (aggregation of clusters and formation of a separate phase enriched in some particular component). In order to distinguish between these two phenomena, a model lipid membrane system containing ion channels, formed by a negatively charged peptide, O-pyromellitylgramicidin, and polycations of different structures was used. Microelectrophoresis of liposomes, changes in boundary potential of planar bilayers, the shape of compression curves and potentials of lipid and lipid/peptide monolayers were used to monitor the electrostatic factors in polymer adsorption to the membrane and peptide-polymer interactions. The synthesized PEO-grafted polylysine, PLL-PEO20000, did not induce peptide demixing monitored by stabilization of the gramicidin channels, in contrast to parent polylysine (PLL). Both polymers were shown to bind effectively to negatively charged liposomes and lipid monolayers, suggesting that the ineffectiveness of PLL-PEO20000 was not due to reduction of its binding. It was hypothesized that PLL-PEO20000 could not induce domain formation due to steric hindrance of long PEO chains preventing lateral fusion of clusters. Another copolymer, PLL-PEO4000, having four PEO chains of 4000 Da, exhibited intermediate effect between PLL and PLL-PEO20000, which shows the importance of the copolymer architecture for the effect on the lateral distribution of OPg channels. The model system can be relevant to regulation of lateral organization of ion channels and other components in natural membrane systems.