Spermidine-Induced Attraction of Like-Charged Surfaces Is Correlated with the pH-Dependent Spermidine Charge: Force Spectroscopy Characterization.

The ubiquitous molecule spermidine is known for its pivotal roles in the contact mediation, fusion, and reorganization of biological membranes and DNA. In our model system, borosilicate beads were attached to atomic force microscopy cantilevers and used to probe mica surfaces to study the details of the spermidine-induced attractions. The negative surface charges of both materials were largely constant over the measured pH range of pH 7.8 to 12. The repulsion observed between the surfaces turned into attraction after the addition of spermidine. The attractive force was correlated with the degree of spermidine protonation, which changed from +3 to +1 over the measured pH range. The force was maximal at pH 7.8. To explain the observed pH and spermidine concentration dependence, two different theoretical approaches were used: a chemical model of the charge equilibrium of spermidine and Monte-Carlo simulations of the orientation of the rodlike spermidine molecules in the gap between the borosilicate and mica surfaces. Monte-Carlo simulations of the orientational ordering of the rodlike spermidine molecules suggested the induction of attractive interactions between the surfaces if the gap was bridged by the molecules. For larger gaps, the orientational distribution function of the spermidine molecules predicted a considerable degree of parallel attachment of the molecules to the surfaces, resulting in reduced effective surface charge densities of both surfaces, which reduced their electrostatic repulsion.

Influence of nanoparticle-membrane electrostatic interactions on membrane fluidity and bending elasticity.

The aim of this work is to investigate the effect of electrostatic interactions between the nanoparticles and the membrane lipids on altering the physical properties of the liposomal membrane such as fluidity and bending elasticity. For this purpose, we have used nanoparticles and lipids with different surface charges. Positively charged iron oxide (γ-Fe2O3) nanoparticles, neutral and negatively charged cobalt ferrite (CoFe2O4) nanoparticles were encapsulated in neutral lipid 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine and negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine lipid mixture. Membrane fluidity was assessed through the anisotropy measurements using the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene. Though the interaction of both the types of nanoparticles reduced the membrane fluidity, the results were more pronounced in the negatively charged liposomes encapsulated with positively charged iron oxide nanoparticles due to strong electrostatic attractions. X-ray photoelectron spectroscopy results also confirmed the presence of significant quantity of positively charged iron oxide nanoparticles in negatively charged liposomes. Through thermally induced shape fluctuation measurements of the giant liposomes, a considerable reduction in the bending elasticity modulus was observed for cobalt ferrite nanoparticles. The experimental results were supported by the simulation studies using modified Langevin-Poisson-Boltzmann model.

Influence of ionic strength and beta2-glycoprotein I concentration on agglutination of like-charged phospholipid membranes.

The effect of ionic strength on adhesion between negatively charged giant unilamellar vesicles induced by beta2-glycoprotein I (ß2-GPI) was studied experimentally and theoretically. Measuring the effective angle of contact between adhering vesicles indicated that the strength of adhesion between vesicles decreases with increasing ionic strength, and increases with concentration of ß2-GPI. In the theoretical part we focused on the study of the average orientation of ß2-GPI near the charged membrane and its role in mediating the attractive interactions between the vesicles. ß2-GPI proteins were modelled as rods with internal distribution of electric charge. The predictions of Monte Carlo simulations show orthogonal orientation of some of the membrane attached ß2-GPI in narrow gap between two vesicles. On the contrary, at larger distances between vesicles the proteins are parallelly attached to the membrane surface. A local minimum of the free energy corresponding to ß2-GPI-mediated adhesion of two neighbouring vesicles was predicted. The strength of adhesion was confirmed to decrease at high ionic strength.

Interaction between dipolar lipid headgroups and charged nanoparticles mediated by water dipoles and ions.

In this work, a theoretical model describing the interaction between a positively or negatively charged nanoparticle and neutral zwitterionic lipid bilayers is presented. It is shown that in the close vicinity of the positively charged nanoparticle, the zwitterionic lipid head groups are less extended in the direction perpendicular to the membrane surface, while in the vicinity of the negatively charged nanoparticle, the headgroups are more extended.This result coincides with the calculated increase in the osmotic pressure between the zwitterionic lipid surface and positively charged nanoparticle and the decrease of osmotic pressure between the zwitterionic lipid surface and the negatively charged nanoparticle.Our theoretical predictions agree well with the experimentally determined fluidity of a lipid bilayer membrane in contact with positively or negatively charged nanoparticles. The prospective significance of the present work is mainly to contribute to better understanding of the interactions of charged nanoparticles with a zwitterionic lipid bilayer, which may be important in the efficient design of the lipid/nanoparticle nanostructures (like liposomes with encapsulated nanoparticles), which have diverse biomedical applications, including targeted therapy (drug delivery) and imaging of cancer cells.

Monovalent ions and water dipoles in contact with dipolar zwitterionic lipid headgroups-theory and MD simulations.

The lipid bilayer is a basic building block of biological membranes and can be pictured as a barrier separating two compartments filled with electrolyte solution. Artificial planar lipid bilayers are therefore commonly used as model systems to study the physical and electrical properties of the cell membranes in contact with electrolyte solution. Among them the glycerol-based polar phospholipids which have dipolar, but electrically neutral head groups, are most frequently used in formation of artificial lipid bilayers. In this work the electrical properties of the lipid layer composed of zwitterionic lipids with non-zero dipole moments are studied theoretically. In the model, the zwitterionic lipid bilayer is assumed to be in contact with aqueous solution of monovalent salt ions. The orientational ordering of water, resulting in spatial variation of permittivity, is explicitly taken into account. It is shown that due to saturation effect in orientational ordering of water dipoles the relative permittivity in the zwitterionic headgroup region is decreased, while the corresponding electric potential becomes strongly negative. Some of the predictions of the presented mean-field theoretical consideration are critically evaluated using the results of molecular dynamics (MD) simulation.

Exocytotic fusion pore stability and topological defects in the membrane with orientational degree of ordering.

Regulated exocytosis is a process that strongly depends on the formation and stability of the fusion pore. It was indicated experimentally and theoretically that narrow and highly curved fusion pore may be stabilized by accumulation of anisotropic membrane components possessing orientational ordering. On the other hand, narrow fusion pore may also undergo repetitive opening and closing, disruption in the so called kiss and run process or become completely opened in the process of full fusion of the vesicle with the membrane. In this paper we attempt to elucidate the subtle interplay between the stabilizing and destabilizing processes in the fusion neck. A possible physical mechanism which may lead to disruption of the stable fusion pore or complete fusion of the vesicle with the membrane is discussed. It is indicated that topologically driven defects of the in-plane orientational membrane ordering in the region of the fusion pore may disrupt the fusion. Alternatively, it may facilitate repetitive opening and closing of the fusion pore or induce full fusion of the vesicle with the target membrane.

Elastic deformations in hexagonal phases studied by small-angle X-ray diffraction and simulations.

In this study we present experimental and theoretical results which concern the deviations from circularity of the pivotal plane in the inverse hexagonal phases (H(II)) of phospholipid self-assemblies. Due to packing constraints, the cross-section of the polar/apolar interface deviates from a circle, which we studied in minute detail by analysing small-angle X-ray diffraction data of dioleoyl-phosphatidylethanolamine (DOPE) and stearoyl-oleoyl-phosphatidylethanolamine (SOPE), respectively. On this structural basis, Monte Carlo (MC) simulated annealing variations of the free energy were carried out, both on the formation of the H(II)-phase and on the particular shape of the cross-section in the H(II)-phase. The equilibrium of the H(II)-phase pivotal plane contour and the corresponding values of the mean intrinsic curvature, H(m), and the hydrocarbon chain stiffness, τ, were determined from MC calculations. The results of these calculations were tested by solving the corresponding system of non-linear differential equations derived using variational calculus. Here our main aim is to predict the range of possible values of H(m) and τ. Comparing the measured structural data with predictions from MC calculations including lipid anisotropy, and accounting for the elastic deformations of the pivotal plane allowed us to determine a relationship between the bending deformation and stretching of hydrocarbon chains.

Echinophilic proteins stomatin, sorcin, and synexin locate outside gangliosideM1 (GM1) patches in the erythrocyte membrane.

The detergent (Triton X-100, 4°C)-resistant membrane (DRM)-associated membrane proteins stomatin, sorcin, and synexin (anexin VII) exposed on the cytoplasmic side of membrane were investigated for their lateral distribution in relation to induced ganglioside(M1) (GM1) raft patches in flat (discocytic) and curved (echinocytic) human erythrocyte membrane. In discocytes, no accumulation of stomatin, sorcin, and synexin in cholera toxin subunit B (CTB) plus anti-CTB-induced GM1 patches was detected by fluorescence microscopy. In echinocytes, stomatin, sorcin, and synexin showed a similar curvature-dependent lateral distribution as GM1 patches by accumulating to spiculae induced by ionophore A23187 plus calcium. Stomatin was partly and synexin and sorcin were fully recruited to the spiculae. However, the DRM-associated proteins only partially co-localized with GM1 and were frequently distributed into different spiculae than GM1. The study indicates that stomatin, sorcin, and synexin are echinophilic membrane components that mainly locate outside GM1 rafts in the human erythrocyte membrane. Echinophilicity is suggested to contribute to the DRM association of a membrane component in general.

Interaction between equally charged membrane surfaces mediated by positively and negatively charged macro-ions.

In biological systems, charged membrane surfaces are surrounded by charged molecules such as electrolyte ions and proteins. Our recent experiments in the systems of giant phospholipid vesicles indicated that some of the blood plasma proteins (macro-ions) may promote adhesion between equally charged membrane surfaces. In this work, theory was put forward to describe an IgG antibody-mediated attractive interaction between negatively charged membrane surfaces which was observed in experiments on giant phospholipid vesicles with cardiolipin-containing membranes. The attractive interactions between negatively charged membrane surfaces in the presence of negatively and positively charged spherical macro-ions are explained using functional density theory and Monte Carlo simulations. Both, the rigorous solution of the variational problem within the functional density theory and the Monte Carlo simulations show that spatial and orientational ordering of macro-ions may give rise to an attractive interaction between negatively charged membrane surfaces. It is also shown that the distinctive spatial distribution of the charge within the macro-ions (proteins) is essential in this process.

Mechanical stability of membrane nanotubular protrusions influenced by attachment of flexible rod-like proteins.

It is indicated that nonhomogeneous lateral distribution of membrane attached and flexible rod-like proteins (MRPs) may stabilize nanotubular membrane protrusions. We have shown that curvature induced accumulation of MRPs in the nanotubular membrane protrusion and the corresponding reduction of the membrane free energy are possible if the decrease of the deviatoric free energy of MRPs in the nanotubular protrusions is large enough to overcome the increase of the free energy due to decrease of configurational entropy in the process of lateral sorting of MRPs. The decrease of isotropic curvature energy of MRPs in the region of membrane protrusion is usually not sufficient for substantial MRPs sorting and consequent stabilization of the nanotubular membrane protrusions.

Role of phospholipid asymmetry in the stability of inverted hexagonal mesoscopic phases.

The role of phospholipid asymmetry in the transition from the lamellar (L(alpha)) to the inverted hexagonal (H(II)) phase upon the temperature increase was considered. The equilibrium configuration of the system was determined by the minimum of the free energy including the contribution of the isotropic and deviatoric bending and the interstitial energy of phospholipid monolayers. The shape and local interactions of a single lipid molecule were taken into account. The minimization with respect to the configuration of the lipid layers was performed by a numerical solution of the system of the Euler-Lagrange differential equations and by the Monte Carlo simulated annealing method. At high enough temperature, the lipid molecules attain a shape exhibiting higher intrinsic mean and deviatoric curvatures, which fits better into the H(II) phase than into the L(alpha) phase. Furthermore, the orientational ordering of lipid molecules in the curvature field expressed as the deviatoric bending provides a considerable negative contribution to the free energy, which stabilizes the nonlamellar H(II) phase. The nucleation configuration for the L(alpha)-H(II) phase transition is tuned by the isotropic and deviatoric bending energies and the interstitial energy.