Svetlana Baoukina

Computer simulation study of fullerene translocation through lipid membranes.

Recent toxicology studies suggest that nanosized aggregates of fullerene molecules can enter cells and alter their functions, and also cross the blood-brain barrier. However, the mechanisms by which fullerenes penetrate and disrupt cell membranes are still poorly understood. Here we use computer simulations to explore the translocation of fullerene clusters through a model lipid membrane and the effect of high fullerene concentrations on membrane properties. The fullerene molecules rapidly aggregate in water but disaggregate after entering the membrane interior. The permeation of a solid-like fullerene aggregate into the lipid bilayer is thermodynamically favoured and occurs on the microsecond timescale. High concentrations of fullerene induce changes in the structural and elastic properties of the lipid bilayer, but these are not large enough to mechanically damage the membrane. Our results suggest that mechanical damage is an unlikely mechanism for membrane disruption and fullerene toxicity.

The molecular mechanism of lipid monolayer collapse

Lipid monolayers at an air–water interface can be compressed laterally and reach high surface density. Beyond a certain threshold, they become unstable and collapse. Lipid monolayer collapse plays an important role in the regulation of surface tension at the air–liquid interface in the lungs. Although the structures of lipid aggregates formed upon collapse can be characterized experimentally, the mechanism leading to these structures is not fully understood. We investigate the molecular mechanism of monolayer collapse using molecular dynamics simulations. Upon lateral compression, the collapse begins with buckling of the monolayer, followed by folding of the buckle into a bilayer in the water phase. Folding leads to an increase in the monolayer surface tension, which reaches the equilibrium spreading value. Immediately after their formation, the bilayer folds have a flat semielliptical shape, in agreement with theoretical predictions. The folds undergo further transformation and form either flat circular bilayers or vesicles. The transformation pathway depends on macroscopic parameters of the system: the bending modulus, the line tension at the monolayer–bilayer connection, and the line tension at the bilayer perimeter. These parameters are determined by the system composition and temperature. Coexistence of the monolayer with lipid aggregates is favorable at lower tensions of the monolayer–bilayer connection. Transformation into a vesicle reduces the energy of the fold perimeter and is facilitated for softer bilayers, e.g., those with a higher content of unsaturated lipids, or at higher temperatures.

Lipid Nanoparticles Containing siRNA Synthesized by Microfluidic Mixing Exhibit an Electron-Dense Nanostructured Core

Lipid nanoparticles (LNP) containing ionizable cationic lipids are the leading systems for enabling therapeutic applications of siRNA; however, the structure of these systems has not been defined. Here we examine the structure of LNP siRNA systems containing DLinKC2-DMA(an ionizable cationic lipid), phospholipid, cholesterol and a polyethylene glycol (PEG) lipid formed using a rapid microfluidic mixing process. Techniques employed include cryo-transmission electron microscopy, 31P NMR, membrane fusion assays, density measurements, and molecular modeling. The experimental results indicate that these LNP siRNA systems have an interior lipid core containing siRNA duplexes complexed to cationic lipid and that the interior core also contains phospholipid and cholesterol. Consistent with experimental observations, molecular modeling calculations indicate that the interior of LNP siRNA systems exhibits a periodic structure of aqueous compartments, where some compartments contain siRNA. It is concluded that LNP siRNA systems formulated by rapid mixing of an ethanol solution of lipid with an aqueous medium containing siRNA exhibit a nanostructured core. The results give insight into the mechanism whereby LNP siRNA systems are formed, providing an understanding of the high encapsulation efficiencies that can be achieved and information on methods of constructing more sophisticated LNP systems.

Martini straight: Boosting performance using a shorter cutoff and GPUs

Abstract In molecular dynamics simulations, sufficient sampling is of key importance and a continuous challenge in the field. The coarse grain Martini force field has been widely used to enhance sampling. In its original implementation, this force field applied a shifted Lennard-Jones potential for the non-bonded van der Waals interactions, to avoid problems related to a relatively short cutoff. Here we investigate the use of a straight cutoff Lennard-Jones potential with potential modifiers. Together with a Verlet neighbor search algorithm, the modified potential allows the use of GPUs to accelerate the computations in Gromacs. We find that this alternative potential has little influence on most of the properties studied, including partitioning free energies, bulk liquid properties and bilayer properties. At the same time, energy conservation is kept within reasonable bounds. We conclude that the newly proposed straight cutoff approach is a viable alternative to the standard shifted potentials used in Martini, offering significant speedup even in the absence of GPUs.

Molecular View of Phase Coexistence in Lipid Monolayers

We used computer simulations to study the effect of phase separation on the properties of lipid monolayers. This is important for understanding the lipid-lipid interactions underlying lateral heterogeneity (rafts) in biological membranes and the role of domains in the regulation of surface tension by lung surfactant. Molecular dynamics simulations with the coarse-grained MARTINI force field were employed to model large length (~80 nm in lateral dimension) and time (tens of microseconds) scales. Lipid mixtures containing saturated and unsaturated lipids and cholesterol were investigated under varying surface tension and temperature. We reproduced compositional lipid demixing and the coexistence of liquid-expanded and liquid-condensed phases as well as liquid-ordered and liquid-disordered phases. Formation of the more ordered phase was induced by lowering the surface tension or temperature. Phase transformations occurred via either nucleation or spinodal decomposition. In nucleation, multiple domains formed initially and subsequently merged. Using cluster analysis combined with Voronoi tessellation, we characterized the partial areas of the lipids in each phase, the phase composition, the boundary length, and the line tension under varying surface tension. We calculated the growth exponents for nucleation and spinodal decomposition using a dynamical scaling hypothesis. At low surface tensions, liquid-ordered domains manifest spontaneous curvature. Lateral diffusion of lipids is significantly slower in the more ordered phase, as expected. The presence of domains increased the monolayer surface viscosity, in particular as a result of domain reorganization under shear.

Direct simulation of protein-mediated vesicle fusion: lung surfactant protein B.

We simulated spontaneous fusion of small unilamellar vesicles mediated by lung surfactant protein B (SP-B) using the MARTINI force field. An SP-B monomer triggers fusion events by anchoring two vesicles and facilitating the formation of a lipid bridge between the proximal leaflets. Once a lipid bridge is formed, fusion proceeds via a previously described stalk - hemifusion diaphragm - pore-opening pathway. In the absence of protein, fusion of vesicles was not observed in either unbiased simulations or upon application of a restraining potential to maintain the vesicles in close proximity. The shape of SP-B appears to enable it to bind to two vesicles at once, forcing their proximity, and to facilitate the initial transfer of lipids to form a high-energy hemifusion intermediate. Our results may provide insight into more general mechanisms of protein-mediated membrane fusion, and a possible role of SP-B in the secretory pathway and transfer of lung surfactant to the gas exchange interface.

Molecular Structure of Membrane Tethers

Membrane tethers are nanotubes formed by a lipid bilayer. They play important functional roles in cell biology and provide an experimental window on lipid properties. Tethers have been studied extensively in experiments and described by theoretical models, but their molecular structure remains unknown due to their small diameters and dynamic nature. We used molecular dynamics simulations to obtain molecular-level insight into tether formation. Tethers were pulled from single-component lipid bilayers by application of an external force to a lipid patch along the bilayer normal or by lateral compression of a confined bilayer. Tether development under external force proceeded by viscoelastic protrusion followed by viscous lipid flow. Weak forces below a threshold value produced only a protrusion. Larger forces led to a crossover to tether elongation, which was linear at a constant force. Under lateral compression, tethers formed from undulations of unrestrained bilayer area. We characterized in detail the tether structure and its formation process, and obtained the material properties of the membrane. To our knowledge, these results provide the first molecular view of membrane tethers.

Computer simulations of the phase separation in model membranes

We used computer simulations to investigate the properties of model lipid membranes with coexisting phases. This is relevant for understanding lipid-lipid interactions underlying lateral organization in biological membranes. Molecular dynamics simulations with the MARTINI coarse-grained force field were employed to study lipid bilayers -40 nm in lateral dimension on a 20 micros time scale. The simulations retain near atomic-level detail and lipid chemical specificity, and allow formation of multiple domains of tens of nanometers in size. Using ternary lipid mixtures of saturated and unsaturated lipids and cholesterol, we reproduced the coexistence of the Lalpha/gel phases and the Lo/Ld phases. Phase transformation proceeded by either nucleation or spinodal decomposition. The properties of coexisting phases were characterized in detail, including partial lipid areas, composition, phase boundary and domain registry, based on Voronoi tessellation. We investigated variations of these properties with temperature and surface tension, and compared them to our recent simulations of lipid monolayers of the same size and composition. We found substantial overlap in bilayer and monolayer properties. Increasing the temperature in bilayers produced similar effects as increasing the surface tension in monolayers. This information can be used for interpreting experimental data on model membranes.

Lung Surfactant Protein SP-B Promotes Formation of Bilayer Reservoirs from Monolayer and Lipid Transfer between the Interface and Subphase

We investigated the possible role of SP-B proteins in the function of lung surfactant. To this end, lipid monolayers at the air/water interface, bilayers in water, and transformations between them in the presence of SP-B were simulated. The proteins attached bilayers to monolayers, providing close proximity of the reservoirs with the interface. In the attached aggregates, SP-B mediated establishment of the lipid-lined connection similar to the hemifusion stalk. Via this connection, a lipid flow was initiated between the monolayer at the interface and the bilayer in water in a surface-tension-dependent manner. On interface expansion, the flow of lipids to the monolayer restored the surface tension to the equilibrium spreading value. SP-B induced formation of bilayer folds from the monolayer at positive surface tensions below the equilibrium. In the absence of proteins, lipid monolayers were stable at these conditions. Fold nucleation was initiated by SP-B from the liquid-expanded monolayer phase by local bending, and the proteins lined the curved perimeter of the growing fold. No effect on the liquid-condensed phase was observed. Covalently linked dimers resulted in faster kinetics for monolayer folding. The simulation results are in line with existing hypotheses on SP-B activity in lung surfactant and explain its molecular mechanism.

Lateral pressure profiles in lipid monolayers

We have used molecular dynamics simulations with coarse-grained and atomistic models to study the lateral pressure profiles in lipid monolayers. We first consider simple oillair and oil/water interfaces, and then proceed to lipid monolayers at air/water and oil/water interfaces. The results are qualitatively similar in both atomistic and coarse-grained models. The lateral pressure profile in a monolayer is characterized by a headgroup/water pressure-interfacial tension-chain pressure pattern. In contrast to lipid bilayers, the pressure decreases towards the chain free ends. An additional chain/air tension peak is present in monolayers at the air/water interface. Lateral pressure profiles are calculated for monolayers of different lipid composition under varying surface tension. Increasing the surface tension suppresses both pressure peaks and widens the interfacial tension in monolayers at the oil/water interface, and mainly suppresses the chain pressure in monolayers at the air/water interface. In monolayers in the liquid-condensed phase, the pressure peaks split due to ordering. Variation of lipid composition leads to noticeable changes in all regions of the pressure profile at a fixed surface tension.