Is the cholesterol bilayer domain a barrier to oxygen transport into the eye lens?

In the eye lens, the oxygen partial pressure is very low and the cholesterol (Chol) content in cell membranes is very high. Disturbance of these quantities results in cataract development. In human lens membranes, both bulk phospholipid-Chol domains and the pure Chol bilayer domains (CBDs) were experimentally detected. It is hypothesized that the CBD constitutes a significant barrier to oxygen transport into the lens. Transmembrane profiles of the oxygen diffusion-concentration product, obtained with electron paramagnetic resonance spin-labeling methods, allow evaluation of the oxygen permeability (PM) of phospholipid membranes but not the CBD. Molecular dynamics simulation can independently provide components of the product across any bilayer domain, thus allowing evaluation of the PM across the CBD. Therefore, to test the hypothesis, MD simulation was used. Three bilayers containing palmitoyl-oleoyl-phosphorylcholine (POPC) and Chol were built. The pure Chol bilayer modeled the CBD, the 1:1 POPC-Chol bilayer modeled the bulk membrane in which the CBD is embedded, and the POPC bilayer was a reference. To each model, 200 oxygen molecules were added. After equilibration, the oxygen concentration and diffusion profiles were calculated for each model and multiplied by each other. From the respective product profiles, the PM of each bilayer was calculated. Favorable comparison with experimental data available only for the POPC and POPC-Chol bilayers validated these bilayer models and allowed the conclusion that oxygen permeation across the CBD is ~10 smaller than across the bulk membrane, supporting the hypothesis that the CBD is a barrier to oxygen transport into the eye lens.

Computer modelling studies of the bilayer/water interface.

This review summarises high resolution studies on the interface of lamellar lipid bilayers composed of the most typical lipid molecules which constitute the lipid matrix of biomembranes. The presented results were obtained predominantly by computer modelling methods. Whenever possible, the results were compared with experimental results obtained for similar systems. The first and main section of the review is concerned with the bilayer-water interface and is divided into four subsections. The first describes the simplest case, where the interface consists only of lipid head groups and water molecules and focuses on interactions between the lipid heads and water molecules; the second describes the interface containing also mono- and divalent ions and concentrates on lipid-ion interactions; the third describes direct inter-lipid interactions. These three subsections are followed by a discussion on the network of direct and indirect inter-lipid interactions at the bilayer interface. The second section summarises recent computer simulation studies on the interactions of antibacterial membrane active compounds with various models of the bacterial outer membrane. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg.

Saturation with cholesterol increases vertical order and smoothes the surface of the phosphatidylcholine bilayer: a molecular simulation study.

Molecular dynamics (MD) simulations of a mono-cis-unsaturated 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC) bilayer and a POPC bilayer containing 50mol% cholesterol (POPC-Chol50) were carried out for 200ns to compare the spatial organizations of the pure POPC bilayer and the POPC bilayer saturated with Chol. The results presented here indicate that saturation with Chol significantly narrows the distribution of vertical positions of the center-of-mass of POPC molecules and POPC atoms in the bilayer. In the POPC-Chol50 bilayer, the same moieties of the lipid molecules are better aligned at a given bilayer depth, forming the following clearly separated membrane regions: the polar headgroup, the rigid core consisting of steroid rings and upper fragments of the acyl chains, and the fluid hydrocarbon core consisting of Chol chains and the lower fragments of POPC chains. The membrane surface of the POPC-Chol50 bilayer is smooth. The results have biological significance because the POPC-Chol50 bilayer models the bulk phospholipid portion of the fiber-cell membrane in the eye lens. It is hypothesized that in the eye lens cholesterol-induced smoothing of the membrane surface decreases light-scattering and helps to maintain lens transparency.

Protecting the Eye Lens from Oxidative Stress through Oxygen Regulation.

Molecular oxygen is a primary oxidant that is involved in the formation of active oxygen species and in the oxidation of lipids and proteins. Thus, controlling oxygen partial pressure (concentration) in the human organism, tissues, and organs can be the first step in protecting them against oxidative stress. However, it is not an easy task because oxygen is necessary for ATP synthesis by mitochondria and in many biochemical reactions taking place in all cells in the human body. Moreover, the blood circulatory system delivers oxygen to all parts of the body. The eye lens seems to be the only organ that is protected from the oxidative stress through the regulation of oxygen partial pressure. The basic mechanism that developed during evolution to protect the eye lens against oxidative damage is based on the maintenance of a very low concentration of oxygen within the lens. This antioxidant mechanism is supported by the resistance of both the lipid components of the lens membrane and cytosolic proteins to oxidation. Any disturbance, continuous or acute, in the working of this mechanism increases the oxygen concentration, in effect causing cataract development. Here, we describe the biophysical basis of the mechanism and its correlation with lens transparency.

Stacks of monogalactolipid bilayers can transform into a lattice of water channels.

The lipid matrix of thylakoid membranes is a lamellar bilayer, but under a certain condition it can convert locally into a nonlamellar structure. This is possible because one of the main membrane lipids, MGDG, promotes the formation of an inverse hexagonal phase. Here, the spontaneous transformation of aligned hydrated MGDG bilayers into nonlamellar structures is investigated using all-atom molecular dynamics simulation. Previous studies have demonstrated that MGDG polar head groups connect vertically across the interface. In this study, the evolution of the system's initial structure into a lattice of water channels and contacted surfaces created by numerous vertical MGDG connections depended on the width of the hydrating water layers. These widths controlled the bilayers' ability to bend, which was a prerequisite for channel formation. Locally, an intensive exchange of MGDG molecules between apposing bilayer leaflets occurred, although a stable semi-toroidal stalk did not develop.

Strong preferences of dopamine and l-dopa towards lipid head group: importance of lipid composition and implication for neurotransmitter metabolism.

The interactions of the neurotransmitter dopamine, and its precursor l-dopa, with membrane lipids were investigated through a set of molecular dynamic simulations with all atom resolution. The results obtained indicate that both dopamine and l-dopa have a pronounced association with the lipid head groups, predominantly mediated through H-bonds. As a result the molecules are anchored to the interfacial region of the membrane. The strength of this interaction is dependent on lipid composition - the presence of phosphatidylserine leads to an increase in the strength of this interaction, resulting in an H-bond network with a lifetime much longer than the timescale of our simulations. Also, bilayers that include sphingomieline and cholesterol interact strongly with dopamine and l-dopa. We postulate that the high membrane association that we have observed for both dopamine and l-dopa could have the following effects: 1) when on the plasma membrane exterior, favour the availability of these compounds for cell membrane uptake processes and, 2) when on an internal membrane surface, accentuate the importance of membrane-bound metabolizing enzymes over their soluble counterparts.

Lipid/water interface of galactolipid bilayers in different lyotropic liquid-crystalline phases.

In this study, carried out using computational methods, the organisation of the lipid/water interface of bilayers composed of galactolipids with both α-linolenoyl acyl chains is analysed and compared in three different lyotropic liquid-crystalline phases. These systems include the monogalactosyldiglyceride (MGDG) and digalactosyldiglyceride (DGDG) bilayers in the lamellar phase, the MGDG double bilayer during stalk phase formation and the inverse hexagonal MGDG phase. For each system, lipid-water and direct and water-mediated lipid-lipid interactions between the lipids of one bilayer leaflet and those of two apposing leaflets at the onset of new phase (stalk) formation, are identified. A network of interactions between DGDG molecules and its topological properties are derived and compared to those for the MGDG bilayer.

Molecular oxygen as a probe molecule in EPR spin-labeling studies of membrane structure and dynamics.

Molecular oxygen (O2) is the perfect probe molecule for membrane studies carried out using the saturation recovery EPR technique. O2 is a small, paramagnetic, hydrophobic enough molecule that easily partitions into a membrane's different phases and domains. In membrane studies, the saturation recovery EPR method requires two paramagnetic probes: a lipid-analog nitroxide spin label and an oxygen molecule. The experimentally derived parameters of this method are the spin-lattice relaxation times (T 1s) of spin labels and rates of bimolecular collisions between O2 and the nitroxide fragment. Thanks to the long T 1 of lipid spin labels (from 1 to 10 µs), the approach is very sensitive to changes of the local (around the nitroxide fragment) O2 diffusion-concentration product. Small variations in the lipid packing affect O2 solubility and O2 diffusion, which can be detected by the shortening of T 1 of spin labels. Using O2 as a probe molecule and a different lipid spin label inserted into specific phases of the membrane and membrane domains allows data about the lateral arrangement of lipid membranes to be obtained. Moreover, using a lipid spin label with the nitroxide fragment attached to its head group or a hydrocarbon chain at different positions also enables data about molecular dynamics and structure at different membrane depths to be obtained. Thus, the method can be used to investigate not only the lateral organization of the membrane (i.e., the presence of membrane domains and phases), but also the depth-dependent membrane structure and dynamics, and, hence, the membrane properties in three dimensions.

Comparative model studies of gastric toxicity of nonsteroidal anti-inflammatory drugs.

A high percentage of people treated with a long-term nonsteroidal anti-inflammatory drug (NSAID) therapy suffer NSAID-induced gastrointestinal-tract-related side effects. A current hypothesis states that the side effects are related to the topical action of NSAID molecules on gastric mucus that lowers its resistance to luminal acid. The main lipids in human mucus are palmitoyloleoylphosphatidylcholine (POPC) and cholesterol (Chol). In this study, both X-ray diffraction and molecular dynamics (MD) simulation methods were employed to investigate the effects of selected NSAIDs in protonated and deprotonated states on the structural parameters of a POPC-Chol bilayer. The drugs were three commonly used NSAIDs with apparently different gastric toxicity: ketoprofen (KET), aspirin (ASP), and piroxicam (PXM). Both methods revealed that the effects of the NSAIDs on the POPC-Chol bilayer parameters were moderate and only slightly differentiated among the drugs. Much larger differences among the drugs were noticed in their interactions with interfacial water and Na(+) as well as with the polar groups of POPC and Chol, mainly via H-bonds. Of the three NSAIDs, KET interacted with POPC and water the most extensively, whereas ASP interacted with Chol and Na(+) more than did the other two. Interactions of PXM with POPC and Chol polar groups as well as with water and Na(+) were limited.

Study of PEGylated lipid layers as a model for PEGylated liposome surfaces: molecular dynamics simulation and Langmuir monolayer studies.

We have combined Langmuir monolayer film experiments and all-atom molecular dynamics (MD) simulation of a bilayer to study the surface structure of a PEGylated liposome and its interaction with the ionic environment present under physiological conditions. Lipids that form both gel and liquid-crystalline membranes have been used in our study. By varying the salt concentration in the Langmuir film experiment and including salt at the physiological level in the simulation, we have studied the effect of salt ions present in the blood plasma on the structure of the poly(ethylene glycol) (PEG) layer. We have also studied the interaction between the PEG layer and the lipid bilayer in both the liquid-crystalline and gel states. The MD simulation shows two clear results: (a) The Na(+) ions form close interactions with the PEG oxygens, with the PEG chains forming loops around them and (b) PEG penetrates the lipid core of the membrane for the case of a liquid-crystalline membrane but is excluded from the tighter structure of the gel membrane. The Langmuir monolayer results indicate that the salt concentration affects the PEGylated lipid system, and these results can be interpreted in a fashion that is in agreement with the results of our MD simulation. We conclude that the currently accepted picture of the PEG surface layer acting as a generic neutral hydrophilic polymer entirely outside the membrane, with its effect explained through steric interactions, is not sufficient. The phenomena we have observed may affect both the interaction between the liposome and bloodstream proteins and the liquid-crystalline-gel transition and is thus relevant to nanotechnological drug delivery device design.

Communication: consistent picture of lateral subdiffusion in lipid bilayers: molecular dynamics simulation and exact results.

This communication presents a molecular dynamics simulation study of a bilayer consisting of 128 dioleoyl-sn-glycero-3-phosphocholine molecules, which focusses on the center-of-mass diffusion of the lipid molecules parallel to the membrane plane. The analysis of the simulation results is performed within the framework of the generalized Langevin equation and leads to a consistent picture of subdiffusion. The mean square displacement of the lipid molecules evolves as ∝ t(α), with α between 0.5 and 0.6, and the fractional diffusion coefficient is close to the experimental value for a similar system obtained by fluorescence correlation spectroscopy. We show that the long-time tails of the lateral velocity autocorrelation function and the associated memory function agree well with exact results which have been recently derived by asymptotic analysis [G. Kneller, J. Chem. Phys. 134, 224106 (2011)]. In this context, we define characteristic time scales for these two quantities.

Data for molecular dynamic simulations in the OPLSAA force field: Partial charges of cholesterol, C7-hydroxycholesterol and C7-hydroperoxycholesterol, torsional parameters for the hydroperoxy group of C7-hydroperoxycholesterol.

This data article contains partial charges generated for cholesterol, C7-hydroxycholesterol and C7-hydroperoxycholesterol and torsional parameters for hydroperoxy of C7-hydroperoxycholesterol for molecular dynamics simulations in the OPLSAA force field [1] using the package Gromacs [2]. The hydroperoxy group remained unparameterized in the OPLSAA force field and the parameters obtained have the potential for re-use in similar simulations. The atom-centred point charges on each sterol molecule were derived using the restrained electrostatic potential (RESP) approach [3]. The parameters for the C7-OET-OH-HO and C8-C7-OET-OH torsion angles were derived by fitting the parameters of the torsional term (Ryckaert-Bellemans function) of the OPLSAA potential energy function to the quantum mechanical rotational energy profile calculated at CCSD(T)/cc-pVQZ level of theory. This article presents data used in the research article "Chirality affects cholesterol-oxysterol association in water, a computational study" [4].

Lutein and Zeaxanthin in the Lipid Bilayer-Similarities and Differences Revealed by Computational Studies.

Lutein and zeaxanthin are two similar carotenoids of the xanthophyll subgroup. Carotenoids are synthesized almost entirely by plants but are also present in significant amounts in animals. They are essential components of the lipid matrix of biomembranes, and one of their functions is to protect cells from light radiation, free radicals and oxidative stress. Carotenoids, depending on their chemical structure, can locate at various positions and in different orientations in the bilayer. Xanthophylls (XAN) are polar and in the bilayer are positionally restricted. In the case of lutein and zeaxanthin, whose both ionone rings are hydroxy-substituted and as such are anchored in the lipid bilayer interfaces, the position is generally transmembrane. However, both experimental and computer modelling studies indicate that lutein can also locate horizontally below the bilayer interface. This location has never been observed for zeaxanthin. To find a molecular-level explanation for the difference in the orientations of the XAN molecules in the bilayer, a number of phosphatidylcholine-XAN bilayers were constructed and molecular dynamics (MD) simulated for 1.1 µs each. The all-trans XAN molecules were initially placed either parallel or perpendicular to the bilayer surface. With the exception of one lutein, the horizontally placed molecules adopted the transmembrane orientation within 100-600 ns. On the basis of detailed analyses of the XAN orientations and the numbers and lifetimes of their interactions in the bilayer, a plausible explanation is offered as to why a lutein molecule may remain in the horizontal orientation while zeaxanthin does not. Contrary to common believe, lutein horizontal orientation is not related to the ε-ring rotation around the C6'-C7' bond.

Chirality affects cholesterol-oxysterol association in water, a computational study.

Cholesterol (Chol) is the most prevalent sterol in the animal kingdom and an indispensable component of mammalian cell membranes. Chol content in the membrane is strictly controlled, although the oxidation of phospholipids may change the relative content of membrane Chol. An excess of it results in the formation of pure Chol microdomains in the membrane. It is likely that some Chol molecules detach from the domains and self-assemble in the aqueous environment. This may promote Chol microcrystallisation, which initiates the development of gallstones and atherosclerotic plaque. In this study, the molecular dynamics, free energy perturbation, umbrella sampling and Voronoi diagram methods are used to reveal the details of self-association of Chol and its oxidised forms (oxChol), namely 7α,ß-hydroxycholesterol and 7α,ß-hydroperoxycholesterol, in water. In the first part of the study the interactions between a sterol monomer and water over a short and longer timescale as well as the energy of hydration of each sterol are analysed. This helps one to understand Chol-Chol and Chol-OxChol with different chirality self-association in water better, which is analysed in the second part of the study. The Voronoi diagram approach is used to determine the relative arrangement of molecules in the dimer and, most importantly, to analyse the dehydration of the contacting surfaces of the assembling molecules. Free energy calculations indicate that Chol and 7ß-hydroxycholesterol associate into the most stable dimer and that Chol-Chol is the next most stable of the five dimers studied. Employing different computational methods enables us to obtain an adequate picture of Chol-sterol self-association in water, which includes dynamic, energetic and temporal aspects of the process.

Ordering effects of cholesterol and its analogues.

Without any exaggeration, cholesterol is one of the most important lipid species in eukaryotic cells. Its effects on cellular membranes and functions range from purely mechanistic to complex metabolic ones, besides which it is also a precursor of the sex hormones (steroids) and several vitamins. In this review, we discuss the biophysical effects of cholesterol on the lipid bilayer, in particular the ordering and condensing effects, concentrating on the molecular level or inter-atomic interactions perspective, starting from two-component systems and proceeding to many-component ones e.g., modeling lipid rafts. Particular attention is paid to the roles of the methyl groups in the cholesterol ring system, and their possible biological function. Although our main research methodology is computer modeling, in this review we make extensive comparisons between experiments and different modeling approaches.

Hypothetical Pathway for Formation of Cholesterol Microcrystals Initiating the Atherosclerotic Process.

Major factors leading to the development of atherosclerosis are a high cholesterol (Chol) level in the blood and oxidative stress. Both promote the formation of Chol microcrystals in blood vessel walls. Deposition of Chol microcrystals in arterial intima causes inflammation, which initiates and accompanies the atherosclerotic process in all its phases. One of the possible sources of Chol in the blood vessel walls is oxidized low-density lipoproteins-this atherosclerotic plaque formation pathway has already been described in the literature. Here, we hypothesize that initiation of the atherosclerotic process may involve Chol domains in the plasma membranes of arterial cells. Increased Chol content and the presence of polyunsaturated phospholipids in these membranes together with oxidative stress (phospholipid peroxidation) may lead to the formation of pure Chol bilayer domains that, with further peroxidation and increased Chol content, may collapse in the form of Chol seed crystals. Independent of their origin, Chol microcrystals activate inflammasomes, thereby stimulate immune responses, and initiate inflammation that may lead to the development of atherosclerosis. This new, hypothetical pathway has not yet been investigated in depth; however, data from the literature and our own results support its feasibility.

Formation of cholesterol Bilayer Domains Precedes Formation of Cholesterol Crystals in Membranes Made of the Major Phospholipids of Human Eye Lens Fiber Cell Plasma Membranes.

Purpose/Aim: The goal of this study is to reveal how age-related changes in phospholipid (PL) composition in the fiber cell plasma membranes of the human eye lens affect the cholesterol (Chol) content at which Chol bilayer domains (CBDs) and Chol crystals start to form.Materials and Methods: Saturation-recovery electron paramagnetic resonance with spin-labeled cholesterol analogs and differential scanning calorimetry were used to determine the Chol contents at which CBDs and cholesterol crystals, respectively, start to form in in membranes made of the major PL constituents of the plasma membrane of the human eye lens fiber cells. To preserve compositional homogeneity throughout the membrane suspension, the lipid multilamellar dispersions investigated in this work were prepared using a rapid solvent exchange method. The cholesterol content changed from 0 to 75 mol%.Results: The saturation recovery electron paramagnetic resonance results show that CBDs start to form at 33, 50, 46, and 48 mol% Chol in the phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, and sphingomyelin bilayers, respectively. The differential scanning calorimetry results show that Chol crystals start to form at 50, 66, 70, and 66 mol% Chol in the phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, and sphingomyelin bilayers, respectively.Conclusions: These results, as well those of our previous studies, indicate that the formation of CBDs precedes the formation of Chol crystals in all of the studied systems, and the appearance of each depends on the type of PL forming the bilayer. These findings contribute to a better understanding of the molecular mechanisms involved in the regulation of Chol-dependent processes in eye lens fiber cell membranes.

Water isotope effect on the phosphatidylcholine bilayer properties: a molecular dynamics simulation study.

Physicochemical properties of heavy water (D2O) differ to some extent from those of normal water. Substituting D2O for H2O has been shown to affect the structural and dynamic properties of proteins, but studies of its effects on lipid bilayers are scarce. In this paper, the atomic level molecular dynamics (MD) simulation method was used to determine the effects of this substitution on the properties of a dipalmitoylphosphatidylcholine (DPPC) bilayer and its hydrating water. MD simulations of two DPPC bilayers, one fully hydrated with H2O and the other with D2O, were carried out for over 50 ns. For H2O, the simple point charge (SPC) model was used, and for D2O, the extended SPC-HW model was employed. Analyses of the simulation trajectories indicate that several properties of the membrane core and the membrane/water interface are affected by replacing H2O by D2O. However, the time-averaged properties, such as membrane compactness, acyl chain order, and numbers of PC-water H (D)-bonds and PC-PC water bridges, are much less affected than time-resolved properties. In particular, the lifetimes of these interactions are much longer for D2O molecules than for H2O ones. These longer lifetimes results in a slightly better ordering of the D2O molecules and average self-diffusion, which is 50% slower compared with the H2O molecules. This large isotope effect has been assigned to the repercussions of the longer lived D-bonding to DPPC headgroups insofar as all water molecules sense the presence of the DPPC bilayer.

Asymmetric Spontaneous Intercalation of Lutein into a Phospholipid Bilayer, a Computational Study.

Lutein, a hydroxylated carotenoid, is a pigment synthesised by plants and bacteria. Animals are unable to synthesise lutein, nevertheless, it is present in animal tissues, where its only source is dietary intake. Both in plants and animals, carotenoids are associated mainly with membranes where they carry out important physiological functions. Their trafficking to and insertion into membranes are not well recognised due to experimental difficulties. In this paper, a computational approach is used to elucidate details of the dynamics and energetics of lutein intercalation from the water to the phospholipid bilayer phase. The dynamics is studied using molecular dynamics simulation, and the energetics using umbrella sampling. Lutein spontaneous insertion into the bilayer and translocation across it proceed via formation of hydrogen bonds between its hydroxyl groups and water and/or phospholipid oxygen atoms as well as desolvation of its polyene chain. As lutein molecule is asymmetric, its bilayer intercalation is also asymmetric. The course of events and timescale of the intercalation are different from those of helical peptides. The time of full lutein intercalation ranges from 20 to 100 ns and its final orientation is predominately vertical. Nevertheless, some lutein molecules are in the final horizontal position and some aggregate in the water phase and remain there for the whole simulation time. The highest energy barrier for the intercalation process is ~2.2 kcal/mol and the energy gain is ~18 kcal/mol. The results obtained for lutein can be applied to other xanthophylls and molecules of a similar structure.

Interplay of unsaturated phospholipids and cholesterol in membranes: effect of the double-bond position.

The structural and dynamical properties of lipid membranes rich in phospholipids and cholesterol are known to be strongly affected by the unsaturation of lipid acyl chains. We show that not only unsaturation but also the position of a double bond has a pronounced effect on membrane properties. We consider how cholesterol interacts with phosphatidylcholines comprising two 18-carbon long monounsaturated acyl chains, where the position of the double bond is varied systematically along the acyl chains. Atomistic molecular dynamics simulations indicate that when the double bond is not in contact with the cholesterol ring, and especially with the C18 group on its rough beta-side, the membrane properties are closest to those of the saturated bilayer. However, any interaction between the double bond and the ring promotes membrane disorder and fluidity. Maximal disorder is found when the double bond is located in the middle of a lipid acyl chain, the case most commonly found in monounsaturated acyl chains of phospholipids. The results suggest a cholesterol-mediated lipid selection mechanism in eukaryotic cell membranes. With saturated lipids, cholesterol promotes the formation of highly ordered raft-like membrane domains, whereas domains rich in unsaturated lipids with a double bond in the middle remain highly fluid despite the presence of cholesterol.