Cholesterol effects on the phosphatidylcholine bilayer polar region: a molecular simulation study

A molecular dynamics (MD) simulation of a fully hydrated, liquid-crystalline dimyristoylphosphatidylcholine (DMPC)-Chol bilayer membrane containing approximately 22 mol% Chol was carried out for 4.3 ns. The bilayer reached thermal equilibrium after 2.3 ns of MD simulation. A 2.0-ns trajectory generated during 2.3-4.3 ns of MD simulation was used for analyses to determine the effects of Chol on the membrane/water interfacial region. In this region, 70% of Chol molecules are linked to DMPC molecules via short-distance interactions, where the Chol hydroxyl group (OH-Chol) is 1) charge paired to methyl groups of the DMPC choline moiety ( approximately 34%), via the hydroxyl oxygen atom (Och); 2) water bridged to carbonyl ( approximately 19%) and nonester phosphate ( approximately 14%) oxygen atoms, via both Och and the hydroxyl hydrogen atom (Hch); and 3) directly hydrogen (H) bonded to carbonyl ( approximately 11%) and nonester phosphate ( approximately 5%) oxygen atoms, via Hch ( approximately 17% of DMPC-Chol links are multiple). DMPC's gamma-chain carbonyl oxygen atom is involved in 44% of water bridges and 51% of direct H bonds formed between DMPC and Chol. On average, a Chol molecule forms 0.9 links with DMPC molecules, while a DMPC molecule forms 2.2 and 0.3 links with DMPC and Chol molecules, respectively. OH-Chol makes hydrogen bonds with 1.1 water molecules, preferentially via Hch. The average number of water molecules H bonded to the DMPC headgroup is increased by 7% in the presence of Chol. These results indicate that inclusion of Chol decreases interlipid links and increases hydration in the polar region of the membrane.     

A Molecular Dynamics Study of DMPC Lipid Bilayers Interacting with Dimethylsulfoxide–Water Mixtures

The diffusion process of dimethylsulfoxide (DMSO) through zwitterionic dimyristoylphosphatidylcholine (DMPC) lipid bilayer was studied by means of molecular dynamics (MD) simulations. To account for the cryoprotectant concentration difference between the inside and the outside of the cell, dual DMPC lipid bilayers which separate two aqueous reservoirs with and without DMSO were modeled. The initial configuration of the simulation model had DMSO molecules present in one of the aqueous phases (outside the cell) at two different concentrations of ~3 and ~6?mol%. MD simulations were performed on the systems for 50?ns at 323?K and 1?bar. Although the simulation time considered in the study was insufficient for the DMSO molecules to reach the other aqueous phase and equilibrium, early stages of the diffusion process indicated that DMSO molecules had a tendency to diffuse towards the other aqueous phase. The effects of DMSO on bilayer structural characteristics during the diffusion process were investigated. Simulations were analyzed to correlate the following properties of lipid bilayers in the presence of two different aqueous phases: area per lipid, lipid thickness, mass density profiles, lipid tail order parameter and water dipole orientation. Area per lipid calculated for the leaflet facing the aqueous DMSO?Cwater mixture did not show any significant difference compared to area per lipid for the DMSO-free pure DMPC bilayer. Mass density profiles revealed that DMSO molecules had a strong tendency to diffuse toward the aqueous phase with pure water. The lipid tail order parameter calculated for the sn-1 tail of the leaflet facing the aqueous DMSO?Cwater mixture showed that the ordering of lipid tails decreased compared to the leaflet exposed to pure water. However, the ordering of lipid tails in a system where a single bilayer is hydrated by an aqueous DMSO?Cwater mixture is far lower.     

An artificial lipid bilayer formed on an agarose-coated glass for simultaneous electrical and optical measurement of single ion channels

The purpose of this study is to develop an apparatus for simultaneous measurement of electrical and spectroscopic parameters of single ion channels. We have combined the single channel recording apparatus with an artificial lipid bilayer and a fluorescence microscope designed to detect single fluorescent molecules. The artificial membranes were formed on an agarose-coated glass and observed with an objective-type total internal reflection fluorescence microscope (TIRFM). The lateral motion of a single lipid molecule (beta-BODIPY 530/550 HPC) was recorded. The lateral diffusion constant of the lipid molecule was calculated from the trajectories of single molecules as D = 8.5 +/- 4.9 x 10(-8) cm(2)/s. Ionic channels were incorporated into the membrane and current fluctuations were recorded at the single-channel level. After incorporation of Cy3-labeled alametithin molecules into the membrane, bright spots were observed moving rather slowly (D = 4.0 +/- 1.6 x 10(-8) cm(2)/s) in the membrane, simultaneously with the alametithin-channel current. These data show the possibility of the present technique for simultaneous measurement of electrical and spectroscopic parameters of single-channel activities.     

Single-molecule anisotropy imaging

A novel method, single-molecule anisotropy imaging, has been employed to simultaneously study lateral and rotational diffusion of fluorescence-labeled lipids on supported phospholipid membranes. In a fluid membrane composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, in which the rotational diffusion time is on the order of the excited-state lifetime of the fluorophore rhodamine, a rotational diffusion constant, D(rot) = 7 x 10(7) rad(2)/s, was determined. The lateral diffusion constant, measured by direct analysis of single-molecule trajectories, was D(lat) = 3.5 x 10(-8) cm(2)/s. As predicted from the free-volume model for diffusion, the results exhibit a significantly enhanced mobility on the nanosecond time scale. For membranes of DPPC lipids in the L(beta) gel phase, the slow rotational mobility permitted the direct observation of the rotation of individual molecules characterized by D(rot) = 1.2 rad(2)/s. The latter data were evaluated by a mean square angular displacement analysis. The technique developed here should prove itself profitable for imaging of conformational motions of individual proteins on the time scale of milliseconds to seconds.     

Molecular aspects of the interaction between amphotericin B and a phospholipid bilayer: molecular dynamics studies

Amphotericin B (AmB) is a polyene macrolide antibiotic used to treat systemic fungal infections. The molecular mechanism of AmB action is still only partly characterized. AmB interacts with cell-membrane components and forms membrane channels that eventually lead to cell death. The interaction between AmB and the membrane surface can be regarded as the first (presumably crucial) step on the way to channel formation. In this study molecular dynamics simulations were performed for an AmB–lipid bilayer model in order to characterize the molecular aspects of AmB–membrane interactions. The system studied contained a box of 200 dimyristoylphosphatidylcholine (DMPC) molecules, a single AmB molecule placed on the surface of the lipid bilayer and 8,065 water molecules. Two molecular dynamics simulations (NVT ensemble), each lasting 1 ns, were performed for the model studied. Two different programs, CHARMM and NAMD2, were used in order to test simulation conditions. The analysis of MD trajectories brought interesting information concerning interactions between polar groups of AmB and both DMPC and water molecules. Our studies show that AmB preferentially took a vertical position, perpendicular to the membrane surface, with no propensity to enter the membrane. Our finding may suggest that a single AmB molecule entering the membrane is very unlikely.Figure The figure presents the whole structure of the system simulated—starting point. AmB is presented as a space-filling model, DMPC molecules—green sticks, water molecules—red sticks     

Molecular dynamics simulations of local anesthetic articaine in a lipid bilayer

In order to investigate structural and dynamical properties of local anesthetic articaine in a model lipid bilayer, a series of molecular dynamics simulations have been performed. Simulations were carried out for neutral and charged (protonated) forms of articaine inserted in fully hydrated dimyristoylphosphatidylcholine (DMPC) lipid bilayer. For comparison purpose, a fully hydrated DMPC bilayer without articaine was also simulated. The length of each simulation was 200 ns. Various properties of the lipid bilayer systems in the presence of both charged and uncharged forms of articaine taken at two different concentrations have been examined: membrane area per lipid, mass density distributions, order parameters, radial distribution functions, head group tilt, diffusion coefficients, electrostatic potential, etc, and compared with results of previous simulations of DMPC bilayer in the presence of lidocaine. It was shown that addition of both charged and neutral forms of articaine causes increase of the dipole electrostatic potential in the membrane interior.     

The influence of cholesterol on interactions and dynamics of ibuprofen in a lipid bilayer

In this work, molecular dynamics (MD) simulations with atomistic details were performed to examine the influence of the cholesterol on the interactions and the partitioning of the hydrophobic drug ibuprofen in a fully hydrated 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayer. Analysis of MD simulations indicated that ibuprofen molecules prefer to be located in the hydrophobic acyl chain region of DMPC/cholesterol bilayers. This distribution decreases the lateral motion of lipid molecules. The presence of ibuprofen molecules in the bilayers with 0 and 25 mol% cholesterol increases the ordering of hydrocarbon tails of lipids whereas for the bilayers with 50 mol% cholesterol, ibuprofen molecules perturb the flexible chains of DMPC lipids which leads to the reduction of the acyl chain order parameter. The potential of the mean force (PMF) method was used to calculate the free energy profile for the transferring of an ibuprofen molecule from the bulk water into the DMPC/cholesterol membranes. The PMF studies indicated that the presence of 50 mol% cholesterol in the bilayers increases the free energy barrier and slows down the permeation of the ibuprofen drug across the DMPC bilayer. This can be due to the condensing and ordering effects of the cholesterol on the bilayer.     

Lateral diffusion of small compounds in human stratum corneum and model lipid bilayer systems.

An image-based technique of fluorescence recovery after photobleaching (video-FRAP) was used to measure the lateral diffusion coefficients of a series of nine fluorescent probes in two model lipid bilayer systems, dimyristoylphosphatidylcholine (DMPC) and DMPC/cholesterol (40 mol%), as well as in human stratum corneum-extracted lipids. The probes were all lipophilic, varied in molecular weight from 223 to 854 Da, and were chosen to characterize the lateral diffusion of small compounds in these bilayer systems. A clear molecular weight dependence of the lateral diffusion coefficients in DMPC bilayers was observed. Values ranged from 6.72 x 10(-8) to 16.2 x 10(-8) cm2/s, with the smaller probes diffusing faster than the larger ones. Measurements in DMPC/cholesterol bilayers, which represent the most thorough characterization of small-solute diffusion in this system, exhibited a similar molecular weight dependence, although the diffusion coefficients were lower, ranging from 1.62 x 10(-8) to 5.60 x 10(-8) cm2/s. Lateral diffusion measurements in stratum corneum-extracted lipids, which represent a novel examination of diffusion in this unique lipid system, also exhibited a molecular weight dependence, with values ranging from 0.306 x 10(-8) to 2.34 x 10(-8) cm2/s. Literature data showed that these strong molecular weight dependencies extend to even smaller compounds than those examined in this study. A two-parameter empirical expression is presented that describes the lateral diffusion coefficient in terms of the solute's molecular weight and captures the size dependence over the range examined. This study illustrates the degree to which small-molecule lateral diffusion in stratum corneum-extracted lipids can be represented by diffusion in DMPC and DMPC/cholesterol bilayer systems, and may lead to a better understanding of small-solute transport across human stratum corneum.     

Flexibility of ras lipid modifications studied by 2H solid-state NMR and molecular dynamics simulations

Human posttranslationally modified N-ras oncogenes are known to be implicated in numerous human cancers. Here, we applied a combination of experimental and computational techniques to determine structural and dynamical details of the lipid chain modifications of an N-ras heptapeptide in 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) membranes. Experimentally, 2H NMR spectroscopy was used to study oriented membranes that incorporated ras heptapeptides with two covalently attached perdeuterated hexadecyl chains. Atomistic molecular dynamics simulations of the same system were carried out over 100 ns including 60 DMPC and 4 ras molecules. Several structural and dynamical experimental parameters could be directly compared to the simulation. Experimental and simulated 2H NMR order parameters for the methylene groups of the ras lipid chains exhibited a systematic difference attributable to the absence of collective motions in the simulation and to geometrical effects. In contrast, experimental 2H NMR spin-lattice relaxation rates for Zeeman order were well reproduced in the simulation. The lack of slower collective motions in the simulation did not appreciably influence the relaxation rates at a Larmor frequency of 115.1 MHz. The experimental angular dependence of the 2H NMR relaxation rates with respect to the external magnetic field was also relatively well simulated. These relaxation rates showed a weak angular dependence, suggesting that the lipid modifications of ras are very flexible and highly mobile in agreement with the low order parameters. To quantify these results, the angular dependence of the 2H relaxation rates was calculated by an analytical model considering both molecular and collective motions. Peptide dynamics in the membrane could be modeled by an anisotropic diffusion tensor with principal values of Dparallel=2.1x10(9) s(-1) and Dperpendicular=4.5x10(5) s(-1). A viscoelastic fitting parameter describing the membrane elasticity, viscosity, and temperature was found to be relatively similar for the ras peptide and the DMPC host matrix. Large motional amplitudes and relatively short correlation times facilitate mixing and dispersal with the lipid bilayer matrix, with implications for the role of the full-length ras protein in signal transduction and oncogenesis.     

Bilayer thickness and lipid interface area in unilamellar extruded 1,2-diacylphosphatidylcholine liposomes: a small-angle neutron scattering study

Small-angle neutron scattering (SANS) experiments have been performed on large unilamellar liposomes prepared from 1,2-dilauroylphosphatidylcholine (DLPC), 1,2-dimyristoyl-phosphatidylcholine (DMPC) and 1,2-distearoylphosphatidylcholine (DSPC) in heavy water by extrusion through polycarbonate filters with 500 A pores. The neutron scattering intensity I(Q) in the region of scattering vectors Q corresponding to 0.0015 A(-2) < or = Q(2) < or = 0.0115 A(-2) was fitted using a step function model of bilayer neutron scattering length density and supposing that the liposomes are spherical and have a Gaussian distribution of radii. Using the lipid volumetric data, and supposing that the thickness of bilayer polar region equals to d(H) = 9+/-1 A and the water molecular volume intercalated in the bilayer polar region is the same as in the aqueous bulk aqueous phase, the steric bilayer thickness d(L), the lipid surface area A(L) and the number of water molecules per lipid molecule N intercalated in the bilayer polar region were obtained: d(L) = 41.58+/-1.93 A, A(L) = 57.18+/-1.00 A(2) and N = 6.53+/-1.93 in DLPC at 20 degrees C, d(L) = 44.26+/-1.42 A, A(L) = 60.01+/-0.75 A(2) and N = 7.37+/-1.94 in DMPC at 36 degrees C, and d(L) = 49.77+/-1.52 A, A(L) = 64.78+/-0.46 A(2) and N = 8.67+/-1.97 in DSPC at 60 degrees C. After correcting for area thermal expansivity alpha approximately 0.00417 K(-1), the lipid surface area shows a decrease with the lipid acyl chain length at 60 degrees C: A(L) = 67.56+/-1.18 A(2) in DLPC, A(L) = 66.33+/-0.83 A(2) in DMPC and A(L) = 64.78+/-0.46 A(2) in DSPC. It is also shown that a joint evaluation of SANS and small-angle X-ray scattering on unilamellar liposomes can be used to obtain the value of d(H) and the distance of the lipid phosphate group from the bilayer hydrocarbon region d(H1).     

Molecular Dynamics Simulation of Conformational Flexibility of Alamethicin Fragments in Aqueous and Membranous Environment

Abstract

We present here results on molecular dynamics (MD) simulation on two fragments of channel forming antibiotic peptide Alamethicin, containing isoamino butyric acid (Aib). Simulations are carried out in aqueous and membranous environment in a bilayer of 39 molecules of Dimyristoyl phosphatidyl choline (DMPC). The peptides Boc—;Pro-Aib-Ala-Aib- OBzl (Alam 1) and Boc-Leu-Aib-Pro-OBzl (Alam 2) were simulated from their crystallography coordinates. The bilayers were built from two different conformations (A and B) of DMPC reported in crystal data. The P-N dipoles were arranged hexagonally with surface area per lipid molecule 66.5 A°² and P-P separation across the bilayer 34 A°. They were hydrated by 28.6 and 25.5 water molecules per DMPC molecule. Simulations are done using AMBER 4.0 package in constant number volume temperature (NVT) condition for 100 pico seconds (ps) in aqueous environment and 250 ps of equilibrated bilayer. Geometric parameters of lipids as: bilayer thickness, order parameter of the chains, transfraction of chain torsional angles were monitored. We also monitored geometric parameters of the peptides as backbone torsional angles, distances amongst Ca atoms, angles between Cα atoms, movement of center of gravity (CG) along and perpendicular to bilayer normal. We find that membrane bilayer is slightly disturbed due to the presence of peptides. In case of alam 2 in water angles ψ1 and ψ3 showed larger variation in water compared to same in the bilayer. The peptide conformation is more stable in DMPC bilayer. However the peptides showed movement along and perpendicular to bilayer normal. This we believe is due to hydrophobic nature of these peptides.     

Phosphatidylinositol induces fluid phase formation and packing defects in phosphatidylcholine model membranes

Liposomes consisted of phosphatidylinositol (PI) and phosphatidylcholine (PC) have been utilized as delivery vehicle for drugs and proteins. In the present work, we studied the effect of soy PI on physical properties of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) liposomes such as phase state of lipid bilayer, lipid packing and phase properties using multiple orthogonal biophysical techniques. The 6-dodecanoyl-2-dimethylamino naphthalene (Laurdan) fluorescence studies showed that presence of PI induces the formation of fluid phases in DMPC. Differential scanning calorimetry (DSC), temperature dependent fluorescence anisotropy measurements, and generalized polarization values for Laurdan showed that the presence of as low as 10mol% of PI induces substantial broadening and shift to lower temperature of phase transition of DMPC. The fluorescence emission intensity of DPH labeled, PI containing DMPC lipid bilayer decreased possibly due to deeper penetration of water molecules in lipid bilayer. In order to further delineate the effect of PI on the physico chemical properties of DMPC is due to either significant hydrophobic mismatch between the acyl chains of the DMPC and that of soy PI or due to the inositol head group, we systematically replaced soy PI with PC species of similar acyl chain composition (DPPC and 18:2 (Cis) PC) or with diacylglycerol (DAG), respectively. The anisotropy of PC membrane containing soy PI showed largest fluidity change compared to other compositions. The data suggests that addition of PI alters structure and dynamics of DMPC bilayer in that it promotes deeper water penetration in the bilayer, induces fluid phase characteristics and causes lipid packing defects that involve its inositol head group.     

Interaction of PEGylated anti-hypertensive drugs,amlodipine, atenolol and lisinopril with lipid bilayer membrane: A molecular dynamics simulation study

The interaction of PEGylated anti-hypertensive drugs, amlodipine, atenolol and lisinopril with lipid bilayer membrane dimyristoylphosphatidylcholine (DMPC) has been studied in nine different simulation systems consisting of 128 lipid molecules and appropriate number of water molecules by molecular dynamics method and by utilizing GROMACS software. The influences of PEGylation on the mentioned drugs and the differences in application of two types of spacer molecules on the performance of drugs and DMPC membrane have been evaluated and mass density of the components in the simulation box, mean square displacement (MSD), electrostatic potential, hydrogen bonding, radial distribution function (RDF), area per lipid, order parameter, and angle distribution of the component molecules including drug, DMPC and PEG has been investigated. Furthermore, umbrella sampling analysis indicated that, PEGylation of the drugs made amlodipine to behave more hydrophilic, whereas in case of lisinopril and atenolol, PEGylation made these drugs to behave more hydrophobic. In almost all of the simulated systems, PEGylation increased the diffusion coefficient of the drugs.     

Monte Carlo simulation of two-component bilayers: DMPC/DSPC mixtures

In this paper, we describe a relatively simple lattice model of a two-component, two-state phospholipid bilayer. Application of Monte Carlo methods to this model permits simulation of the observed excess heat capacity versus temperature curves of dimyristoylphosphatidylcholine (DMPC)/distearoylphosphatidylcholine (DSPC) mixtures as well as the lateral distributions of the components and properties related to these distributions. The analysis of the bilayer energy distribution functions reveals that the gel-fluid transition is a continuous transition for DMPC, DSPC, and all DMPC/DSPC mixtures. A comparison of the thermodynamic properties of DMPC/DSPC mixtures with the configurational properties shows that the temperatures characteristics of the configurational properties correlate well with the maxima in the excess heat capacity curves rather than with the onset and completion temperatures of the gel-fluid transition. In the gel-fluid coexistence region, we also found excellent agreement between the threshold temperatures at different system compositions detected in fluorescence recovery after photobleaching experiments and the temperatures at which the percolation probability of the gel clusters is 0.36. At every composition, the calculated mole fraction of gel state molecules at the fluorescence recovery after photobleaching threshold is 0.34 and, at the percolation threshold of gel clusters, it is 0.24. The percolation threshold mole fraction of gel or fluid lipid depends on the packing geometry of the molecules and the interchain interactions. However, it is independent of temperature, system composition, and state of the percolating cluster.     

Interaction of aspirin (acetylsalicylic acid) with lipid membranes

We studied the interaction of Aspirin (acetylsalicylic acid) with lipid membranes using x-ray diffraction for bilayers containing up to 50 mol% of aspirin. From 2D x-ray intensity maps that cover large areas of reciprocal space we determined the position of the ASA molecules in the phospholipid bilayers and the molecular arrangement of the molecules in the plane of the membranes. We present direct experimental evidence that ASA molecules participate in saturated lipid bilayers of DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine) and preferably reside in the head group region of the membrane. Up to 50 mol% ASA molecules can be dissolved in this type of bilayer before the lateral membrane organization is disturbed and the membranes are found to form an ordered, 2D crystal-like structure. Furthermore, ASA and cholesterol were found to co-exist in saturated lipid bilayers, with the ASA molecules residing in the head group region and the cholesterol molecules participating in the hydrophobic membrane core.     

Charge pairing of headgroups in phosphatidylcholine membranes: A molecular dynamics simulation study

Molecular dynamics simulation of the hydrated dimyristoylphosphatidylcholine (DMPC) bilayer membrane in the liquid-crystalline phase was carried out for 5 ns to study the interaction among DMPC headgroups in the membrane/water interface region. The phosphatidylcholine headgroup contains a positively charged choline group and negatively charged phosphate and carbonyl groups, although it is a neutral molecule as a whole. Our previous study (Pasenkiewicz-Gierula, M., Y. Takaoka, H. Miyagawa, K. Kitamura, and A. Kusumi. 1997. J. Phys. Chem. 101:3677-3691) showed the formation of water cross-bridges between negatively charged groups in which a water molecule is simultaneously hydrogen bonded to two DMPC molecules. Water bridges link 76% of DMPC molecules in the membrane. In the present study we show that relatively stable charge associations (charge pairs) are formed between the positively and negatively charged groups of two DMPC molecules. Charge pairs link 93% of DMPC molecules in the membrane. Water bridges and charge pairs together form an extended network of interactions among DMPC headgroups linking 98% of all membrane phospholipids. The average lifetimes of DMPC-DMPC associations via charge pairs, water bridges and both, are at least 730, 1400, and over 1500 ps, respectively. However, these associations are dynamic states and they break and re-form several times during their lifetime.     

The effects of Ca2+ on lipid diffusion

The effects of Ca2+ on rotational and translational diffusion of lipids in multilamellar dimyristoylphosphatidylcholine (DMPC)-water systems were investigated by time-resolved phosphorescence anisotropy steady-state fluorescence polarization and fluorescence recovery after photobleaching (FRAP) experiments. Above the phase transition temperature (Tm), addition of Ca2+ caused a steady increase in the segmental motion of the phosphorescent probe, but resulted in slower diffusion of the fluorescent and lateral diffusion probes. The former result is attributed to changes in the structure of the lipid/water interface that affects the chromophore mobility on the phosphorescence time scale but does not reflect lipid motion. Below the phase transition temperature, slower diffusion of all probes were observed with increasing concentrations of Ca2+, with sudden large changes occurring at [Ca2+] approximately 500 mM. This behaviour is attributed to association of Ca2+ with the lipid phosphate groups and the exclusion of water molecules which results in tighter packing of lipids and smaller segmental motion, leading eventually to the immobilization of lipid molecules.     

Effects of epicholesterol on the phosphatidylcholine bilayer: a molecular simulation study

Epicholesterol (Echol) is an epimeric form of cholesterol (Chol). A molecular dynamics simulation of the fully hydrated dimyristoylphosphatidylcholine-Echol (DMPC-Echol) bilayer membrane containing approximately 22 mol % of Echol was carried out for 5 ns. A 3-ns trajectory generated between 2 and 5 ns of molecular dynamics simulation was used for analyses to determine the effects of Echol on the membrane properties. As reference systems, pure DMPC and mixed DMPC-Chol bilayers were used. The study shows that Echol, like Chol, changes the organization of the bilayer/water interface and increases membrane order and condensation, but to a lesser degree. Effects of both sterols are based on the same atomic level mechanisms; their different strength arises from different vertical localizations of Echol and Chol hydroxyl groups in the membrane/water interface.     

Transmembrane helix structure, dynamics, and interactions: multi-nanosecond molecular dynamics simulations.

To probe the fundamentals of membrane/protein interactions, all-atom multi-nanosecond molecular dynamics simulations were conducted on a single transmembrane poly(32)alanine helix in a fully solvated dimyristoyphosphatidylcholine (DMPC) bilayer. The central 12 residues, which interact only with the lipid hydrocarbon chains, maintained a very stable helical structure. Helical regions extended beyond these central 12 residues, but interactions with the lipid fatty-acyl ester linkages, the lipid headgroups, and water molecules made the helix less stable in this region. The C and N termini, exposed largely to water, existed as random coils. As a whole, the helix tilted substantially, from perpendicular to the bilayer plane (0 degree) to a 30 degrees tilt. The helix experienced a bend at its middle, and the two halves of the helix at times assumed substantially different tilts. Frequent hydrogen bonding, of up to 0.7 ns in duration, occurred between peptide and lipid molecules. This resulted in correlated translational diffusion between the helix and a few lipid molecules. Because of the large variation in lipid conformation, the lipid environment of the peptide was not well defined in terms of "annular" lipids and on average consisted of 18 lipid molecules. When compared with a "neat" bilayer without peptide, no significant difference was seen in the bilayer thickness, lipid conformations or diffusion, or headgroup orientation. However, the lipid hydrocarbon chain order parameters showed a significant decrease in order, especially in those methylene groups closest to the headgroup.     

The rotational diffusion of chloroplast phosphate translocator and of lipid molecules in bilayer membranes

The rotational mobility of the phosphate translocator from the chloroplast envelope and of lipid molecules in the membrane of unilamellar azolectin liposomes has been investigated. The rotational dynamics of the liposome membrane were investigated by measuring the rotational diffusion of eosin-5-isothiocyanate(EITC)-labeled L-alpha-dipalmitoylglycerophosphoethanolamine (Pam2 GroPEtn) in the lipid phase of the vesicles, either in the presence or absence of the reconstituted phosphate translocator. The temperature dependence of the anisotropy decay showed that above 25 degrees C the main contribution to the anisotropy decay was caused by uniaxial anisotropic rotation of the labelled lipid molecules around the axis normal to the membrane plane. The rate of rotation of the labelled lipid molecules was strongly dependent on the viscosity of the medium (eta 1). Extrapolation to eta 1 = 0 Pa.s yielded a correlation time of phi = 20 +/- 5 ns, t = 30 degrees C, for lipid rotation with respect to the membrane normal. The rotational diffusion coefficient of the lipid molecules was calculated to be Dr = 2.0 x 10(9) rad2.s-1 and the apparent microviscosity in the vesicle membrane, as derived from the rotational correlation time, was eta 2 approximately 12 mPa.s. The rotational correlation time of the phosphate translocator in the membrane was only slightly dependent on the viscosity of the medium. The temperature dependence of the protein rotation also indicated that the rotation of the protein in the membrane was largely restricted and occurred mainly about the axis normal to the membrane plane. Measurements at a medium viscosity of eta 1 = 1 mPa.s yielded a value of phi r approximately 450 ns corresponding to Dr = 8.8 x 10(7) rad2.s-1 for protein rotation with respect to the membrane normal. From this value and the data of the lipid rotation, the cross-sectional area of the protein part embedded in the membrane was calculated to be approximately 9 nm2. This cross-sectional area is large enough to include at most 14 membrane-spanning helices. Our results also indicated that at lipid/protein molar ratios greater than or equal to 1.5 x 10(4): 1 aggregation occurred in the model membranes below 30 degrees C. However, above 30 degrees C and at a high dilution of the protein in the membrane it appeared that the membrane viscosity monitored by lipid and protein rotational diffusion were identical.