Interactions of cationic-hydrophobic peptides with lipid bilayers: a Monte Carlo simulation method

We present a computational model of the interaction between hydrophobic cations, such as the antimicrobial peptide, Magainin2, and membranes that include anionic lipids. The peptide's amino acids were represented as two interaction sites: one corresponds to the backbone alpha-carbon and the other to the side chain. The membrane was represented as a hydrophobic profile, and its anionic nature was represented by a surface of smeared charges. Thus, the Coulombic interactions between the peptide and the membrane were calculated using the Gouy-Chapman theory that describes the electrostatic potential in the aqueous phase near the membrane. Peptide conformations and locations near the membrane, and changes in the membrane width, were sampled at random, using the Metropolis criterion, taking into account the underlying energetics. Simulations of the interactions of heptalysine and the hydrophobic-cationic peptide, Magainin2, with acidic membranes were used to calibrate the model. The calibrated model reproduced structural data and the membrane-association free energies that were measured also for other basic and hydrophobic-cationic peptides. Interestingly, amphipathic peptides, such as Magainin2, were found to adopt two main membrane-associated states. In the first, the peptide resided mostly outside the polar headgroups region. In the second, which was energetically more favorable, the peptide assumed an amphipathic-helix conformation, where its hydrophobic face was immersed in the hydrocarbon region of the membrane and the charged residues were in contact with the surface of smeared charges. This dual behavior provides a molecular interpretation of the available experimental data.     

Interactions of the M2delta segment of the acetylcholine receptor with lipid bilayers: a continuum-solvent model study

M2delta, one of the transmembrane segments of the nicotinic acetylcholine receptor, is a 23-amino-acid peptide, frequently used as a model for peptide-membrane interactions. In this and the companion article we describe studies of M2delta-membrane interactions, using two different computational approaches. In the present work, we used continuum-solvent model calculations to investigate key thermodynamic aspects of its interactions with lipid bilayers. M2delta was represented in atomic detail and the bilayer was represented as a hydrophobic slab embedded in a structureless aqueous phase. Our calculations show that the transmembrane orientation is the most favorable orientation of the peptide in the bilayer, in good agreement with both experimental and computational data. Moreover, our calculations produced the free energy of association of M2delta with the lipid bilayer, which, to our knowledge, has not been reported to date. The calculations included 10 structures of M2delta, determined by nuclear magnetic resonance in dodecylphosphocholine micelles. All the structures were found to be stable inside the lipid bilayer, although their water-to-membrane transfer free energies differed by as much as 12 kT. Although most of the structures were roughly linear, a single structure had a kink in its central region. Interestingly, this structure was found to be the most stable inside the lipid bilayer, in agreement with molecular dynamics simulations of the peptide and with the recently determined structure of the intact receptor. Our analysis showed that the kink reduced the polarity of the peptide in its central region by allowing the electrostatic masking of the Gln13 side chain in that area. Our calculations also showed a tendency for the membrane to deform in response to peptide insertion, as has been previously found for the membrane-active peptides alamethicin and gramicidin. The results are compared to Monte Carlo simulations of the peptide-membrane system, as presented in the accompanying article.     

Molecular dynamics simulations of model trans-membrane peptides in lipid bilayers: a systematic investigation of hydrophobic mismatch

Hydrophobic mismatch, which is the difference between the hydrophobic length of trans-membrane segments of a protein and the hydrophobic width of the surrounding lipid bilayer, is known to play a role in membrane protein function. We have performed molecular dynamics simulations of trans-membrane KALP peptides (sequence: GKK(LA)nLKKA) in phospholipid bilayers to investigate hydrophobic mismatch alleviation mechanisms. By varying systematically the length of the peptide (KALP15, KALP19, KALP23, KALP27, and KALP31) and the lipid hydrophobic length (DLPC, DMPC, and DPPC), a wide range of mismatch conditions were studied. Simulations of durations of 50-200 ns show that under positive mismatch, the system alleviates the mismatch predominantly by tilting the peptide and to a smaller extent by increased lipid ordering in the immediate vicinity of the peptide. Under negative mismatch, alleviation takes place by a combination of local bilayer bending and the snorkeling of the lysine residues of the peptide. Simulations performed at a higher peptide/lipid molar ratio (1:25) reveal slower dynamics of both the peptide and lipid relative to those at a lower peptide/lipid ratio (1:128). The lysine residues have favorable interactions with specific oxygen atoms of the phospholipid headgroups, indicating the preferred localization of these residues at the lipid/water interface.     

Monte Carlo studies of the hydrocarbon region of lipid bilayers

We present the results of a Monte Carlo study of systems of hydrocarbon chains attached to a plane interface and interacting through hard core repulsive forces only. The chain-order parameters which we find in our studies are compared to experimental results (NMR and ESR). The role of “kink” states and the relevance of our studies to theoretical models are also discussed.     

Interactions of hexachlorocyclohexanes with lipid bilayers

Quenching of the fluorescence of a hydrophobic analogue of tryptophan incorporated into lipid bilayers has been used to measure partition coefficients for lindane and the α- and δ-isomers of hexachlorocyclohexane. Partition coefficients between water and lipid are comparable to those between water and octanol and exhibit a negative temperature coefficient. Binding to the lipid phase is limited by saturation of the aqueous phase rather than of the lipid phase. The binding of lindane has no detectable effect on membrane fluidity as measured by fluorescence polarisation of diphenylhexatriene, or on the permeability properties of the membrane, as measured by the leak of carboxyfluorescein.     

Grazers and grass: Monte Carlo simulations

To illustrate the interplay between grazers and grass, we present a novel Monte Carlo model including grass-island growth, consumption of grass by grazers, and birth, migration and death of grazers. The rates of the former and three latter processes are assumed to depend on the environment so that the conventional mean-field approximation does not hold (in particular, the model takes into account that grass grows on the grass-island boundaries, and grazers are mobile and prefer to stay on the areas covered by grass). Due to the feedback between various processes, as expected, the model predicts stable regimes and irregular oscillations of the area of the grass islands and grazer population. The patterns observed are however different compared to those predicted by conventional Monte Carlo prey-predator models. Specifically, there is no tendency for grazers and grass to segregate. The mean-field version of the model is briefly discussed as well.     

Probing the interaction forces between hydrophobic peptides and supported lipid bilayers using AFM

Despite the vast body of literature that has accumulated on tilted peptides in the past decade, direct information on the forces that drive their interaction with lipid membranes is lacking. Here, we attempted to use atomic force microscopy (AFM) to explore the interaction forces between the Simian immunodeficiency virus peptide and phase-separated supported bilayers composed of various lipids, i.e. dipalmitoylphosphatidylcholine, dioleoylphosphatidylcholine, dioleoylphosphatidic acid and dipalmitoylphosphatidylethanolamine. Histidine-tagged peptides were attached onto AFM tips terminated with nitrilotriacetate and tri(ethylene glycol) groups, an approach expected to ensure optimal exposure of the C-terminal hydrophobic domain. Force-distance curves recorded between peptide-tips and the different bilayer domains always showed a long-range repulsion upon approach and a lack of adhesion upon retraction, in marked contrast with the hydrophobic nature of the peptide. To explain this unexpected behaviour, we suggest a mechanism in which lipids are pulled out from the bilayer due to strong interactions with the peptide-tip, in agreement with the very low force needed to extract lipids from supported bilayers.     

Flexible charged macromolecules on mixed fluid lipid membranes: theory and Monte Carlo simulations

Fluid membranes containing charged lipids enhance binding of oppositely charged proteins by mobilizing these lipids into the interaction zone, overcoming the concomitant entropic losses due to lipid segregation and lower conformational freedom upon macromolecule adsorption. We study this energetic-entropic interplay using Monte Carlo simulations and theory. Our model system consists of a flexible cationic polyelectrolyte, interacting, via Debye-Hückel and short-ranged repulsive potentials, with membranes containing neutral lipids, 1% tetravalent, and 10% (or 1%) monovalent anionic lipids. Adsorption onto a fluid membrane is invariably stronger than to an equally charged frozen or uniform membrane. Although monovalent lipids may suffice for binding rigid macromolecules, polyvalent counter-lipids (e.g., phosphatidylinositol 4,5 bisphosphate), whose entropy loss upon localization is negligible, are crucial for binding flexible macromolecules, which lose conformational entropy upon adsorption. Extending Rosenbluth's Monte Carlo scheme we directly simulate polymer adsorption on fluid membranes. Yet, we argue that similar information could be derived from a biased superposition of quenched membrane simulations. Using a simple cell model we account for surface concentration effects, and show that the average adsorption probabilities on annealed and quenched membranes coincide at vanishing surface concentrations. We discuss the relevance of our model to the electrostatic-switch mechanism of, e.g., the myristoylated alanine-rich C kinase substrate protein.     

Monte Carlo simulations of peptide solvation

To increase our understanding of peptide–water interactions, we are simulating the behavior of water molecules in the intermolecular channels of [Phe⁴Val⁶]antamanide dododecahydrate crystals. There is good overall agreement between the positions predicted using two alternative potential functions and those that have been observed by x-ray diffraction. Detailed differences between the predictions for the two potential functions are discussed.     

Interaction of small peptides with lipid bilayers.

Molecular dynamics simulations of the tripeptide Ala-Phe-Ala-O-tert-butyl interacting with dimyristoylphosphatidylcholine lipid bilayers have been carried out. The lipid and aqueous environments of the peptide, the alkyl chain order, and the lipid and peptide dynamics have been investigated with use of density profiles, radial distribution functions, alkyl chain order parameter profiles, and time correlation functions. It appears that the alkyl chain region accommodates the peptides in the bilayer with minimal perturbation to this region. The peptide dynamics in the bilayer bound form has been compared with that of the free peptide in water. The peptide structure does not vary on the simulation time scale (of the order of hundreds of picoseconds) compared with the solution structure in which a random structure is observed.     

Component and state separation in DMPC/DSPC lipid bilayers: a Monte Carlo simulation study

In this paper a two-state, two-component, Ising-type model is used to simulate the lateral distribution of the components and gel/fluid state acyl chains in dimyristoylphosphatidylcholine/distearoylphosphatidylcholine (DMPC/DSPC) lipid bilayers. The same model has been successful in calculating the excess heat capacity curves, the fluorescence recovery after photobleaching (FRAP) threshold temperatures, the most frequent center-to-center distances between DSPC clusters, and the fractal dimensions of gel clusters (Sugar, I. P., T. E. Thompson, and R. L. Biltonen, 1999. Biophys. J. 76:2099-2110). Depending on the temperature and mole fraction the population of the cluster size is either homogeneous or inhomogeneous. In the inhomogeneous population the size of the largest cluster scales with the size of the system, while the rest of the clusters remain small with increasing system size. In a homogeneous population, however, every cluster remains small with increasing system size. For both compositional and fluid/gel state clusters, threshold temperatures-the so-called percolation threshold temperatures-are determined where change in the type of the population takes place. At a given mole fraction, the number of percolation threshold temperatures can be 0, 1, 2, or 3. By plotting these percolation threshold temperatures on the temperature/mole fraction plane, the diagrams of component and state separation of DMPC/DSPC bilayers are constructed. In agreement with the small-angle neutron scattering measurements, the component separation diagram shows nonrandom lateral distribution of the components not only in the gel-fluid mixed phase region, but also in the pure gel and pure fluid regions. A combined diagram of component and state separation is constructed to characterize the lateral distribution of lipid components and gel/fluid state acyl chains in DMPC/DSPC mixtures. While theoretical phase diagrams of two component mixtures can be constructed only in the case of first-order transitions, state and component separation diagrams can be constructed whether or not the system is involved in first-order transition. The effects of interchain interactions on the component and state separation diagrams are demonstrated on three different models. The influences of state and component separation on the in-plane and off-plane membrane reactions are discussed.     

Clustering of tetrameric influenza M2 peptides in lipid bilayers investigated by 19F solid-state NMR

The influenza M2 protein forms a drug-targeted tetrameric proton channel to mediate virus uncoating, and carries out membrane scission to enable virus release. While the proton channel function of M2 has been extensively studied, the mechanism by which M2 catalyzes membrane scission is still not well understood. Previous fluorescence and electron microscopy studies indicated that M2 tetramers concentrate at the neck of the budding virus in the host plasma membrane. However, molecular evidence for this clustering is scarce. Here, we use ¹⁹F solid-state NMR to investigate M2 clustering in phospholipid bilayers. By mixing equimolar amounts of 4F-Phe47 labeled M2 peptide and CF₃-Phe47 labeled M2 peptide and measuring F-CF₃ cross peaks in 2D ¹⁹F¹⁹F correlation spectra, we show that M2 tetramers form nanometer-scale clusters in lipid bilayers. This clustering is stronger in cholesterol-containing membranes and phosphatidylethanolamine (PE) membranes than in cholesterol-free phosphatidylcholine and phosphatidylglycerol membranes. The observed correlation peaks indicate that Phe47 sidechains from different tetramers are less than ~2 nm apart. ¹H¹⁹F correlation peaks between lipid chain protons and fluorinated Phe47 indicate that Phe47 is more deeply inserted into the lipid bilayer in the presence of cholesterol than in its absence, suggesting that Phe47 preferentially interacts with cholesterol. Static ³¹P NMR spectra indicate that M2 induces negative Gaussian curvature in the PE membrane. These results suggest that M2 tetramers cluster at cholesterol- and PE-rich regions of cell membranes to cause membrane curvature, which in turn can facilitate membrane scission in the last step of virus budding and release.     

Interactions of Folch-Lees proteolipid apoprotein with planar lipid bilayers

Summary Water-soluble Folch-Lees proteolipid apoprotein from bovine CNS white matter induces a voltage-dependent conductance in black lipid membranes. Na⁺ is required for the induced conductance change but the established conductance has very low ionic selectivity. The induced conductance fluctuates with a minimum amplitude of 10^–11–10^–10 mho. The magnitude of the conductivity change is dependent on protein concentration and on the composition of lipid bilayers. At a fixed voltage the induced conductance of a phosphatidylcholine-cholesterol membrane is proportional to the sixth power of the protein concentration and the first power of Na⁺ concentration. The interactions between the apoprotein and the lipids are both electrostatic and hydrophobic, but the interaction leading to the conductance increase appears to be mainly hydrophobic. Both the increase in conductance and the current fluctuations remain after extensive washing of the chambers to remove the protein. Furthermore, pronase or glutaraldehyde added to either the cis or trans side of the membrane does not affect the apoprotein-established conductance. However, if the bilayer is formed in the presence of both the apoprotein and pronase or if the apoprotein is treated with pronase prior to its addition to the chamber, no conductance change is observed. The association of the apoprotein with the membrane thus appears to render the protein inaccessible to proteolytic digestion, suggesting that the apoprotein is at least partially imbedded in the membrane interior.     

Monte Carlo simulations of ionic channel selectivity

Monte Carlo simulations have been developed to study the selectivityof ionic channels in biological membranes. The channel is consideredto be in either of two possible states: (i) densely packed withions, the ions moving in single file in one direction, or alternatively,(ii) sparsely packed, where holes could appear at any particulartime thereby allowing bidirectional movement of ions. The twomodels enable us to envisage a quantitative flux of permeableions in the presence of smaller sized ions, taking into considerationtheir concentrations in the bulk solutions, the ion-channelinteractions and probability with which they fill up the channel.The programs are written in FORTRAN-77 (MS-FORTRAN) for an IBM-compatiblepersonal computer. From the simulation results we observe anenzymatic function of the channel and also note that the smallersized ions tend to block the movement of permeable ions. Thesimulations represent a technique for visualization of the factorsthat decide ionic permeability and help in manipulating theireffects with ease and speed which would otherwise involve intricateexperimental setups. Received on September 7, 1990; accepted on January 14, 1991     

Wave-guide spectroscopy on planar lipid bilayers doped with hydrophobic ions

Wave-guide spectroscopy exploits the light pipe properties of planar lipid bilayers by propagating a light wave along the plane of the bilayer. Applying this technique to the optical absorption of chromophore in the membrane, results in an enhanced sensitivity when compared to normal incidence spectroscopy. This gain factor is of the order of 100 per mm optical path along the bilayer, thus transforming the weak absorbances in lipid membranes into easily measurable quantities. Wave-guide spectroscopy has been used to measure the adsorption isotherm of hydrophobic dipicrylamine ions in a phosphatidylcholine membrane. The adsorption isotherm is linear for low aqueous concentrations, in the micromolar range however, it changes into a sublinear dependence. The addition of an inert alkali salt to the electrolyte favours the adsorption of hydrophobic ions. Current saturation is observed with the transition to the sublinear isotherm. When using the time constant for current relaxation as an indicator of changes in the magnitude of the surface potential, it does not seem to vary with the additional dipicrylamine which adsorbs in the presence of high concentrations of alkali salt in the electrolyte. A compensation of hydrophobic charge by the alkali ions from the inert electrolyte is proposed.     

The transport of hydrophobic ions across lipid bilayers

The three-capacitor model for hydrophobic ion adsorption in lipid membranes (Andersen, O.S., Feldberg, S., Nakadomari, H., Levy, S. and McLaughlin, S. (1978) Biophys. J. 21, 35-70) is extended to ion transport whereby electrostatic effects from the interfacially adsorbed hydrophobic space charge have been encountered. The phenomena of current saturation with increasing concentrations of hydrophobic ions in the bulk electrolyte and the associated increase of the time constant of current relaxation can be quantitatively understood on the basis of space charge-limited currents as well as the nonexponential current decay.     

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.     

Combined Monte Carlo and molecular dynamics simulation of hydrated lipid-cholesterol lipid bilayers at low cholesterol concentration

We have applied a hybrid equilibration and sampling procedure for the atomic level simulation of a hydrated lipid bilayer to systems consisting of dipalmitoyl phosphatidylcholine (DPPC) and cholesterol, and palmitoyl-oleyl phosphatidylcholine (POPC) at low (approximately 6%) cholesterol concentration. The procedure is applied to bilayers of 94 molecules of DPPC, 6 molecules of cholesterol, and 3205 water molecules, and to bilayers of 120 molecules of POPC, 8 molecules of cholesterol, and 4268 water molecules, at a temperature of 325 K. After equilibration, three separate 400-ps continuous molecular dynamics runs, separated by 10,000 configurational bias Monte Carlo steps, were carried out for each system. Properties of the systems were calculated and averaged over the three separate runs. Results of the simulations are presented and compared with experimental data and with other recent simulations of DPPC and cholesterol, and of pure DPPC, and pure POPC. Certain properties of the bilayers are indistinguishable from cholesterol-free bilayers, including lateral diffusion and electron density. Other properties, most notably the order parameter profile, show the effect of cholesterol even at low concentrations.