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This review summarizes recent investigations on the association of macromolecules on lipid bilayers. Hydrophilic and flexible polymers can form soft coronae tenuously adsorbed or anchored on the lipid membrane. Other synthetic macromolecules are embedded in the apolar region of the membrane. Recent experimental and theoretical works focus on the perturbation of lipid properties achieved depending on the nature and strength of binding. Of importance to biomimicry, to tethered model membranes, and drug carriers, the effects achievable include modulation of the lateral diffusivity of lipids, shape distortions, lateral segregations, formation of well-defined nanopores and ultimately the stimuli responsive disruption of the membrane.

The pH response of pH-responsive cell-penetrating peptides in cell membrane is directly associated with many potential applications and cell activities such as drug delivery, membrane fusion, and protein folding, but it is still poorly understood. In this study, we used GALA as a model and applied sum frequency generation vibrational spectroscopy to systematically investigate the pH response of GALA in lipid bilayers with different hydrophobic length and lipid head groups. We determined the GALA structures in lipid bilayers by combining second-ordered amide I and amide III spectral signals, which can accurately differentiate the loop and α-helical structures at the interface. It is found that GALA can insert into fluid-phase lipid bilayers even at neutral pH, while lies down on the gel-phase lipid bilayer surface. Under acidic conditions, GALA inserts into both fluid-phase and gel-phase lipid bilayers. GALA adopts a mixed loop and α-helical structures in lipid bilayers. Besides, the reversible activation ...

Supported lipid bilayers have the potential to deliver a breakthrough in separation processes, e.g. , in desalination. Yet, formation of macroscopically large lipid membranes for use in separations and understanding of their long-term mechanical stability, especially, when they host membrane proteins, is still a challenge. Arrays of microscopic pore-spanning lipid membranes are a promising upscalable configuration, in which lipid membranes do not contact support directly, making it attractive for studying their function and stability. Here we report on (1) the formation mechanism of an array of pore-spanning phospholipid membranes via ‘vesicle fusion’, and (2) a microfluidic device that is used to assess the stability of the pore-spanning lipid membrane under flow and osmotic gradient. It is shown that the formation of pore-spanning lipid membranes via ‘vesicles fusion’ proceeds in three steps: first, small vesicles merge into giant ones of about the size of the substrate pore size. The giant vesicles then settle at the pore mouths and flatten. Last, the flattened giant vesicles rupture and form a lipid membrane that closes the pore. Exposing the membrane to combined transmembrane osmotic and tangential shear flows in a microfluidic device, which simulates common osmotic process conditions, shows that, in addition to remaining open pores, a fraction of pore-spanning membranes ruptures. Possible ways to avoid such rupture and minimize fraction of open pores are discussed.

We study the formation of membrane budding in model lipid bilayers with the budding assumed to be driven by means of diffusion of trans-membrane proteins over a composite membrane surface. The theo...

The stochastic protrusion of hydrophobic lipid tails into solution, a subclass of hydrophobic membrane defects, has recently been shown to be a critical step in a number of biological processes like membrane fusion. Understanding the factors that govern the appearance of lipid tail protrusions is critical for identifying membrane features that affect the rate of fusion or other processes that depend on contact with solvent-exposed lipid tails. In this work, we utilize atomistic molecular dynamics simulations to characterize the likelihood of tail protrusions in phosphotidylcholine lipid bilayers of varying composition, curvature, and hydration. We distinguish two protrusion modes corresponding to atoms near the end of the lipid tail or near the glycerol group. Through potential of mean force calculations, we demonstrate that the thermodynamic cost for inducing a protrusion depends on tail saturation but is insensitive to other bilayer structural properties or hydration above a threshold value. Similarly, highly curved vesicles or micelles increase both the overall frequency of lipid tail protrusions as well as the preference for splay protrusions, both of which play an important role in driving membrane fusion. In multi-component bilayers, however, the incidence of protrusion events does not clearly depend on the mismatch between tail length or tail saturation of the constituent lipids. Together, these results provide significant physical insight into how system components might affect the appearance of protrusions in biological membranes, and help explain the roles of composition or curvature-modifying proteins in membrane fusion., National Science Foundation (U.S.). MRSEC Program (award number DMR-0819762), National Science Foundation (U.S.). Faculty Early Career Development Program (Award No. DMR-1054671), United States. Department of Energy. Computational Science Graduate Fellowship Program (grant number DE-FG02-97ER25308), National Science Foundation (U.S.) (grant number OCI-1053575)

Biomembrane interfaces create regions of slowed water dynamics in their vicinity. When two lipid bilayers come together, this effect is further accentuated, and the associated slowdown can affect the dynamics of larger-scale processes such as membrane fusion. We have used molecular dynamics simulations to examine how lipid and water dynamics are affected as two lipid bilayers approach each other. These two interacting fluid systems, lipid and water, both slow and become coupled when the lipid membranes are separated by a thin water layer. We show in particular that the water dynamics become glassy, and diffusion of lipids in the apposed leaflets becomes coupled across the water layer, while the "outer" leaflets remain unaffected. This dynamic coupling between bilayers appears mediated by lipid-water-lipid hydrogen bonding, as it occurs at bilayer separations where water-lipid hydrogen bonds become more common than water-water hydrogen bonds. We further show that such coupling occurs in simulations of vesicle-vesicle fusion prior to the fusion event itself. Such altered dynamics at membrane-membrane interfaces may both stabilize the interfacial contact and slow fusion stalk formation within the interface region.

Understanding of nanoparticle-membrane interactions is useful for various applications of nanoparticles like drug delivery and imaging. Here we report on the studies of interaction between hydrophilic charged polymer coated semiconductor quantum dot nanoparticles with model lipid membranes. Atomic force microscopy and X-ray reflectivity measurements suggest that cationic nanoparticles bind and penetrate bilayers of zwitterionic lipids. Penetration and binding depend on the extent of lipid packing and result in the disruption of the lipid bilayer accompanied by enhanced lipid diffusion. On the other hand, anionic nanoparticles show minimal membrane binding although, curiously, their interaction leads to reduction in lipid diffusivity. It is suggested that the enhanced binding of cationic QDs at higher lipid packing can be understood in terms of the effective surface potential of the bilayers which is tunable through membrane lipid packing. Our results bring forth the subtle interplay of membrane lipid packing and electrostatics which determine nanoparticle binding and penetration of model membranes with further implications for real cell membranes.

Recent literature has shown that buffers affect the interaction between lipid bilayers through a mechanism that involves van der Waals forces, electrostatics, hydration forces and membrane bending rigidity. This letter shows an additional peculiar effect of buffers on the mixed chain 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid bilayers, namely phase coexistence similar to what was reported by Rappolt et al. for alkali chlorides. The data presented suggest that one phase appears to dehydrate below the value in pure water, while the other phase swells as the concentration of buffer is increased. However, since the two phases must be in osmotic equilibrium with one another, this behavior challenges theoretical models of lipid interactions.

Interaction force-distance profiles between substrate-supported membranes composed of equimolar ternary mixtures of unsaturated phosphotidylcholine (PC) lipid, saturated PC lipid, and cholesterol were determined using the surface force apparatus. Both double and single unsaturated PC lipids were studied. In all cases, the membranes were slightly negatively charged, resulting in a weak, long-range electrostatic repulsion. Corroborative atomic force microscopy, zeta potential, and fluorescence microscopy measurements were used to establish that a small level of charged lipid impurities (∼1/400 lipid molecules) were responsible for the repulsive electrostatic interaction between the membranes. At contact, the membranes were adhesive. The magnitude of the adhesion was greater than the van der Waals interaction between pure PC membranes without cholesterol. The enhanced adhesion was primarily attributed to hydrophobic attraction due to the presence of nanoscopic membrane defects which exposed the underlying membrane leaflet. The interaction force-distance profiles also demonstrated that the nanoscopic defects enabled membrane restructuring in the contact region.

Supported lipid bilayers (SLB) comprise a very important set of model systems of biomembranes. In particular, they can be prepared on metallic surfaces such as gold electrodes, which allow a number of electrochemical studies and applications that could not be undertaken with other types of model systems, such as liposome suspensions. Also of special relevance is lipid bilayer composition, especially those combinations of lipids which permit the formation of biologically relevant membrane domains such as lipid rafts. Indeed, membrane domain organization is a crucial feature concerning not only the biophysical properties of the bilayer itself, but also the behavior and bioactivity of membrane-interacting biomolecules. In spite of its relevance, the presence of bilayer domains has not been central in many investigations involving lipid bilayers supported on conductive surfaces. Moreover, air exposed gold surface is hydrophobic, which is not ideally suitable to establish a proper interaction with the lipids polar head group. To overcome such limitation different strategies may be adopted, encompassing the fine tuning of the buffer conditions and previous surface modification by hydrophilic self-assembled monolayer (SAM) or thiolipids. This review will be focused on the formation and characterization of SLB and lipid rafts on gold substrates, illustrating the range of applications of such platforms, with examples of the study of electroactive molecules and the development of new biosensing interfaces.

The composition of natural lipid membranes varies greatly depending on the type of organism, and it has been observed that small variations in lipid composition affect dramatically the membrane properties, such as structural stability and solute permeability. One of the variations is the methyl-branched lipids commonly found in archaea and bacteria. Using molecular dynamics simulations, we studied the influence of methyl branching on the electric-field induced formation of water channels in lipid bilayers and ion transports through them. We employed a double lipid bilayer setup to create within a periodic box two water compartments separated by those two bilayers. One of the compartments contains an excess of cations, while the other an equal excess of anions. This setup creates an initial transmembrane potential controlled by the ion concentration in each water compartment. We compared the response of diphytanoylphosphatidylcholine (DPhPC) lipid bilayers that have multiple methyl-branches with that of the straight-chain dipalmitoylphosphatidylcholine (DPPC) lipids. We found that compared to the straight-chain DPPC lipids, branched DPhPC lipids require a higher critical transmembrane potential and a longer time for the membrane to break down, followed by water channel formation, and transport of anions and cations through the channel. We demonstrated that while adding methyl branches reduces the lateral diffusion of the lipids, different transport properties of branched lipids are mainly due to the bulkiness of the branched lipid tails resulting in different water channel morphologies. The transmembrane potential creates toroidal pores in the straight-chain lipid bilayers, but barrel stave pores in the branched-lipid bilayers the formation of which requires a higher transmembrane potential. Our results provided a deeper understanding of the ion transport process through lipid bilayer membranes and shed light on the transport of various molecules across the lipid membranes.

Insertion of short peptides into the cell membrane is energetically unfavorable and challenges the commonly accepted hydrophobic matching principle. Yet there has been evidence that many short peptides can penetrate into the cells to perform the biological functions in salt solution. On the basis of the previous study (J. Phys. Chem. C 2013, 117, 11095−11103), here we further performed a systematic study on the interaction of mastoparan with various neutral lipid bilayers with different lipid chain lengths in situ to examine the hydrophobic matching principle in different aqueous salt environments using sum frequency generation vibrational spectroscopy. It is found that the hydrophobic matching is the dominant driving force for the association of MP with a lipid bilayer in a pure water environment. However, in a kosmotropic ion environment, the hydration of ions can overcome the hydrophobic mismatching effects, leading to the insertion of MP into lipid bilayers with much longer hydrophobic lengths. When t...

Pore formation of lipid bilayers under mechanical stress is critical to biological processes. A series of coarse grained molecular dynamics simulations of lipid bilayers with carbon nanoparticles different in size have been performed. Surface tension was applied to study the disruption of lipid bilayers by nanoparticles and the formation of pores inside the bilayers. The presence of small nanoparticles enhances the probability of water penetration thus decreasing the membrane rupture tension, while big nanoparticles have the opposite effect. Nanoparticle volume affects bilayer strength indirectly, and particle surface density can complicate the interaction. The structural, dynamic, elastic properties and lateral densities of lipid bilayers with nanoparticles under mechanical stress were analyzed. The results demonstrate the ability of nanoparticles to adjust the structural and dynamic properties of a lipid membrane, and to efficiently regulate the pore formation behavior and hydrophobicity of membranes.

Lipid flip-flop and its associated transient pore formation are key thermodynamic properties of living cell membranes. However, there is a lack of understanding of whether ionic imbalance that exists ubiquitously across cell membranes affects lipid flip-flop and its associated functions. Potential of mean force calculations show that the free-energy barrier of lipid flip-flop on the extracellular leaflet reduces with the presence of ionic imbalance, whereas the barrier on the intracellular leaflet is generally not affected. The linear decrease of the activation energy of lipid flip-flop on the extracellular leaflet is consistent with the experimentally measured conductance-voltage relationship of zwitterionic lipid bilayers. This suggests: 1) lipid flip-flop has a directionality under physiological conditions and phospholipids accumulate at a rate on the order of 105 µm-2 h-1 on the cytoplasmic side of cell membranes; 2) ion permeation across a lipid membrane is moderated by lipid flip-flop; 3) the energy barrier of pore formation is aligned with the weaker leaflet that has a lower energy of lipid flip-flop. The asymmetry of lipid flip-flop and pore nucleation may have substantial implications for protein translocation, signaling, enzymatic activities, vesicle fusion, and transportation of biomolecules on cell membranes.

The complement system is an important part of the innate immune response, and there is great interest in understanding how complement proteins interact with lipid membrane interfaces, especially in the context of recognizing foreign particulates (e.g., liposomal nanomedicines). Herein, a supported lipid bilayer platform was employed in order to investigate the effect of membrane surface charge (positive, negative, or neutral) on the adsorption of three complement proteins. Quartz crystal microbalance-dissipation (QCM-D) experiments measured the real-time kinetics and total uptake of protein adsorption onto supported lipid bilayers. The results demonstrate that all three proteins exhibit preferential, mainly irreversible adsorption onto negatively charged lipid bilayers, yet there was also significant variation in total uptake and the relative degree of adsorption onto negatively charged bilayers versus neutral and positively charged bilayers. The total uptake was also observed to strongly depend on the bulk protein concentration. Taken together, our findings contribute to a broader understanding of the factors which influence adsorption of complement proteins onto lipid membranes and offer guidance towards the design of synthetic lipid bilayers with immunocompetent features.

To understand the effects of lipid composition on membrane protein function in a mixture as complex as a biomembrane, one must know whether the lipid composition local to the protein differs from the mean lipid composition. In this study, we simulated the transmembrane domain of a β-barrel protein, OmpA, in mixtures of lipids of different tail lengths under conditions of negative hydrophobic mismatch, i.e., local bilayer thinning. We modeled the influence of OmpA on the local lipid composition both at a coarse-grained (CG) resolution using conventional molecular dynamics, and at an atomistic resolution within the semi-grand canonical ensemble using mutation moves to rapidly approach an equilibrium lateral distribution of lipids. Moderate enrichment of the shorter tail component (either DDPC in DDPC/DMPC mixtures or DMPC in DMPC/DSPC mixtures) extending 2–3 nm away from the protein surface was observed with both the atomistic and CG models. The similarity in trends suggests that the more computationally economical CG models capture the essential features of lipid sorting induced by hydrophobic mismatch.

High-surface-area nanoparticles often cluster, with unknown effects on their cellular uptake and environmental impact. In the presence of vesicles or cell membranes, lipid adsorption can occur on the nanoparticles, resulting in the formation of supported lipid bilayers (SLBs), which tend to resist cellular uptake. When the amount of lipid available is in excess compared with that required to form a single-SLB, large aggregates of SLBs enclosed by a close-fitting lipid bilayer sheath are shown to form. The proposed mechanism for this process is one where small unilamellar vesicles (SUVs) adsorb to aggregates of SLBs just above the gel-to-liquid phase transition temperature, T(m) , of the lipids (as observed by dynamic light scattering), and then fuse with each other (rather than to the underlying SLBs) upon cooling below T(m) . The sacks of SLB nanoparticles that are formed are encapsulated by the contiguous close-fitting lipid sheath, and precipitate below T(m) , due to reduced hydration repulsion and the absence of undulation/protrusion forces for the lipids attached to the solid support. The single-SLBs can be released above T(m) , where these forces are restored by the free lipid vesicles. This mechanism may be useful for encapsulation/release of drugs/DNA, and has implications for the toxic effects of nanoparticles, which may be mitigated by lipid sequestration.

Investigation of lateral heterogeneities (clusters) in cell membranes is an important step toward understanding the physical processes that lead to the formation of lipid domains and rafts. Computer modeling methods represent a powerful tool to solve the problem, since they can detect clusters containing only a few lipid molecules—the situation that still resists characterization with modern experimental techniques. Parameters of clustering depend on lipid composition of a membrane. In this work, we propose a computational method to detect and analyze parts of membrane with different packing densities. Series of one- and two-component fluid systems containing lipids with the same polar heads and different acyl chains, dioleoylphosphatidylcholine (18 : 1) and dipalmitoylphosphatidylcholine (16 : 0), were chosen as the objects under study. The developed algorithm is based on molecular dynamics simulation of hydrated lipid bilayers in all-atom mode. The method is universal and could be applied to any other membrane system with arbitrary lipid composition. Here, we demonstrated that the studied lipid bilayers reveal small lateral dynamic clusters composed of just several (most often, three) lipid molecules. This seems to be one of the most important reasons determining the “mosaic” nature of the membrane-water interface.

Lipid membranes, particularly under nonequilibrium conditions, have recently been investigated ever more vigorously because of their relevance in the biological context. We survey our recent approaches to the theoretical study of lipid bilayers that are perturbed in different ways. Self-organization phenomena involving curvature and/or composition spatiotemporal organization are investigated in membrane systems subjected to externally induced chemical reactions, transversal mass transport and insertion of proteins. The outcomes of these studies are expected to be applicable to different curvature and lateral organization phenomena in synthetic lipid bilayers and also in plasmatic cell membranes.

The host lipid bilayer is increasingly being recognized as an important non-specific regulator of membrane protein function. Despite considerable progress the interplay between hydrophobic coupling and lipid ordering is still elusive. We use electron spin resonance (ESR) to study the interaction between the model protein gramicidin and lipid bilayers of varying thickness. The free energy of the interaction is up to -6kJ/mol; thus not strongly favored over lipid-lipid interactions. Incorporation of gramicidin results in increased order parameters with increased protein concentration and hydrophobic mismatch. Our findings also show that at high protein:lipid ratios the lipids are motionally restricted but not completely immobilized. Both exchange on and off rate values for the lipid--gramicidin interaction are lowest at optimal hydrophobic matching. Hydrophobic mismatch of few A results in up to 10-fold increased exchange rates as compared to the 'optimal' match situation pointing to the regulatory role of hydrophobic coupling in lipid-protein interactions.

A set of 49 protein nanopore-lipid bilayer systems was explored by means of coarse-grained molecular-dynamics simulations to study the interactions between nanopores and the lipid bilayers in which they are embedded. The seven nanopore species investigated represent the two main structural classes of membrane proteins (alpha-helical and beta-barrel), and the seven different bilayer systems range in thickness from approximately 28 to approximately 43 A. The study focuses on the local effects of hydrophobic mismatch between the nanopore and the lipid bilayer. The effects of nanopore insertion on lipid bilayer thickness, the dependence between hydrophobic thickness and the observed nanopore tilt angle, and the local distribution of lipid types around a nanopore in mixed-lipid bilayers are all analyzed. Different behavior for nanopores of similar hydrophobic length but different geometry is observed. The local lipid bilayer perturbation caused by the inserted nanopores suggests possible mechanisms for both lipid bilayer-induced protein sorting and protein-induced lipid sorting. A correlation between smaller lipid bilayer thickness (larger hydrophobic mismatch) and larger nanopore tilt angle is observed and, in the case of larger hydrophobic mismatches, the simulated tilt angle distribution seems to broaden. Furthermore, both nanopore size and key residue types (e.g., tryptophan) seem to influence the level of protein tilt, emphasizing the reciprocal nature of nanopore-lipid bilayer interactions.

We investigate the interactions between lipid bilayers and amphiphilic peptides using a solvent-free coarse-grained simulation technique. In our model, each lipid is represented by one hydrophilic and three hydrophobic beads. The amphiphilic peptide is modeled as a hydrophobic-hydrophilic cylinder with hydrophilic caps. We find that with increasing peptide-lipid attraction the preferred state of the peptide changes from desorbed, to adsorbed, to inserted. A single peptide with weak attraction binds on the bilayer surface, while one with strong attraction spontaneously inserts into the bilayer. We show how several peptides, which individually bind only to the bilayer surface, cooperatively insert. Furthermore, hydrophilic strips along the peptide cylinder induce the formation of multipeptide pores, whose size and morphology depend on the peptides’ overall hydrophilicity, the distribution of hydrophilic residues, and the peptide-peptide interactions. Strongly hydrophilic peptides insert less readily, but prove to be more destructive to bilayer integrity.

The creation of three-dimensional structures in supported lipid bilayers has been examined. In bilayers, shape transformations can be triggered by adjusting a variety of parameters. Here, it is shown that bilayers composed of phosphatidylcholine and phosphatidic acid can be induced to reversibly form cap structures when exposed to an asymmetry in ionic strength. The structures that form depend on the asymmetry in the ionic strength and the amount of anionic lipid. Other factors that may be of importance in the creation of the structures, expansion forces, osmotic forces, and the bilayer-support interaction are discussed. The cap structures have the potential to be of considerable utility in examining the effect that curvature has on membrane processes.

Supported lipid bilayers have been in use for over 30 years. They have been employed to study the structure, composition, and dynamics of lipid bilayer phases, the binding and distribution of soluble, integral, and lipidated proteins in membranes, membrane fusion, and interactions of membranes with elements of the cytoskeleton. This review focuses on the unique ability of supported lipid bilayers to study liquid-ordered and liquid-disordered domains in membranes. We highlight methods to produce asymmetric lipid bilayers with lipid compositions that mimic those of the extracellular and cytoplasmic leaflets of cell membranes and the functional reconstitution of membrane proteins into such systems. Questions related to interleaflet domain coupling and membrane protein activation have been addressed and answered using advanced reconstitution and imaging procedures in symmetric and asymmetric supported membranes with and without coexisting lipid phase domains. Previously controversial topics regarding anomalous and anisotropic diffusion in membranes have been resolved by using supported membrane approaches showing that the propensity of certain lipid compositions to form “rafts” are important but overlaid with “picket-fence” interactions that are imposed by a subtended cytoskeletal network.

Membrane proteins perform relevant physiological functions by means of intricate conformational changes in a hydrophobic environment. Lipid bilayers and embedded proteins, therefore, play a functional role in biomembranes, where the interplay of interactions keeps a delicate balance between cell barriers and selective transducers, transporters, pores, channels, etc. Molecular dynamics and experimental methods (e.g. X-ray diffraction, neutron scattering, nuclear magnetic resonance, infrared spectroscopy, dielectric relaxation spectroscopy, among others) encompass a set of tools to determine the relevant properties that make biomembranes so efficient for preserving life. In this chapter, I provide a perspective on studies that combine experimental methods and molecular dynamics approaches to decipher couplings of membrane proteins and lipid bilayers.

Water is crucial to the structure and function of biological membranes. In fact, the membrane's basic structural unit, i.e. the lipid bilayer, is self-assembled and stabilized by the so-called hydrophobic effect, whereby lipid molecules unable to hydrogen bond with water aggregate in order to prevent their hydrophobic portions from being exposed to water. However, this is just the beginning of the lipid-bilayer-water relationship. This mutual interaction defines vesicle stability in solution, controls small molecule permeation, and defines the spacing between lamella in multi-lamellar systems, to name a few examples. This chapter will describe the structural and dynamical properties central to these, and other water- lipid bilayer interactions.

Understanding cellular membrane processes is critical for the study of events such as viral entry, neurotransmitter exocytosis, and immune activation. Supported lipid bilayers are commonly used to model these membrane processes experimentally. Despite the relative simplicity of such a system, many important structural and dynamic parameters are not experimentally observable with current techniques. Computational approaches allow the development of a high-resolution model of bilayer processes. We have performed molecular dynamics simulations of dimyristoylphosphatidylcholine (DMPC) bilayers to model the creation of bilayer gaps—a common process in bilayer patterning—and to analyze their structure and dynamics. We propose a model for gap formation in which the bilayer edges form metastable micelle-like structures on a nanosecond timescale. Molecules near edges structurally resemble lipids in ungapped bilayers but undergo small-scale motions more rapidly. These data suggest that lipids may undergo rapid local rearrangements during membrane fusion, facilitating the formation of fusion intermediates thought key to the infection cycle of viruses such as influenza, Ebola, and HIV.

Asymmetric lipid distribution between the two leaflets of bilayer is a physiologically important feature of biological membranes. Dissipation of lipid asymmetry by lipid scrambling is a ubiquitous cellular mechanism essential for membrane biogenesis, cell signaling, and bacterial cell wall assembly. The recently crystallized structure of a TMEM16 family member nhTMEM16 provided the first structural insight into the mechanism of lipid scramblases, which facilitate the passive bidirectional transport of diverse lipids between the two leaflets of the membrane. It identified a conserved Ca2+-binding site harbored in a hydrophilic crevice predicted to face the hydrophobic core of the membrane and involve in lipid scrambling, although no lipids were resolved in the structure. In order to investigate the Ca2+-mediated scrambling mechanism, identify the lipid transport pathway, and characterize the role of Ca2+, we have performed extended equilibrium molecular dynamics (MD) simulations with nhTMEM16 embedded in various lipid bilayers (POPC, POPE, POPS, and mixtures thereof) in the presence or absence of Ca2+. Our simulations reveal a previously unidentified spiral membrane-traversing hydrophilic track around the protein surface, connecting the inner and outer leaflets. Especially in the Ca2+-bound systems, lipids from the inner and outer leaflets spontaneously bind to the hydrophilic track with their headgroups and propagate toward the center of the membrane. In the Ca2+-free systems, on the other hand, the hydrophilic track is observed to substantially narrow allowing only smaller species such as water and ions to bind. Furthermore, umbrella sampling (US) simulations are used to characterize the complete lipid transport pathway and to identify key residues and energetics of the lipid translocation. Our study provides crucial atomic details of the scrambling pathway for the first time, and uncovers the nature of Ca2+-dependence by revealing the structural rearrangements of the hydrophilic track for lipid translocation.

The synthetic peptide acetyl-K(2)-G-L(24)-K(2)-A-amide (P(24)) and its analogs have been successfully utilized as models of the hydrophobic transmembrane alpha-helical segments of integral membrane proteins. The central polyleucine region of these peptides was designed to form a maximally stable, very hydrophobic alpha-helix which will partition strongly into the hydrophobic environment of the lipid bilayer core, while the dilysine caps were designed to anchor the ends of these peptides to the polar surface of the lipid bilayer and to inhibit the lateral aggregation of these peptides. Moreover, the normally positively charged N-terminus and the negatively charged C-terminus have both been blocked in order to provide a symmetrical tetracationic peptide, which will more faithfully mimic the transbilayer region of natural membrane proteins and preclude favorable electrostatic interactions. In fact, P(24) adopts a very stable alpha-helical conformation and transbilayer orientation in lipid model membranes. The results of our recent studies of the interaction of this family of alpha-helical transmembrane peptides with phospholipid bilayers are summarized here.

Interactions between lipid bilayers are critical in many biological processes in which membrane surfaces come close together. Recent X-ray diffraction analyses of bilayers subjected to known osmotic pressures have provided critical information on the magnitude of both the repulsive and the attractive forces that exist between phospholipid and glycolipid membranes.

Structural perturbations of fluid lipid bilayers can be caused by various processes, such as the insertion of transmembrane inclusions, protein and peptide adsorption, the formation of membrane pores, or even by the transition to the inverse hexagonal phase. Theoretical concepts for their energetic description are often based on membrane elasticity theory. New insights have emerged from both applying this theory to inclusion-containing membranes and from relating membrane elasticity theory to microscopic models of perturbed lipid layers.

Disparate biological fusion reactions and fusion of purely lipid bilayers are similarly influenced by 'non-bilayer' lipids (lipids which do not form lipid bilayers in water by themselves). Lipid composition of membranes affects biological fusion at a stage downstream of activation of fusion proteins and prior to fusion pore formation. These data suggest that actual merger of membrane lipid bilayers in different fusion reactions proceeds via the same pathway. The effects of non-bilayer lipids specifically correlate with their ability to bend lipid monolayers in different directions, and appear to be consistent with the specific hypothesis of membrane fusion suggesting that fusion proceeds through highly bent intermediates--stalks, local connections between contacting monolayers of fusing membranes.

Lipid membrane elastic properties play an important role in the membrane deformations and dynamic morphological transitions necessary for cell function. A key elastic property underlying these dynamics is the bending rigidity, motivating significant research efforts to quantify the effects of lipid structure and additives on the membrane bending modulus. To date, the majority of experimental research has focused on the dynamics of model membrane systems composed of zwitterionic lipids; however, most biomembranes are negatively charged at physiological conditions due to the presence of charged lipid headgroups. Here we study the bending dynamics of negatively-charged phosphatidylglycerol (PG) bilayers using neutron spin echo spectroscopy. Our results show that the charged lipid bilayers are softer than analogous zwitterionic phosphatidylcholine (PC) bilayers in both the gel and fluid phases at low ionic strength conditions. Interestingly, theoretical predictions indicate that the opposite should be true and that the bending rigidity should increase with increasing surface charge. We propose that this discrepancy is due to charged-related differences in the area per headgroup and lipid hydration between PG and PC bilayers that are not considered by existing theoretical models. Our results provide new insights into the influence of charge on the membrane elastic properties and demonstrate how these dynamics are coupled to charge effects on the membrane structure.

The hydrophobic interaction arises from the ordered structure of water around nonpolar groups of molecules in an aqueous solvent. Because biological systems are made of various macromolecules and amphiphiles which are suspended in aqueous solution, the hydrophobic interaction plays a very important role in the formation of higher-order structure and phase transitions in biological systems. Considering the hydrophobic interaction, the van der Waals interaction and the entropic effect, an equation of state of a lipid membrane was obtained which was analogous to the van der Waals equation. The characteristics of the lipid bilayer phase transition as well as the phase behaviors of a lipid monolayer were explained by this equation of state. Experimental evidence was obtained from ultrasonic measurements which indicated that its phase transition accompanys significant critical phenomena. Analysis of the hydrophobicity of amino acid sequences revealed that the morphology of the proteins was determined b...

A model recently used to study lipid-protein interactions in one-component lipid bilayers (Sperotto and Mouritsen, 1991 a, b) has been extended in order to include two different lipid species characterized by different acyl-chain lengths. The model, which is a statistical mechanical lattice model, assumes that hydrophobic matching between lipid-bilayer hydrophobic thickness and hydrophobic length of the integral protein is an important aspect of the interactions. By means of Monte Carlo simulation techniques, the lateral distribution of the two lipid species near the hydrophobic protein-lipid interface in the fluid phase of the bilayer has been derived. The results indicate that there is a very structured and heterogeneous distribution of the two lipid species near the protein and that the protein-lipid interface is enriched in one of the lipid species. Out of equilibrium, the concentration profiles of the two lipid species away from the protein interface are found to develop a long-range oscillatory behavior. Such dynamic membrane heterogeneity may be of relevance for determining the physical factors involved in lipid specificity of protein function.

Lipid bilayers can be induced to adhere to each other by molecular mediators, and, depending on the lipid composition, such adhesion can lead to merging of the contacting monolayers in a process known as hemifusion. Such bilayer-bilayer reactions have never been systematically studied. In the course of our studies of membrane-active molecules, we encountered such reactions. We believe that they need to be understood whenever bilayer-bilayer interactions take place, such as during membrane fusion. For illustration, we discuss three examples: spontaneous adhesion between phospholipid bilayers induced by low pH, polymer-induced osmotic depletion attraction between lipid bilayers, and anionic lipid bilayers cross-bridged by multicationic peptides. Our purpose here is to describe a general method for studying such interactions. We used giant unilamellar vesicles, each of which was aspirated in a micropipette so that we could monitor the tension of the membrane and the membrane area changes during the bilayer-bilayer interaction. We devised a general method for measuring the free energy of adhesion or hemifusion. The results show that the energies of adhesion or hemifusion of lipid bilayers could vary over 2 orders of magnitude from −1 to −50 × 10−5 J/m2 in these examples alone. Our method can be used to measure the energy of transition in each step of lipid transformation during membrane fusion. This is relevant for current research on membrane fusion, which focuses on how fusion proteins induce lipid transformations.

Phospholipid molecules form bilayers in water due to their hydrophilic heads and hydrophobic tails. The electroporation of lipid bilayers (cell membranes) is a phenomenon where membranes are permeabilized by the application of electric fields. At some critical voltage, a dramatic increase in conductivity across the membranes is observed. This phenomenon is widely used in DNA and RNA transfer as well as targeted drug delivery systems. However, the membrane ruptures with a continuous increase in voltage where interaction between lipid and water molecules is an important factor in electroporation behavior. This study characterizes the wettability, of both the head and tail groups of lipid molecules, by calculating the contact angle of a water droplet on a planar phospholipid monolayer using molecular dynamics simulations. The water droplet completely spreads on the hydrophilic heads of the lipid, while forming an average contact angle of 136.05° on the hydrophobic tails. An analysis using the Young’s equation suggests that a difference in free energy of 116 mJ/m2 contributes to the overall energy barrier for water penetration across the lipid monolayer. We aim to control this permeabilization phenomenon to achieve water desalination.Copyright © 2010 by ASME

The mechanism of neurodegeneration caused by β-amyloid (Aβ) in Alzheimer's disease is still controversial. Neuronal toxicity is exerted mostly by various species of soluble Aβ oligomers. Recent data depict membranes as the main sites where proteins/peptides are recruited and concentrated, misfold, and nucleate amyloids; at the same time, membranes are considered key triggers of amyloid toxicity.We demonstrated the capability of Aβ to penetrate and destabilize stacked lipid bilayers in a previous work. In this study, in order to maintain the natural fluidity of the membrane, polymer cushioned lipid bilayers have been used as a model for neuronal membrane. Layer-by-layer technique was used for the fabrication of the polymer cushion of charged poly-electrolytes, the lipid membrane is built on the polymer film by unilamellar vesicle fusion. Neutron reflectivity was used to monitor the kinetics of adsorption of the lipid bilayer onto the polymer surface; the conditions for the best surface coverage have been determined. The structure of the lipid bilayers is modified by the interaction with Aβ1-42; Neutron reflectivity showed a change of the scattering density profile in the direction perpendicular to the membrane plane, suggesting penetration of Aβ in the double layer. Atomic force microscope (AFM) has been used to test the lipid packing of the membrane through film rupture experiments and to compare the bilayer morphology in the presence or in the absence of Aβ. We demonstrated that the presence of Aβ weakens the lipid packing in the model membranes.We compared the results obtained on polymer cushioned lipid bilayers with those obtained using a rigid substrate (freshly cleaved mica) for membrane preparation.

The canonical Singer–Nicolson model of lipid bilayers with embedded proteins emphasize the passive barrier properties of the bilayer component. It is, however, becoming increasingly clear that the notion of bilayers as essentially inert two-dimensional sheets of liquid hydrocarbon is problematic. There is increasing evidence for regulation of integral membrane protein function by the molecular composition of the bilayer matrix. In many cases, the regulation is nonspecific in the sense that the regulation is coupled to a physical property of the bilayer such as equilibrium thickness and spontaneous monolayer curvature. This regulation can be rationalized by considering the hydrophobic matching between proteins and the lipid bilayer. Thus, whenever an integral membane protein undergoes a conformational change involving its hydrophobic transmembrane moiety, part of the free energy associated with the conformational change is determined by the physical properties of the lipid bilayer. This chapter first reviews the basic structural properties of lipid bilayers and the two main structural classes of integral membrane proteins: alpha-helical bundles and beta-barrels. Then, examples of recent advances in electron paramagnetic resonance (EPR) and nuclear magnetic resonance (NMR) spectroscopy revealing structural and orientational information about integral membrane protein in fluid lipid bilayers are presented. Finally, a continuum elastic description of protein–lipid interactions is presented with emphasis on how to quantify the energetics of protein–lipid interactions. Keywords: protein–lipid interaction; fluid-mosaic; Singer–Nicolson model; hydrophobic matching principle

While most membrane protein helices are clearly hydrophobic, recent experiments have indicated that it is possible to insert marginally hydrophobic helices into bilayers and have suggested apparent in vivo free energies of insertion for charged residues that are low, e.g., a few kcals for arginine. In contrast, a number of biophysical simulation studies have predicted that the bilayer interior is close to a pure hydrophobic environment with large penalties for hydrophilic amino acids--and yet the experimental scales do significantly better at predicting actual membrane proteins from sequence. Here, we have systematically studied the dependence of the free energy profiles on lipid properties, including tail length, saturation, headgroup hydrogen bond strength, and charge, both to see to whether the in vivo insertion can be explained in whole or part from lipid composition of the endoplasmic reticulum (ER) membranes, and if the solvation properties can help interpret how protein function depends on the lipids. We find that lipid charge is important to stabilize charged amino acids inside the bilayer (with implications, e.g., for ion channels), that thicker bilayers have higher solvation costs for hydrophilic side chains, and that headgroup hydrogen bond strength determines how adaptive the lipids are as a hydrophobic/hydrophilic solvent. None of the different free energy profiles are even close to the low apparent in vivo insertion cost, which suggests that regardless of the specific ER membrane composition the current experimental results cannot be explained by normal lipid-type variation.

The fusion of lipid bilayers is studied with dissipative particle dynamics simulations. First, to achieve control over membrane properties, the effects of individual simulation parameters are studied and optimized. Then, a large number of fusion events for a vesicle and a planar bilayer are simulated using the optimized parameter set. In the observed fusion pathway, configurations of individual lipids play an important role. Fusion starts with individual lipids assuming a splayed tail configuration with one tail inserted in each membrane. To determine the corresponding energy barrier, we measure the average work for interbilayer flips of a lipid tail, i.e., the average work to displace one lipid tail from one bilayer to the other. This energy barrier is found to depend strongly on a certain dissipative particle dynamics parameter, and, thus, can be adjusted in the simulations. Overall, three subprocesses have been identified in the fusion pathway. Their energy barriers are estimated to lie in the range 8–15 kBT. The fusion probability is found to possess a maximum at intermediate tension values. As one decreases the tension, the fusion probability seems to vanish before the tensionless membrane state is attained. This would imply that the tension has to exceed a certain threshold value to induce fusion.

The cell membrane forms a selectively permeable barrier that enables cells to maintain a distinct chemical environment from their surroundings, making cell membranes one of the key components required for life. In this chapter we review the interactions between small molecules and lipid bilayers from a computational perspective, focusing on the permeation of small molecules and how small molecules affect the properties of lipid bilayers. We organize the molecules based on broad chemical characteristics such as hydrophobicity, polarity, and charge. We also discuss three specific classes of interest: anesthetics, sugars, and carbon nanoparticles.

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.

The mechanistic details of processes occurring in and during contact between lipid bilayers are still poorly understood due to their complexity on the mesoscopic scale. Here, we analyze lipid exchange during contact of oppositely charged lipid bilayers. Specifically, we explore a generic mechanism, where this process occurs via diffusion of individual lipids between the layers. Our estimates indicate that this scenario is feasible on the time scale of conventional experiments and also on the time scale of biochemical processes in cells.

A thorough understanding of the structure of fusion domains of enveloped viruses in changing lipid environments helps us to formulate mechanistic models on how they might function in mediating viral entry by membrane fusion. We have expressed the N-terminal fusion domain of HIV-1 gp41 as a construct that is water-soluble in the absence of membranes, but that also binds with high affinity to lipid micelles and bilayers in their presence. We have solved the structure and studied the dynamics of this domain bound to dodecylphosphocholine micelles by homo- and heteronuclear NMR spectroscopy. The fusion peptide forms a stable hydrophobic helix from Ile(4) to Ala(14), but is increasingly more disordered and dynamic in a segment of intermediate polarity that stretches from Ala(15) to Ser(23). When bound to lipid bilayers at low concentration, the HIV fusion domain is also largely alpha-helical, as determined by CD and FTIR spectroscopy. However, at higher protein/lipid ratios, the domain is partially converted to form beta-structures in lipid bilayers. Controlled lipid mixing occurs at concentrations that support the alpha-helical, but not the beta-strand conformation.

Biological membranes are complex and highly cooperative structures. To relate biomembrane structure to their biological function it is often necessary to consider simpler systems. Lipid bilayers composed of one or two lipid species, and with embedded proteins, provide a model system for biological membranes. Here we present a mesoscopic model for lipid bilayers with embedded proteins, which we have studied with the help of the dissipative particle dynamics simulation technique. Because hydrophobic matching is believed to be one of the main physical mechanisms regulating lipid-protein interactions in membranes, we considered proteins of different hydrophobic length (as well as different sizes). We studied the cooperative behavior of the lipid-protein system at mesoscopic time- and lengthscales. In particular, we correlated in a systematic way the protein-induced bilayer perturbation, and the lipid-induced protein tilt, with the hydrophobic mismatch (positive and negative) between the protein hydrophobic length and the pure lipid bilayer hydrophobic thickness. The protein-induced bilayer perturbation was quantified in terms of a coherence length, xi(P), of the lipid bilayer hydrophobic thickness profile around the protein. The dependence on temperature of xi(P), and the protein tilt-angle, were studied above the main-transition temperature of the pure system, i.e., in the fluid phase. We found that xi(P) depends on mismatch, i.e., the higher the mismatch is, the longer xi(P) becomes, at least for positive values of mismatch; a dependence on the protein size appears as well. In the case of large model proteins experiencing extreme mismatch conditions, in the region next to the so-called lipid annulus, there appears an undershooting (or overshooting) region where the bilayer hydrophobic thickness is locally lower (or higher) than in the unperturbed bilayer, depending on whether the protein hydrophobic length is longer (or shorter) than the pure lipid bilayer hydrophobic thickness. Proteins may tilt when embedded in a too-thin bilayer. Our simulation data suggest that, when the embedded protein has a small size, the main mechanism to compensate for a large hydrophobic mismatch is the tilt, whereas large proteins react to negative mismatch by causing an increase of the hydrophobic thickness of the nearby bilayer. Furthermore, for the case of small, peptidelike proteins, we found the same type of functional dependence of the protein tilt-angle on mismatch, as was recently detected by fluorescence spectroscopy measurements.

This review describes the effects caused by synthetic polymers on model lipid membranes. Among a large variety of synthetic polymers, we focused on two quite different groups. One of them consists of polyelectrolytes, whose adsorption on lipid membranes is driven by Coulomb forces, and the other group consisting of Pluronics and polyethoxylated surfactants that have amphiphilic structure and interact with lipid bilayers by hydrophobic forces. This choice was determined by the continuously increasing interest to the application of these polymers in biomedical research. It was shown that interactions of cationic polymers with oppositely charged lipid bilayers result in the formation of lipid domains in the membranes, influence the lateral diffusion of lipid molecules and proteins incorporated into the bilayer, modulate functional activity of membrane proteins and in some cases even favor asymmetrical distribution of lipids. Incorptions of amphiphilic copolymers into lipid bilayers may result in changes in the rate of flip-flop and membranes microviscosity. Polymer-membrane interactions in both cases cause changes in the membrane premeability, obviously resulting from formation of disturbances in the membrane packing. In case of Pluronics, non-covalent interactions between the membrane-bound polymer and the permeant may also contribute to the transportation of the latter through lipid bilayer. Special attention in this review is paid to the relatioship between the polymer structure and its ability to modulate membrane properties. A qualitative thermodynamic treatment of the polymer-membrane interactions allowed to explain the polymer effects on the bilayer structure and led to a list of properties that the polymer structure must possess in order to affect membrane permeability.

Depending on their hydrophobicity, peptides can interact differently with lipid membranes inducing dramatic modifications into their host systems. In the present paper, the interaction of a synthetic peptide with a scrambled hydrophobic/hydrophilic sequence (Pro-Asp-Ala-Asp-Ala-His-Ala- His-Ala-His-Ala-Ala-Ala-His-Gly) (PADH) with 1,2-dipalmitoyl-sn-glycero- 3phosphocholine (DPPC) model membranes has been investigated by differential scanning calorimetry (DSC), adopting three different experimental approaches. In the first, the peptide is forced to be included into the hydrocarbon region of the lipid bilayer, by codissolving it with the lipid giving rise to mixed multilamellar vesicles-peptide systems; in the second, this system is passed through an extruder, thus producing large unilamellar vesicles-peptide systems; in the third, it is allowed to interact with the external surface of the membrane. The whole of the DSC results obtained have shown that the incorporation of the peptide into the lipid bilayer by means of the first method induces a decrease in the enthalpy of the gel-liquid crystal transition of the membrane and a shift of the transition to the lower temperatures, thus resembling, in spite of its prevalently hydrophilic nature, the behavior of transbilayer hydrophobic peptides. The extrusion of these systems creates unilamellar vesicles free of peptides but of smaller size as evidenced by the decreased cooperativity of the transition. The peptide, added externally to the DPPC model membrane, has no effect on the phase behavior of the bilayer. These findings suggest that the effect of the interaction of scrambled hydrophobic/hydrophilic peptides into lipid bilayers strongly affects the thermotropic behavior of the host membrane depending on the preparation method of the lipid/peptide systems. The whole of the results obtained in the present paper can be useful in approaching studies of bioactive peptides/lipids systems. © 2002 Elsevier Science B.V. All rights reserved.

Many prokaryotic organisms (archaea and bacteria) are covered by a regularly ordered surface layer (S-layer) as the outermost cell wall component. S-layers are built up of a single protein or glycoprotein species and represent the simplest biological membrane developed during evolution. Pores in S-layers are of regular size and morphology, and functional groups on the protein lattice are aligned in well-defined positions and orientations. Due to the high degree of structural regularity S-layers represent unique systems for studying the structure, morphogenesis, and function of layered supramolecular assemblies. Isolated S-layer subunits of numerous organisms are able to assemble into monomolecular arrays either in suspension, at air/water interfaces, on planar mono- and bilayer lipid films, on liposomes and on solid supports (e.g. silicon wafers). Detailed studies on composite S-layer/lipid structures have been performed with Langmuir films, freestanding bilayer lipid membranes, solid supported lipid membranes, and liposomes. Lipid molecules in planar films and liposomes interact via their head groups with defined domains on the S-layer lattice. Electrostatic interactions are the most prevalent forces. The hydrophobic chains of the lipid monolayers are almost unaffected by the attachment of the S-layer and no impact on the hydrophobic thickness of the membranes has been observed. Upon crystallization of a coherent S-layer lattice on planar and vesicular lipid membranes, an increase in molecular order is observed, which is reflected in a decrease of the membrane tension and an enhanced mobility of probe molecules within an S-layer-supported bilayer. Thus, the terminology 'semifluid membrane' has been introduced for describing S-layer-supported lipid membranes. The most important feature of composite S-layer/lipid membranes is an enhanced stability in comparison to unsupported membranes.