Fusogenic tilted peptides induce nanoscale holes in supported phosphatidylcholine bilayers

Tilted peptides are known to insert in lipid bilayers with an oblique orientation, thereby destabilizing membranes and facilitating membrane fusion processes. Here, we report the first direct visualization of the interaction of tilted peptides with lipid membranes using in situ atomic force microscopy (AFM) imaging. Phase-separated supported dioleoylphosphatidylcholine/dipalmitoylphosphatidylcholine (DOPC/DPPC) bilayers were prepared by fusion of small unilamellar vesicles and imaged in buffer solution, in the absence and in the presence of the simian immunodeficiency virus (SIV) peptide. The SIV peptide was shown to induce the rapid appearance of nanometer scale bilayer holes within the DPPC gel domains, while keeping the domain shape unaltered. We attribute this behavior to a local weakening and destabilization of the DPPC domains due to the oblique insertion of the peptide molecules. These results were directly correlated with the fusogenic activity of the peptide as determined using fluorescently labeled DOPC/DPPC liposomes. By contrast, the nontilted ApoE peptide did not promote liposome fusion and did not induce bilayer holes but caused slight erosion of the DPPC domains. In conclusion, this work provides the first direct evidence for the production of stable, well-defined nanoholes in lipid bilayer domains by the SIV peptide, a behavior that we have shown to be specifically related to the tilted character of the peptide. A molecular mechanism underlying spontaneous insertion of the SIV peptide within lipid bilayers and the subsequent removal of bilayer patches is proposed, and its relevance to membrane fusion processes is discussed.     

Spectroscopic analysis of the interaction of a peptide sequence of Hepatitis G virus with bilayers

Merocyanine 540 (MC540) has been used as external probe to determine the interaction of the peptide sequence 125-139 corresponding to the E2 protein of Hepatitis G virus, with lipid bilayers. The probe was incorporated into large unilamellar vesicles (LUVs) or small unilamellar vesicles (SUVs) of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). When incorporated into bilayers, MC540 shows two absorption maxima corresponding to the monomer (570 nm) and dimer (530 nm) form of the probe. Changes in the probe microenvironment are reflected by a modification in the position and/or intensity of these maxima. Addition of increasing amounts of peptide resulted in a slight decrease of the ratio A570/A530 thus indicating a change in MC540 partition into the membrane, going from a hydrophobic to a more hydrophilic environment. This effect was concomitant with an increase in dimer formation as stated from the values of the apparent dimerization constant (K(app)) obtained. Fluorescence spectra as well as steady state anisotropy measurements were in agreement with the above results indicating that the peptide was able to relocate the probe and displacing MC540 from its initial location into the bilayer. Results with SUVs or LUVs were similar for what curvature does not seem to play any role on peptide activity. These results reflect the ability of peptide to interact with biomimetic membranes in the lipid head group region.     

Modulating membrane properties: the effect of trehalose and cholesterol on a phospholipid bilayer

The protective properties of trehalose on cholesterol-containing lipid dipalmitoylphosphatidylcholine (DPPC) bilayers are studied through molecular simulations. The ability of the disaccharide to interact with the phospholipid headgroups and stabilize the membrane persists even at high cholesterol concentrations and restricts some of the changes to the structure that would otherwise be imposed by cholesterol molecules. Predictions of bilayer properties such as area per lipid, tail ordering, and chain conformation support the notion that the disaccharide decreases the main melting transition in these multicomponent model membranes, which correspond more closely to common biological systems than pure bilayers. Molecular simulations indicate that the membrane dynamics are slowed considerably by the presence of trehalose, indicating that high sugar concentrations would serve to avert possible phase separations that could arise in mixed phospholipid systems. Various time correlation functions suggest that the character of the modifications in lipid dynamics induced by trehalose and cholesterol is different in the hydrophilic and hydrophobic regions of the membrane.     

Methotrexate interaction with a lipid membrane model of DPPC

Dipalmitoylphosphatidylcholine (DPPC) liposomes were employed as membrane models for the investigation of the interaction occurring between methotrexate (MTX) and bilayer lipid matrix. Liposomes were obtained by hydrating a lipid film with 50 mM Tris buffer (pH 7.4). The differential scanning calorimetry (DSC) evaluation of the thermotropic parameters associated with the phase transitions of DPPC liposomes gave useful information about the kind of drug-membrane interaction. The results showed an electrostatic interaction taking place with the negatively charged molecules of MTX and the phosphorylcholine head groups, constituting the outer part of DPPC bilayers. No interaction with the hydrophobic phospholipid bilayer domains was detected, revealing a poor capability of MTX to cross through lipid membranes to reach the interior compartment of a lipid bounded structure. These findings correlate well within vitro biological experiments on MTX cell susceptibility.     

Interaction of propyl paraben with dipalmitoyl phosphatidylcholine bilayer: a differential scanning calorimetry and nuclear magnetic resonance study

The influence of the preservative, propyl paraben (PPB) on the biophysical properties of dipalmitoyl phosphatidyl choline (DPPC) vesicles, both in multilamellar vesicle (MLV) and unilamellar vesicle (ULV) forms, has been studied using DSC and (¹H and ³¹P) NMR. The mechanism by which PPB interacts with DPPC bilayers was found to be independent of the morphological organization of the lipid bilayer. Incorporation of PPB in DPPC vesicles causes a significant depression in the transition temperature and enthalpy of both the pre-transition (PT) and the gel to liquid crystalline transition. The presence of the PPB also reduces the co-operativity of these transitions. However, at high PPB concentration the PT disappears. DSC and NMR findings indicate that: (i) PPB is bound strongly to the lipid bilayer leading to increased headgroup fluidity due to reduced headgroup–headgroup interaction and (ii) the PPB molecules are intercalated between the DPPC polar headgroups with its alkyl chain penetrate into the co-operative region. MLV incorporated with high PPB concentration shows additional transitions whose intensity increases with increasing PPB concentration. This phase segregation observed could probably be due to co-existence of PPB-rich and PPB-poor phospholipid domains within the bilayers. The effect of inclusion of cholesterol in the PPB-free and PPB-doped DPPC dispersion was also studied. Equilibration studies suggest that PPB molecules are very strongly bound and remain intercalated between the polar headgroup for prolonged time.     

Rationalizing the membrane interactions of cationic amphipathic antimicrobial peptides by their molecular shape

Biophysical and structural studies of cationic amphipathic antimicrobial peptides have revealed new mechanistic details concerning their membrane interactions. In interfacial environments the peptides adopt amphipathic conformations and the resulting distribution of polar, charged and hydrophobic residues allows them to partition into the bilayer interface. For several helical peptides it was found that their long axis is oriented parallel to the membrane surface, an arrangement which results in considerable perturbations in the packing of the lipid bilayer. Within the molecular shape concept the peptides act as wedge-like structures which impose positive curvature strain on the membrane. As a consequence a wide variety of morphologies are observed of peptide–lipid mixtures which strongly depend on the detailed peptide sequence, the membrane lipid composition, buffer, temperature and other environmental parameters. Therefore, the peptide–lipid systems are best described by phase diagrams, similar to the ones of detergent–lipid mixtures, encompassing on the one extreme regions where the peptide stabilizes the bilayer and on the other extreme regions where membrane lysis occurs. The effects of peptide sequence, membrane penetration depth, lipid composition and membrane surface charge density on membrane-association, -morphology and the resulting phase boundaries are discussed.     

Effects of lipid chain unsaturation and headgroup type on molecular interactions between paclitaxel and phospholipid within model biomembrane

Molecular interactions between paclitaxel, an anticancer drug, and phospholipids of various chain unsaturations and headgroup types were investigated in the present study by Langmuir film balance and differential scanning calorimetry. Both the lipid monolayer at the air-water interface and the lipid bilayer vesicles (liposomes) were employed as model cell membranes. It was found that, regardless of the difference in molecular structure of the lipid chains and headgroup, the drug can form nonideal, miscible systems with the lipids at the air-water interface over a wide range of paclitaxel mole fractions. The interaction between paclitaxel and phospholipid within the monolayer was dependent on the molecular area of the lipids at the interface and can be explained by intermolecular forces or geometric accommodation. Paclitaxel is more likely to form thermodynamically stable systems with 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC) and 1,2-dielaidoyl-sn-glycero-3-phosphocholine (DEPC) than with 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) and 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC). Investigation of the drug penetration into the lipid monolayer showed that DPPC and DEPC have higher incorporation abilities for the drug than DPPE and DSPC. A similar trend was also evidenced by DSC investigation with liposomes. While little change of DSC profiles was observed for the DPPE/paclitaxel and DSPC/paclitaxel liposomes, paclitaxel caused noticeable changes in the thermographs of DPPC and DEPC liposomes. Paclitaxel was found to cause broadening of the main phase transition without significant change in the peak melting temperature of the DPPC bilayers, which demonstrates that paclitaxel was localized in the outer hydrophobic cooperative zone of the bilayer, i.e., in the region of the C1-C8 carbon atoms of the acyl chain or binding at the polar headgroup site of the lipids. However, it may penetrate into the deeper hydrophobic zone of the DEPC bilayers. These findings provide useful information for liposomal formulation of anticancer drugs as well as for understanding drug-cell membrane interactions.     

Impact on lipid membrane organization by free branched-chain fatty acids

Here, we exploit the non-invasive techniques of solid-state NMR (nuclear magnetic resonance) and differential scanning calorimetry (DSC) to study the effect of free iso and ante-iso branched chain fatty acids (BCFAs) on the physicochemical properties of lipid membranes. Free fatty acids are present in biological membranes at low abundance, but can influence the cellular function by modulating the membrane organization. Solid state NMR spectra of dimyristoylphosphatidylcholine (DMPC) lipid membranes containing either free 12-methyltetradecanoic acid (a15:0) or free 13-methyltetradecanoic acid (i15:0), show significant differences in their impact on the lipid bilayer. Chain order profiles obtained by deuterium NMR on fully deuterated DMPC-d(67) bilayers revealed an ordering effect induced by both fatty acids on the hydrophobic membrane core. This behavior was also visible in the corresponding DSC thermograms where the main phase transition of DMPC bilayers-indicative of the hydrophobic membrane region-was shifted to higher temperatures, with the iso isomer triggering more pronounced changes as compared to the ante-iso isomer. This is probably due to a higher packing density in the core of the lipid bilayer, which causes reduced diffusion across membranes. By utilizing the naturally occurring spin reporters nitrogen-14 and phosphorus-31 present in the hydrophilic DMPC headgroup region, even fatty acid induced changes at the membrane interface could be detected, an observation reflecting changes in the lipid headgroup dynamics.     

Membrane interactions of host-defense peptides studied in model systems

Host-defense, antibiotic peptides are believed to generate their cytolytic effects by interacting with the membranes of bacterial cells. Direct analyses of peptide interactions with real cellular membranes are difficult, however, due to the high complexity of physiological membranes. This review summarizes experimental work aiming to understand peptide-membrane interactions and their relationships with the peptides' biological actions using specific model systems. Varied model assemblies have been constructed that generally aim to mimic the fundamental lipid bilayer organization of the membrane. The model systems we will describe include multilamellar and unilamellar vesicles, planar lipid bilayers, lipid monolayers and micelles, and colorimetric biomimetic membranes. The different artificial models have facilitated examination of specific biological or chemical parameters affecting peptide action, for example the effect of membrane lipid composition on peptide affinities and membrane penetration, the relationship between membrane fluidity and peptide interactions, the conformations of active peptides, and other factors. We evaluate the strengths and limitations of the various approaches, and point to future directions in the field.     

Freeze-fracture electron microscopic and calorimetric studies on microscopic states of surface-modified liposomes with poly(ethylene glycol) chains

The differential scanning calorimetry (DSC) and the freeze-fracture electron microscopy of dipalmitoyl phosphatidylcholine (DPPC) liposomes containing distearoyl-N-monomethoxy poly(ethylene glycol)-succinyl-phosphatidylethanolamines (PEG-DSPE) were carried out. The DSC peak of DPPC liposomes containing PEG-DSPE had a shoulder. The main phase transition temperature of DPPC bilayer membranes containing PEG-DSPE whose molecular weight of PEG is less than 3000 was slightly shifted to a higher temperature, while that containing PEG-DSPE whose molecular weight of PEG is more than 5000 was slightly shifted to a lower temperature. The electron micrographs of freeze-fracture replicas of DPPC liposomes containing PEG-DSPE quenched from 37±2°C exhibited banded and planar textures, suggesting the lateral phase separation in the bilayer membranes.     

Membrane orientation of MSI-78 measured by sum frequency generation vibrational spectroscopy

Antimicrobial peptides (AMPs) selectively disrupt bacterial cell membranes to kill bacteria whereas they either do not or weakly interact with mammalian cells. The orientations of AMPs in lipid bilayers mimicking bacterial and mammalian cell membranes are related to their antimicrobial activity and selectivity. To understand the role of AMP-lipid interactions in the functional properties of AMPs better, we determined the membrane orientation of an AMP (MSI-78 or pexiganan) in various model membranes using sum frequency generation (SFG) vibrational spectroscopy. A solid-supported single 1,2-dipalmitoyl-an-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) bilayer or 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) bilayer was used as a model bacterial cell membrane. A supported 1,2-dipalmitoyl-an-glycero-3-phosphocholine (DPPC) bilayer or a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayer was used as a model mammalian cell membrane. Our SFG results indicate that the helical MSI-78 molecules are associated with the bilayer surface with ～70° deviation from the bilayer normal in the negatively charged gel-phase DPPG bilayer at 400 nM peptide concentration. However, when the concentration was increased to 600 nM, MSI-78 molecules changed their orientation to make a 25° tilt from the lipid bilayer normal whereas multiple orientations were observed for an even higher peptide concentration in agreement with toroidal-type pore formation as reported in a previous solid-state NMR study. In contrary, no interaction between MSI-78 and a zwitterionic DPPC bilayer was observed even at a much higher peptide concentration (～12,000 nM). These results demonstrate that SFG can provide insights into the antibacterial activity and selectivity of MSI-78. Interestingly, the peptide exhibits a concentration-dependent membrane orientation in the lamellar-phase POPG bilayer and was also found to induce toroidal-type pore formation. The deduced lipid flip-flop from SFG signals observed from lipids also supports MSI-78-induced toroidal-type pore formation.     

Solubilization of supported lipid membranes by octyl glucoside observed by time-lapse atomic force microscopy

Detergents are very useful for the purification of membrane proteins. A good detergent for protein extraction has to prevent denaturation by unfolding, and to avoid aggregation. Therefore, gaining access to the mechanism of biomembranes’ solubilization by detergents is crucial in biochemical research. Among the wide range of detergents used to purify membrane proteins, n-octyl β-d-glucopyranoside (OG) is one of the most important as it can be easily removed from final protein extracts.

Here, we used real-time atomic force microscopy (AFM) imaging to visualize the behavior of a model supported lipid bilayer in the presence of OG. Two kinds of supported model membranes were prepared by fusion of unilamellar vesicles: with an equimolar mixing of dioleoylphosphatidylcholine/dipalmitoylphosphatidylcholine (DOPC/DPPC) or with DPPC alone. Time-lapse AFM experiments evidenced that below its critical micelle concentration (CMC), OG was not able to solubilize the bilayer but the gel DPPC domains were instantly dissolved into the DOPC matrix. This result was interpreted as a disorganization of the DPPC molecular packing induced by OG. When membranes were incubated with OG at concentrations higher than CMC, the detergent immediately provoked the complete and immediate desorption of the whole bilayer for both compositions: DPPC and DOPC/DPPC. After a while, some patches appeared onto the bare mica surface. This redeposition activity, together with fusion events, progressively led to the recovery of a continuous bilayer. These results provide a new insight on the unique properties of OG employed in membrane reconstitution protocols.     

Relationship between lipid peroxidation and rigidity in L-alpha-phosphatidylcholine-DPPC vesicles

DPPC incorporation into egg-PC unilamellar vesicles reduces their oxidation rate beyond that expected from the unsaturated lipid dilution. Addition of the unsaturated lipids produces changes in the physical properties of the inner parts of the lipid bilayer, as sensed by fluorescence anisotropy of DPH, and in the hydrophilic/hydrophobic region, as sensed by the generalized polarization of laurdan. DPPC (30 mol%) incorporation into egg-PC vesicles produces a decrease in alkyl chain mobility in the inner part of the bilayer, evaluated by the increase of DPH fluorescence anisotropy, and a rise of the generalized polarization value of laurdan in the bilayer interface. It also leads to a decrease in the rate of water efflux promoted by a hypertonic shock. Oxidation of PC LUVs, promoted by AAPH, as sensed by oxygen uptake and MDA formation, leads to qualitatively similar results than DPPC addition: rigidification at the inner part and the surface of the liposomes, and a lower rate of water permeation. It is suggested that these changes could contribute to the observed decrease in oxidation rate with conversion.     

Effect of pressure on the Prodan fluorescence in bilayer membranes of phospholipids with varying acyl chain lengths

The fluorescence spectra of 6-propionyl-2-(dimethylamino)naphthalene (Prodan) were observed as a function of pressure for the bilayer membrane systems of dilauroylphosphatidylcholine (DLPC), dimyristoylphosphatidylcholine (DMPC), dipalmitoylphosphatidylcholine (DPPC), and distearoylphosphatidylcholine (DSPC). The wavelength of the emission maximum, lambdamax, was found to be 480, 430, and 500 nm for the liquid crystalline (Lalpha), ripple gel (P'beta), and pressure-induced interdigitated gel (LbetaI) phase, respectively. Since the lambdamax reflects the solvent property around the probe molecules, we could speculate on the location of the Prodan molecules in the bilayer membranes; in the Lalpha phase of the lipid bilayer, the Prodan molecules distribute around the phosphate of the lipids (i.e. the polar region). The Lalpha/P'beta phase transition caused the Prodan molecules to move into the less polar region near the glycerol backbone. The fluorescence intensity of the Prodan in the P'beta phase was dependent on the chain length of the lipids and on pressure; the shorter the chain length of the lipid, the stronger the fluorescence intensity of the Prodan. Moreover, for the DLPC bilayer membrane system, the fluorescence intensity at 430 nm increased with increasing pressure, indicating that the partition of Prodan into the DLPC bilayer membrane is promoted by applying pressure. In the case of the DPPC and DSPC bilayers, as the pressure increased further, the pressure-induced interdigitation caused the Prodan molecules to squeeze out of the glycerol backbone region and to move the hydrophilic region near the bilayer surface. The ratio of fluorescence intensity at 480 nm to that at 430 nm, F480/F430, showed a sharp change at the phase-transition pressure. In the case of the DPPC and DSPC bilayers, the values of F480/F430 showed an abrupt increase above a certain pressure higher than the Lalpha/P'beta transition pressure, which corresponds to the interdigitation from the P'beta to the LbetaI phase. The plot of F480/F430 versus pressure is available for recognition of the bilayer phase transitions, especially the bilayer interdigitation.     

Porphinato Zinc Complexes Incorporated into the Bilayer Membrane of Lipid Liposomes

3,8,13,17-Tetramethyl-7,12-divinyl-2,18-bis(18-hydroxyoctadecyl propionate) porphinato zinc (BHPZn) and a model compound, dimethyl ester of protoporphinato zinc (DMPZn), were synthesized and incorporated into the hydrophobic region of bilayer membrane of dipalmitoylphosphatidylcholine (DPPC) liposome. The introduction of long alkyl groups onto the porphyrin ring is effective for restriction of porphyrin aggregation in the bilayer membrane of DPPC liposome. When the molar ratio of DPPC lipid to porphyrin is above 100, the spectrum of BHPZn in the liposome suggests that it is in a typically monomeric state. Quenching of BHPZn fluorescence in the hydrophobic bilayer membrane by hydrophilic quenchers is slow and shows smaller Stern-Volmer constants, while the quenching by hydrophobic quenchers shows much larger Stern-Volmer constants than that of the model compound, DMPZn. These results suggest that the location of the porphyrin ring of BHPZn is fixed at a certain depth in the hydrophobic bilayer membrane of DPPC liposome, and that that of DMPZn is widely distributed in the whole hydrophobic region.     

Mechanisms of the interaction of alpha-helical transmembrane peptides with phospholipid bilayers

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.     

Influence of lipidation of GBV-C/HGV NS3 (513-522) and (505-514) peptide sequences on its interaction with mono and bilayers

Two decapeptide fragments of the non-structural hepatitis G NS3 protein (GBV-C/HGV), 513-522 (RGRTGRGRSG) and 505-514 (SAELSMQRRG), as well as their palmitoylated derivatives were synthesized. The physico-chemical properties of the peptides were analyzed in both the absence and presence of the zwitterionic 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC), the negative 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) and the positive 1,2-dioeloyl-3-trimethylammonium-propane (DOTAP) lipid monolayers. Based on their high hydrophilic properties, neither parent peptide presented surface activity and their incorporation into lipid monolayers was low. In contrast, their palmitoylated derivatives showed concentration-dependent surface activity and could be inserted into lipid monolayers to varying degrees depending on their sequence. Compression isotherms showed that the presence of palmitoylated peptides in the subphase resulted in a molecular arrangement less condensed than that corresponding to the pure phospholipid. In concordance with the monolayer results, differential scanning calorimetry (DSC) demonstrated that the parent peptides did not have any effect on the thermograms, while the palmitoylated derivatives affected the thermotropic properties of DPPC bilayers.     

Physicochemical properties of a muramyldipeptide derivative (B30-MDP) related to membrane formation

We investigated the physicochemical properties of B30-MDP [6-O-(2-tetradecylhexadecanoyl)-N-acetyl-muramyl-L-alanyl-D-isoglutamine], a muramyldipeptide derivative having immunoadjuvant activity [1], using polarizing optical microscopy, differential scanning calorimetry (DSC), and electron spin resonance (ESR) spectroscopy. Microscopic observations showed that B30-MDP molecules form myelin figures in phosphate buffered saline (PBS). It was revealed that B30-MDP forms membranous structure because of an increase in the hydrophobicity. In the DSC measurements, the B30-MDP membrane in PBS gave no endothermic peak between 5° to 50°C. Enthalpy change upon the phase transition from the gel to liquid crystalline state or dipalmitoylphosphatidylcholine (DPPC) membrane and its phase transition temperature decreased by the addition of B30-MDP. ESR measurements using 5 doxyl stearic acid showed that the fluidity of the B30-MDP membrane was almost comparable to that of DPPC membrane at the temperature below the phase transition temperature of DPPC, while it was lower than that of DPPC at the temperature higher than this point. The fluidity of DPPC membrane increased upon the addition of B30-MDP. These results indicate that B30-MDP forms membranous structure and that the bulky hydrophilic region of B30-MDP influences its membrane structures, thermal behavior, and membrane fluidity.     

Phase separation of saturated and mono-unsaturated lipids as determined from a microscopic model

A molecular model is proposed of a bilayer consisting of fully saturated dipalmitoylphosphatidylcholine (DPPC) and mono-unsaturated dioleoylphosphatidylcholine (DOPC). The model not only encompasses the constant density within the hydrophobic core of the bilayer, but also the tendency of chain segments to align. It is solved within self-consistent field theory. A model bilayer of DPPC undergoes a main-chain transition to a gel phase, while a bilayer of DOPC does not do so above zero degrees centigrade because of the double bond which disrupts order. We examine structural and thermodynamic properties of these membranes and find our results in reasonable accord with experiment. In particular, order-parameter profiles are in good agreement with NMR experiments. A phase diagram is obtained for mixtures of these lipids in a membrane at zero tension. The system undergoes phase separation below the main-chain transition temperature of the saturated lipid. Extensions to the ternary DPPC, DOPC, and cholesterol system are outlined.     

HIV fusion inhibitor peptide T-1249 is able to insert or adsorb to lipidic bilayers. Putative correlation with improved efficiency

T-1249 is a HIV fusion inhibitor peptide under clinical trials. Its interaction with biological membrane models (large unilamellar vesicles) was studied using fluorescence spectroscopy. A gp41 peptide that includes one of the hydrophobic terminals of T-1249 was also studied. Both peptides partition extensively to liquid-crystalline POPC (1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine) (DeltaG = -7.0 kcal/mol and -8.7 kcal/mol, for T-1249 and terminal peptide, respectively) and are located at the interface of the membrane. T-1249 is essentially in a random coil conformation in this lipidic medium, although a small alpha-helix contribution is present. When other lipid compositions are used (DPPC, POPG + POPC, and POPC + cholesterol) (DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) and POPG (1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)), partition decreases, the most severe effect being the presence of cholesterol. Partition experiments and fluorescence resonance energy transfer analysis show that T-1249 adsorbs to cholesterol-rich membranes. The improved clinical efficiency of T-1249 relative to enfuvirtide (T20) may be related to its bigger partition coefficient and ability to adsorb to rigid lipidic areas on the cell surface, where most receptors are inserted. Moreover, adsorption to the sterol-rich viral membrane helps to increase the local concentration of the inhibitor peptide at the fusion site.