Bilayer Interior Research Articles

A Fluorescent Probe for C24 Fatty Acids to Report the Deep Interior of Lipid Bilayers

AbstractTo investigate the interdigitation and the resulting fluctuation of lipid chains in the deep part of biomembranes, we aimed to develop a new fluorescent probe based on the well-known trans-parinaric acid (t-PA), but with a tetraene near the end of the C24 chain. The probe, tetracosa-16,18,20,22-tetraenoic acid (TCTEA), synthesized mainly by means of reported strategies, was found to better reflect the behavior of lipids in the deeper interior of bilayers than did t-PA. TCTEA could serve as a useful probe for studying the plasma-membrane dynamics of very-long-chain fatty acids, which have recently elicited significant attention in relation to necroptosis.

Delineating effect of headgroup and preparation method on transfection versus toxicity of DNA-loaded lipid nanocarriers.

Aim: To perform a parallel comparison of key parameters affecting the safety and efficiency of lipid-based nanovectors (i.e., complexing headgroups, composition and preparation method). Materials & methods: Various cationic and ionizable headgroups were screened for formulating lipoplexes with GFP-plasmid DNA. Ethanol injection and microfluidics were used to prepare nanoparticles with GFP-plasmid DNA complexed on the surface or within the interior of lipid bilayers. Results: Lipoplexes composed of sphingomyelin 102 exhibited the highest transfection efficiency given their higher cellular uptake in BRAF inhibitor-resistant melanoma cells. Lipid nanoparticlesdemonstrated acceptable transfection efficiency and high spheroid penetration while protecting plasmid DNA under simulated physiological conditions. Conclusion: Selecting the right complexing lipid and preparation method is critical for developing lipid nanocarriers to treat intractable diseases.

Probing the Hydrophobic Region of a Lipid Bilayer at Specific Depths Using Vibrational Spectroscopy.

A novel spectroscopic approach for studying the flexibility and mobility in the hydrophobic interior of lipid bilayers at specific depths is proposed. A set of test compounds featuring an azido moiety and a cyano or carboxylic acid moiety, connected by an alkyl chain of different lengths, was synthesized. FTIR data and molecular dynamics calculations indicated that the test compounds in a bilayer are oriented so that the cyano or carboxylic acid moiety is located in the lipid head-group region, while the azido group stays inside the bilayer at the depth determined by its alkyl chain length. We found that the asymmetric stretching mode of the azido group (νN3) can serve as a reporter of the membrane interior dynamics. FTIR and two-dimensional infrared (2DIR) studies were performed at different temperatures, ranging from 22 to 45 °C, covering the Lβ-Lα phase transition temperature of dipalmitoylphosphatidylcholine (∼41 °C). The width of the νN3 peak was found to be very sensitive to the phase transition and to the temperature in general. We introduced an order parameter, SN3, which characterizes restrictions to motion inside the bilayer. 2DIR spectra of νN3 showed different extents of inhomogeneity at different depths in the bilayer, with the smallest inhomogeneity in the middle of the leaflet. The spectral diffusion dynamics of the N3 peak was found to be dependent on the depth of the N3 group location in the bilayer. The obtained results enhance our understanding of the bilayer dynamics and can be extended to investigate membranes with more complex compositions.

The effects of adsorbed benzo(a)pyrene on dynamic behavior of polystyrene nanoplastics through phospholipid membrane: A molecular simulation study

Nanoplastics (NPs) are mainly generated from the decomposition of plastic waste and industrial production, which have attracted much attention due to the potential risk for humans. The ability of NPs to penetrate different biological barriers has been proved, but the understanding of molecular details is very limited, especially for organic pollutant-NP combinations. Here, we investigated the uptake process of polystyrene NPs (PSNPs) combined with benzo(a)pyrene (BAP) molecules by dipalmitoylphosphatidylcholine (DPPC) bilayers by molecular dynamics (MD) simulations. The results showed that the PSNPs can adsorb and accumulate BAP molecules in water phase and then carried BAP molecules to enter DPPC bilayers. At the same time, the adsorbed BAP promoted the penetration of PSNPs into DPPC bilayers effectively by hydrophobic effect. The process of BAP-PSNP combinations penetrating into DPPC bilayers can be summarized into four steps including adhesion on the DPPC bilayer surface, uptake by the DPPC bilayer, BAP molecules detached from the PSNPs, and the PSNPs depolymerized in the bilayer interior. Furthermore, the amount of adsorbed BAP on PSNPs affected the properties of DPPC bilayers directly, especially the fluidity of DPPC bilayers that determine the physiologic function. Obviously, the combined effect of PSNPs and BAP enhanced the cytotoxicity. This work not only presented a vivid transmembrane process of BAP-PSNP combinations and revealed the nature of the effects of adsorbed benzo(a)pyrene on the dynamic behavior of polystyrene nanoplastics through phospholipid membrane, but also provide some necessary information of the potential damage for organic pollutant-nanoplastic combinations on human health at a molecular level.

Effect of oxidation on POPC lipid bilayers: anionic carboxyl group plays a major role.

Phospholipids with unsaturated acyl chains are major targets of reactive oxygen species leading to formation of oxidized lipids. Oxidized phospholipids have a pronounced role in cell membrane damage. We investigated the effect of oxidation on physiological properties of phospholipid bilayers using atomistic molecular dynamics simulations. We studied phospholipid bilayer systems of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and its two stable oxidized products, 1-palmitoyl-2-(9'-oxo-nonanoyl)-sn-glycero-3-phosphocholine (PoxnoPC) and 1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine (PazePC). Structural properties of the POPC lipid bilayer upon the addition of PoxnoPC or PazePC with concentration ranging from 10% to 30% were described. The key finding is that PazePC lipids bend their polar tails toward the bilayer-water interface whereas PoxnoPC lipids orient their tail toward the bilayer interior. The bilayer thickness decreases such that the thickness reduction in bilayers containing PazePC is stronger than in bilayers containing PoxnoPC. The average area per lipid decreases with a stronger effect in bilayers containing PoxnoPC. The addition of PoxnoPC makes both POPC acyl chains slightly more ordered whereas the addition of PazePC reduces the order in the two POPC acyl chains. These structural changes lead to an enhancement in the permeabilities of the bilayers containing these two oxidized products depending on the type, and the amount of oxidation. This enhancement can be achieved with a lower concentration of PazePC (10% or 15%), whereas a higher concentration of PoxnoPC (20%) is required to achieve an apparent enhancement in permeability. While the permeability of bilayers containing PazePC is higher than bilayers containing PoxnoPC in the 10-20% concentration range, by increasing the concentration of the oxidized products to higher than 20%, permeability of the bilayers containing PazePC is reduced such that it is slightly smaller than those containing PoxnoPC.

Elucidating collective translocation of nanoparticles across the skin lipid matrix: a molecular dynamics study.

The top layer of skin, the stratum corneum, provides a formidable barrier to the skin. Nanoparticles are utilized and further explored for personal and health care applications related to the skin. In the past few years, several researchers have studied the translocation and permeation of nanoparticles of various shapes, sizes, and surface chemistry through cell membranes. Most of these studies focused on a single nanoparticle and a simple bilayer system, whereas skin has a highly complex lipid membrane architecture. Moreover, it is highly unlikely that a nanoparticle formulation applied on the skin will not have multiple nanoparticle-nanoparticle and skin-nanoparticle interactions. In this study, we have utilized coarse-grained MARTINI molecular dynamics simulations to assess the interactions of two types (bare and dodecane-thiol coated) of nanoparticles with two models (single bilayer and double bilayer) of skin lipid membranes. The nanoparticles were found to be partitioned from the water layer to the lipid membrane as an individual entity as well as in the cluster form. It was discovered that each nanoparticle reached the interior of both single bilayer and double bilayer membranes irrespective of the nanoparticle type and concentration, though coated particles were observed to efficiently traverse across the bilayer when compared with bare particles. The coated nanoparticles also created a single large cluster inside the membrane, whereas the bare nanoparticles were found in small clusters. Both the nanoparticles exhibited preferential interactions with cholesterol molecules present in the lipid membrane as compared to other lipid components of the membrane. We have also observed that the single membrane model exhibited unrealistic instability at moderate to higher concentrations of nanoparticles, and hence for translocation study, a minimum double bilayer model should be employed.

Delivery mechanism of doxorubicin by PEG-DPPE micelles on membrane invasion by dynamic simulations.

Exploiting micelles of polyethylene glycol-dipalmitoylglycerophosphoethanolamine (PEG-DPPE) as a drug delivery approach is of great promise for improving therapeutic targeting and the half-lives of drugs. To optimize the micelle carriers, pending issues concerning the kinetics underlying the carrier-membrane interplay and the specific contributions of the micelle hydrophobic/hydrophilic components remain to be addressed. Relying on MARTINI coarse-grain (CG) molecular dynamics simulations, we explored the carrier-membrane fusion dynamics of PEG-DPPE micelles with different PEG repetitions in delivering doxorubicin (DOX). A bilayer model composed of 20% phosphatidylglycerol (POPG) and 80% phosphatidylcholine (POPC) was constructed to mimic anionic cancer cell membranes. The CG model of DOX was pioneeringly constructed herein, and it was found to distribute at the hydrophilic/hydrophobic interface of the PEGylated micelles, in agreement with experimental results. The free DOXs cause insignificant disorder of the membrane organization, whereas the PEG-DPPE micelles encapsulating DOX lead to a remarkable membrane invasion supported by the order parameter of the lipid acyl carbon tails and the membrane permeation free energy of DOX. The carrier-bilayer interaction shows a stepwise form attributed to the rearrangement of the zwitterionic/anionic lipids upon the absorption of the DOX-micelle complex on a membrane locality, which initiates the rapid release of DOX to the bilayer interior. Benefiting from the enhanced micelle-membrane interplay, the PEG1250-DPPE micelles result in severe bilayer breakage and deeper membrane insertion of DOX compared to the PEG2000-DPPE micelles. This study provides new theoretical insights into the mechanism of PEG-DPPE micelles in delivering drugs through membranes, which is of benefit for further optimization of PEGylated delivery systems.

Self-Uptake Mechanism of Polymyxin-Based Lipopeptide against Gram-Negative Bacterial Membrane: Role of the First Adsorbed Lipopeptide.

Research in the continuously increasing threat of polymyxin-resistant multidrug-resistant Pseudomonas aeruginosa, which causes severe infection in immunocompromised patients, has resulted in the development of several polymyxin-derived cyclic lipopeptides containing l-α-γ- diamino butyric acid-like FADDI-019 (F19). In this work, F19's insertion into a minimal model of the asymmetric outer membrane of the bacterium, which contained only penta-acylated lipid A (LipA) and lacked keto-d-octulosonic acid and O-antigens, in the top leaflet and phospholipids in the bottom leaflet, was studied. F19 exhibited all of the hallmarks of the self-uptake mechanism into the asymmetric bilayer. While a single monomer of the lipopeptide did not get partitioned into the inside of the bilayer, it competitively displaced Ca2+ from the membrane surface, observed as a decrease in Ca2+ coordination number with phosphate groups (1.89 vs 1.718), resulting in membrane destabilization. This resulted in an increment of the average defect size and the probability of interplay between lipid tails and hydrophobic residues of another F19. When more than one monomer was present in the system, the first monomer remained docked on the surface, while other monomers intercalated into the bilayer interior with their hydrophobic moieties "sleeved" by lipid acyl chains. The free energy barrier for partial insertion of the lipopeptide into a bilayer in the presence of surface-docked second F19 was recorded at ∼1.3 kcal/mol using two-dimensional (2D) well-tempered metadynamics, making it a low barrier process at 300 K. This study is an attempt to demonstrate the self-uptake mechanism of F19 during intercalation process into the bilayer interior, which may help in the design of better alternates for polymyxins to work against polymyxin resistance.

Amyloid fibrils act as a reservoir of soluble oligomers, the main culprits in protein deposition diseases.

Amyloid fibril formation plays a central role in the pathogenesis of a number of neurodegenerative diseases, including Alzheimer and Parkinson diseases. Transient prefibrillar oligomers forming during the aggregation process, exhibiting a small size and a large hydrophobic surface, can aberrantly interact with a number of molecular targets on neurons, including the lipid bilayer of plasma membranes, resulting in a fatal outcome for the cells. By contrast, the mature fibrils, despite presenting generally a high hydrophobic surface, are endowed with a low diffusion rate and poorly penetrate the interior of the lipid bilayer. However, increasing evidence shows that both intracellular α-synuclein fibrils, as well and as extracellular amyloid-β and β2-microglobulin fibrils, can release oligomers over time that quickly diffuse to reach the membrane of the neighboring cells. The persistent leakage of harmful oligomers from fibrils triggers an ongoing cascade of events resulting in a sustained injury to neurons and glia and also provides aggregates with the ability to cross biological membranes and diffuse between cells or cellular compartments.

Synthesis of Spin-Labeled Ibuprofen and Its Interaction with Lipid Membranes.

Ibuprofen is a non-steroidal anti-inflammatory drug possessing analgesic and antipyretic activity. Electron paramagnetic resonance (EPR) spectroscopy could be applied to study its interaction with biological membranes and proteins if its spin-labeled analogs were synthesized. Here, a simple sequence of ibuprofen transformations—nitration, esterification, reduction, Sandmeyer reaction, Sonogashira cross-coupling, oxidation and saponification—was developed to attain this goal. The synthesis resulted in spin-labeled ibuprofen (ibuprofen-SL) in which the spin label TEMPOL is attached to the benzene ring. EPR spectra confirmed interaction of ibuprofen-SL with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayers. Using 2H electron spin echo envelope modulation (ESEEM) spectroscopy, ibuprofen-SL was found to be embedded into the hydrophobic bilayer interior.

THE INFLUENCE OF CHLORPROMAZINE HYDROCHLORIDE ON THE THERMOTROPIC BEHAVIOR OF DIMYRISTOYL PHOSPHATIDYLCHOLINE LIPOSOMES AS REVEALED BY DIFFERENTIAL SCANNING CALORIMETRY

Objective: The aim of this study is to investigate the influence of the model cationic, amphiphilic, drug chlorpromazine hydrochloride (CPZ-HCl) on the thermotropic behavior of dimyristoyl phosphatidylcholine (DMPC) liposomes, using differential scanning calorimetry (DSC). The effect of sonication, charged lipids and CPZ-HCl at concentrations known to cause anesthesia on the enthalpy (ΔHt), entropy (ΔSt), phase transition (Tc), pre-transition (pre-TC) and half-height width (HHW) of DSC thermograms were examined. Methods: The experiments conducted, using the Perkin Elmer (DSC-2C), include the effect of a wide range of CPZ-HCl concentrations on ΔHt, ΔSt, Tc, pre-Tc and HHW of DSC thermograms of DMPC liposomes. The effect of sonication on ΔHt, ΔSt, Tc and HHW of DSC thermograms of DMPC/CPZ-HCl liposomes as a function of sonication time. The effect of both positively charged stearyl amine (ST) and negatively charged diacetyl phosphate (DCP) lipids on ΔHt, ΔSt and Tc of DMPC/CPZ-HCl liposomes. In addition, the effect of CPZ-HCl at concentrations known to cause anesthesia on ΔHt, ΔSt. and Tc of DMPC liposomes in the presence and absence of ST and DCP in phosphate buffer (pH 7.4), was also carried out. Results: Using DSC, CPZ-HCl concentrations as low as 1×10-7M were observed to alter the gel-liquid crystalline phase transition and thus to possess a membrane destabilizing effect. CPZ-HCl reduces ΔHt, ΔSt, TC, the pre-TC and increases HHW of DMPC liposomes. ΔHt and ΔSt of DMPC liposomes were observed to decrease with increasing CPZ-HCl concentrations, exhibiting an inflection point at 5×10-5M. ΔHt of DMPC liposomes was observed to decrease linearly in the absence and presence of and CPZ-HCl as a function of sonication time. Both ΔHt and ΔSt of DMPC liposomes were observed to increase in the presence of cationic lipid (ST) and to decrease in the presence of anionic lipid (DCP). ΔSt and Tc of DMPC, DMPC/ST, DMPC/DCP liposomes, were found to decrease as a function of CPZ-HCl concentrations known to cause anesthesia. Conclusion: Using DSC, CPZ-HCl concentrations, as low as 1×10-7 M were observed to influence the enthalpy, entropy, phase transition, pre-transition and half-height width of DSC thermograms of DMPC liposomes, altering the gel-liquid crystalline phase transition and thus possessing a membrane destabilizing effect. It can also be inferred that CPZ-HCl interacts with both the polar head group and the hydrophobic interior of the phospholipid bilayer. These results could support the hypothesis that the addition of local anesthetics might trigger a change in the lipid surrounding the sodium channel from the gel to the liquid crystalline state, allowing the sodium channel to close with the resulting anesthesia.

3,4,5-Trimethoxybenzoate of Catechin, an Anticarcinogenic Semisynthetic Catechin, Modulates the Physical Properties of Anionic Phospholipid Membranes

3,4,5-Trimethoxybenzoate of catechin (TMBC) is a semisynthetic catechin which shows strong antiproliferative activity against malignant melanoma cells. The amphiphilic nature of the molecule suggests that the membrane could be a potential site of action, hence the study of its interaction with lipid bilayers is mandatory in order to gain information on the effect of the catechin on the membrane properties and dynamics. Anionic phospholipids, though being minor components of the membrane, possess singular physical and biochemical properties that make them physiologically essential. Utilizing phosphatidylserine biomimetic membranes, we study the interaction between the catechin and anionic bilayers, bringing together a variety of experimental techniques and molecular dynamics simulation. The experimental data suggest that the molecule is embedded into the phosphatidylserine bilayers, where it perturbs the thermotropic gel to liquid crystalline phase transition. In the gel phase, the catechin promotes the formation of interdigitation, and in the liquid crystalline phase, it decreases the bilayer thickness and increases the hydrogen bonding pattern of the interfacial region of the bilayer. The simulation data agree with the experimental ones and indicate that the molecule is located in the interior of the anionic bilayer as monomer and small clusters reaching the carbonyl region of the phospholipid, where it also disturbs the intermolecular hydrogen bonding between neighboring lipids. Our observations suggest that the catechin incorporates well into phosphatidylserine bilayers, where it produces structural changes that could affect the functioning of the membrane.

Ultrafast energy transfer between self-assembled fluorophore and photosynthetic light-harvesting complex 2 (LH2) in lipid bilayer.

Photosynthetic light-harvesting (LH) systems consist of photosynthetic pigments, which are non-covalently self-assembled with protein scaffolds in many phototrophs and attain highly efficient excitation energy transfer via ultrafast dynamics. In this study, we constructed a biohybrid LH system composed of an LH complex (LH2) from Rhodoblastus acidophilus strain 10050 and a hydrophobic fluorophore ATTO647N (ATTO) as an extrinsic antenna in the lipid bilayer. Through the addition of ATTOs into a solution of LH2-reconstituted lipid vesicles, ATTOs were incorporated into the hydrophobic interior of the lipid bilayer to configure the non-covalently self-assembled biohybrid LH. Steady-state fluorescence spectroscopy clearly showed efficient energy transfer from ATTO to B850 bacteriochlorophylls in LH2. Femtosecond transient absorption spectroscopy revealed that the energy transfer took place in the time range of 3-13ps, comparable to that of the covalently linked LH2-ATTO that we previously reported. In addition, the biohybrid LH system exhibited a much higher antenna effect than the LH2-ATTO system because of the higher loading level of ATTO in the membrane. These findings suggest that the facile self-assembled biohybrid LH system is a promising system for constructing LH for solar-energy conversion.

Effect of phospholipid/flaxseed oil ratio on characteristics, structure change, and storage stability of liposomes

The flaxseed oil, a rich source of α-linolenic acid, was loaded in liposomes. The vesicle characteristics, encapsulation efficiency, and loading content with different amounts of flaxseed oil addition were investigated. At the ratio of phospholipid to flaxseed oil 5:1, the liposomes displayed an almost spherical shape and a homogeneous size distribution (observed by transmission electron microscopy and atomic force microscopy), with an average particle size 156 nm and an encapsulation efficiency of approximately 85%. The addition of flaxseed oil could change the phospholipid arrangement by Fourier-transform infrared spectroscopy and Raman spectra. The hydrophobicity of membrane surface and the micropolarity of the bilayer interior decreased with the incorporation of flaxseed oil by fluorescence probe analysis. Liposomes showed higher oxidative stability and retention rate during the storage at the ratio of 5:1. In addition, the encapsulation of flaxseed oil in liposomes showed the ability of slower release. It comprehensively illustrated the flaxseed oil could affect the membrane structure of liposomes and provided useful information regarding the preparation of liposomes for hydrophobic compounds.

Folding and Insertion of Transmembrane Helices at the ER.

In eukaryotic cells, the endoplasmic reticulum (ER) is the entry point for newly synthesized proteins that are subsequently distributed to organelles of the endomembrane system. Some of these proteins are completely translocated into the lumen of the ER while others integrate stretches of amino acids into the greasy 30 Å wide interior of the ER membrane bilayer. It is generally accepted that to exist in this non-aqueous environment the majority of membrane integrated amino acids are primarily non-polar/hydrophobic and adopt an α-helical conformation. These stretches are typically around 20 amino acids long and are known as transmembrane (TM) helices. In this review, we will consider how transmembrane helices achieve membrane integration. We will address questions such as: Where do the stretches of amino acids fold into a helical conformation? What is/are the route/routes that these stretches take from synthesis at the ribosome to integration through the ER translocon? How do these stretches ‘know’ to integrate and in which orientation? How do marginally hydrophobic stretches of amino acids integrate and survive as transmembrane helices?

Influence of hydration on segmental chain librations and dynamical transition in lipid bilayers

Continuous wave electron paramagnetic resonance spectroscopy of chain-labeled phospholipids is used to investigate the effects of hydration on the librational oscillations and the dynamical transition of phospholipid membranes in the low-temperature range 120–270 K. Bilayers of dipalmitoylphostatidiycholine (DPPC) spin-labeled at the first acyl chain segments and at the methyl ends and prepared at full, low, and very low hydration are considered. The segmental mean-square angular amplitudes of librations, 〈α2〉, are larger in the bilayer interior than at the polar/apolar interface and larger in the fully and low hydrated than in the very low hydrated membranes. For chain segments at the beginning of the hydrocarbon region, 〈α2〉-values are markedly restricted and temperature independent in DPPC with the lowest water content, whereas they increase with temperature in the low and fully hydrated bilayers, particularly at the highest temperatures. For chain segments at the chain termini, the librational amplitudes increase progressively, first slowly and then more rapidly with temperature in bilayers at any level of hydration. From the temperature dependence of the mean-square librational amplitude, the dynamical transition is detected around 240 K at the polar/apolar interface in fully and low hydrated DPPC and at around 225 K at the inner hydrocarbon region for bilayers at any hydration condition. At the dynamical transition the bilayers cross low energy barriers of activation energy in the range 10–20 kJ/mol. The results highlight biophysical properties of DPPC bilayers at low-temperature and provide evidence of the effects of the hydration on the dynamical transition in bilayers.

Molecular Simulation Studies on the Interactions of Bilirubin at Different States with a Lipid Bilayer.

The unconjugated bilirubin (BR) may penetrate through the cell membrane and cause a severe cytotoxicity. However, the molecular mechanism underlying the penetration of BR into the cell membrane is still largely unknown. In this work, we systematically investigate the interaction of BR and a lipid bilayer under different conditions by using all-atom molecular dynamics simulations. It is found that BR at the Z,Z conformation can easily enter into the interior of the lipid bilayer due to its hydrophobicity. However, when BR transforms from the Z,Z conformation to the E,E conformation (after the blue-light emission), its penetration ability is greatly reduced (especially at its ionized state). This study may offer useful physical insights into the effect of phototherapy on the penetration behavior and the cytotoxicity of the unconjugated BR.

Amino acids change solute affinity for lipid bilayers

Time-resolved fluorescence and differential scanning calorimetry (DSC) were used to examine how two amino acids, L-phenylalanine (L-PA) and N-acetyl-DL-tryptophan (NAT), affect the temperature-dependent membrane affinity of two structurally similar coumarin solutes for 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) vesicles. The 7-aminocoumarin solutes, coumarin 151 (C151) and coumarin 152 (C152), differ in their substitution at amine position—C151 is a primary amine, and C152 is a tertiary amine—and both solutes show different tendencies to associate with lipid bilayers consistent with differences in their respective log-P-values. Adding L-PA to the DPPC vesicle solution did not change C151’s propensity to remain freely solvated in aqueous solution, but C152 showed a greater tendency to partition into the hydrophobic bilayer interior at temperatures below DPPC’s gel-liquid crystalline transition temperature (Tgel-lc). This finding is consistent with L-PA’s ability to enhance membrane permeability by disrupting chain-chain interactions. Adding NAT to DPPC-vesicle-containing solutions changed C151 and C152 affinity for the DPPC membranes in unexpected ways. DSC data show that NAT interacts strongly with the lipid bilayer, lowering Tgel-lc by up to 2°C at concentrations of 10 mM. These effects disappear when either C151 or C152 is added to solution at concentrations below 10 μM, and Tgel-lc returns to a value consistent with unperturbed DPPC bilayers. Together with DSC results, fluorescence data imply that NAT promotes coumarin adsorption to the vesicle bilayer surface. NAT’s effects diminish above Tgel-lc and imply that unlike L-PA, NAT does not penetrate into the bilayer but instead remains adsorbed to the bilayer’s exterior. Taken in their entirety, these discoveries suggest that amino acids—and by inference, polypeptides and proteins—change solute affinity for lipid bilayers with specific effects that depend on individualized amino-acid-lipid-bilayer interactions.

Capacity for increased surface area in the hydrophobic core of β-sheet peptide bilayer nanoribbons.

Amphipathic peptides with amino acids arranged in alternating patterns of hydrophobic and hydrophilic residues efficiently self‐assemble into β‐sheet bilayer nanoribbons. Hydrophobic side chain functionality is effectively buried in the interior of the putative bilayer of these nanoribbons. This study investigates consequences on self‐assembly of increasing the surface area of aromatic side chain groups that reside in the hydrophobic core of nanoribbons derived from Ac‐(XKXE)2‐NH2 peptides (X = hydrophobic residue). A series of Ac‐(XKXE)2‐NH2 peptides incorporating aromatic amino acids of increasing molecular volume and steric profile (X = phenylalanine [Phe], homophenylalanine [Hph], tryptophan [Trp], 1‐naphthylalanine [1‐Nal], 2‐naphthylalanine [2‐Nal], or biphenylalanine [Bip]) were assessed to determine substitution effects on self‐assembly propensity and on morphology of the resulting nanoribbon structures. Additional studies were conducted to determine the effects of incorporating amino acids of differing steric profile in the hydrophobic core (Ac‐X1KFEFKFE‐NH2 and Ac‐(X1,5KFE)‐NH2 peptides, X = Trp or Bip). Spectroscopic analysis by circular dichroism (CD) and Fourier transform infrared (FT‐IR) spectroscopy indicated β‐sheet formation for all variants. Self‐assembly rate increased with peptide hydrophobicity; increased molecular volume of the hydrophobic side chain groups did not appear to induce kinetic penalties on self‐assembly rates. Transmission electron microscopy (TEM) imaging indicated variation in fibril morphology as a function of amino acid in the X positions. This study confirms that hydrophobicity of amphipathic Ac‐(XKXE)2‐NH2 peptides correlates to self‐assembly propensity and that the hydrophobic core of the resulting nanoribbon bilayers has a significant capacity to accommodate sterically demanding functional groups. These findings provide insight that may be used to guide the exploitation of self‐assembled amphipathic peptides as functional biomaterials.

Molecular organization in hydroperoxidized POPC bilayers

Lipid hydroperoxides are the primary reaction products of lipid oxidation, a natural outcome of life under oxygen. While playing a major role in cell metabolism, the microscopic origins of the effects of lipid hydroperoxidation on biomembranes remain elusive. Here we probe the polar structure of partially to fully hydroperoxidized bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) by a combination of environment-sensitive fluorescent probes and coarse-grained Martini numerical simulations. We find that the inserted organic hydroperoxide group -OOH migrates preferentially to the surface for bilayers with small fractions of hydroperoxidized lipids, but populates also significantly the bilayer interior for larger fractions. Our findings suggest that by modifying the intimate polarity of biomembranes, lipid peroxidation will have a significant impact on the activity of transmembrane proteins and on the bio-medical efficiency of membrane active molecules such as cell-penetrating and antimicrobial peptides.