Hydrophobic Core Of Lipid Bilayer Research Articles

Membrane contact probability: An essential and predictive character for the structural and functional studies of membrane proteins.

One of the unique traits of membrane proteins is that a significant fraction of their hydrophobic amino acids is exposed to the hydrophobic core of lipid bilayers rather than being embedded in the protein interior, which is often not explicitly considered in the protein structure and function predictions. Here, we propose a characteristic and predictive quantity, the membrane contact probability (MCP), to describe the likelihood of the amino acids of a given sequence being in direct contact with the acyl chains of lipid molecules. We show that MCP is complementary to solvent accessibility in characterizing the outer surface of membrane proteins, and it can be predicted for any given sequence with a machine learning-based method by utilizing a training dataset extracted from MemProtMD, a database generated from molecular dynamics simulations for the membrane proteins with a known structure. As the first of many potential applications, we demonstrate that MCP can be used to systematically improve the prediction precision of the protein contact maps and structures.

Effect of Essential Oils on Growth Inhibition, Biofilm Formation and Membrane Integrity of Escherichia coli and Staphylococcus aureus.

Biofilm as a cellular conformation confers survival properties to microbial populations and favors microbial resistance. Here, we investigated the antimicrobial, antibiofilm, antimotility, antihemolytic activity, and the interaction with synthetic membranes of 15 essential oils (EOs) on E. coli ATCC 25922 and S. aureus ATCC 29213. Antimicrobial activity of EOs was determined through microdilution method; development of the biofilm was assessed using the crystal violet assay and SEM microscopy. Results indicate that Lippia origanoides thymol–carvacrol II chemotype (LTC II) and Thymus vulgaris (TV) exhibited a significant antibacterial activity, with MIC values of 0.45 and 0.75 mg/mL, respectively. The percentage of biofilm formation inhibition was greater than 70% at subinhibitory concentrations (MIC50) for LTC II EO. The results demonstrate that these two oils had significantly reduced the hemolytic effect of S. aureus by 54% and 32%, respectively, and the mobility capacity by swimming in E. coli with percentages of decrease of 55% and 47%, respectively. The results show that LTC II and TV EOs can interact with the hydrophobic core of lipid bilayers and alter the physicochemical properties of membranes. The findings suggest that LTC II and TV oils may potentially be used to aid in the treatment of S. aureus and E. coli infections.

Can Nanoplastics Alter Cell Membranes?

Whilst the formation of plastic nanoparticles (nanoplastics) from plastic wastes has been unequivocally evidenced, little is known about the effects of these materials on living organisms at the subcellular or molecular levels. In the present contribution we show through molecular dynamics simulations that polyethylene nanoparticles dissolve in the hydrophobic core of lipid bilayers into a network of disentangled, single polymeric chains. The thereby induced structural and dynamic changes in the bilayer alter vital functions of the cell membrane, which if lacking a mechanism to decompose the polymer chains may result in the death of the cell.

Membrane cholesterol delays cellular apoptosis induced by ginsenoside Rh2, a steroid saponin

Saponins exhibit several biological and pharmacological activities, such as antibacterial, anti-inflammatory and anticancer effects. Many studies attribute their activities to their interactions with cholesterol. In this study, we focus on the steroid saponin ginsenoside Rh2, one of the active principles of Panax ginseng root. Some evidence suggests that lipid rafts, defined as nanodomains enriched in cholesterol and sphingolipids, could be involved in the Rh2-induced apoptosis. However, the role of membrane lipids, especially cholesterol, in this process is still poorly understood. Here, we demonstrate that (i) A549, THP-1 and U937 cells are all susceptible to the Rh2-induced apoptosis but to a differential extent and (ii) the cytotoxic effect inversely correlates with the cell membrane cholesterol content. Upon cholesterol depletion via methyl-β-cyclodextrin, those three cells lines become more sensitive to Rh2-induced apoptosis. Then, focusing on the cholesterol-auxotroph U937 cell line, we showed that Rh2 alters plasma membrane fluidity by compacting the hydrophobic core of lipid bilayer (DPH anisotropy) and relaxing the interfacial packaging of the polar head of phospholipids (TMA-DPH anisotropy). The treatment with Rh2 conducts to the dephosphorylation of Akt and the activation of the intrinsic pathway of apoptosis (loss of mitochondrial membrane potential, caspase-9 and -3 activation). All these features are induced faster in cholesterol-depleted cells, which could be explained by faster cell accumulation of Rh2 in these conditions. This work is the first reporting that membrane cholesterol could delay the activity of ginsenoside Rh2, renewing the idea that saponin cytotoxicity is ascribed to an interaction with membrane cholesterol.

LOCALIZATION AND ORIENTATION OF METHOXY FLAVONOIDS IN DPPC BILAYERS: EFFECT ON THEIR ANTI-PROLIFERATIVE ACTIVITY

Objective: Flavones and flavonols are an important class of naturally occurring flavonoids. They are well known for their pharmacological activity. This activity is associated with the ability of flavones and flavonols to influence membrane–dependent processes. In this paper, we have reported localization, orientation and interaction, of four synthesized flavone/flavonols with 1, 2–dipalmitoyl–sn–glycero–3–phosphocholine (DPPC) bilayers. These are compared with standard flavone; chrysin (CHY) and flavonol Quercetin (QUE).Methods: The molecules studied are 4ʹ–methoxy flavone (MF), 3ʹ,4ʹ–dimethoxyflavone (DMF), 4ʹ–methoxyflavonol (MF–ol) and 3ʹ,4ʹ–dimethoxyflavonol (DMF–ol). The techniques used are Differential Scanning Calorimetry (DSC) and multi–nuclear NMR.Results: Highest binding to lipid bilayers is shown by DMF, followed by QUE. Based on DSC studies it is seen, that maximum interaction of MF and DMF, takes place with the hydrophobic core of lipid bilayers. DMF–ol shows formation of a heterogeneous system at higher concentrations. The 1H NMR spectra of unilamellar vesicles of DPPC, incorporated with MF, DMF and MF–ol shows significant interaction of these compounds with the alkyl chain of the hydrophobic core. MF, DMF and MF–ol acquire parallel orientation in bilayers with the B–ring pointing towards hydrophobic core, while DMF–ol acquire mixed orientation. This may be ascribed to the presence of two methoxy and one hydroxyl group on the B–ring of DMF–ol which hinders its partitioning inside the hydrophobic core of lipid bilayer. Multi–lamellar vesicles (MLV) of DPPC incorporated with flavones, show maximum increase in Chemical Shift Anisotropy in 31P spectrum of DMF. This is followed by MF. DSC.Conclusion: NMR and binding studies indicate that DMF is partitioned deeply inside the hydrophobic core, while MF, MF–ol and DMF–ol are mostly located in the vicinity of sn–glycero region. Therefore, we conclude that DMF which penetrates deepest inside the hydrophobic core also shows the highest anti–proliferative activity against K562 and MCF–7 cancer cell lines. Its activity is also better than CHY.

Implicit membrane treatment of buried charged groups: Application to peptide translocation across lipid bilayers

The energetic cost of burying charged groups in the hydrophobic core of lipid bilayers has been controversial, with simulations giving higher estimates than certain experiments. Implicit membrane approaches are usually deemed too simplistic for this problem. Here we challenge this view. The free energy of transfer of amino acid side chains from water to the membrane center predicted by IMM1 is reasonably close to all-atom free energy calculations. The shape of the free energy profile, however, for the charged side chains needs to be modified to reflect the all-atom simulation findings (IMM1-LF). Membrane thinning is treated by combining simulations at different membrane widths with an estimate of membrane deformation free energy from elasticity theory. This approach is first tested on the voltage sensor and the isolated S4 helix of potassium channels. The voltage sensor is stably inserted in a transmembrane orientation for both the original and the modified model. The transmembrane orientation of the isolated S4 helix is unstable in the original model, but a stable local minimum in IMM1-LF, slightly higher in energy than the interfacial orientation. Peptide translocation is addressed by mapping the effective energy of the peptide as a function of vertical position and tilt angle, which allows identification of minimum energy pathways and transition states. The barriers computed for the S4 helix and other experimentally studied peptides are low enough for an observable rate. Thus, computational results and experimental studies on the membrane burial of peptide charged groups appear to be consistent. This article is part of a Special Issue entitled: Interfacially Active Peptides and Proteins. Guest Editors: William C. Wimley and Kalina Hristova.

Efficiency of the monofunctionalized C60fullerenes as membrane targeting agents studied by all-atom molecular dynamics simulations

Transmembrane translocation of C60 fullerenes functionalized by the single amino-derivative in neutral and charged forms was studies by extensive all-atom molecular dynamics simulations. It is shown that these complexes exhibit very strong affinity to the membrane core, but their spontaneous translocation through the membrane is not possible at practical time scale. In contrast, free amino derivatives translocate through the membrane much easier than their complexes with fullerenes, but do not have pronounced affinity to the membrane interior. Our results suggest that monofunctionalized C60 could be extremely efficient membrane targeting agents, which facilitate accumulation of the water-soluble compounds in the hydrophobic core of lipid bilayer.

Solid-State NMR Reveals the Hydrophobic-Core Location of Poly(amidoamine) Dendrimers in Biomembranes

Poly(amidoamine) (PAMAM) dendrimer nanobiotechnology shows great promise in targeted drug delivery and gene therapy. Because of the involvement of cell membrane lipids with the pharmacological activity of dendrimer nanomedicines, the interactions between dendrimers and lipids are of particular relevance to the pharmaceutical applications of dendrimers. In this study, solid-state NMR was used to obtain a molecular image of the complex of generation-5 (G5) PAMAM dendrimer with the lipid bilayer. Using (1)H radio frequency driven dipolar recoupling (RFDR) and (1)H magic angle spinning (MAS) nuclear Overhauser effect spectroscopy (NOESY) techniques, we show that dendrimers are thermodynamically stable when inserted into zwitterionic lipid bilayers. (14)N and (31)P NMR experiments on static samples and measurements of the mobility of C-H bonds using a 2D proton detected local field protocol under MAS corroborate these results. The localization of dendrimers in the hydrophobic core of lipid bilayers restricts the motion of bilayer lipid tails, with the smaller G5 dendrimer having more of an effect than the larger G7 dendrimer. Fragmentation of the membrane does not occur at low dendrimer concentrations in zwitterionic membranes. Because these results show that the amphipathic dendrimer molecule can be stably incorporated in the interior of the bilayer (as opposed to electrostatic binding at the surface), they are expected to be useful in the design of dendrimer-based nanobiotechnologies.

Perturbation of the hydrophobic core of lipid bilayers by the human antimicrobial peptide LL-37.

LL-37 is a cationic, amphipathic alpha-helical antimicrobial peptide found in humans that kills cells by disrupting the cell membrane. To disrupt membranes, antimicrobial peptides such as LL-37 must alter the hydrophobic core of the bilayer. Differential scanning calorimetry and deuterium ((2)H) NMR experiments on acyl chain perdeuterated lipids demonstrate that LL-37 inserts into the hydrophobic region of the bilayer and alters the chain packing and cooperativity. The results show that hydrophobic interactions between LL-37 and the hydrophobic acyl chains are as important for the ability of this peptide to disrupt lipid bilayers as its electrostatic interactions with the polar headgroups. The (2)H NMR data are consistent with the previously determined surface orientation of LL-37 (Henzler Wildman, K. A., et al. (2003) Biochemistry 42, 6545) with an estimated 5-6 A depth of penetration of the hydrophobic face of the amphipathic helix into the hydrophobic interior of the bilayer. LL-37 also alters the material properties of lipid bilayers, including the area per lipid, hydrophobic thickness, and coefficient of thermal expansion in a manner that varies with lipid type and temperature. Comparison of the effect of LL-37 on 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC-d(31)) and 1,2-dimyristoyl-phosphatidylcholine (DMPC-d(54)) at different temperatures demonstrates the importance of bilayer order in determining the type and extent of disordering and disruption of the hydrophobic core by LL-37. One possible explanation, which accounts for both the (2)H NMR data presented here and the known surface orientation of LL-37 under identical conditions, is that bilayer order influences the depth of insertion of LL-37 into the hydrophobic/hydrophilic interface of the bilayer, altering the balance of electrostatic and hydrophobic interactions between the peptide and the lipids.

Interaction of DnaK with native proteins and membrane proteins correlates with their accessible hydrophobicity.

Molecular chaperones are involved in protein folding, protein targeting to membranes, and protein renaturation after stress. They interact specifically with hydrophobic sequences that are exposed in unfolded proteins, and buried in native proteins. We have studied the interaction of DnaK with native water-soluble proteins and membrane proteins. DnaK-native protein interactions are characterized by dissociation constants between 1 and 50 microM (compared with 0.01-1 microM for unfolded proteins). This affinity is within the range of most intracellular protein concentrations, suggesting that DnaK interacts with a greater number of native proteins than previously suspected. We found a correlation between the affinity of native proteins for DnaK and their affinity for hydrophobic-interaction chromatography adsorbents, suggesting that DnaK interacts with exposed hydrophobic groups in native proteins. The interaction between DnaK and membrane proteins is characterized by DnaK's high affinity for detergent-solubilized membrane proteins, and its lower affinity for membrane proteins inserted in lipid bilayers, suggesting that the chaperone can interact with the hydrophobic sequences of the former, while it cannot penetrate the hydrophobic core of lipid bilayers. Thus, the specificity of DnaK for hydrophobic sequences is involved in its interaction with not only unfolded proteins, but also native water-soluble proteins and membrane proteins. All proteins interact with DnaK according to their exposed hydrophobicity.

Zeaxanthin (dihydroxy-β-carotene) but not β-carotene rigidifies lipid membranes: a 1H-NMR study of carotenoid-egg phosphatidylcholine liposomes

1H-NMR technique was applied to study liposomes formed with egg-yolk phosphatidylcholine containing as an additional component two carotenoid pigments: β-carotene or zeaxanthin (dihydroxy-β-carotene). A strong rigidifying effect of zeaxanthin but not of β-carotene with respect to hydrophobic core of lipid bilayer was concluded from the carotenoid-dependent broadening of the NMR lines assigned to -CH 2- groups and terminal -CH 3 groups of lipid alkyl chains. A similar effect of zeaxanthin with respect to polar headgroups was concluded on the basis of the effect of the pigment on the shape of NMR lines attributed to -N +(CH 3) 3 groups. In contrast, β-carotene increases motional freedom of lipid polar headgroups. The inclusion of both carotenoids to liposomes resulted in the enhanced penetration of Pr 3+ ions to the polar zone of the external layer of a membrane monitored by the splitting of the -N +(CH 3) 3 signal, the effect of β-carotene being much more pronounced. Differences in the effect on membrane structure and molecular dynamics observed for β-carotene and its polar derivative are discussed in terms of organization of a carotenoid-containing lipid membrane.

Interaction of Humic Substances with Liposomes

The effects of marine and terrestrial humic substances on the K+-permeability of lipid bilayer of dimyristoyl phosphatidylcholine (DMPC) liposomes were investigated for their physiological activity on biological membranes. Commercial humic acid and humic and fulvic acids extracted from soil and marine sediment were used for the experiment.Potassium ions inside liposomes were released through the DMPC bilayer following the addition of humic acid. The effect of humic acid was conspicuous above the phase transition temperature of DMPC. Humic acids containing many carboxyl groups but few phenolic and alcoholic hydroxyl groups had little effect on the K+-permeability of the bilayer. From the relation between the K+-permeability and the degree of humification determined by optical spectroscopy, humic acids humified to a lesser extent were found to interact strongly with liposomes. The K+-permeability enhancing effect was stronger in the case of highly aliphatic marine humic acids than highly aromatic terrestrial acids. Hydrophilic fulvic acid exerted virtually no effect on K+-permeability. The hydrophobic chain of the acid would thus appear to be incorporated into the hydrophobic core of lipid bilayer, enhancing the fluidity of the lipid.The lower molecular weight fractions of marine humic acids promoted K+-release more than the original humic acids. The commercial humic acids with smaller molecular size affected strongly K+-permeability compared to marine humic acids having essentially the same humification and functional group content. This molecular size effect indicates the rates of adsorption of humic acid molecules onto and their incorporation into the lipid bilayer to possibly contribute to interactions with liposomes.

Electrostatic and hydrophobic interactions of the intermediate filament protein vimentin and its amino terminus with lipid bilayers.

Immunofluorescence and electron microscopical studies on the intracellular distribution of intermediate filaments (IFs) have demonstrated a close proximity of these cytoskeletal structures to cellular membranes. Moreover, nonepithelial IF (protein)s have been shown to exhibit high affinities for lipids, especially for negatively charged and nonpolar lipids. Here, using hydrophobic labeling with the photoactivatable phosphatidylcholine analogue [3H]1-palmitoyl-2-[11-[4-(trifluoromethyldiazirinyl]undecanoyl+ ++]-sn- glycero-3-phosphorylcholine or with 1-azidopyrene at low and physiological ionic strength, it is demonstrated that the IF subunit protein vimentin can interact with the hydrophobic core of lipid bilayers, in addition to strong ionic relationships between both reactants. Whereas the presence of acidic phospholipids in the lipid vesicles was absolutely essential for efficient vimentin labeling, cholesterol played a synergistic role in this reaction. Proteolytic degradation of photolabeled vimentin localized the derivatization exclusively to the non-alpha-helical, highly positively charged N-terminal domain of the filament protein. Furthermore, circular dichroism studies performed on the isolated N terminus of vimentin revealed a significant increase in the alpha-helical content of the polypeptide upon its interaction with vesicles containing negatively charged phospholipids. These results indicate an amphiphilic character of the N terminus and suggest that the cationic arginine residues of the N-terminal domain react with the negatively charged head groups of acidic phospholipids prior or parallel to interaction of the polypeptide with hydrophobic regions of the lipid bilayer.

Analysis of the mechanism of histamine release induced by substance P

Substance P causes release of histamine from rat peritoneal mast cells; the structure-activity relationship shows that N-terminal residue is essential and the hydrophobic region of C-terminal plays an important role. Electrical conductivity of black lipid membrane containing phosphatidic acid was augmented by substance P. In this case, N-terminal residues and C-terminal hydrophobicity were also unavoidable. The partitioning of substance P into the organic phase increased in the presence of phosphatidic acid. The CD spectrum of substance P was changed from the unordered form to β-form by coexistence of phosphatidic acid/PC liposomes in the medium. The addition of calcium or magnesium in the test solution is effective to prevent either of these phenomena. These findings indicate that substance P probably binds to negatively charged sites of membrane lipids, and subsequent penetration of C-terminal into the hydrophobic core of lipid bilayer may induce an increase of membrane permeability and the following histamine release.

Alamethicin-induced changes in lipid bilayer morphology

We have found that alamethicin, in the absence of an electric field, modifies both the hydrophilic surface and hydrophobic core of lipid bilayers. As shown by freeze-fracture and X-ray diffraction experiments with multiwalled vesicles, alamethicin increases the fluid space between bilayers by as much as 50 nm, and at the same time perturbs the hydrocarbon regions of the bilayers. For suspensions of gel-state lipid treated with alamethicin, uniformly spaced rows of particles cover the fracture faces and corresponding linear arrays of stain-collecting depressions cover the hydrophilic surfaces. In the liquid-crystalline state, alamethicin induces an irregular granular texture on the fracture faces.

Statistical Mechanical Theory for Hydrophobic Core of Lipid Bilayer

Abstract A partition function is proposed for the molten hydrocarbon chains in the hydrophobic region of a lipid bilayer. In the construction, constancy of the volume of the bilayer is accepted as well as constancy of the density within the hydrophobic range when increasing the arqa of the bilayer. MarEelja's method1 for determining the inter- and intramolecular interactions was used. The distribution of the lateral pressure along the thickness of the bilayer and the elasticity modulusof the bilayer (without taking into account the electrostatic interactions between the heads) were determined. Explicite calculations were made for lipids with one and two saturatedalkyl chains with 12 carbon atoms per chain.