Interaction modes between nanosized graphene flakes and liposomes: Adsorption,insertion and membrane fusion

BackgroundUnderstanding the effects of graphene-based nanomaterials on lipid membranes is critical to determine their environmental impact and their efficiency in the biomedical context. Graphene has been reported to favourably interact with biological and model lipid membranes.MethodsWe report on a systematic coarse-grained molecular dynamics study of the interaction modes of graphene nanometric flakes with POPC/cholesterol liposome membranes. We have simulated graphene layers with a variety of sizes and oxidation degrees, and we have analyzed the trajectories, the interaction modes, and the energetics of the observed phenomena.ResultsThree interaction modes are reported. Graphene can be transiently adsorbed onto the liposome membrane and/or inserted in its hydrophobic region. Inserted nanosheets prefer a perpendicular orientation, and tilt in order to maximize the contact with phospholipid tails while avoiding the contact with cholesterol molecules. When placed between two liposomes, graphene facilitates their fusion in a single vesicle.ConclusionsGraphene can be temporary adsorbed on the liposome before insertion. Bilayer curvature has an influence on the orientation of inserted graphene particles. Cholesterol molecules are depleted from the surrounding of graphene particles. Graphene layers may catalyse membrane fusion by bypassing the energy barrier required in stalk formation.General significanceNanometric graphene layers can be adsorbed/inserted in lipid-based membranes in different manners and affect the cholesterol distribution in the membrane, implying important consequences on the structure and functionality of biological cell membranes, and on the bioaccumulation of graphene in living organisms. The graphene-mediated mechanism opens new possibilities for vesicle fusion in the experimental context.     

Zwitterionic polymers functionalised nanoporous graphene for water desalination: a molecular dynamics study

In this paper, one nanoporous graphene grafting several zwitterionic polymer chains was designed as the osmosis membrane for seawater desalination. Using molecular dynamics simulation, the efficiency and mechanism of salt rejection were discussed. The simulated results showed that the zwitterionic polymer chains on nanoporous graphene can form the charge channel to block Na⁺ and Cl^? ions pass through, and the slat rejection efficiency of functionalised graphene can reach to about 90%. In the simulation, the steric hindrance and electrostatic interaction are the main factors for the salt rejection. With time evolution, the charge channel formed by the soft polymer chains can decrease the effective pore area of membrane, leading to the increase of steric hindrance; the positive and negative centres of polymer chains can adsorb Na⁺ and Cl^? ions under electrostatic interaction in the solution, contributing into the increase of charge density above the membrane. These conclusions are consistent with experimental report. Our designed osmosis membrane about the graphene is helpful for improving the potential application of defect graphene in water desalination and reducing the trouble of obtaining appropriate graphene sheet with small aperture.     

Movement and possible metabolism of ethylene in dormant Picea abies

Ethylene or ethane was added to and sampled from the base, middle and top of the stem of intact Picea abies (L.) Karst. using microdialysis probes. The endogenous concentrations of ethylene and ethane in all positions were below 5 × 10^–9 mol l^–1 throughout the experimental period. Applied ethylene or ethane was always detected in probes above the probe used for the application, never below. If increased ethylene or ethane concentrations were detected, the ethylene concentrations initially increased but thereafter decreased to the basal level, whereas levels of ethane steadily increased. It is proposed that the maintainance of low ethylene levels is important in the regulation of wood formation.     

Ethylene and ethane production in response to salinity stress

Abstract Ethylene and ethane production in mung bean hypocotyl sections were evaluated as possible indicators of stress due to contact with four salts that are common in natural sites. Ethylene production decreased with increasing concentrations of applied NaCl and KCl. When CaCl₂ was applied, the ethylene evolution was greater. However, when MgCl₂ was applied, ethylene evolution remained high then decreased and at higher salt concentrations again showed an increase. NaCl (up to 0.1 kmol m^?1) and KCl (up to 0.5 kmol m^?3) caused a concentration-dependent increase in ethane production. The ethane production with CaCl₂ was the lowest among the salts tested and only a minute increase was noticed with the increase of concentration from 0.01 to 1 kmol m^?3. Ethane production showed a distinct maximum at 0.2 kmol m^?3 MgCl₂. The introduction of 0.01 kmol m^?3 CaCl₂, as well as anaerobic conditions obtained by purging vials with N₂, eliminated that high ethane production. Respiratory activity of the mung bean hypocotyl sections in MgCl₂ concentrations from 0 to 0.5 kmol m^?3 was correlated with ethane but not with ethylene production. The ethane/ethylene ratio showed three patterns for the four salts tested.     

A Lack of Specificity for Ethylene-induced Mitochondrial Changes

A critical evaluation was made of the hypothesis that the primary mode of action of ethylene in inducing physiological responses is by changing the permeability of cell organelles. The parameter investigated was the evaluation of the influence of ethylene and other gases on mitochondrial oxidation and swelling. Spectrometric evidence demonstrated that mitochondria prepared with good respiratory control can be induced to swell more rapidly with ethylene and other aliphatic gases (ethane, propene, propane, I-butene) in test solutions of 0.125 m KCl. The fact that saturated as well as unsaturated hydrocarbon gases elicited similar changes provides evidence that ethylene does not directly alter membrane permeability as its mechanism of action.     

Molecular simulations of adsorption and separation of ethylene/ethane and propylene/propane mixtures on Ni2(dobdc) and Ni2(m-dobdc) metal-organic frameworks

Porous solid adsorbents have received considerable attention as a promising alternative to the traditional cryogenic distillation for separating olefin/paraffin mixtures. In this work, we studied pure components as well as ethylene/ethane and propylene/propane binary mixtures uptakes and selectivities at 318 K and 1 bar into metal-organic frameworks Ni₂(dobdc) and Ni₂(m-dobdc) using GCMC simulations. We used DFT method to modify the potential model of carbon–carbon double bond in unsaturated hydrocarbons. GCMC results show that ethylene and ethane uptakes on Ni₂(m-dobdc) are higher than that of Ni₂(dobdc) but propylene and propane uptakes are equal in Ni₂(m-dobdc) and Ni₂(dobdc). Also, Ni₂(m-dobdc) has higher selectivity than Ni₂(dobdc) for separation of ethylene/ethane and propylene/propane mixtures.     

Patterns of peroxidative ethane emission from submerged rice seedlings indicate that damage from reactive oxygen species takes place during submergence and is not necessarily a post-anoxic phenomenon

Using ethane as a marker for peroxidative damage to membranes by reactive oxygen species (ROS) we examined the injury of rice seedlings during submergence in the dark. It is often expressed that membrane injury from ROS is a post-submergence phenomenon occurring when oxygen is re-introduced after submergence-induced anoxia. We found that ethane production, from rice seedlings submerged for 24–72 h, was stimulated to 4–37 nl gFW⁻¹, indicating underwater membrane peroxidation. When examined a week later the seedlings were damaged or had died. On de-submergence in air, ethane production rates rose sharply, but fell back to less than 0.1 nl gFW⁻¹ h⁻¹ after 2 h. We compared submergence-susceptible and submergence-tolerant cultivars, submergence starting in the morning (more damage) and in the afternoon (less damage) and investigated different submergence durations. The seedlings showed extensive fatality whenever total ethane emission exceeded about 15 nl gFW⁻¹. Smaller amounts of ethane emission were linked to less extensive injury to leaves. Partial oxygen shortage (O₂ levels <1%) imposed for 2 h in gas phase mixtures also stimulated ethane production. In contrast, seedlings under anaerobic gas phase conditions produced no ethane until re-aerated: then a small peak was observed followed by a low, steady ethane production. We conclude that damage during submergence is not associated with extensive anoxia. Instead, injury is linked to membrane peroxidation in seedlings that are partially oxygen deficient while submerged. On return to air, further peroxidation is suppressed within about 2 h indicating effective control of ROS production not evident during submergence itself.     

Ethane exhalation as an index of in vivo lipid peroxidation: Concentrating ethane from a breath collection chamber

A simple method is described for concentrating ethane from large volumes of air; the sensitivity limits for detecting exhaled ethane are increased by 100-fold or more. The method is intended for monitoring low-level ethane exhalation resulting from peroxidative damage to tissue lipids in vivo. Various adsorbents for concentrating ethane and ethylene were tested; in addition, the permeability characteristics of various types of tubing were studied in order to construct a suitable, inexpensive breath collection chamber for laboratory animals. Recommended adsorbents are activated charcoal and molecular sleve; activated alumina is a poor adsorbent. Polyvinyl and Teflon tubing are impermeable, whereas latex and silicone tubing are permeable to ethane. When ethane and ethylene are injected intraperitoneally into mice, the exhalation of these volatile hydrocarbons is readily monitored in the breath collection chamber. Whereas ethylene and the C₃–C₅ alkanes probably are metabolized by mice, ethane apparently is not; however, a portion of the ethane appears to be trapped by absorption within body tissues.     

Stimulation of ethylene production in asparagus bean leaves by cupric ion

Asparagus bean (Vigna sesquipedalis Fruhw.) cuttings responded to Cu²⁺ treatments by a rapid and marked increase in ethylene production. This response showed a transient nature but was only partially reversed by returning cuttings to water after prolonged Cu²⁺ treatments suggesting that relative stable links between Cu²⁺ and the ethylene forming system may occur. Cu²⁺ treatments did not induce any visible necrosis nor affect ethane release from cuttings. The Cu²⁺-stimulated ethylene production was not attributable to a light-driven peroxidation of membrane lipids, because it was not affected by a quencher of singlet oxygen nor by a free radical scavenger, whereas it was strongly depressed by light and entirely suppressed by aminoethoxy-vinylglycine. These results support previous finding on the so-called “stress ethylene” and suggest that the Cu²⁺-stimulated ethylene production may be attributable to an enhanced conversion of S-adenosylmethionine to l-aminocyclopropane-l-carboxylie acid. Research work supported by CNR, Italy. Special grant IPRA-Subproject 1. Paper No. 1614.     

基于免疫性PTR的血小板膜糖蛋白及T/B淋巴细胞表达水平研究

摘要 目的：探究血小板膜糖蛋白及T/B淋巴细胞免疫在免疫性血小板输注无效发生机制中的重要作用。方法：采用血小板特异性抗体检测试剂盒及流式细胞技术检测免疫性PTR患者血小板特异性抗体（抗GPIIb/IIIa、抗GPIa/IIa、抗GPIb/IX、抗GPIV）、血小板膜糖蛋白（CD36、CD61、CD41a）表达情况及外周血T/B淋巴细胞数量,运用SPSS19.0软件及R软件分析免疫性PTR与血小板膜糖蛋白、外周血T/B淋巴细胞的相关性。结果：免疫性PTR与血小板输注有效者比较,血小板特异性抗体的产生无显著差异,血小板膜糖蛋白CD36和CD61的表达具有显著差异,CD36对免疫性PTR发生风险具有极大的预测价值,外周血CD8⁺T细胞比例增高,而CD4⁺T细胞比例减低。结论：免疫性PTR患者产生血小板抗体具体机制不明确,了解患者细胞免疫状态有助于明确免疫性PTR的发生机制,为患者提供更优质的诊疗策略。     

Isolation and Identification of the Precursor of Ethane in Phaseolus vulgaris L

Ethane production by homogenates of Phaseolus vulgaris L. cv. Harvester was studied. The precursor of ethane was identified as linolenic acid. The liberation of ethane was optimum at pH 4.2 and was highest from homogenates of leaves and apical buds. When roots were homogenized in linolenic acid solution, ethane and ethylene production were stimulated. In corn root homogenates, ethylene biosynthesis was stimulated nearly 8-fold by linolenic acid. The enzyme responsible for ethane production from oat root homogenates was soluble and had a high molecular weight.     

Studies on the possible role of protein phosphorylation in the transduction of the ethylene signal

Previous work in our laboratory has demonstrated the existence of high affinity binding sites for the plant growth regulator ethylene. The ethylene binding protein (EBP), from Phaseolus cotyledons, shows many of the characteristics of a functional receptor for ethylene, has been purified on SDS-PAGE and polyclonal antibodies raised in rabbits. Current work involves the investigation of the ethylene transduction signal in a number of ethylene-responsive tissues.In peas, it has been shown that ethylene promotes the phosphorylation of specific proteins of similar molecular weight to the EBP from Phaseolus. Such ethylene-induced phosphorylation can be inhibited by the ethylene antagonist, 2,5-NBD. The antibodies raised to the EBP from Phaseolus have been shown to immunoprecipitate ³²P-labelled proteins from membrane protein preparations obtained from pea tissue. Studies on ethylene binding in pea have also shown that the binding of ethylene may be regulated by phosphorylation. Thus, under conditions which promote phosphorylation, binding is inhibited, whereas the reverse is true under conditions which enhance dephosphorylation.Further work is described which examines the effect of protein kinase, protein phosphatase and calcium channel inhibitors on ethylene-induced phosphorylation in peas, together with wild-type (WT) and ethylene insensitive (eti) mutants of Arabidopsis thaliana. The effects of these treatments can be monitored in vivo using the ethylene-induced triple response as a screen. Furthermore, the protein profiles of such treated seedlings can then be compared by labelling protein extracts with ³²P and subjecting the samples to SDS-PAGE followed by autoradiography.     

Mechanism of local anesthetic-induced disruption of raft-like ordered membrane domains

BackgroundBecause ordered membrane domains, called lipid rafts, regulate activation of ion channels related to the nerve pulse, lipids rafts are thought to be a possible target for anesthetic molecules. To understand the mechanism of anesthetic action, we examined influence of representative local anesthetics (LAs); dibucaine, tetracaine, and lidocaine, on raft-like liquid-ordered (L_o)/non-raft-like liquid-disordered (L_d) phase separation.MethodsImpact of LAs on the phase separation was observed by fluorescent microscopy. LA-induced perturbation of the L_o and L_d membranes was examined by DPH anisotropy measurements. Incorporation of LAs to the membranes was examined by fluorescent anisotropy of LAs. The biding location of the LAs was indicated by small angle x-ray diffraction (SAXD).ResultsFluorescent experiments showed that dibucaine eliminated the phase separation the most effectively, followed by tetracaine and lidocaine. The disruption of the phase separation can be explained by their disordering effects on the L_o membrane. SAXD and other experiments further suggested that dibucaine's most potent perturbation of the L_o membrane is attributable to its deeper immersion and bulky molecular structure. Tetracaine, albeit immersed in the L_o membrane as deeply as dibucaine, less perturbs the L_o membrane probably because of its smaller bulkiness. Lidocaine hardly reaches the hydrophobic region, resulting in the weakest L_o membrane perturbation.ConclusionDibcaine perturbs the L_o membrane the most effectively, followed by tetracaine and lidocaine. This ranking correlates with their anesthetic potency.General significanceThis study suggests a possible mechanistic link between anesthetic action and perturbation of lipid rafts.     

Melatonin Alters Fluid Phase Coexistence in POPC/DPPC/Cholesterol Membranes

The structure and biophysical properties of lipid membranes are important for cellular functions in health and disease. In Alzheimer’s disease, the neuronal membrane is a target for toxic amyloid-β (Aβ). Melatonin is an important pineal gland hormone that has been shown to protect against Aβ toxicity in cellular and animal studies, but the molecular mechanism of this protection is not fully understood. Melatonin is a small membrane-active molecule that has been shown to interact with model lipid membranes and alter the membrane biophysical properties, such as membrane molecular order and dynamics. This effect of melatonin has been previously studied in simple model bilayers with one or two lipid components. To make it more relevant to neuronal membranes, we used a more complex ternary lipid mixture as our membrane model. In this study, we used ²H-NMR to investigate the effect of melatonin on the phase behavior of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and cholesterol lipid membranes. We used deuterium-labeled POPC-d₃₁ and DPPC-d₆₂,separately to probe the changes in hydrocarbon chain order as a function of temperature and melatonin concentration. We find that POPC/DPPC/cholesterol at molar proportions of 3:3:2 is close to liquid-disordered/liquid-ordered phase separation and that melatonin can induce phase separation in these ternary mixtures by preferentially incorporating into the disordered phase and increasing its level of disorder. At 5 mol% melatonin, we observed phase separation in samples with POPC-d₃₁, but not with DPPC-d₆₂, whereas at 10 mol% melatonin, phase separation was observed in both samples with either POPC-d₃₁ or DPPC-d₆₂. These results indicate that melatonin can have a strong effect on membrane structure and physical properties, which may provide some clues to understanding how melatonin protects against Aβ, and that choice of chain perdeuteration is an important consideration from a technical point of view.     

The effect of solubilisation on the character of an ethylene-binding site from Phaseolus vulgaris L. cotyledons

The ethylene-binding site (EBS) from Phaseolus vulgaris cv. Canadian Wonder cotyledons can be solubilised from 96,000 g pelleted material by Triton X-100 or sodium cholate. Extraction of 96,000 g pellets with acetone, butanol or butanol and ether results in a total loss of ethylene-binding activity. Like the membrane-bound form, the solubilised EBS has an apparent K_D(liquid) of 10^-10 M at a concentration of 32 pmol EBS per gram tissue fresh weight. Propylene and acetylene act as competitive inhibitors, carbon dioxide appears to promote ethylene binding and ethane has no significant effect. The solubilised EBS is completely denatured affect. The solubilised EBS is completely denatured after 10 min at 70°C, by 1 mM mercaptoethanol and 0.1 mM dithiothreitol, but not by trypsin or chymotrypsin. However, solubilisation decreases the rate constant of association from 10³ M^-1 s^-1 to 10¹–10² M^-1 s^-1 and hence does not permit experimental determination of the rate constant of dissociation. The pH optimum for ethylene binding is altered from the range pH 7–10 in the membrane-bound form to the pH range 4–7 in the solubilised form. The EBS appears to be a hydrophobic, intergral membrane protein, which requires a hydrophobic environment to retain its activity. Partitioning of the EBS into polymer phases is determined by the detergent used for solubilisation indicating that when solubilised, the EBS forms a complex with detergent molecules.Abbreviations EBS ethylene-binding site - PEG polyethylene glycol     

Mechanisms of pressure-induced water infiltration process through graphene nanopores

Understanding the mechanism of water infiltration through nanopores is essential for wide applications ranging from membrane separation to gene therapy. In this paper, the molecular dynamics simulation method is used to investigate the pressure-assisted water transport process through graphene nanopores. Various factors including the hydrophobicity of nanopore surface, nanopore dimension, temperature as well as external electric field that affect water in permeation into graphene nanopores are discussed. It is found that classic Laplace-Young equation fails and the relationship between pressure and diameter (D) does not follow the 1/D dependence as the characteristic dimension of a nanopore is sufficiently small (smaller than 1?nm). The critical pressure significantly depends on both the pore length and electric field as D is smaller than 5?nm. Besides, enhancing temperature and electric field intensity are obviously beneficial for water infiltration through those nanopores with a diameter smaller than 5?nm.     

Ethylene and Ethane Production from Sulfur Dioxide-injured Plants

After alfalfa (Medicago sativa) seedlings were exposed to approximately 0.7 microliter per liter SO₂ for 8 hours, elevated ethylene and ethane production was observed. Ethylene production peaked about 6 hours and returned to control levels by about 24 hours following the fumigation, while ethane production peaked about 36 hours and was still above control levels 48 hours after the fumigation. Light had an opposite effect upon the production of the two gases: ethane production rates were higher from plants held in light, whereas ethylene production rates were higher from those held in the dark. Peak ethylene and ethane production rates from SO₂-treated plants were about 10 and 4 to 5 times greater, respectively, than those of the control plants. Ethylene appeared to be formed primarily from stressed yet viable leaves and ethane from visibly damaged leaves. The different time courses and light requirements for ethylene and ethane production suggest that these two gases were formed via different mechanisms. Light appears to have a dual role. It enhances SO₂-induced cellular damage and plays a role for repairs.     

How the imidazole ring modulates amyloid formation of islet amyloid polypeptide: A chemical modification study

BackgroundThe misfolding of human islet amyloid polypeptide (hIAPP) is an important pathological factor on the onset of type 2 diabetes. A number of studies have been focused on His¹⁸, the only histidine of hIAPP, whose imidazole ring and the protonation state might impact hIAPP amyloid formation, but the exact mechanism remains unclear.MethodsWe used diethylpyrocarbonate (DEPC) to specifically modify His¹⁸ and obtained mono-ethyloxyformylated hIAPP (DMI). Thioflavin T based fluorescence, transmission electronic microscopy, circular dichroism spectroscopy, fluorescence dye leakage, Fourier transform infrared spectroscopy and replica-exchange molecular dynamics (REMD) simulation were applied to study the impact of DEPC-modification on hIAPP amyloid formation.ResultsAfter an ethyl-acetate group was introduced to the His¹⁸ of hIAPP by diethylpyrocarbonate (DEPC) modification, the pH dependent hIAPP fibrillation went to the opposite order and the number of intra-molecular hydrogen bonds decreased, while the possibility of His¹⁸ participating in the formation of α-helical structures increased. Furthermore, the membrane–peptide interaction and ion–peptide interaction were both impaired.ConclusionsThe intramolecular hydrogen bond formation by His¹⁸ and the possibility of His¹⁸ participating in the formation of α-helical structures greatly modulated the manner of hIAPP amyloid formation. The imidazole ring directly participates in the hIAPP–membrane/ion interaction.General significanceDEPC modification is an alternative approach to investigate the role of the imidazole ring during amyloid formation.     

The Groovy TMEM16 Family: Molecular Mechanisms of Lipid Scrambling and Ion Conduction

The TMEM16 family of membrane proteins displays a remarkable functional dichotomy – while some family members function as Ca²⁺-activated anion channels, the majority of characterized TMEM16 homologs are Ca²⁺-activated lipid scramblases, which catalyze the exchange of phospholipids between the two membrane leaflets. Furthermore, some TMEM16 scramblases can also function as channels. Due to their involvement in important physiological processes, the family has been actively studied ever since their molecular identity was unraveled. In this review, we will summarize the recent advances in the field and how they influenced our view of TMEM16 family function and evolution. Structural, functional and computational studies reveal how relatively small rearrangements in the permeation pathway are responsible for the observed functional duality: while TMEM16 scramblases can adopt both ion- and lipid conductive conformations, TMEM16 channels can only populate the former. Recent data further provides the molecular details of a stepwise activation mechanism, which is initiated by Ca²⁺ binding and modulated by various cellular factors, including lipids. TMEM16 function and the surrounding membrane properties are inextricably intertwined, with the protein inducing bilayer deformations associated with scrambling, while the surrounding lipids modulate TMEM16 conformation and activity.     

A simple method for the determination of resistance to gas diffusion in plant organs

A simple method was developed for the determination of resistance coefficients for ethylene diffusion in plant tissues based on the kinetic analysis of the efflux of preloaded ethane gas. Efflux curves were analyzed to obtain first-order rate constants and resistance coefficients. Resistance coefficients determined by the ethane efflux and steady-state methods were found to agree well. Employing the ethane efflux method, it was shown that over 97% of gas exchange of tomato (Lycopersicum esculentum Mill., cv. `Ace') fruits occurs through the stem scar. The resistances to diffusion of tomato skin and stem scar were found to be 280,000 and 300 seconds per centimeter, respectively; the combined resistance of intact tomato fruits was approximately 7,800 seconds per centimeter. The ethane efflux method was employed to show that plastic shrink-wrapping of English cucumbers (Cucumis sativus L. var anglicus Bailey) increased the resistance to ethane diffusion from 1.1 × 10³ to 23 × 10³ seconds per centimeter.