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Sample records for activate membrane fusion

  1. Membrane fusion

    Bendix, Pól Martin

    2015-01-01

    At Stanford University, Boxer lab, I worked on membrane fusion of small unilamellar lipid vesicles to flat membranes tethered to glass surfaces. This geometry closely resembles biological systems in which liposomes fuse to plasma membranes. The fusion mechanism was studied using DNA zippering...... between complementary strands linked to the two apposing membranes closely mimicking the zippering mechanism of SNARE fusion complexes....

  2. Sendai Virus Fusion Activity as Modulated by Target Membrane Components

    Nunes-Correia, Isabel; Ramalho-Santos, João; Maria C Pedroso de Lima

    1998-01-01

    We have studied the differences between erythrocytes and erythrocyte ghosts as target membranes for the study of Sendai virus fusion activity. Fusion was monitored continuously by fluorescence dequenching of R18-labeled virus. Experiments were carried out either with or without virus/target membrane prebinding. When Sendai virus was added directly to a erythrocyte/erythrocyte ghost suspension, fusion was always lower than that obtained when experiments were carried out with virus already boun...

  3. Membrane-mobility agent-promoted fusion of erythrocytes: fusibility is correlated with attack by calcium-activated cytoplasmic proteases on membrane proteins.

    Kosower, N S; Glaser, T; Kosower, E M

    1983-01-01

    Rat, but not human, erythrocytes undergo fusion promoted by the membrane-mobility agent 2-(2-methoxyethoxy)-ethyl cis-8-(2-octylcyclopropyl)octanoate (A2C). The difference in behavior is correlated with rat erythrocyte membrane protein degradation caused by Ca2+-activated proteases. The human erythrocyte is deficient in such protease activity. Membrane protein degradation is a necessary, but not sufficient, requirement for membrane fusion. Membrane protein degradation probably releases membra...

  4. Fusion-Triggered Switching of Enzymatic Activity on an Artificial Cell Membrane

    Jun-ichi Kikuchi

    2012-05-01

    Full Text Available A nanosensory membrane device was constructed for detecting liposome fusion through changes in an enzymatic activity. Inspired by a biological signal transduction system, the device design involved functionalized liposomal membranes prepared by self-assembly of the following molecular components: a synthetic peptide lipid and a phospholipid as matrix membrane components, a Schiff’s base of pyridoxal 5’-phosphate with phosphatidylethanolamine as a thermo-responsive artificial receptor, NADH-dependent L-lactate dehydrogenase as a signal amplifier, and Cu2+ ion as a signal mediator between the receptor and enzyme. The enzymatic activity of the membrane device was adjustable by changing the matrix lipid composition, reflecting the thermotropic phase transition behavior of the lipid membranes, which in turn controlled receptor binding affinity toward the enzyme-inhibiting mediator species. When an effective fusogen anionic polymer was added to these cationic liposomes, membrane fusion occurred, and the functionalized liposomal membranes responded with changes in enzymatic activity, thus serving as an effective nanosensory device for liposome fusion detection.

  5. Viral membrane fusion

    Harrison, Stephen C., E-mail: harrison@crystal.harvard.edu

    2015-05-15

    Membrane fusion is an essential step when enveloped viruses enter cells. Lipid bilayer fusion requires catalysis to overcome a high kinetic barrier; viral fusion proteins are the agents that fulfill this catalytic function. Despite a variety of molecular architectures, these proteins facilitate fusion by essentially the same generic mechanism. Stimulated by a signal associated with arrival at the cell to be infected (e.g., receptor or co-receptor binding, proton binding in an endosome), they undergo a series of conformational changes. A hydrophobic segment (a “fusion loop” or “fusion peptide”) engages the target-cell membrane and collapse of the bridging intermediate thus formed draws the two membranes (virus and cell) together. We know of three structural classes for viral fusion proteins. Structures for both pre- and postfusion conformations of illustrate the beginning and end points of a process that can be probed by single-virion measurements of fusion kinetics. - Highlights: • Viral fusion proteins overcome the high energy barrier to lipid bilayer merger. • Different molecular structures but the same catalytic mechanism. • Review describes properties of three known fusion-protein structural classes. • Single-virion fusion experiments elucidate mechanism.

  6. Viral membrane fusion

    Membrane fusion is an essential step when enveloped viruses enter cells. Lipid bilayer fusion requires catalysis to overcome a high kinetic barrier; viral fusion proteins are the agents that fulfill this catalytic function. Despite a variety of molecular architectures, these proteins facilitate fusion by essentially the same generic mechanism. Stimulated by a signal associated with arrival at the cell to be infected (e.g., receptor or co-receptor binding, proton binding in an endosome), they undergo a series of conformational changes. A hydrophobic segment (a “fusion loop” or “fusion peptide”) engages the target-cell membrane and collapse of the bridging intermediate thus formed draws the two membranes (virus and cell) together. We know of three structural classes for viral fusion proteins. Structures for both pre- and postfusion conformations of illustrate the beginning and end points of a process that can be probed by single-virion measurements of fusion kinetics. - Highlights: • Viral fusion proteins overcome the high energy barrier to lipid bilayer merger. • Different molecular structures but the same catalytic mechanism. • Review describes properties of three known fusion-protein structural classes. • Single-virion fusion experiments elucidate mechanism

  7. Fusion of biological membranes

    K Katsov; M Müller; M Schick

    2005-06-01

    The process of membrane fusion has been examined by Monte Carlo simulation, and is found to be very different than the conventional picture. The differences in mechanism lead to several predictions, in particular that fusion is accompanied by transient leakage. This prediction has recently been verified. Self-consistent field theory is applied to examine the free energy barriers in the different scenarios.

  8. Poxvirus entry and membrane fusion

    The study of poxvirus entry and membrane fusion has been invigorated by new biochemical and microscopic findings that lead to the following conclusions: (1) the surface of the mature virion (MV), whether isolated from an infected cell or by disruption of the membrane wrapper of an extracellular virion, is comprised of a single lipid membrane embedded with non-glycosylated viral proteins; (2) the MV membrane fuses with the cell membrane, allowing the core to enter the cytoplasm and initiate gene expression; (3) fusion is mediated by a newly recognized group of viral protein components of the MV membrane, which are conserved in all members of the poxvirus family; (4) the latter MV entry/fusion proteins are required for cell to cell spread necessitating the disruption of the membrane wrapper of extracellular virions prior to fusion; and furthermore (5) the same group of MV entry/fusion proteins are required for virus-induced cell-cell fusion. Future research priorities include delineation of the roles of individual entry/fusion proteins and identification of cell receptors

  9. Membrane fusion during phage lysis.

    Rajaure, Manoj; Berry, Joel; Kongari, Rohit; Cahill, Jesse; Young, Ry

    2015-04-28

    In general, phages cause lysis of the bacterial host to effect release of the progeny virions. Until recently, it was thought that degradation of the peptidoglycan (PG) was necessary and sufficient for osmotic bursting of the cell. Recently, we have shown that in Gram-negative hosts, phage lysis also requires the disruption of the outer membrane (OM). This is accomplished by spanins, which are phage-encoded proteins that connect the cytoplasmic membrane (inner membrane, IM) and the OM. The mechanism by which the spanins destroy the OM is unknown. Here we show that the spanins of the paradigm coliphage lambda mediate efficient membrane fusion. This supports the notion that the last step of lysis is the fusion of the IM and OM. Moreover, data are provided indicating that spanin-mediated fusion is regulated by the meshwork of the PG, thus coupling fusion to murein degradation by the phage endolysin. Because endolysin function requires the formation of μm-scale holes by the phage holin, the lysis pathway is seen to require dramatic dynamics on the part of the OM and IM, as well as destruction of the PG. PMID:25870259

  10. Physical Aspects of Viral Membrane Fusion

    Laura Wessels; Keith Weninger

    2009-01-01

    Enveloped viruses commonly employ membrane fusion during cell penetration in order to deliver their genetic material across the cell boundary. Large conformational changes in the proteins embedded in the viral membrane play a fundamental role in the membrane fusion process. Despite the tremendously wide variety of viruses that contain membranes, it appears that they all contain membrane fusion protein machinery with a remarkably conserved mechanism of action. Much of our current biochemical u...

  11. Membrane lysis during biological membrane fusion: collateral damage by misregulated fusion machines

    Engel, Alex; Walter, Peter

    2008-01-01

    In the canonical model of membrane fusion, the integrity of the fusing membranes is never compromised, preserving the identity of fusing compartments. However, recent molecular simulations provided evidence for a pathway to fusion in which holes in the membrane evolve into a fusion pore. Additionally, two biological membrane fusion models—yeast cell mating and in vitro vacuole fusion—have shown that modifying the composition or altering the relative expression levels of membrane fusion comple...

  12. Rabies Virus-Induced Membrane Fusion Pathway

    Gaudin, Yves

    2000-01-01

    Fusion of rabies virus with membranes is triggered at low pH and is mediated by the viral glycoprotein (G). The rabies virus-induced fusion pathway was studied by investigating the effects of exogenous lipids having various dynamic molecular shapes on the fusion process. Inverted cone-shaped lysophosphatidylcholines (LPCs) blocked fusion at a stage subsequent to fusion peptide insertion into the target membrane. Consistent with the stalk-hypothesis, LPC with shorter alkyl chains inhibited fus...

  13. The role of membrane fusion activity of a whole inactivated influenza virus vaccine in (re)activation of influenza-specific cytotoxic T lymphocytes.

    Budimir, Natalija; Meijerhof, Tjarko; Wilschut, Jan; Huckriede, Anke; de Haan, Aalzen

    2010-12-01

    Induction of cytotoxic T lymphocyte (CTL) activity against conserved influenza antigens, e.g. nucleoprotein (NP) could be a step towards cross-protective influenza vaccine. The major challenge for non-replicating influenza vaccines aiming for activation of CTLs is targeting of antigen to the MHC class I processing and presentation pathway of professional antigen presenting cells, in particular dendritic cells (DCs). Intrinsic fusogenic properties of the vaccine particle itself can enable direct cytosolic delivery of the antigen by enhancing release of the antigen from the endosome to the cytosol. Alternatively, the vaccine particle would need to possess the capacity to activate DCs thereby triggering cell-intrinsic mechanisms of cross-presentation, processes that do not require fusion. Here, using fusion-active and fusion-inactive whole inactivated virus (WIV) as a vaccine model, we studied the relative contribution of these two pathways on priming and reactivation of influenza NP-specific CTLs in a murine model. We show that activation of bone marrow-derived DCs by WIV, as well as reactivation of NP-specific CTLs in vitro and in vivo were not affected by inactivation of membrane fusion of the WIV particles. However, in vivo priming of naive CTLs was optimal only upon vaccination with fusion-active WIV. Thus, DC-intrinsic mechanisms of cross-presentation are involved in the activation of CTLs upon vaccination with WIV. However, for optimal priming of naive CTLs these mechanisms should be complemented by delivery of antigen to the cytosol mediated by the membrane fusion capacity of the WIV particles. PMID:20965298

  14. Sphingolipids activate membrane fusion of Semliki Forest virus in a stereospecific manner

    Moesby, Lise; Corver, J; Erukulla, R K; Bittman, R; Wilschut, J

    1995-01-01

    assessed by flotation on sucrose density gradients, was not dependent on the presence of fusion-competent or fusion-incompetent sphingolipids in the liposomes. The results of this study support the notion that a stereospecific interaction of the viral fusion protein with D-erythro sphingolipids in the...

  15. Fusion of the endoplasmic reticulum and mitochondrial outer membrane in rats brown adipose tissue: activation of thermogenesis by Ca2+.

    de Meis, Leopoldo; Ketzer, Luisa A; da Costa, Rodrigo Madeiro; de Andrade, Ivone Rosa; Benchimol, Marlene

    2010-01-01

    Brown adipose tissue (BAT) mitochondria thermogenesis is regulated by uncoupling protein 1 (UCP 1), GDP and fatty acids. In this report, we observed fusion of the endoplasmic reticulum (ER) membrane with the mitochondrial outer membrane of rats BAT. Ca(2+)-ATPase (SERCA 1) was identified by immunoelectron microscopy in both ER and mitochondria. This finding led us to test the Ca(2+) effect in BAT mitochondria thermogenesis. We found that Ca(2+) increased the rate of respiration and heat production measured with a microcalorimeter both in coupled and uncoupled mitochondria, but had no effect on the rate of ATP synthesis. The Ca(2+) concentration needed for half-maximal activation varied between 0.08 and 0.11 microM. The activation of respiration was less pronounced than that of heat production. Heat production and ATP synthesis were inhibited by rotenone and KCN. Liver mitochondria have no UCP1 and during respiration synthesize a large amount of ATP, produce little heat, GDP had no effect on mitochondria coupling, Ca(2+) strongly inhibited ATP synthesis and had little or no effect on the small amount of heat released. These finding indicate that Ca(2+) activation of thermogenesis may be a specific feature of BAT mitochondria not found in other mitochondria such as liver. PMID:20209153

  16. Investigation of SNARE-Mediated Membrane Fusion Mechanism Using Atomic Force Microscopy

    Abdulreda, Midhat H.; Moy, Vincent T.

    2009-01-01

    Membrane fusion is driven by specialized proteins that reduce the free energy penalty for the fusion process. In neurons and secretory cells, soluble N-ethylmaleimide-sensitive factor-attachment protein (SNAP) receptors (SNAREs) mediate vesicle fusion with the plasma membrane during vesicular content release. Although, SNAREs have been widely accepted as the minimal machinery for membrane fusion, the specific mechanism for SNARE-mediated membrane fusion remains an active area of research. Her...

  17. Fusion of the Endoplasmic Reticulum and Mitochondrial Outer Membrane in Rats Brown Adipose Tissue: Activation of Thermogenesis by Ca2+

    de Meis, Leopoldo; Ketzer, Luisa A.; da Costa, Rodrigo Madeiro; de Andrade, Ivone Rosa; Benchimol, Marlene

    2010-01-01

    Brown adipose tissue (BAT) mitochondria thermogenesis is regulated by uncoupling protein 1 (UCP 1), GDP and fatty acids. In this report, we observed fusion of the endoplasmic reticulum (ER) membrane with the mitochondrial outer membrane of rats BAT. Ca2+-ATPase (SERCA 1) was identified by immunoelectron microscopy in both ER and mitochondria. This finding led us to test the Ca2+ effect in BAT mitochondria thermogenesis. We found that Ca2+ increased the rate of respiration and heat production ...

  18. Minimum Membrane Bending Energies of Fusion Pores

    Jackson, Meyer B.

    2009-01-01

    Membranes fuse by forming highly curved intermediates, culminating in structures described as fusion pores. These hourglass-like figures that join two fusing membranes have high bending energies, which can be estimated using continuum elasticity models. Fusion pore bending energies depend strongly on shape, and the present study developed a method for determining the shape that minimizes bending energy. This was first applied to a fusion pore modeled as a single surface and then extended to a...

  19. The role of cholesterol in membrane fusion.

    Yang, Sung-Tae; Kreutzberger, Alex J B; Lee, Jinwoo; Kiessling, Volker; Tamm, Lukas K

    2016-09-01

    Cholesterol modulates the bilayer structure of biological membranes in multiple ways. It changes the fluidity, thickness, compressibility, water penetration and intrinsic curvature of lipid bilayers. In multi-component lipid mixtures, cholesterol induces phase separations, partitions selectively between different coexisting lipid phases, and causes integral membrane proteins to respond by changing conformation or redistribution in the membrane. But, which of these often overlapping properties are important for membrane fusion?-Here we review a range of recent experiments that elucidate the multiple roles that cholesterol plays in SNARE-mediated and viral envelope glycoprotein-mediated membrane fusion. PMID:27179407

  20. A high throughput Cre-lox activated viral membrane fusion assay identifies pharmacological inhibitors of HIV entry.

    Esposito, Anthony M; Cheung, Pamela; Swartz, Talia H; Li, Hongru; Tsibane, Tshidi; Durham, Natasha D; Basler, Christopher F; Felsenfeld, Dan P; Chen, Benjamin K

    2016-03-01

    Enveloped virus entry occurs when viral and cellular membranes fuse releasing particle contents into the target cell. Human immunodeficiency virus (HIV) entry occurs by cell-free virus or virus transferred between infected and uninfected cells through structures called virological synapses. We developed a high-throughput cell-based assay to identify small molecule inhibitors of cell-free or virological synapse-mediated entry. An HIV clone carrying Cre recombinase as a Gag-internal gene fusion releases active Cre into cells upon viral entry activating a recombinatorial gene switch changing dsRed to GFP-expression. A screen of a 1998 known-biological profile small molecule library identified pharmacological HIV entry inhibitors that block both cell-free and cell-to-cell infection. Many top hits were noted as HIV inhibitors in prior studies, but not previously recognized as entry antagonists. Modest therapeutic indices for simvastatin and nigericin were observed in confirmatory HIV infection assays. This robust assay is adaptable to study HIV and heterologous viral pseudotypes. PMID:26803470

  1. Mechanical tension drives cell membrane fusion

    Kim, Ji Hoon; Ren, Yixin; Ng, Win Pin; Li, Shuo; Son, Sungmin; Kee, Yee-Seir; Zhang, Shiliang; Zhang, Guofeng; Fletcher, Daniel A.; Robinson, Douglas N.; Chen, Elizabeth H.

    2015-01-01

    Membrane fusion is an energy-consuming process that requires tight juxtaposition of two lipid bilayers. Little is known about how cells overcome energy barriers to bring their membranes together for fusion. Previously, we have shown that cell-cell fusion is an asymmetric process in which an “attacking” cell drills finger-like protrusions into the “receiving” cell to promote cell fusion. Here we show that the receiving cell mounts a Myosin II (MyoII)-mediated mechanosensory response to its inv...

  2. Separate fusion of outer and inner mitochondrial membranes

    Malka, Florence; Guillery, Olwenn; Cifuentes-Diaz, Carmen; Guillou, Emmanuelle; Belenguer, Pascale; Lombès, Anne; Rojo, Manuel

    2005-01-01

    Mitochondria are enveloped by two closely apposed boundary membranes with different properties and functions. It is known that they undergo fusion and fission, but it has remained unclear whether outer and inner membranes fuse simultaneously, coordinately or separately. We set up assays for the study of inner and outer membrane fusion in living human cells. Inner membrane fusion was more sensitive than outer membrane fusion to inhibition of glycolysis. Fusion of the inner membrane, but not of...

  3. SARS-coronavirus spike S2 domain flanked by cysteine residues C822 and C833 is important for activation of membrane fusion

    The S2 domain of the coronavirus spike (S) protein is known to be responsible for mediating membrane fusion. In addition to a well-recognized cleavage site at the S1-S2 boundary, a second proteolytic cleavage site has been identified in the severe acute respiratory syndrome coronavirus (SARS-CoV) S2 domain (R797). C-terminal to this S2 cleavage site is a conserved region flanked by cysteine residues C822 and C833. Here, we investigated the importance of this well conserved region for SARS-CoV S-mediated fusion activation. We show that the residues between C822-C833 are well conserved across all coronaviruses. Mutagenic analysis of SARS-CoV S, combined with cell-cell fusion and pseudotyped virion infectivity assays, showed a critical role for the core-conserved residues C822, D830, L831, and C833. Based on available predictive models, we propose that the conserved domain flanked by cysteines 822 and 833 forms a loop structure that interacts with components of the SARS-CoV S trimer to control the activation of membrane fusion.

  4. Proteolytic cleavage of Opa1 stimulates mitochondrial inner membrane fusion and couples fusion to oxidative phosphorylation

    Mishra, Prashant; Carelli, Valerio; Manfredi, Giovanni; Chan, David C.

    2014-01-01

    Mitochondrial fusion is essential for maintenance of mitochondrial function. The mitofusin GTPases control mitochondrial outer membrane fusion, whereas the dynamin-related GTPase Opa1 mediates inner membrane fusion. We show that mitochondrial inner membrane fusion is tuned by the level of oxidative phosphorylation (OXPHOS), whereas outer membrane fusion is insensitive. Consequently, cells from patients with pathogenic mtDNA mutations show a selective defect in mitochondrial inner membrane fus...

  5. Palaeontological evidence of membrane relationship in step-by-step membrane fusion

    Wang, Xin; Liu, Wenzhe; DU, KAIHE

    2010-01-01

    Studies on membrane fusion in living cells indicate that initiation of membrane fusion is a transient and hard to capture process. Despite previous research, membrane behaviour at this point is still poorly understood. Recent palaeobotanical research has revealed snapshots of membrane fusion in a 15-million-year-old fossil pinaceous cone. To reveal the membrane behaviour during the fusion, we conducted more observations on the same fossil material. Several discernible steps of membrane fusion...

  6. Defensins promote fusion and lysis of negatively charged membranes.

    Fujii, G; Selsted, M E; Eisenberg, D.

    1993-01-01

    Defensins, a family of cationic peptides isolated from mammalian granulocytes and believed to permeabilize membranes, were tested for their ability to cause fusion and lysis of liposomes. Unlike alpha-helical peptides whose lytic effects have been extensively studied, the defensins consist primarily of beta-sheet. Defensins fuse and lyse negatively charged liposomes but display reduced activity with neutral liposomes. These and other experiments suggest that fusion and lysis is mediated prima...

  7. Pseudorabies Virus Glycoprotein M Inhibits Membrane Fusion

    Klupp, Barbara G.; Nixdorf, Ralf; Mettenleiter, Thomas C.

    2000-01-01

    A transient transfection-fusion assay was established to investigate membrane fusion mediated by pseudorabies virus (PrV) glycoproteins. Plasmids expressing PrV glycoproteins under control of the immediate-early 1 promoter-enhancer of human cytomegalovirus were transfected into rabbit kidney cells, and the extent of cell fusion was quantitated 27 to 42 h after transfection. Cotransfection of plasmids encoding PrV glycoproteins B (gB), gD, gH, and gL resulted in formation of polykaryocytes, as...

  8. Induction of heterosubtypic cross-protection against influenza by a whole inactivated virus vaccine: the role of viral membrane fusion activity.

    Natalija Budimir

    Full Text Available BACKGROUND: The inability of seasonal influenza vaccines to effectively protect against infection with antigenically drifted viruses or newly emerging pandemic viruses underlines the need for development of cross-reactive influenza vaccines that induce immunity against a variety of virus subtypes. Therefore, potential cross-protective vaccines, e.g., whole inactivated virus (WIV vaccine, that can target conserved internal antigens such as the nucleoprotein (NP and/or matrix protein (M1 need to be explored. METHODOLOGY/PRINCIPAL FINDINGS: In the current study we show that a WIV vaccine, through induction of cross-protective cytotoxic T lymphocytes (CTLs, protects mice from heterosubtypic infection. This protection was abrogated after depletion of CD8+ cells in vaccinated mice, indicating that CTLs were the primary mediators of protection. Previously, we have shown that different procedures used for virus inactivation influence optimal activation of CTLs by WIV, most likely by affecting the membrane fusion properties of the virus. Specifically, inactivation with formalin (FA severely compromises fusion activity of the virus, while inactivation with β-propiolactone (BPL preserves fusion activity. Here, we demonstrate that vaccination of mice with BPL-inactivated H5N1 WIV vaccine induces solid protection from lethal heterosubtypic H1N1 challenge. By contrast, vaccination with FA-inactivated WIV, while preventing death after lethal challenge, failed to protect against development of disease and severe body weight loss. Vaccination with BPL-inactivated WIV, compared to FA-inactivated WIV, induced higher levels of specific CD8+ T cells in blood, spleen and lungs, and a higher production of granzyme B in the lungs upon H1N1 virus challenge. CONCLUSION/SIGNIFICANCE: The results underline the potential use of WIV as a cross-protective influenza vaccine candidate. However, careful choice of the virus inactivation procedure is important to retain membrane

  9. The Flocculating Cationic Polypetide from Moringa oleifera Seeds Damages Bacterial Cell Membranes by Causing Membrane Fusion.

    Shebek, Kevin; Schantz, Allen B; Sines, Ian; Lauser, Kathleen; Velegol, Stephanie; Kumar, Manish

    2015-04-21

    A cationic protein isolated from the seeds of the Moringa oleifera tree has been extensively studied for use in water treatment in developing countries and has been proposed for use in antimicrobial and therapeutic applications. However, the molecular basis for the antimicrobial action of this peptide, Moringa oleifera cationic protein (MOCP), has not been previously elucidated. We demonstrate here that a dominant mechanism of MOCP antimicrobial activity is membrane fusion. We used a combination of cryogenic electron microscopy (cryo-EM) and fluorescence assays to observe and study the kinetics of fusion of membranes in liposomes representing model microbial cells. We also conducted cryo-EM experiments on E. coli cells where MOCP was seen to fuse the inner and outer membranes. Coarse-grained molecular dynamics simulations of membrane vesicles with MOCP molecules were used to elucidate steps in peptide adsorption, stalk formation, and fusion between membranes. PMID:25845029

  10. Dissipative Particle Dynamics of Tension-induced Membrane Fusion

    Grafmueller, Andrea; Lipowsky, Reinhard; Shillcock, Julian C.

    2009-01-01

    Abstract Recent studies of tension-induced membrane fusion using dissipative particle dynamics (DPD) simulations are briefly reviewed. The stochastic nature of the fusion process makes it necessary to simulate a large number of fusion attempts in order to obtain reliable fusion statistics and to extract meaningful values for the fusion probability and the average fusion times. All successful fusion events follow the same pathway. In this fusion pathway, configurations of individual...

  11. IM30 triggers membrane fusion in cyanobacteria and chloroplasts.

    Hennig, Raoul; Heidrich, Jennifer; Saur, Michael; Schmüser, Lars; Roeters, Steven J; Hellmann, Nadja; Woutersen, Sander; Bonn, Mischa; Weidner, Tobias; Markl, Jürgen; Schneider, Dirk

    2015-01-01

    The thylakoid membrane of chloroplasts and cyanobacteria is a unique internal membrane system harbouring the complexes of the photosynthetic electron transfer chain. Despite their apparent importance, little is known about the biogenesis and maintenance of thylakoid membranes. Although membrane fusion events are essential for the formation of thylakoid membranes, proteins involved in membrane fusion have yet to be identified in photosynthetic cells or organelles. Here we show that IM30, a conserved chloroplast and cyanobacterial protein of approximately 30 kDa binds as an oligomeric ring in a well-defined geometry specifically to membranes containing anionic lipids. Triggered by Mg(2+), membrane binding causes destabilization and eventually results in membrane fusion. We propose that IM30 establishes contacts between internal membrane sites and promotes fusion to enable regulated exchange of proteins and/or lipids in cyanobacteria and chloroplasts. PMID:25952141

  12. Dissipative Particle Dynamics of tension-induced membrane fusion

    Shillcock, Julian C.

    2009-01-01

    Recent studies of tension-induced membrane fusion using dissipative particle dynamics (DPD) simulations are briefly reviewed. The stochastic nature of the fusion process makes it necessary to simulate a large number of fusion attempts in order to obtain reliable fusion statistics and to extract...

  13. Expansion of the fusion stalk and its implication for biological membrane fusion

    Risselada, H.; Bubnis, G.; Grubmüller, H.

    2014-01-01

    Over the past 20 years, it has been widely accepted that membrane fusion proceeds via a hemifusion step before opening of the productive fusion pore. An initial hourglass-shaped lipid structure, the fusion stalk, is formed between the adjacent membrane leaflets (cis leaflets). It remains controversial if and how fusion proteins drive the subsequent transition (expansion) of the stalk into a fusion pore. Here, we propose a comprehensive and consistent thermodynamic understanding in terms of th...

  14. Stalk model of membrane fusion: solution of energy crisis.

    Kozlovsky, Yonathan; Kozlov, Michael M.

    2002-01-01

    Membrane fusion proceeds via formation of intermediate nonbilayer structures. The stalk model of fusion intermediate is commonly recognized to account for the major phenomenology of the fusion process. However, in its current form, the stalk model poses a challenge. On one hand, it is able to describe qualitatively the modulation of the fusion reaction by the lipid composition of the membranes. On the other, it predicts very large values of the stalk energy, so that the related energy barrier...

  15. Oligomerization of Fusogenic Peptides Promotes Membrane Fusion by Enhancing Membrane Destabilization

    Lau, Wai Leung; Ege, David S.; Lear, James D.; Hammer, Daniel A.; DeGrado, William F.

    2004-01-01

    A key element of membrane fusion reactions in biology is the involvement of specific fusion proteins. In many viruses, the proteins that mediate membrane fusion usually exist as homotrimers. Furthermore, they contain extended triple-helical coiled-coil domains and fusogenic peptides. It has been suggested that the coiled-coil domains present the fusogenic peptide in a conformation or geometry favorable for membrane fusion. To test the hypothesis that trimerization of fusogenic peptide is rela...

  16. Membrane fusion machines of paramyxoviruses: capture of intermediates of fusion

    Charles J Russell; Theodore S Jardetzky; Lamb, Robert A.

    2001-01-01

    Peptides derived from heptad repeat regions adjacent to the fusion peptide and transmembrane domains of many viral fusion proteins form stable helical bundles and inhibit fusion specifically. Paramyxovirus SV5 fusion (F) protein-mediated fusion and its inhibition by the peptides N-1 and C-1 were analyzed. The temperature dependence of fusion by F suggests that thermal energy, destabilizing proline residues and receptor binding by the hemagglutinin–neuraminidase (HN) protein collectively contr...

  17. Fusion Pore Diameter Regulation by Cations Modulating Local Membrane Anisotropy

    Doron Kabaso

    2012-01-01

    Full Text Available The fusion pore is an aqueous channel that is formed upon the fusion of the vesicle membrane with the plasma membrane. Once the pore is open, it may close again (transient fusion or widen completely (full fusion to permit vesicle cargo discharge. While repetitive transient fusion pore openings of the vesicle with the plasma membrane have been observed in the absence of stimulation, their frequency can be further increased using a cAMP-increasing agent that drives the opening of nonspecific cation channels. Our model hypothesis is that the openings and closings of the fusion pore are driven by changes in the local concentration of cations in the connected vesicle. The proposed mechanism of fusion pore dynamics is considered as follows: when the fusion pore is closed or is extremely narrow, the accumulation of cations in the vesicle (increased cation concentration likely leads to lipid demixing at the fusion pore. This process may affect local membrane anisotropy, which reduces the spontaneous curvature and thus leads to the opening of the fusion pore. Based on the theory of membrane elasticity, we used a continuum model to explain the rhythmic opening and closing of the fusion pore.

  18. Observations of membrane fusion in a liposome dispersion: the missing fusion intermediate?

    Marianna Foldvari

    2015-01-01

    Early intermediate structures of liposome-liposome fusion events were captured by freeze-fracture electron microscopic (EM) technique. The images show the morphology of the fusion interface at several different stages of the fusion event. One of the intermediates was captured at a serendipitous stage of two vesicles’ membranes (both leaflets) merging and their contents starting to intermix clearly showing the fusion interface with a previously unseen fusion rim. From the morphological informa...

  19. Membrane Fusion Induced by Small Molecules and Ions

    Sutapa Mondal Roy; Munna Sarkar

    2011-01-01

    Membrane fusion is a key event in many biological processes. These processes are controlled by various fusogenic agents of which proteins and peptides from the principal group. The fusion process is characterized by three major steps, namely, inter membrane contact, lipid mixing forming the intermediate step, pore opening and finally mixing of inner contents of the cells/vesicles. These steps are governed by energy barriers, which need to be overcome to complete fusion. Structural reorganizat...

  20. Acidification triggers Andes hantavirus membrane fusion and rearrangement of Gc into a stable post-fusion homotrimer.

    Acuña, Rodrigo; Bignon, Eduardo A; Mancini, Roberta; Lozach, Pierre-Yves; Tischler, Nicole D

    2015-11-01

    The hantavirus membrane fusion process is mediated by the Gc envelope glycoprotein from within endosomes. However, little is known about the specific mechanism that triggers Gc fusion activation, and its pre- and post-fusion conformations. We established cell-free in vitro systems to characterize hantavirus fusion activation. Low pH was sufficient to trigger the interaction of virus-like particles with liposomes. This interaction was dependent on a pre-fusion glycoprotein arrangement. Further, low pH induced Gc multimerization changes leading to non-reversible Gc homotrimers. These trimers were resistant to detergent, heat and protease digestion, suggesting characteristics of a stable post-fusion structure. No acid-dependent oligomerization rearrangement was detected for the trypsin-sensitive Gn envelope glycoprotein. Finally, acidification induced fusion of glycoprotein-expressing effector cells with non-susceptible CHO cells. Together, the data provide novel information on the Gc fusion trigger and its non-reversible activation involving lipid interaction, multimerization changes and membrane fusion which ultimately allow hantavirus entry into cells. PMID:26310672

  1. A role for a TIMP-3-sensitive, Zn(2+)-dependent metalloprotease in mammalian gamete membrane fusion.

    Correa, L M; Cho, C; Myles, D G; Primakoff, P

    2000-09-01

    During fertilization, sperm and egg plasma membranes adhere and then fuse by a mechanism that is not well understood. Zinc metalloproteases are necessary for some intercellular fusion events, for instance, cell-cell fusion in yeast. In this study we tested the effects of class-specific and family-specific protease inhibitors on mouse gamete fusion. Capacitated, acrosome-reacted sperm and zona-free eggs were used in assays designed to define the effects of inhibitors on sperm-egg plasma membrane binding or fusion. Inhibitors of the aspartic, cysteine, and serine protease classes had no effect on sperm-egg binding or fusion. Both a synthetic metalloprotease substrate (succinyl-Ala-Ala-Phe-amidomethylcoumarin) and the zinc chelator 1,10-phenanthroline inhibited sperm-egg fusion but did not decrease sperm-egg binding. The fusion-inhibition effect of phenanthroline was reversible and activity of the inhibitable zinc metalloprotease was shown to be required during a short time window, the first 15 min after insemination. Tissue inhibitor of metalloprotease-3 and Ro 31-9790, specific inhibitors of zinc metalloproteases in the matrixin and adamalysin families, also inhibited sperm-egg fusion but not sperm-egg binding. These data indicate a role in gamete fusion for one or more zinc metalloproteases of the matrixin and/or adamalysin families that act after plasma membrane binding and before sperm-egg membrane fusion. PMID:10964469

  2. A compensatory mutation provides resistance to disparate HIV fusion inhibitor peptides and enhances membrane fusion.

    Matthew P Wood

    Full Text Available Fusion inhibitors are a class of antiretroviral drugs used to prevent entry of HIV into host cells. Many of the fusion inhibitors being developed, including the drug enfuvirtide, are peptides designed to competitively inhibit the viral fusion protein gp41. With the emergence of drug resistance, there is an increased need for effective and unique alternatives within this class of antivirals. One such alternative is a class of cyclic, cationic, antimicrobial peptides known as θ-defensins, which are produced by many non-human primates and exhibit broad-spectrum antiviral and antibacterial activity. Currently, the θ-defensin analog RC-101 is being developed as a microbicide due to its specific antiviral activity, lack of toxicity to cells and tissues, and safety in animals. Understanding potential RC-101 resistance, and how resistance to other fusion inhibitors affects RC-101 susceptibility, is critical for future development. In previous studies, we identified a mutant, R5-tropic virus that had evolved partial resistance to RC-101 during in vitro selection. Here, we report that a secondary mutation in gp41 was found to restore replicative fitness, membrane fusion, and the rate of viral entry, which were compromised by an initial mutation providing partial RC-101 resistance. Interestingly, we show that RC-101 is effective against two enfuvirtide-resistant mutants, demonstrating the clinical importance of RC-101 as a unique fusion inhibitor. These findings both expand our understanding of HIV drug-resistance to diverse peptide fusion inhibitors and emphasize the significance of compensatory gp41 mutations.

  3. Cell-based analysis of Chikungunya virus E1 protein in membrane fusion

    Kuo Szu-Cheng

    2012-04-01

    Full Text Available Abstract Background Chikungunya fever is a pandemic disease caused by the mosquito-borne Chikungunya virus (CHIKV. E1 glycoprotein mediation of viral membrane fusion during CHIKV infection is a crucial step in the release of viral genome into the host cytoplasm for replication. How the E1 structure determines membrane fusion and whether other CHIKV structural proteins participate in E1 fusion activity remain largely unexplored. Methods A bicistronic baculovirus expression system to produce recombinant baculoviruses for cell-based assay was used. Sf21 insect cells infected by recombinant baculoviruses bearing wild type or single-amino-acid substitution of CHIKV E1 and EGFP (enhanced green fluorescence protein were employed to investigate the roles of four E1 amino acid residues (G91, V178, A226, and H230 in membrane fusion activity. Results Western blot analysis revealed that the E1 expression level and surface features in wild type and mutant substituted cells were similar. However, cell fusion assay found that those cells infected by CHIKV E1-H230A mutant baculovirus showed little fusion activity, and those bearing CHIKV E1-G91D mutant completely lost the ability to induce cell-cell fusion. Cells infected by recombinant baculoviruses of CHIKV E1-A226V and E1-V178A mutants exhibited the same membrane fusion capability as wild type. Although the E1 expression level of cells bearing monomeric-E1-based constructs (expressing E1 only was greater than that of cells bearing 26S-based constructs (expressing all structural proteins, the sizes of syncytial cells induced by infection of baculoviruses containing 26S-based constructs were larger than those from infections having monomeric-E1 constructs, suggesting that other viral structure proteins participate or regulate E1 fusion activity. Furthermore, membrane fusion in cells infected by baculovirus bearing the A226V mutation constructs exhibited increased cholesterol-dependences and lower pH thresholds

  4. Membrane fusion by VAMP3 and plasma membrane t-SNAREs

    Pairing of SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins on vesicles (v-SNAREs) and SNARE proteins on target membranes (t-SNAREs) mediates intracellular membrane fusion. VAMP3/cellubrevin is a v-SNARE that resides in recycling endosomes and endosome-derived transport vesicles. VAMP3 has been implicated in recycling of transferrin receptors, secretion of α-granules in platelets, and membrane trafficking during cell migration. Using a cell fusion assay, we examined membrane fusion capacity of the ternary complexes formed by VAMP3 and plasma membrane t-SNAREs syntaxin1, syntaxin4, SNAP-23 and SNAP-25. VAMP3 forms fusogenic pairing with t-SNARE complexes syntaxin1/SNAP-25, syntaxin1/SNAP-23 and syntaxin4/SNAP-25, but not with syntaxin4/SNAP-23. Deletion of the N-terminal domain of syntaxin4 enhanced membrane fusion more than two fold, indicating that the N-terminal domain negatively regulates membrane fusion. Differential membrane fusion capacities of the ternary v-/t-SNARE complexes suggest that transport vesicles containing VAMP3 have distinct membrane fusion kinetics with domains of the plasma membrane that present different t-SNARE proteins

  5. Herpesvirus glycoproteins undergo multiple antigenic changes before membrane fusion.

    Daniel L Glauser

    Full Text Available Herpesvirus entry is a complicated process involving multiple virion glycoproteins and culminating in membrane fusion. Glycoprotein conformation changes are likely to play key roles. Studies of recombinant glycoproteins have revealed some structural features of the virion fusion machinery. However, how the virion glycoproteins change during infection remains unclear. Here using conformation-specific monoclonal antibodies we show in situ that each component of the Murid Herpesvirus-4 (MuHV-4 entry machinery--gB, gH/gL and gp150--changes in antigenicity before tegument protein release begins. Further changes then occurred upon actual membrane fusion. Thus virions revealed their final fusogenic form only in late endosomes. The substantial antigenic differences between this form and that of extracellular virions suggested that antibodies have only a limited opportunity to block virion membrane fusion.

  6. The dengue virus type 2 envelope protein fusion peptide is essential for membrane fusion

    The flaviviral envelope (E) protein directs virus-mediated membrane fusion. To investigate membrane fusion as a requirement for virus growth, we introduced 27 unique mutations into the fusion peptide of an infectious cDNA clone of dengue 2 virus and recovered seven stable mutant viruses. The fusion efficiency of the mutants was impaired, demonstrating for the first time the requirement for specific FP AAs in optimal fusion. Mutant viruses exhibited different growth kinetics and/or genetic stabilities in different cell types and adult mosquitoes. Virus particles could be recovered following RNA transfection of cells with four lethal mutants; however, recovered viruses could not re-infect cells. These viruses could enter cells, but internalized virus appeared to be retained in endosomal compartments of infected cells, thus suggesting a fusion blockade. Mutations of the FP also resulted in reduced virus reactivity with flavivirus group-reactive antibodies, confirming earlier reports using virus-like particles.

  7. The effect of acute microgravity on mechanically-induced membrane damage and membrane-membrane fusion events

    Clarke, M. S.; Vanderburg, C. R.; Feeback, D. L.; McIntire, L. V. (Principal Investigator)

    2001-01-01

    Although it is unclear how a living cell senses gravitational forces there is no doubt that perturbation of the gravitational environment results in profound alterations in cellular function. In the present study, we have focused our attention on how acute microgravity exposure during parabolic flight affects the skeletal muscle cell plasma membrane (i.e. sarcolemma), with specific reference to a mechanically-reactive signaling mechanism known as mechanically-induced membrane disruption or "wounding". Both membrane rupture and membrane resealing events mediated by membrane-membrane fusion characterize this response. We here present experimental evidence that acute microgravity exposure can inhibit membrane-membrane fusion events essential for the resealing of sarcolemmal wounds in individual human myoblasts. Additional evidence to support this contention comes from experimental studies that demonstrate acute microgravity exposure also inhibits secretagogue-stimulated intracellular vesicle fusion with the plasma membrane in HL-60 cells. Based on our own observations and those of other investigators in a variety of ground-based models of membrane wounding and membrane-membrane fusion, we suggest that the disruption in the membrane resealing process observed during acute microgravity is consistent with a microgravity-induced decrease in membrane order.

  8. Molecular View of the Role of Fusion Peptides in Promoting Positive Membrane Curvature

    Fuhrmans, Marc; Marrink, Siewert J.

    2012-01-01

    Fusion peptides are moderately hydrophobic segments of viral and nonviral membrane fusion proteins that enable these proteins to fuse two closely apposed biological membranes. In vitro assays furthermore show that even isolated fusion peptides alone can support membrane fusion in model systems. In a

  9. ITER activities and fusion technology

    At the 21st IAEA Fusion Energy Conference, 68 and 67 papers were presented in the categories of ITER activities and fusion technology, respectively. ITER performance prediction, results of technology R and D and the construction preparation provide good confidence in ITER realization. The superconducting tokamak EAST achieved the first plasma just before the conference. The construction of other new experimental machines has also shown steady progress. Future reactor studies stress the importance of down sizing and a steady-state approach. Reactor technology in the field of blanket including the ITER TBM programme and materials for the demonstration power plant showed sound progress in both R and D and design activities

  10. Two coiled-coil domains of Chlamydia trachomatis IncA affect membrane fusion events during infection.

    Erik Ronzone

    Full Text Available Chlamydia trachomatis replicates in a parasitophorous membrane-bound compartment called an inclusion. The inclusions corrupt host vesicle trafficking networks to avoid the degradative endolysosomal pathway but promote fusion with each other in order to sustain higher bacterial loads in a process known as homotypic fusion. The Chlamydia protein IncA (Inclusion protein A appears to play central roles in both these processes as it participates to homotypic fusion and inhibits endocytic SNARE-mediated membrane fusion. How IncA selectively inhibits or activates membrane fusion remains poorly understood. In this study, we analyzed the spatial and molecular determinants of IncA's fusogenic and inhibitory functions. Using a cell-free membrane fusion assay, we found that inhibition of SNARE-mediated fusion requires IncA to be on the same membrane as the endocytic SNARE proteins. IncA displays two coiled-coil domains showing high homology with SNARE proteins. Domain swap and deletion experiments revealed that although both these domains are capable of independently inhibiting SNARE-mediated fusion, these two coiled-coil domains cooperate in mediating IncA multimerization and homotypic membrane interaction. Our results support the hypothesis that Chlamydia employs SNARE-like virulence factors that positively and negatively affect membrane fusion and promote infection.

  11. Molecular View of the Role of Fusion Peptides in Promoting Positive Membrane Curvature

    Fuhrmans, Marc; Marrink, Siewert J.

    2012-01-01

    Fusion peptides are moderately hydrophobic segments of viral and nonviral membrane fusion proteins that enable these proteins to fuse two closely apposed biological membranes. In vitro assays furthermore show that even isolated fusion peptides alone can support membrane fusion in model systems. In addition, the fusion peptides have a distinct effect on the phase diagram of lipid mixtures. Here, we present molecular dynamics simulations investigating the effect of a particular fusion peptide, ...

  12. Flavivirus cell entry and membrane fusion

    Smit, Jolanda M.; Moesker, Bastiaan; Rodenhuis-Zybert, Izabela; Wilschut, Jan

    2011-01-01

    Flaviviruses, such as dengue virus and West Nile virus, are enveloped viruses that infect cells through receptor-mediated endocytosis and fusion from within acidic endosomes. The cell entry process of flaviviruses is mediated by the viral E glycoprotein. This short review will address recent advance

  13. Protection of immunocompromised mice against lethal infection with Pseudomonas aeruginosa by active or passive immunization with recombinant P. aeruginosa outer membrane protein F and outer membrane protein I fusion proteins.

    von Specht, B U; Knapp, B.; Muth, G; Bröker, M.; Hungerer, K D; Diehl, K D; Massarrat, K; Seemann, A; Domdey, H

    1995-01-01

    Recombinant outer membrane proteins (Oprs) of Pseudomonas aeruginosa were expressed in Escherichia coli as glutathione S-transferase (GST)-linked fusion proteins. GST-linked Oprs F and I (GST-OprF190-350 [GST linked to OprF spanning amino acids 190 to 350] and GST-OprI21-83, respectively) and recombinant hybrid Oprs (GST-OprF190-342-OprI21-83 and GST-OprI21-83-OprF190-350) were isolated and tested for their efficacy as vaccines in immunodeficient mice. GST-OprF-OprI protected the mice against...

  14. The destructive effect of botulinum neurotoxins on the SNARE protein: SNAP-25 and synaptic membrane fusion

    Bin Lu

    2015-06-01

    Full Text Available Synaptic exocytosis requires the assembly of syntaxin 1A and SNAP-25 on the plasma membrane and synaptobrevin 2 (VAMP2 on the vesicular membrane to bridge the two opposite membranes. It is believed that the three SNARE proteins assemble in steps along the dynamic assembly pathway. The C-terminus of SNAP-25 is known to be the target of botulinum neurotoxins (BoNT/A and BoNT/E that block neurotransmitters release in vivo. In this study, we employed electron paramagnetic resonance (EPR spectroscopy to investigate the conformation of the SNAP-25 C-terminus in binary and ternary SNARE complexes. The fluorescence lipid mixing assay shows that the C-terminal of SNAP-25 is essential for membrane fusion, and that the truncated SNAP-25 mutants cleaved by BoNT/A and BoNT/E display different inhibition effects on membrane fusion: SNAP-25E (Δ26 abolishes the fusion activity of the SNARE complex, while SNAP-25A (Δ9 loses most of its function, although it can still form a SDS-resistant SNARE complex as the wild-type SNAP-25. CW-EPR spectra validate the unstable structures of the SNARE complex formed by SNAP-25 mutants. We propose that the truncated SNAP-25 mutants will disrupt the assembly of the SNARE core complex, and then inhibit the synaptic membrane fusion accordingly.

  15. Membrane Bending Energy and Fusion Pore Kinetics in Ca2+-Triggered Exocytosis

    Zhang, Zhen; Jackson, Meyer B.

    2010-01-01

    A fusion pore composed of lipid is an obligatory kinetic intermediate of membrane fusion, and its formation requires energy to bend membranes into highly curved shapes. The energetics of such deformations in viral fusion is well established, but the role of membrane bending in Ca2+-triggered exocytosis remains largely untested. Amperometry recording showed that during exocytosis in chromaffin and PC12 cells, fusion pores formed by smaller vesicles dilated more rapidly than fusion pores formed...

  16. Amino Acid Sequence Requirements of the Transmembrane and Cytoplasmic Domains of Influenza Virus Hemagglutinin for Viable Membrane Fusion

    Melikyan, Grigory B.; Lin, Sasa; Roth, Michael G.; Cohen, Fredric S.

    1999-01-01

    The amino acid sequence requirements of the transmembrane (TM) domain and cytoplasmic tail (CT) of the hemagglutinin (HA) of influenza virus in membrane fusion have been investigated. Fusion properties of wild-type HA were compared with those of chimeras consisting of the ectodomain of HA and the TM domain and/or CT of polyimmunoglobulin receptor, a nonviral integral membrane protein. The presence of a CT was not required for fusion. But when a TM domain and CT were present, fusion activity w...

  17. Importin β Negatively Regulates Nuclear Membrane Fusion and Nuclear Pore Complex Assembly

    Harel, Amnon; Chan, Rene C.; Lachish-Zalait, Aurelie; Zimmerman, Ella; Elbaum, Michael; Forbes, Douglass J.

    2003-01-01

    Assembly of a eukaryotic nucleus involves three distinct events: membrane recruitment, fusion to form a double nuclear membrane, and nuclear pore complex (NPC) assembly. We report that importin β negatively regulates two of these events, membrane fusion and NPC assembly. When excess importin β is added to a full Xenopus nuclear reconstitution reaction, vesicles are recruited to chromatin but their fusion is blocked. The importin β down-regulation of membrane fusion is Ran-GTP reversible. Inde...

  18. Tri-membrane nanoparticles produced by combining liposome fusion and a novel patchwork of bicelles to overcome endosomal and nuclear membrane barriers to cargo delivery.

    Yamada, Asako; Mitsueda, Asako; Hasan, Mahadi; Ueda, Miho; Hama, Susumu; Warashina, Shota; Nakamura, Takashi; Harashima, Hideyoshi; Kogure, Kentaro

    2016-02-23

    Membrane fusion is a rational strategy for crossing intracellular membranes that present barriers to liposomal nanocarrier-mediated delivery of plasmid DNA into the nucleus of non-dividing cells, such as dendritic cells. Based on this strategy, we previously developed nanocarriers consisting of a nucleic acid core particle coated with four lipid membranes [Akita, et al., Biomaterials, 2009, 30, 2940-2949]. However, including the endosomal membrane and two nuclear membranes, cells possess three intracellular membranous barriers. Thus, after entering the nucleus, nanoparticles coated with four membranes would still have one lipid membrane remaining, and could impede cargo delivery. Until now, coating a core particle with an odd number of lipid membranes was challenging. To produce nanocarriers with an odd number of lipid membranes, we developed a novel coating method involving lipid nano-discs, also known as bicelles, as a material for packaging DNA in a carrier with an odd number of lipid membranes. In this procedure, bicelles fuse to form an outer coating that resembles a patchwork quilt, which allows the preparation of nanoparticles coated with only three lipid membranes. Moreover, the transfection activity of dendritic cells with these three-membrane nanoparticles was higher than that for nanoparticles coated with four lipid membranes. In summary, we developed novel nanoparticles coated with an odd number of lipid membranes using the novel "patchwork-packaging method" to deliver plasmid DNA into the nucleus via membrane fusion. PMID:26667208

  19. Shear-Induced Membrane Fusion in Viscous Solutions

    Kogan, Maxim

    2014-05-06

    Large unilamellar lipid vesicles do not normally fuse under fluid shear stress. They might deform and open pores to relax the tension to which they are exposed, but membrane fusion occurring solely due to shear stress has not yet been reported. We present evidence that shear forces in a viscous solution can induce lipid bilayer fusion. The fusion of 1,2-dioleoyl-sn-glycero-3- phosphocholine (DOPC) liposomes is observed in Couette flow with shear rates above 3000 s-1 provided that the medium is viscous enough. Liposome samples, prepared at different viscosities using a 0-50 wt % range of sucrose concentration, were studied by dynamic light scattering, lipid fusion assays using Förster resonance energy transfer (FRET), and linear dichroism (LD) spectroscopy. Liposomes in solutions with 40 wt % (or more) sucrose showed lipid fusion under shear forces. These results support the hypothesis that under suitable conditions lipid membranes may fuse in response to mechanical-force- induced stress. © 2014 American Chemical Society.

  20. Conflicting views on the membrane fusion machinery and the fusion pore

    Sørensen, Jakob B

    2009-01-01

    of the assembly of the fusogenic SNARE-complex. Here, I review conflicting views on the function of the core fusion machinery consisting of the SNAREs, Munc18, complexin, and synaptotagmin. Munc18 controls docking of vesicles to the plasma membrane and initial SNARE-complex assembly, whereas complexin...

  1. Towards fully automated Identification of Vesicle-Membrane Fusion Events in TIRF Microscopy

    Vallotton, Pascal; James, David E.; Hughes, William E.

    2007-11-01

    Total Internal Reflection Fluorescence Microscopy (TIRFM) is imposing itself as the tool of choice for studying biological activity in close proximity to the plasma membrane. For example, the exquisite selectivity of TIRFM allows monitoring the diffusion of GFP-phogrin vesicles and their recruitment to the plasma membrane in pancreatic β-cells. We present a novel computer vision system for automatically identifying the elusive fusion events of GFP-phogrin vesicles with the plasma membrane. Our method is based on robust object tracking and matched filtering. It should accelerate the quantification of TIRFM data and allow the extraction of more biological information from image data to support research in diabetes and obesity.

  2. Reversible Merger of Membranes at the Early Stage of Influenza Hemagglutinin-mediated Fusion

    Leikina, Eugenia; Chernomordik, Leonid V.

    2000-01-01

    Fusion mediated by influenza hemagglutinin (HA), a prototype fusion protein, is commonly detected as lipid and content mixing between fusing cells. Decreasing the surface density of fusion-competent HA inhibited these advanced fusion phenotypes and allowed us to identify an early stage of fusion at physiological temperature. Although lipid flow between membranes was restricted, the contacting membrane monolayers were apparently transiently connected, as detected by the transformation of this ...

  3. The Gaussian Curvature Elastic Energy of Intermediates in Membrane Fusion

    Siegel, David P.

    2008-01-01

    The Gaussian curvature elastic energy contribution to the energy of membrane fusion intermediates has usually been neglected because the Gaussian curvature elastic modulus, κ, was unknown. It is now possible to measure κ for phospholipids that form bicontinuous inverted cubic (QII) phases. Here, it is shown that one can estimate κ for lipids that do not form QII phases by studying the phase behavior of lipid mixtures. The method is used to estimate κ for several lipid compositions in excess w...

  4. Membrane interaction and structure of the transmembrane domain of influenza hemagglutinin and its fusion peptide complex

    Lin Chi-Hui

    2008-01-01

    Full Text Available Abstract Background To study the organization and interaction with the fusion domain (or fusion peptide, FP of the transmembrane domain (TMD of influenza virus envelope glycoprotein for its role in membrane fusion which is also essential in the cellular trafficking of biomolecules and sperm-egg fusion. Results The fluorescence and gel electrophoresis experiments revealed a tight self-assembly of TMD in the model membrane. A weak but non-random interaction between TMD and FP in the membrane was found. In the complex, the central TMD oligomer was packed by FP in an antiparallel fashion. FP insertion into the membrane was altered by binding to TMD. An infrared study exhibited an enhanced membrane perturbation by the complex formation. A model was built to illustrate the role of TMD in the late stages of influenza virus-mediated membrane fusion reaction. Conclusion The TMD oligomer anchors the fusion protein in the membrane with minimal destabilization to the membrane. Upon associating with FP, the complex exerts a synergistic effect on the membrane perturbation. This effect is likely to contribute to the complete membrane fusion during the late phase of fusion protein-induced fusion cascade. The results presented in the work characterize the nature of the interaction of TMD with the membrane and TMD in a complex with FP in the steps leading to pore initiation and dilation during virus-induced fusion. Our data and proposed fusion model highlight the key role of TMD-FP interaction and have implications on the fusion reaction mediated by other type I viral fusion proteins. Understanding the molecular mechanism of membrane fusion may assist in the design of anti-viral drugs.

  5. Regulation of membrane fusion and secretory events in the sea urchin embryo

    Membrane fusion and secretory events play a key role in fertilization and early development in the sea urchin embryo. To investigate the mechanism of membrane fusion, the effect of inhibitors of metalloendoprotease activity was studied on two model systems of cell fusion; fertilization and spiculogenesis by primary mesenchyme cells in the embryo. Both the zinc chelator, 1,10-phenanthroline, and peptide metalloprotease substrates were found to inhibit both fertilization and gamete fusion, while peptides that are not substrates of metalloproteases did not affect either process. Primary mesenchyme cells form the larval skeleton in the embryo by deposition of mineral and an organic matrix into a syncytial cavity formed by fusion of filopodia of these cells. Metalloprotease inhibitors were found to inhibit spiculogenesis both in vivo and in cultures of isolated primary mesenchyme cells, and the activity of a metalloprotease of the appropriate specificity was found in the primary mesenchyme cells. These two studies implicate the activity of a metalloprotease in a necessary step in membrane fusion. Following fertilization, exocytosis of the cortical granules results in the formation of the fertilization envelope and the hyaline layer, that surround the developing embryo. The hatching enzyme is secreted by the blastula stage sea urchin embryo, which proteolyzes the fertilization envelope surrounding the embryo, allowing the embryo to hatch. Using an assay that measures 125I-fertilization envelope degradation, the hatching enzyme was identified as a 33 kDa metalloprotease, and was purified by ion-exchange and affinity chromatography from the hatching media of Strongylocentrotus purpuratus embryos. The hatching enzyme showed a substrate preference for only a minor subset of fertilization envelope proteins

  6. Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

    A.T. Da Poian

    2005-06-01

    Full Text Available Enveloped viruses always gain entry into the cytoplasm by fusion of their lipid envelope with a cell membrane. Some enveloped viruses fuse directly with the host cell plasma membrane after virus binding to the cell receptor. Other enveloped viruses enter the cells by the endocytic pathway, and fusion depends on the acidification of the endosomal compartment. In both cases, virus-induced membrane fusion is triggered by conformational changes in viral envelope glycoproteins. Two different classes of viral fusion proteins have been described on the basis of their molecular architecture. Several structural data permitted the elucidation of the mechanisms of membrane fusion mediated by class I and class II fusion proteins. In this article, we review a number of results obtained by our laboratory and by others that suggest that the mechanisms involved in rhabdovirus fusion are different from those used by the two well-studied classes of viral glycoproteins. We focus our discussion on the electrostatic nature of virus binding and interaction with membranes, especially through phosphatidylserine, and on the reversibility of the conformational changes of the rhabdovirus glycoprotein involved in fusion. Taken together, these data suggest the existence of a third class of fusion proteins and support the idea that new insights should emerge from studies of membrane fusion mediated by the G protein of rhabdoviruses. In particular, the elucidation of the three-dimensional structure of the G protein or even of the fusion peptide at different pH's might provide valuable information for understanding the fusion mechanism of this new class of fusion proteins.

  7. Low activation materials for fusion

    The viability of fusion as a future energy source may eventually be determined by safety and environmental factors. Control of the induced radioactivity characteristics of the materials used in the first wall and blanket could have a major favorable impact on these issues. In the United States, materials program efforts are focused on developing new structural alloys with radioactive decay characteristics which would greatly simplify long-term waste disposal of reactor components. A range of alloy systems is being explored in order to maintain the maximum number of design options. Significant progress has been made, and it now appears probable that reduced-activation engineering alloys with properties at least equivalent to conventional alloys can be successfully developed and commercialized. 10 refs., 1 fig

  8. Mutations in the DI-DII Linker of Human Parainfluenza Virus Type 3 Fusion Protein Result in Diminished Fusion Activity.

    Wenyan Xie

    Full Text Available Human parainfluenza virus type 3 (HPIV3 can cause severe respiratory tract diseases in infants and young children, but no licensed vaccines or antiviral agents are currently available for treatment. Fusing the viral and target cell membranes is a prerequisite for its entry into host cells and is directly mediated by the fusion (F protein. Although several domains of F are known to have important effects on regulating the membrane fusion activity, the roles of the DI-DII linker (residues 369-374 of the HPIV3 F protein in the fusogenicity still remains ill-defined. To facilitate our understanding of the role of this domain might play in F-induced cell-cell fusion, nine single mutations were engineered into this domain by site-directed mutagenesis. A vaccinia virus-T7 RNA polymerase transient expression system was employed to express the wild-type or mutated F proteins. These mutants were analyzed for membrane fusion activity, cell surface expression, and interaction between F and HN protein. Each of the mutated F proteins in this domain has a cell surface expression level similar to that of wild-type F. All of them resulted in a significant reduction in fusogenic activity in all steps of membrane fusion. Furthermore, all these fusion-deficient mutants reduced the amount of the HN-F complexes at the cell surface. Together, the results of our work suggest that this region has an important effect on the fusogenic activity of F.

  9. Membrane technologies for tritium recovering in the fusion fuel cycle

    Palladium and palladium-silver permeators have been obtained by coating porous ceramic tubes with a thin metal layer. Three coating techniques have been studied and characterized: sputtering, chemical electroless deposition and cold-rolling. The Pd-Ag membranes obtained by cold-rolling and annealing of thin metal foils have shown complete hydrogen selectivity and chemical and physical stability meeting the requirements of the fuel cycle applications. These rolled membranes have been tested at 300-400 deg. C with a hydrogen transmembrane pressure in the range of 100-280 kPa and hydrogen flow rates up to 2.5x10-6m3/s. By filling the Pd-Ag membranes with a catalyst selective for the water gas shift reaction, membrane reactors have been obtained for recovering hydrogen isotopes in elemental form from tritiated water. Particularly, a closed-loop process based on a Pd-Ag membrane reactor has been studied for the tritium recovery system of an ITER scale fusion reactor. (author)

  10. An Ion Switch Regulates Fusion of Charged Membranes

    Siepi, Evgenios; Lutz, Silke; Meyer, Sylke; Panzner, Steffen

    2011-01-01

    Here we identify the recruitment of solvent ions to lipid membranes as the dominant regulator of lipid phase behavior. Our data demonstrate that binding of counterions to charged lipids promotes the formation of lamellar membranes, whereas their absence can induce fusion. The mechanism applies to anionic and cationic liposomes, as well as the recently introduced amphoteric liposomes. In the latter, an additional pH-dependent lipid salt formation between anionic and cationic lipids must occur, as indicated by the depletion of membrane-bound ions in a zone around pH 5. Amphoteric liposomes fuse under these conditions but form lamellar structures at both lower and higher pH values. The integration of these observations into the classic lipid shape theory yielded a quantitative link between lipid and solvent composition and the physical state of the lipid assembly. The key parameter of the new model, κ(pH), describes the membrane phase behavior of charged membranes in response to their ion loading in a quantitative way. PMID:21575575

  11. β2 Adrenergic Receptor Fluorescent Protein Fusions Traffic to the Plasma Membrane and Retain Functionality

    Bubnell, Jaclyn; Pfister, Patrick; Sapar, Maria L.; Rogers, Matthew E.; Feinstein, Paul

    2013-01-01

    Green fluorescent protein (GFP) has proven useful for the study of protein interactions and dynamics for the last twenty years. A variety of new fluorescent proteins have been developed that expand the use of available excitation spectra. We have undertaken an analysis of seven of the most useful fluorescent proteins (XFPs), Cerulean (and mCerulean3), Teal, GFP, Venus, mCherry and TagRFP657, as fusions to the archetypal G-protein coupled receptor, the β2 adrenergic receptor (β2AR). We have characterized these β2AR::XFP fusions in respect to membrane trafficking and G-protein activation. We noticed that in the mouse neural cell line, OP 6, that membrane bound β2AR::XFP fusions robustly localized in the filopodia identical to gap::XFP fusions. All β2AR::XFP fusions show responses indistinguishable from each other and the non-fused form after isoprenaline exposure. Our results provide a platform by which G-protein coupled receptors can be dissected for their functionality. PMID:24086401

  12. Insulin-stimulated plasma membrane fusion of Glut4 glucose transporter-containing vesicles is regulated by phospholipase D1.

    Huang, Ping; Altshuller, Yelena M; Hou, June Chunqiu; Pessin, Jeffrey E; Frohman, Michael A

    2005-06-01

    Insulin stimulates glucose uptake in fat and muscle by mobilizing Glut4 glucose transporters from intracellular membrane storage sites to the plasma membrane. This process requires the trafficking of Glut4-containing vesicles toward the cell periphery, docking at exocytic sites, and plasma membrane fusion. We show here that phospholipase D (PLD) production of the lipid phosphatidic acid (PA) is a key event in the fusion process. PLD1 is found on Glut4-containing vesicles, is activated by insulin signaling, and traffics with Glut4 to exocytic sites. Increasing PLD1 activity facilitates glucose uptake, whereas decreasing PLD1 activity is inhibitory. Diminished PA production does not substantially hinder trafficking of the vesicles or their docking at the plasma membrane, but it does impede fusion-mediated extracellular exposure of the transporter. The fusion block caused by RNA interference-mediated PLD1 deficiency is rescued by exogenous provision of a lipid that promotes fusion pore formation and expansion, suggesting that the step regulated by PA is late in the process of vesicle fusion. PMID:15772157

  13. The SI strain of measles virus derived from a patient with subacute sclerosing panencephalitis possesses typical genome alterations and unique amino acid changes that modulate receptor specificity and reduce membrane fusion activity.

    Seki, Fumio; Yamada, Kentaro; Nakatsu, Yuichiro; Okamura, Koji; Yanagi, Yusuke; Nakayama, Tetsuo; Komase, Katsuhiro; Takeda, Makoto

    2011-11-01

    Subacute sclerosing panencephalitis (SSPE) is a fatal sequela associated with measles and is caused by persistent infection of the brain with measles virus (MV). The SI strain was isolated in 1976 from a patient with SSPE and shows neurovirulence in animals. Genome nucleotide sequence analyses showed that the SI strain genome possesses typical genome alterations for SSPE-derived strains, namely, accumulated amino acid substitutions in the M protein and cytoplasmic tail truncation of the F protein. Through the establishment of an efficient reverse genetics system, a recombinant SI strain expressing a green fluorescent protein (rSI-AcGFP) was generated. The infection of various cell types with rSI-AcGFP was evaluated by fluorescence microscopy. rSI-AcGFP exhibited limited syncytium-forming activity and spread poorly in cells. Analyses using a recombinant MV possessing a chimeric genome between those of the SI strain and a wild-type MV strain indicated that the membrane-associated protein genes (M, F, and H) were responsible for the altered growth phenotype of the SI strain. Functional analyses of viral glycoproteins showed that the F protein of the SI strain exhibited reduced fusion activity because of an E300G substitution and that the H protein of the SI strain used CD46 efficiently but used the original MV receptors on immune and epithelial cells poorly because of L482F, S546G, and F555L substitutions. The data obtained in the present study provide a new platform for analyses of SSPE-derived strains as well as a clear example of an SSPE-derived strain that exhibits altered receptor specificity and limited fusion activity. PMID:21917959

  14. Point-Like Protrusion as a Prestalk Intermediate in Membrane Fusion Pathway

    Efrat, Avishay; Chernomordik, Leonid V; Kozlov, Michael M.

    2007-01-01

    The widely accepted pathway of membrane fusion begins with the fusion stalk representing the initial intermediate of hemifusion. The lipid structures preceding hemifusion and their possible influence on fusion kinetics were not addressed. Here, we suggest the point-like protrusion as a prestalk fusion intermediate, which has energy lower than that of stalk and, therefore, does not limit the fusion rate. We demonstrate that by calculating the energy of the point-like protrusion, which depends ...

  15. Insulin-stimulated Plasma Membrane Fusion of Glut4 Glucose Transporter-containing Vesicles Is Regulated by Phospholipase D1D⃞

    Huang, Ping; Altshuller, Yelena M.; Hou, June Chunqiu; Jeffrey E Pessin; Frohman, Michael A.

    2005-01-01

    Insulin stimulates glucose uptake in fat and muscle by mobilizing Glut4 glucose transporters from intracellular membrane storage sites to the plasma membrane. This process requires the trafficking of Glut4-containing vesicles toward the cell periphery, docking at exocytic sites, and plasma membrane fusion. We show here that phospholipase D (PLD) production of the lipid phosphatidic acid (PA) is a key event in the fusion process. PLD1 is found on Glut4-containing vesicles, is activated by insu...

  16. Pathway of membrane fusion with two tension-dependent energy barriers

    Shillcock, Julian C.

    2007-01-01

    Fusion of bilayer membranes is studied via dissipative particle dynamics (DPD) simulations. A new set of DPD parameters is introduced which leads to an energy barrier for flips of lipid molecules between adhering membranes. A large number of fusion events is monitored for a vesicle in contact...

  17. Trans-complex formation by proteolipid channels in the terminal phase of membrane fusion

    Peters, C; Bayer, M J; Bühler, S;

    2001-01-01

    SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) and Rab-GTPases, together with their cofactors, mediate the attachment step in the membrane fusion of vesicles. But how bilayer mixing--the subsequent core process of fusion--is catalysed remains unclear. Ca2+/calmodu......SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) and Rab-GTPases, together with their cofactors, mediate the attachment step in the membrane fusion of vesicles. But how bilayer mixing--the subsequent core process of fusion--is catalysed remains unclear. Ca2......+/calmodulin controls this terminal process in many intracellular fusion events. Here we identify V0, the membrane-integral sector of the vacuolar H+-ATPase, as a target of calmodulin on yeast vacuoles. Between docking and bilayer fusion, V0 sectors from opposing membranes form complexes. V0 trans...

  18. A host–guest system to study structure–function relationships of membrane fusion peptides

    Han, Xing; Tamm, Lukas K.

    2000-01-01

    We designed a host–guest fusion peptide system, which is completely soluble in water and has a high affinity for biological and lipid model membranes. The guest sequences are those of the fusion peptides of influenza hemagglutinin, which are solubilized by a highly charged unstructured C-terminal host sequence. These peptides partition to the surface of negatively charged liposomes or erythrocytes and elicit membrane fusion or hemolysis. They undergo a conformational ...

  19. Protein secretion by hybrid bacterial ABC-transporters: specific functions of the membrane ATPase and the membrane fusion protein.

    Binet, R; Wandersman, C

    1995-01-01

    The Erwinia chrysanthemi metalloprotease C and the Serratia marcescens haem acquisition protein HasA are both secreted from Gram-negative bacteria by a signal peptide-independent pathway which requires a C-terminal secretion signal and a specific ABC-transporter made up of three proteins: a membrane ATPase (the ABC-protein), a second inner membrane component belonging to the membrane fusion protein family and an outer membrane polypeptide. HasA and protease C transporters are homologous altho...

  20. Inhibition of HIV-1 endocytosis allows lipid mixing at the plasma membrane, but not complete fusion

    de la Vega Michelle

    2011-12-01

    Full Text Available Abstract Background We recently provided evidence that HIV-1 enters HeLa-derived TZM-bl and lymphoid CEMss cells by fusing with endosomes, whereas its fusion with the plasma membrane does not proceed beyond the lipid mixing step. The mechanism of restriction of HIV-1 fusion at the cell surface and/or the factors that aid the virus entry from endosomes remain unclear. Results We examined HIV-1 fusion with a panel of target cells lines and with primary CD4+ T cells. Kinetic measurements of fusion combined with time-resolved imaging of single viruses further reinforced the notion that HIV-1 enters the cells via endocytosis and fusion with endosomes. Furthermore, we attempted to deliberately redirect virus fusion to the plasma membrane, using two experimental strategies. First, the fusion reaction was synchronized by pre-incubating the viruses with cells at reduced temperature to allow CD4 and coreceptors engagement, but not the virus uptake or fusion. Subsequent shift to a physiological temperature triggered accelerated virus uptake followed by entry from endosomes, but did not permit fusion at the cell surface. Second, blocking HIV-1 endocytosis by a small-molecule dynamin inhibitor, dynasore, resulted in transfer of viral lipids to the plasma membrane without any detectable release of the viral content into the cytosol. We also found that a higher concentration of dynasore is required to block the HIV-endosome fusion compared to virus internalization. Conclusions Our results further support the notion that HIV-1 enters disparate cell types through fusion with endosomes. The block of HIV-1 fusion with the plasma membrane at a post-lipid mixing stage shows that this membrane is not conducive to fusion pore formation and/or enlargement. The ability of dynasore to interfere with the virus-endosome fusion suggests that dynamin could be involved in two distinct steps of HIV-1 entry - endocytosis and fusion within intracellular compartments.

  1. Influenza viral membrane fusion is sensitive to sterol concentration but surprisingly robust to sterol chemical identity.

    Zawada, Katarzyna E; Wrona, Dominik; Rawle, Robert J; Kasson, Peter M

    2016-01-01

    Influenza virions are enriched in cholesterol relative to the plasma membrane from which they bud. Previous work has shown that fusion between influenza virus and synthetic liposomes is sensitive to the amount of cholesterol in either the virus or the target membrane. Here, we test the chemical properties of cholesterol required to promote influenza fusion by replacing cholesterol with other sterols and assaying viral fusion kinetics. We find that influenza fusion with liposomes is surprisingly robust to sterol chemical identity, showing no significant dependence on sterol identity in target membranes for any of the sterols tested. In the viral membrane, lanosterol slowed fusion somewhat, while polar sterols produced a more pronounced slowing and inhibition of fusion. No other sterols tested showed a significant perturbation in fusion rates, including ones previously shown to alter membrane bending moduli or phase behavior. Although fusion rates depend on viral cholesterol, they thus do not require cholesterol's ability to support liquid-liquid phase coexistence. Using electron cryo-microscopy, we further find that sterol-dependent changes to hemagglutinin spatial patterning in the viral membrane do not require liquid-liquid phase coexistence. We therefore speculate that local sterol-hemagglutinin interactions in the viral envelope may control the rate-limiting step of fusion. PMID:27431907

  2. SNARE-mediated rapid lysosome fusion in membrane raft clustering and dysfunction of bovine coronary arterial endothelium

    Han, Wei-Qing; Xia, Min; Zhang, Chun; Zhang, Fan; Xu, Ming; Li, Ning-Jun; Li, Pin-Lan

    2011-01-01

    The present study attempted to evaluate whether soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) mediate lysosome fusion in response to death receptor activation and contribute to membrane raft (MR) clustering and consequent endothelial dysfunction in coronary arterial endothelial cells. By immunohistochemical analysis, vesicle-associated membrane proteins 2 (VAMP-2, vesicle-SNAREs) were found to be abundantly expressed in the endothelium of bovine coronary arte...

  3. The Fusion of Membranes and Vesicles: Pathway and Energy Barriers from Dissipative Particle Dynamics

    Grafmüller, Andrea; Shillcock, Julian; Lipowsky, Reinhard

    2009-01-01

    The fusion of lipid bilayers is studied with dissipative particle dynamics simulations. First, to achieve control over membrane properties, the effects of individual simulation parameters are studied and optimized. Then, a large number of fusion events for a vesicle and a planar bilayer are simulated using the optimized parameter set. In the observed fusion pathway, configurations of individual lipids play an important role. Fusion starts with individual lipids assuming a splayed tail configu...

  4. HIV fusion peptide penetrates, disorders, and softens T-cell membrane mimics.

    Tristram-Nagle, Stephanie; Chan, Rob; Kooijman, Edgar; Uppamoochikkal, Pradeep; Qiang, Wei; Weliky, David P; Nagle, John F

    2010-09-10

    This work investigates the interaction of N-terminal gp41 fusion peptide (FP) of human immunodeficiency virus type 1 (HIV-1) with model membranes in order to elucidate how FP leads to fusion of HIV and T-cell membranes. FP constructs were (i) wild-type FP23 (23 N-terminal amino acids of gp41), (ii) water-soluble monomeric FP that adds six lysines on the C-terminus of FP23 (FPwsm), and (iii) the C-terminus covalently linked trimeric version (FPtri) of FPwsm. Model membranes were (i) LM3 (a T-cell mimic), (ii) 1,2-dioleoyl-sn-glycero-3-phosphocholine, (iii) 1,2-dioleoyl-sn-glycero-3-phosphocholine/30 mol% cholesterol, (iv) 1,2-dierucoyl-sn-glycero-3-phosphocholine, and (v) 1,2-dierucoyl-sn-glycero-3-phosphocholine/30 mol% cholesterol. Diffuse synchrotron low-angle x-ray scattering from fully hydrated samples, supplemented by volumetric data, showed that FP23 and FPtri penetrate into the hydrocarbon region and cause membranes to thin. Depth of penetration appears to depend upon a complex combination of factors including bilayer thickness, presence of cholesterol, and electrostatics. X-ray data showed an increase in curvature in hexagonal phase 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, which further indicates that FP23 penetrates into the hydrocarbon region rather than residing in the interfacial headgroup region. Low-angle x-ray scattering data also yielded the bending modulus K(C), a measure of membrane stiffness, and wide-angle x-ray scattering yielded the S(xray) orientational order parameter. Both FP23 and FPtri decreased K(C) and S(xray) considerably, while the weak effect of FPwsm suggests that it did not partition strongly into LM3 model membranes. Our results are consistent with the HIV FP disordering and softening the T-cell membrane, thereby lowering the activation energy for viral membrane fusion. PMID:20655315

  5. Binding and Fusion of Extracellular Vesicles to the Plasma Membrane of Their Cell Targets.

    Prada, Ilaria; Meldolesi, Jacopo

    2016-01-01

    Exosomes and ectosomes, extracellular vesicles of two types generated by all cells at multivesicular bodies and the plasma membrane, respectively, play critical roles in physiology and pathology. A key mechanism of their function, analogous for both types of vesicles, is the fusion of their membrane to the plasma membrane of specific target cells, followed by discharge to the cytoplasm of their luminal cargo containing proteins, RNAs, and DNA. Here we summarize the present knowledge about the interactions, binding and fusions of vesicles with the cell plasma membrane. The sequence initiates with dynamic interactions, during which vesicles roll over the plasma membrane, followed by the binding of specific membrane proteins to their cell receptors. Membrane binding is then converted rapidly into fusion by mechanisms analogous to those of retroviruses. Specifically, proteins of the extracellular vesicle membranes are structurally rearranged, and their hydrophobic sequences insert into the target cell plasma membrane which undergoes lipid reorganization, protein restructuring and membrane dimpling. Single fusions are not the only process of vesicle/cell interactions. Upon intracellular reassembly of their luminal cargoes, vesicles can be regenerated, released and fused horizontally to other target cells. Fusions of extracellular vesicles are relevant also for specific therapy processes, now intensely investigated. PMID:27517914

  6. International program activities in magnetic fusion energy

    The following areas of our international activities in magnetic fusion are briefly described: (1) policy; (2) background; (3) strategy; (4) strategic considerations and concerns; (5) domestic program inplications, and (6) implementation. The current US activities are reviewed. Some of our present program needs are outlined

  7. Implication des peptides de fusion des glycoprotéines de fusion virales de classe I dans la fusion membranaire

    Brasseur R.; Charloteaux B.; Lins L.; Lorin A.

    2007-01-01

    The implication of fusion peptides of class I viral fusion glycoproteins in the membrane fusion. Viral infection involves fusion between the viral envelope and the target cell plasmic membrane. The fusion is induced by a glycoprotein anchored in the viral envelope. After activation, the glycoprotein undergoes a conformational change inducing the exposure of a region named « fusion peptide » essential for the fusion process. Studies on glycoproteins and on isolated fusion peptides have allowed...

  8. N-(3-Cyanophenyl)-2-phenylacetamide, an effective inhibitor of morbillivirus-induced membrane fusion with low cytotoxicity.

    Singethan, K; Hiltensperger, G; Kendl, S; Wohlfahrt, J; Plattet, P; Holzgrabe, U; Schneider-Schaulies, J

    2010-11-01

    Based on the structural similarity of viral fusion proteins within the family Paramyxoviridae, we tested recently described and newly synthesized acetanilide derivatives for their capacity to inhibit measles virus (MV)-, canine distemper virus (CDV)- and Nipah virus (NiV)-induced membrane fusion. We found that N-(3-cyanophenyl)-2-phenylacetamide (compound 1) has a high capacity to inhibit MV- and CDV-induced (IC(50) μM), but not NiV-induced, membrane fusion. This compound is of outstanding interest because it can be easily synthesized and its cytotoxicity is low [50 % cytotoxic concentration (CC(50)) ≥ 300 μM], leading to a CC(50)/IC(50) ratio of approximately 100. In addition, primary human peripheral blood lymphocytes and primary dog brain cell cultures (DBC) also tolerate high concentrations of compound 1. Infection of human PBMC with recombinant wild-type MV is inhibited by an IC(50) of approximately 20 μM. The cell-to-cell spread of recombinant wild-type CDV in persistently infected DBC can be nearly completely inhibited by compound 1 at 50 μM, indicating that the virus spread between brain cells is dependent on the activity of the viral fusion protein. Our findings demonstrate that this compound is a most applicable inhibitor of morbillivirus-induced membrane fusion in tissue culture experiments including highly sensitive primary cells. PMID:20685931

  9. Mitochondrial DNA mutations provoke dominant inhibition of mitochondrial inner membrane fusion.

    Cécile Sauvanet

    Full Text Available Mitochondria are highly dynamic organelles that continuously move, fuse and divide. Mitochondrial dynamics modulate overall mitochondrial morphology and are essential for the proper function, maintenance and transmission of mitochondria and mitochondrial DNA (mtDNA. We have investigated mitochondrial fusion in yeast cells with severe defects in oxidative phosphorylation (OXPHOS due to removal or various specific mutations of mtDNA. We find that, under fermentative conditions, OXPHOS deficient cells maintain normal levels of cellular ATP and ADP but display a reduced mitochondrial inner membrane potential. We demonstrate that, despite metabolic compensation by glycolysis, OXPHOS defects are associated to a selective inhibition of inner but not outer membrane fusion. Fusion inhibition was dominant and hampered the fusion of mutant mitochondria with wild-type mitochondria. Inhibition of inner membrane fusion was not systematically associated to changes of mitochondrial distribution and morphology, nor to changes in the isoform pattern of Mgm1, the major fusion factor of the inner membrane. However, inhibition of inner membrane fusion correlated with specific alterations of mitochondrial ultrastructure, notably with the presence of aligned and unfused inner membranes that are connected to two mitochondrial boundaries. The fusion inhibition observed upon deletion of OXPHOS related genes or upon removal of the entire mtDNA was similar to that observed upon introduction of point mutations in the mitochondrial ATP6 gene that are associated to neurogenic ataxia and retinitis pigmentosa (NARP or to maternally inherited Leigh Syndrome (MILS in humans. Our findings indicate that the consequences of mtDNA mutations may not be limited to OXPHOS defects but may also include alterations in mitochondrial fusion. Our results further imply that, in healthy cells, the dominant inhibition of fusion could mediate the exclusion of OXPHOS-deficient mitochondria from

  10. On the Fusion of Confucianism, Buddhism and Taoism in Chinese Cognitive Membrane

    Jiawei Geng; H. J. Cai

    2014-01-01

    The coexistence of Confucianism, Buddhism and Taoism in China is phenomenal through world cultural history, which was explained by putting emphasis on the particularity or complementarity of those three doctrines. We proposed that, Chinese long-term self-assertiveness demands and their evolution lead to enduring competitions among them and eventual fusion of them, within Chinese Cognitive Membrane. It is emphasized that Chinese Cognitive Membrane needs further fusion with spirits of modern sc...

  11. Membrane pumping technology for helium and hydrogen isotope separation in the fusion reactor

    Pistunovich, V.I. [Kurchatov Inst., Moscow (Russian Federation). NFI RRC; Pigarov, A.Yu. [Kurchatov Inst., Moscow (Russian Federation). NFI RRC; Busnyuk, A.O. [Bonch-Bruyevich University, St. Petersburg (Russian Federation); Livshits, A.I. [Bonch-Bruyevich University, St. Petersburg (Russian Federation); Notkin, M.E. [Bonch-Bruyevich University, St. Petersburg (Russian Federation); Samartsev, A.A. [Bonch-Bruyevich University, St. Petersburg (Russian Federation); Borisenko, K.L. [Efremov Institute, St. Petersburg (Russian Federation); Darmogray, V.V. [Efremov Institute, St. Petersburg (Russian Federation); Ershov, B.D. [Efremov Institute, St. Petersburg (Russian Federation); Filippova, L.V. [Efremov Institute, St. Petersburg (Russian Federation); Mudugin, B.G. [Efremov Institute, St. Petersburg (Russian Federation); Odintsov, V.N. [Efremov Institute, St. Petersburg (Russian Federation); Saksagansky, G.L. [Efremov Institute, St. Petersburg (Russian Federation); Serebrennikov, D.V. [Efremov Institute, St. Petersburg (Russian Federation)

    1995-03-01

    A gas pumping system for ITER, improved by implementation of superpermeable membranes for selective hydrogen isotope exhaust, is considered. A study of the pumping capability of a niobium membrane for a hydrogen-helium mixture has been performed.Monte Carlo simulations of gas behaviour for the experimental facility and fusion reactor have been done.The scheme of the ITER pumping system with the membranes and membrane pumping technology was considered. The conceptual study the membrane pump for the ITER was done. This work gives good prospects for the membrane pumping use in ITER to reduce the total inventory of tritium necessary for reactor operation. (orig.).

  12. Mitotic phosphorylation of VCIP135 blocks p97ATPase-mediated Golgi membrane fusion

    Totsukawa, Go; Matsuo, Ayaka; Kubota, Ayano; Taguchi, Yuya; Kondo, Hisao, E-mail: hk228@med.kyushu-u.ac.jp

    2013-04-05

    Highlights: •VCIP135 is mitotically phosphorylated on Threonine-760 and Serine-767 by Cdc2. •Phosphorylated VCIP135 does not bind to p97ATPase. •The phosphorylation of VCIP135 inhibits p97ATPase-mediated Golgi membrane fusion. -- Abstract: In mammals, the Golgi apparatus is disassembled early mitosis and reassembled at the end of mitosis. For Golgi disassembly, membrane fusion needs to be blocked. Golgi biogenesis requires two distinct p97ATPase-mediated membrane fusion, the p97/p47 and p97/p37 pathways. We previously reported that p47 phosphorylation on Serine-140 and p37 phosphorylation on Serine-56 and Threonine-59 result in mitotic inhibition of the p97/p47 and the p97/p37 pathways, respectively [11,14]. In this study, we show another mechanism of mitotic inhibition of p97-mediated Golgi membrane fusion. We clarified that VCIP135, an essential factor in both p97 membrane fusion pathways, is phosphorylated on Threonine-760 and Serine-767 by Cdc2 at mitosis and that this phosphorylated VCIP135 does not bind to p97. An in vitro Golgi reassembly assay revealed that VCIP135(T760E, S767E), which mimics mitotic phosphorylation, caused no cisternal regrowth. Our results indicate that the phosphorylation of VCIP135 on Threonine-760 and Serine-767 inhibits p97-mediated Golgi membrane fusion at mitosis.

  13. The structural dynamics of the flavivirus fusion peptide-membrane interaction.

    Ygara S Mendes

    Full Text Available Membrane fusion is a crucial step in flavivirus infections and a potential target for antiviral strategies. Lipids and proteins play cooperative roles in the fusion process, which is triggered by the acidic pH inside the endosome. This acidic environment induces many changes in glycoprotein conformation and allows the action of a highly conserved hydrophobic sequence, the fusion peptide (FP. Despite the large volume of information available on the virus-triggered fusion process, little is known regarding the mechanisms behind flavivirus-cell membrane fusion. Here, we evaluated the contribution of a natural single amino acid difference on two flavivirus FPs, FLA(G ((98DRGWGNGCGLFGK(110 and FLA(H ((98DRGWGNHCGLFGK(110, and investigated the role of the charge of the target membrane on the fusion process. We used an in silico approach to simulate the interaction of the FPs with a lipid bilayer in a complementary way and used spectroscopic approaches to collect conformation information. We found that both peptides interact with neutral and anionic micelles, and molecular dynamics (MD simulations showed the interaction of the FPs with the lipid bilayer. The participation of the indole ring of Trp appeared to be important for the anchoring of both peptides in the membrane model, as indicated by MD simulations and spectroscopic analyses. Mild differences between FLA(G and FLA(H were observed according to the pH and the charge of the target membrane model. The MD simulations of the membrane showed that both peptides adopted a bend structure, and an interaction between the aromatic residues was strongly suggested, which was also observed by circular dichroism in the presence of micelles. As the FPs of viral fusion proteins play a key role in the mechanism of viral fusion, understanding the interactions between peptides and membranes is crucial for medical science and biology and may contribute to the design of new antiviral drugs.

  14. Class II fusion protein of alphaviruses drives membrane fusion through the same pathway as class I proteins

    Zaitseva, Elena; Mittal, Aditya; Griffin, Diane E.; Chernomordik, Leonid V.

    2005-01-01

    Viral fusion proteins of classes I and II differ radically in their initial structures but refold toward similar conformations upon activation. Do fusion pathways mediated by alphavirus E1 and influenza virus hemagglutinin (HA) that exemplify classes II and I differ to reflect the difference in their initial conformations, or concur to reflect the similarity in the final conformations? Here, we dissected the pathway of low pH–triggered E1-mediated cell–cell fusion by reducing the numbers of a...

  15. Synthesis of CDP-diacylglycerol by rat liver rough microsomes as visualized by electron microscopic autoradiography: Relationship to GTP-stimulated membrane fusion

    Following conditions of incubation for the analysis of liponucleotide synthesis, we compared GTP-dependent formation of CDP-diacylglycerol (CDP-DG) and membrane fusion in RNA-depleted rough microsomes from rat liver. After incubation of stripped rough microsomes (SRM) in the presence of GTP and [5-3H]-CTP, radioactivity was recovered in lipid extracts and identified by thin-layer chromatography as a single spot which co-migrated with CDP-DG. The nucleotide requirement for CDP-DG synthesis and that for membrane fusion were observed to be identical. We next carried out an electron microscopic autoradiographic analysis on incubated membranes to determine the site of incorporation of [5-3H]-CTP. Silver grains were observed directly over the unilamellar membranes of natural vesicles. In confirmation of the biochemical data, quantitation of silver grain density indicated more grains over membranes incubated in the presence of GTP than over those incubated in the absence of this nucleotide. For membranes incubated in the presence of GTP, the grain density was similar over fused and unfused membranes in the same preparation. When SRM were incubated with the enzyme co-factors required for synthesis of phosphatidylinositol, a GTP-independent membrane fusion was observed by both transmission and freeze-fracture electron microscopy. Together with the biochemical and autoradiographic data, this suggests that phospholipid metabolism may be activated by GTP and lead to the fusion of RER membrane

  16. Geometry of the Contact Zone between Fused Membrane-Coated Beads Mimicking Cell-Cell Fusion.

    Savić, Filip; Kliesch, Torben-Tobias; Verbeek, Sarah; Bao, Chunxiao; Thiart, Jan; Kros, Alexander; Geil, Burkhard; Janshoff, Andreas

    2016-05-24

    The fusion of lipid membranes is a key process in biology. It enables cells and organelles to exchange molecules with their surroundings, which otherwise could not cross the membrane barrier. To study such complex processes we use simplified artificial model systems, i.e., an optical fusion assay based on membrane-coated glass spheres. We present a technique to analyze membrane-membrane interactions in a large ensemble of particles. Detailed information on the geometry of the fusion stalk of fully fused membranes is obtained by studying the diffusional lipid dynamics with fluorescence recovery after photobleaching experiments. A small contact zone is a strong obstruction for the particle exchange across the fusion spot. With the aid of computer simulations, fluorescence-recovery-after-photobleaching recovery times of both fused and single-membrane-coated beads allow us to estimate the size of the contact zones between two membrane-coated beads. Minimizing delamination and bending energy leads to minimal angles close to those geometrically allowed. PMID:27224487

  17. HIV Fusion Peptide Penetrates, Disorders, and Softens T-Cell Membrane Mimics

    Tristram-Nagle, Stephanie; Chan, Rob; Kooijman, Edgar; Uppamoochikkal, Pradeep; Qiang, Wei; Weliky, David P.; Nagle, John F.

    2010-01-01

    This work investigates the interaction of N-terminal gp41 fusion peptide (FP) of human immunodeficiency virus type 1 (HIV-1) with model membranes in order to elucidate how FP leads to fusion of HIV and T-cell membranes. FP constructs were (i) wild-type FP23 (23 N-terminal amino acids of gp41), (ii) water-soluble monomeric FP that adds six lysines on the C-terminus of FP23 (FPwsm), and (iii) the C-terminus covalently linked trimeric version (FPtri) of FPwsm. Model membranes were (i) LM3 (a T-c...

  18. European Activation File for fusion

    This paper describes the work performed to revise and extend the European Activation File (EAF). Extensions, revisions and improvements made in the first two versions of the file (EAF-1, EAF-2) are discussed as well as the work which is planned for EAF-3. The EAF-2 file contains cross-sections for all stable and unstable nuclides with half-lifes exceeding 0.5 day. Cross-sections leading to isomeric states are given separately and those isomeric states with T1/2>0/5 day are included as targets. The EAF-2 version contains about 11000 reactions on 667 targets. The EAF-2 data file will be further extended and improved and will be supplied with uncertainty information. (author). 30 refs.; 5 figs.; 4 tabs

  19. How Membrane-Active Peptides Get into Lipid Membranes.

    Sani, Marc-Antoine; Separovic, Frances

    2016-06-21

    The structure-function relationship for a family of antimicrobial peptides (AMPs) from the skin of Australian tree frogs is discussed and compared with that of peptide toxins from bee and Australian scorpion venoms. Although these membrane-active peptides induce a similar cellular fate by disrupting the lipid bilayer integrity, their lytic activity is achieved via different modes of action, which are investigated in relation to amino acid sequence, secondary structure, and membrane lipid composition. In order to better understand what structural features govern the interaction between peptides and lipid membranes, cell-penetrating peptides (CPPs), which translocate through the membrane without compromising its integrity, are also discussed. AMPs possess membrane lytic activities that are naturally designed to target the cellular membrane of pathogens or competitors. They are extremely diverse in amino acid composition and often show specificity against a particular strain of microbe. Since our antibiotic arsenal is declining precariously in the face of the rise in multiantibiotic resistance, AMPs increasingly are seen as a promising alternative. In an effort to understand their molecular mechanism, biophysical studies of a myriad of AMPs have been reported, yet no unifying mechanism has emerged, rendering difficult the rational design of drug leads. Similarly, a wide variety of cytotoxic peptides are found in venoms, the best known being melittin, yet again, predicting their activity based on a particular amino acid composition or secondary structure remains elusive. A common feature of these membrane-active peptides is their preference for the lipid environment. Indeed, they are mainly unstructured in solution and, in the presence of lipid membranes, quickly adsorb onto the surface, change their secondary structure, eventually insert into the hydrophobic core of the membrane bilayer, and finally disrupt the bilayer integrity. These steps define the molecular

  20. Activation of interfacial enzymes at membrane surfaces

    Mouritsen, Ole G.; Andresen, Thomas Lars; Halperin, Avi;

    2006-01-01

    A host of water-soluble enzymes are active at membrane surfaces and in association with membranes. Some of these enzymes are involved in signalling and in modification and remodelling of the membranes. A special class of enzymes, the phospholipases, and in particular secretory phospholipase A2 (s......PLA2), are only activated at the interface between water and membrane surfaces, where they lead to a break-down of the lipid molecules into lysolipids and free fatty acids. The activation is critically dependent on the physical properties of the lipid-membrane substrate. A topical review is given of...

  1. Water activation in a fusion environment

    Water is activated in a fusion environment by the 16O(n,p)16N reaction. In this work nuclear responses in the magnets, induced by gamma rays from the activated cooling water, for the current design of the International Thermonuclear Experimental Reactor (ITER), are calculated with a detailed Monte Carlo model of the chimney region through which the cooling pipes leave the machine. It is found that, despite a significant dose, the nuclear responses induced by these gamma-rays do not pose an obvious threat to the operation of the magnets. 8 refs., 3 figs., 1 tab

  2. Activation of interfacial enzymes at membrane surfaces

    Mouritsen, Ole G.; Andresen, Thomas Lars; Halperin, Avi; Hansen, Per Lyngs; Jakobsen, Ask F.; Bernchou Jensen, Uffe; Jensen, Morten Ø.; Jørgensen, Kent; Kaasgaard, Thomas; Leidy, Chad; Simonsen, Adam Cohen; Peters, Günther H.J.; Weiss, Matthias

    2006-01-01

    A host of water-soluble enzymes are active at membrane surfaces and in association with membranes. Some of these enzymes are involved in signalling and in modification and remodelling of the membranes. A special class of enzymes, the phospholipases, and in particular secretory phospholipase A2 (s...

  3. Engineering hybrid exosomes by membrane fusion with liposomes

    Sato, Yuko T.; Kaori Umezaki; Shinichi Sawada; Sada-atsu Mukai; Yoshihiro Sasaki; Naozumi Harada; Hiroshi Shiku; Kazunari Akiyoshi

    2016-01-01

    Exosomes are a valuable biomaterial for the development of novel nanocarriers as functionally advanced drug delivery systems. To control and modify the performance of exosomal nanocarriers, we developed hybrid exosomes by fusing their membranes with liposomes using the freeze–thaw method. Exosomes embedded with a specific membrane protein isolated from genetically modified cells were fused with various liposomes, confirming that membrane engineering methods can be combined with genetic modifi...

  4. Susceptibility to virus-cell fusion at the plasma membrane is reduced through expression of HIV gp41 cytoplasmic domains

    The cytoplasmic tail of the HIV transmembrane protein plays an important role in viral infection. In this study we analyzed the role of retroviral cytoplasmic tails in modulating the cytoskeleton and interfering with virus-cell fusion. HeLaP4 cells expressing different HIV cytoplasmic tail constructs showed reduced acetylated tubulin levels whereas the cytoplasmic tail of MLV did not alter microtubule stability indicating a unique function for the lentiviral cytoplasmic tail. The effect on tubulin is mediated through the membrane proximal region of the HIV cytoplasmic tail and was independent of membrane localization. Site-directed mutagenesis identified three motifs in the HIV-2 cytoplasmic tail required to effect the reduction in acetylated tubulin. Both the YxxΦ domain and amino acids 21 to 45 of the HIV-2 cytoplasmic tail need to be present to change the level of acetylated tubulin in transfected cells. T-cells stably expressing one HIV-2 cytoplasmic tail derived construct showed also a reduction in acetylated tubulin thus confirming the importance of this effect not only for HeLaP4 and 293T cells. Challenge experiments using transiently transfected HeLaP4 cells and T cells stably expressing an HIV cytoplasmic tail construct revealed both reduced virus-cell fusion and replication of HIV-1NL4.3 compared to control cells. In the virus-cell fusion assay only virions pseudotyped with either HIV or MLV envelopes showed reduced fusion efficiency, whereas VSV-G pseudotyped virions where not affected by the expression of HIV derived cytoplasmic tail constructs, indicating that fusion at the plasma but not endosomal membrane is affected. Overexpression of human histone-deacetylase 6 (HDAC6) and constitutively active RhoA resulted in a reduction of acetylated tubulin and reduced virus-cell fusion as significant as that observed following expression of HIV cytoplasmic tail constructs. Inhibition of HDAC6 showed a strong increase in acetylated tubulin and increase of

  5. The fusion-related hydrophobic domain of Sendai F protein can be moved through the cytoplasmic membrane of Escherichia coli.

    Davis, N G; Hsu, M. C.

    1986-01-01

    Recent work on a prokaryotic membrane protein, gene III protein (pIII) of coliphage f1, showed that polypeptide segments of sufficient hydrophobicity functioned to stop transfer of the polypeptide across the cell membrane: strings of 16 or more hydrophobic amino acids sufficed. A fusion-related hydrophobic domain (FRHD) of Sendai F protein, a sequence of 26 consecutive uncharged residues, has been implicated in the fusion of the viral membrane envelope and the target-cell membrane through a h...

  6. Activation analyses for different fusion structural alloys

    The leading candidate structural materials, viz., the vanadium alloys, the nickel or the manganese stabilized austenitic steels, and the ferritic steels, are analysed in terms of their induced activation in the TPSS fusion power reactor. The TPSS reactor has 1950 MW fusion power and inboard and outboard average neutron wall loading of 3.75 and 5.35 MW/m2 respectively. The results shows that, after one year of continuous operation, the vanadium alloys have the least radioactivity at reactor shutdown. The maximum difference between the induced radioactivity in the vanadium alloys and in the other iron-based alloys occurs at about 10 years after reactor shutdown. At this time, the total reactor radioactivity, using the vanadium alloys, is about two orders of magnitude less than the total reactor radioactivity utilizing any other alloy. The difference is even larger in the first wall, the FW-vanadium activation is 3 orders of magnitude less than other alloys' FW activation. 2 refs., 7 figs

  7. Expression and purification of Tat-GFP fusion protein and its cell membrane penetrating activity%Tat-GFP融合蛋白的表达纯化及其穿膜活性

    关新刚; 苏维恒; 于欣; 佟海滨; 孙新

    2014-01-01

    Objective To obtain the Tat-GFP fusion proteins with penetrating activity and labeled with green fluorescence protein (GFP), and to explore the cell membrane penetrating activity of Tat-GFP in MCF-7 cells. Methods The plasmid pET-24a-Tat-GFP was transformed into Escherichia coli BL21 cells. Different concentrations (0.5 and 1.0 mmol · L-1 ) of isopropyl-β-D-thiogalactopyranoside (IPTG ) and cell culture temperatures (22℃ and 37℃)were used to optimize the protein expression.The Tat-GFP proteins in supernatant were purified using Ni-IDA resins. Western blotting analysis was used to identify the Tat-GFP protein, and confocal laser scanning microscope (CLSM ) was used to examine the cell penetration of Tat-GFP protein. Results There was no significant difference in the Tat-GFP protein production induced by 0.5 and 1.0 mmol·L-1 IPTG;however,the low temperature (22℃)-induced BL21 cells expressed more Tat-GFP proteins than that at 37℃ induction.The Western blotting analysis results showed that GFP antibody could specifically recognize the proteins in PVDF membranes in dose-dependent manner;the CLSM results indicated the distribution of green fluorescence in cytoplasm and nucleus of MCF-7 cells.Conclusion The Tat-GFP protein highly expresses in the supenatant of Escherichia coli i BL2 1 cells at low temperature;the obtained Tat-GFP protein with green fluorescence preserves the cell penetrating activity.%目的:获得具备穿膜活性与绿色荧光蛋白(GFP)标记的 Tat-GFP 融合蛋白,探讨 Tat-GFP 在MCF-7细胞中的跨膜转运特性。方法:应用 pET-24a-Tat-GFP质粒转化大肠杆菌 BL21感受态细胞,检测不同异丙基硫代半乳糖苷(IPTG)浓度(0.5和1.0 mmol·L-1)和不同温度(22℃和37℃)诱导融合蛋白的表达情况;利用 Ni-IDA树脂亲和纯化Tat-GFP蛋白,利用 GFP特异性抗体采用 Western blotting法分析洗脱液中的蛋白;激光共聚焦荧光显微镜下检测Tat-GFP融合蛋白

  8. Engineering hybrid exosomes by membrane fusion with liposomes

    Sato, Yuko T.; Umezaki, Kaori; Sawada, Shinichi; Mukai, Sada-atsu; Sasaki, Yoshihiro; Harada, Naozumi; Shiku, Hiroshi; Akiyoshi, Kazunari

    2016-01-01

    Exosomes are a valuable biomaterial for the development of novel nanocarriers as functionally advanced drug delivery systems. To control and modify the performance of exosomal nanocarriers, we developed hybrid exosomes by fusing their membranes with liposomes using the freeze–thaw method. Exosomes embedded with a specific membrane protein isolated from genetically modified cells were fused with various liposomes, confirming that membrane engineering methods can be combined with genetic modification techniques. Cellular uptake studies performed using the hybrid exosomes revealed that the interactions between the developed exosomes and cells could be modified by changing the lipid composition or the properties of the exogenous lipids. These results suggest that the membrane-engineering approach reported here offers a new strategy for developing rationally designed exosomes as hybrid nanocarriers for use in advanced drug delivery systems. PMID:26911358

  9. Canadian Fusion Fuels Technology Project activities report

    The Canadian Fusion Fuels Technology Project was formally established in 1982. The project is directed toward the further development of Canadian capabilities in five major areas: tritium technology, breeder technology, materials technology, equipment development and safety and the environment. The project is funded by three partners - Government of Canada (50%), Ontario Provincial Government (25%) and Ontario Hydro (25%). The fiscal year 1984/85 represents the third year of operation of the project. In 1984/85, 108 contracts were awarded totalling $4 million. Supplementary funding by subcontractors added approximately $1.9 million to the total project value. More than 200 people participated in the technical work involved in the project. Sixteen people were on attachment to foreign facilities for terms ranging from 1 month to 2.5 years. Five patents were applied for including a tritium discrimination monitor, a new radio-chemical tritium separation method, a new variation of fuel cleanup by gas chromatography, a passive tritium permeation system using bimetallic membranes, and a new breeder process using lithium salts dissolved in heavy water

  10. Lysosome fusion to the cell membrane is mediated by the dysferlin C2A domain in coronary arterial endothelial cells

    Han, Wei-Qing; Xia, Min; Xu, Ming; Krishna M Boini; Ritter, Joseph K.; Li, Ning-Jun; Li, Pin-Lan

    2012-01-01

    Dysferlin has recently been reported to participate in cell membrane repair in muscle and other cells through lysosome fusion. Given that lysosome fusion is a crucial mechanism that leads to membrane raft clustering, the present study attempted to determine whether dysferlin is involved in this process and its related signalling, and explores the mechanism underlying dysferlin-mediated lysosome fusion in bovine coronary arterial endothelial cells (CAECs). We found that dysferlin is clustered ...

  11. Reconstitution of Membrane Proteins into Model Membranes: Seeking Better Ways to Retain Protein Activities

    Trevor Lithgow; Lisa Martin; Hsin-Hui Shen

    2013-01-01

    The function of any given biological membrane is determined largely by the specific set of integral membrane proteins embedded in it, and the peripheral membrane proteins attached to the membrane surface. The activity of these proteins, in turn, can be modulated by the phospholipid composition of the membrane. The reconstitution of membrane proteins into a model membrane allows investigation of individual features and activities of a given cell membrane component. However, the activity of mem...

  12. Membrane Requirement for Folding of the Herpes Simplex Virus 1 gB Cytodomain Suggests a Unique Mechanism of Fusion Regulation

    Silverman, Jessica L.; Greene, Neil G.; King, David S.; Heldwein, Ekaterina E.

    2012-01-01

    Herpes simplex virus type 1 (HSV-1) enters cells by fusion of its envelope with a host cell membrane, which requires four viral glycoproteins and a cellular receptor. Viral fusion glycoprotein B (gB) mediates membrane fusion through the action of its ectodomain, while its cytoplasmic domain (cytodomain) regulates fusion from the opposite face of the membrane by an unknown mechanism. The gB cytodomain appears to restrict fusion, because point or truncation mutations within it increase the exte...

  13. A Programmable DNA Origami Platform to Organize SNAREs for Membrane Fusion.

    Xu, Weiming; Nathwani, Bhavik; Lin, Chenxiang; Wang, Jing; Karatekin, Erdem; Pincet, Frederic; Shih, William; Rothman, James E

    2016-04-01

    Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes are the core molecular machinery of membrane fusion, a fundamental process that drives inter- and intracellular communication and trafficking. One of the questions that remains controversial has been whether and how SNAREs cooperate. Here we show the use of self-assembled DNA-nanostructure rings to template uniform-sized small unilamellar vesicles containing predetermined maximal number of externally facing SNAREs to study the membrane-fusion process. We also incorporated lipid-conjugated complementary ssDNA as tethers into vesicle and target membranes, which enabled bypass of the rate-limiting docking step of fusion reactions and allowed direct observation of individual membrane-fusion events at SNARE densities as low as one pair per vesicle. With this platform, we confirmed at the single event level that, after docking of the templated-SUVs to supported lipid bilayers (SBL), one to two pairs of SNAREs are sufficient to drive fast lipid mixing. Modularity and programmability of this platform makes it readily amenable to studying more complicated systems where auxiliary proteins are involved. PMID:26938705

  14. Structure and membrane interaction of the internal fusion peptide of avian sarcoma leukosis virus.

    Cheng, Shu-Fang; Wu, Cheng-Wei; Kantchev, Eric Assen B; Chang, Ding-Kwo

    2004-12-01

    The structure and membrane interaction of the internal fusion peptide (IFP) fragment of the avian sarcoma and leucosis virus (ASLV) envelope glycoprotein was studied by an array of biophysical methods. The peptide was found to induce lipid mixing of vesicles more strongly than the fusion peptide derived from the N-terminal fusion peptide of influenza virus (HA2-FP). It was observed that the helical structure was enhanced in association with the model membranes, particularly in the N-terminal portion of the peptide. According to the infrared study, the peptide inserted into the membrane in an oblique orientation, but less deeply than the influenza HA2-FP. Analysis of NMR data in sodium dodecyl sulfate micelle suspension revealed that Pro13 of the peptide was located near the micelle-water interface. A type II beta-turn was deduced from NMR data for the peptide in aqueous medium, demonstrating a conformational flexibility of the IFP in analogy to the N-terminal FP such as that of gp41. A loose and multimodal self-assembly was deduced from the rhodamine fluorescence self-quenching experiments for the peptide bound to the membrane bilayer. Oligomerization of the peptide and its variants can also be observed in the electrophoretic experiments, suggesting a property in common with other N-terminal FP of class I fusion proteins. PMID:15606759

  15. Membrane support of accelerated fuel capsules for inertial fusion energy reactors

    The use of a thin membrane to suspend an (inertial fusion energy) fuel capsule in a holder for injection into a reactor chamber is investigated. Capsule displacement and membrane deformation angle are calculated for an axisymmetric geometry for a range of membrane strain and capsule size. This information is used to calculate maximum target accelerations. Membranes must be thin (perhaps of order one micron) to minimize their effect on capsule implosion symmetry. For example, a 5 μm thick cryogenic mylar membrane is calculated to allow 1,000 m/s2 acceleration of a 3 mm radius, 100 mg capsule. Vibration analysis (for a single membrane support) shows that if membrane vibration is not deliberately minimized, allowed acceleration may be reduced by a factor of four. A two membrane alternative geometry would allow several times greater acceleration. Therefore, alternative membrane geometry's should be used to provide greater target acceleration potential and reduce capsule displacement within the holder (for a given membrane thickness)

  16. Coarse-grained molecular dynamics study of membrane fusion: Curvature effects on free energy barriers along the stalk mechanism

    The effects of membrane curvature on the free energy barrier for membrane fusion have been investigated using coarse-grained molecular dynamics (CG-MD) simulations, assuming that fusion takes place through a stalk intermediate. Free energy barriers were estimated for stalk formation as well as for fusion pore formation using the guiding potential method. Specifically, the three different geometries of two apposed membranes were considered: vesicle–vesicle, vesicle–planar, and planar–planar membranes. The free energy barriers for the resulting fusion were found to depend importantly on the fusing membrane geometries; the lowest barrier was obtained for vesicular membranes. Further, lipid sorting was observed in fusion of the mixed membranes of dimyristoyl phosphatidylcholine and dioleoyl phosphatidylethanolamine (DOPE). Specifically, DOPE molecules were found to assemble around the stalk to support the highly negative curved membrane surface. A consistent result for lipid sorting was observed when a simple continuum model (CM) was used, where the Helfrich energy and mixing entropy of the lipids were taken into account. However, the CM predicts a much higher free energy barrier than found using CG-MD. This discrepancy originates from the conformational changes of lipids, which were not considered in the CM. The results of the CG-MD simulations reveal that a large conformational change in the lipid takes place around the stalk region, which results in a reduction of free energy barriers along the stalk mechanism of membrane fusion

  17. Identification of a Small Molecular Anti - HIV - 1 Compound that Interferes with Formation of the Fusion - active gp41 Core

    2001-01-01

    The human immunodeficiency virus type 1 (HIV - 1 ) envelope glycoprotein gp41 plays a critical role in the fusion of viral and target cell membranes. The gp41 extracellular domain, which contains fusion peptide (FP), N - and C - terminal hydrophobic heptad repeats (NHR and CHR, respectively). Peptides derived from NHR and CHR regions,designated N- and C- peptides, respectively, can interact with each other to form a six - stranded coiled - coil domain, representing the fusion-active gp41 core. Our previous studies demonstrated that the C- peptides have potent inhibitory activity against HIV- 1 infection.These peptides inhibit HIV- 1 -mediated membrane fusion by binding to NHR regions for preventing the formation of fusion- active gp41 core. One of the C - peptides, T - 20, which is in the phase Ⅲ clinical trails, is expected to become the first peptide HIV fusion inhibitory drug in the near future. However, this peptide HIV fusion inhibitor lacks oral availability and is sensitive to the proteolytic digestion.Therefore, it is essential to develop small molecular non -peptide HIV fusion inhibitors having similar mechanism of action as the C- peptides. We have established an ELISA- based screening assay using a unique monoclonal antibody, NC- 1, which can specifically bind to a conformational epitope on the gp41 core domain. Using this screening assay, we have identified a small molecular anti- HIV- 1 compound,named ADS-Jl, which inhibits HIV- 1- mediated membrane fusion by blocking the interaction between the NHR and CHR regions to form the fusion - active gp41 core. This compound will be used as a lead to design and develop novel HIV fusion inhibitors as new drugs for the treatment of HIV infection and/or AIDS.

  18. Is the optimal pH for membrane fusion in host cells by avian influenza viruses related to host range and pathogenicity?

    Okamatsu, Masatoshi; Motohashi, Yurie; Hiono, Takahiro; Tamura, Tomokazu; Nagaya, Kazuki; Matsuno, Keita; Sakoda, Yoshihiro; Kida, Hiroshi

    2016-08-01

    Influenza viruses isolated from wild ducks do not replicate in chickens. This fact is not explained solely by the receptor specificity of the hemagglutinin (HA) from such viruses for target host cells. To investigate this restriction in host range, the fusion activities of HA molecules from duck and chicken influenza viruses were examined. Influenza viruses A/duck/Mongolia/54/2001 (H5N2) (Dk/MNG) and A/chicken/Ibaraki/1/2005 (H5N2) (Ck/IBR), which replicate only in their primary hosts, were used. The optimal pH for membrane fusion of Ck/IBR was 5.9, higher than that of Dk/MNG at 4.9. To assess the relationship between the optimal pH for fusion and the host range of avian influenza viruses, the optimal pH for fusion of 55 influenza virus strains isolated from ducks and chickens was examined. No correlation was found between the host range and optimal pH for membrane fusion by the viruses, and this finding applied also to the H5N1 highly pathogenic avian influenza viruses. The optimal pH for membrane fusion for avian influenza viruses was shown to not necessarily be correlated with their host range or pathogenicity in ducks and chickens. PMID:27231009

  19. Versatile membrane deformation potential of activated pacsin.

    Shih Lin Goh

    Full Text Available Endocytosis is a fundamental process in signaling and membrane trafficking. The formation of vesicles at the plasma membrane is mediated by the G protein dynamin that catalyzes the final fission step, the actin cytoskeleton, and proteins that sense or induce membrane curvature. One such protein, the F-BAR domain-containing protein pacsin, contributes to this process and has been shown to induce a spectrum of membrane morphologies, including tubules and tube constrictions in vitro. Full-length pacsin isoform 1 (pacsin-1 has reduced activity compared to its isolated F-BAR domain, implicating an inhibitory role for its C-terminal Src homology 3 (SH3 domain. Here we show that the autoinhibitory, intramolecular interactions in pacsin-1 can be released upon binding to the entire proline-rich domain (PRD of dynamin-1, resulting in potent membrane deformation activity that is distinct from the isolated F-BAR domain. Most strikingly, we observe the generation of small, homogenous vesicles with the activated protein complex under certain experimental conditions. In addition, liposomes prepared with different methods yield distinct membrane deformation morphologies of BAR domain proteins and apparent activation barriers to pacsin-1's activity. Theoretical free energy calculations suggest bimodality of the protein-membrane system as a possible source for the different outcomes, which could account for the coexistence of energetically equivalent membrane structures induced by BAR domain-containing proteins in vitro. Taken together, our results suggest a versatile role for pacsin-1 in sculpting cellular membranes that is likely dependent both on protein structure and membrane properties.

  20. Structural criteria for regulation of membrane fusion and virion incorporation by the murine leukemia virus TM cytoplasmic domain

    The cytoplasmic domains of viral glycoproteins influence the trafficking and subcellular localization of the glycoproteins and their incorporation into virions. They also promote correct virus morphology and viral budding. The cytoplasmic domains of murine-leukemia-virus envelope-protein TM subunits regulate membrane fusion. During virion maturation the carboxy-terminal 16 amino acid residues of the TM protein are removed by the retroviral protease. Deletion of these residues activates envelope-protein-mediated membrane fusion. Our quantitative analysis of the effects of Moloney murine leukemia virus TM mutations on envelope-protein function support the proposition that a trimeric coiled coil in the TM cytoplasmic domain inhibits fusion. The data demonstrate that cleavage of the TM cytoplasmic domain is not required for viral entry and provide evidence for a model in which fusogenic and nonfusogenic conformations of the envelope protein exists in an equilibrium that is regulated by the cytoplasmic domain. In addition, a conserved tyrosine residue in the TM cytoplasmic domain was shown to play an important role in envelope-protein incorporation into retroviral particles

  1. The fusion of membranes and vesicles: pathway and energy barriers from Dissipative Particle Dynamics

    Shillcock, Julian C.

    2009-01-01

    The fusion of lipid bilayers is studied with dissipative particle dynamics simulations. First, to achieve control over membrane properties, the effects of individual simulation parameters are studied and optimized. Then, a large number of fusion events for a vesicle and a planar bilayer...... measure the average work for interbilayer flips of a lipid tail, i.e., the average work to displace one lipid tail from one bilayer to the other. This energy barrier is found to depend strongly on a certain dissipative particle dynamics parameter, and, thus, can be adjusted in the simulations. Overall...

  2. Accelerator and Fusion Research Division: Summary of activities, 1986

    1987-04-15

    This report contains a summary of activities at the Lawrence Berkeley Laboratory's Accelerator and Fusion Research Division for the year 1986. Topics and facilities investigated in individual papers are: 1-2 GeV Synchrotron Radiation Source, the Center for X-Ray Optics, Accelerator Operations, High-Energy Physics Technology, Heavy-Ion Fusion Accelerator Research and Magnetic Fusion Energy. Six individual papers have been indexed separately. (LSP)

  3. Accelerator and Fusion Research Division: Summary of activities, 1986

    This report contains a summary of activities at the Lawrence Berkeley Laboratory's Accelerator and Fusion Research Division for the year 1986. Topics and facilities investigated in individual papers are: 1-2 GeV Synchrotron Radiation Source, the Center for X-Ray Optics, Accelerator Operations, High-Energy Physics Technology, Heavy-Ion Fusion Accelerator Research and Magnetic Fusion Energy. Six individual papers have been indexed separately

  4. Activity assay of membrane transport proteins

    Hao Xie

    2008-01-01

    Membrane transport proteins are integral membrane proteins and considered as potential drug targets. Activity assay of transport proteins is essential for developing drugs to target these proteins. Major issues related to activity assessment of transport proteins include availability of transporters,transport activity of transporters, and interactions between ligands and transporters. Researchers need to consider the physiological status of proteins (bound in lipid membranes or purified), availability and specificity of substrates, and the purpose of the activity assay (screening, identifying, or comparing substrates and inhibitors) before choosing appropriate assay strategies and techniques. Transport proteins bound in vesicular membranes can be assayed for transporting substrate across membranes by means of uptake assay or entrance counterflow assay. Alternatively, transport proteins can be assayed for interactions with ligands by using techniques such as isothermal titration calorimetry, nuclear magnetic resonance spectroscopy, or surface plasmon resonance. Other methods and techniques such as fluorometry, scintillation proximity assay, electrophysiological assay, or stopped-flow assay could also be used for activity assay of transport proteins. In this paper the major strategies and techniques for activity assessment of membrane transport proteins are reviewed.

  5. Aqueous extract from a Chaga medicinal mushroom, Inonotus obliquus (higher Basidiomycetes), prevents herpes simplex virus entry through inhibition of viral-induced membrane fusion.

    Pan, Hong-Hui; Yu, Xiong-Tao; Li, Ting; Wu, Hong-Ling; Jiao, Chun-Wei; Cai, Mian-Hua; Li, Xiang-Min; Xie, Yi-Zhen; Wang, Yi; Peng, Tao

    2013-01-01

    Chaga medicinal mushroom, Inonotus obliquus, a popular prescription in traditional medicine in Europe and Asia, was used to reduce inflammation in the nasopharynx and to facilitate breathing. The aqueous extract from I. obliquus (AEIO) exhibited marked decrease in herpes simplex virus (HSV) infection (the 50% inhibitory concentration was 3.82 μg/mL in the plaque reduction assay and 12.29 μg/mL in the HSV-1/blue assay) as well as safety in Vero cells (the 50% cellular cytotoxicity was > 1 mg/mL, and selection index was > 80). Using a time course assay, effective stage analysis, and fusion inhibition assay, the mechanism of anti-HSV activity was found against the early stage of viral infection through inhibition of viral-induced membrane fusion. Therefore, AEIO could effectively prevent HSV-1 entry by acting on viral glycoproteins, leading to the prevention of membrane fusion, which is different from nucleoside analog antiherpetics. PMID:23510282

  6. Viral Membrane Fusion and Nucleocapsid Delivery into the Cytoplasm are Distinct Events in Some Flaviviruses

    Nour, Adel M.; Li, Yue; Wolenski, Joseph; Modis, Yorgo

    2013-01-01

    Author Summary Many viruses package their genetic material into a lipid envelope. In order to deliver their genome into the host-cell cytoplasm, where it can be replicated, viruses must fuse their envelope with a cellular lipid membrane. This fusion event is therefore a critical step in the entry of an enveloped virus into the cell. In this study, we used various cell biological and biochemical approaches to map precisely the cell entry pathway of two major human pathogens from the flavivirus...

  7. Association of the pr Peptides with Dengue Virus at Acidic pH Blocks Membrane Fusion

    Yu, I.-M.; Holdaway, H.A.; Chipman, P.R.; Kuhn, R.J.; Rossmann, M.G.; Chen, J.; Purdue

    2010-07-27

    Flavivirus assembles into an inert particle that requires proteolytic activation by furin to enable transmission to other hosts. We previously showed that immature virus undergoes a conformational change at low pH that renders it accessible to furin (I. M. Yu, W. Zhang, H. A. Holdaway, L. Li, V. A. Kostyuchenko, P. R. Chipman, R. J. Kuhn, M. G. Rossmann, and J. Chen, Science 319:1834-1837, 2008). Here we show, using cryoelectron microscopy, that the structure of immature dengue virus at pH 6.0 is essentially the same before and after the cleavage of prM. The structure shows that after cleavage, the proteolytic product pr remains associated with the virion at acidic pH, and that furin cleavage by itself does not induce any major conformational changes. We also show by liposome cofloatation experiments that pr retention prevents membrane insertion, suggesting that pr is present on the virion in the trans-Golgi network to protect the progeny virus from fusion within the host cell.

  8. Fusion technology activities at JET: Latest results

    Barbier, Dominique, E-mail: dominique.barbier@jet.efda.org [JET- EFDA- Close Support Unit, Culham Science Centre, Abingdon OX14 3DB (United Kingdom); CEA-DSM/IRFM, Centre de Cadarache, 13108 St Paul Lez Durance (France); Batistoni, Paola [ENEA - Fusion Technical Unit Via E. Fermi, 45 I-00044 FRASCATI, Rome (Italy); Coad, Paul [Association Euratom/CCFE, Culham Science Centre, Abingdon OX14 3DB (United Kingdom); Likonen, Jari [Association Euratom/TEKES, VTT, P.O. Box 1000, FIN-02044 VTT (Finland)

    2011-10-15

    The JET Task Force Fusion Technology (TF-FT) was launched in 2000 to use the unique capabilities, facilities and operating experience at JET to provide significant contributions to the research programme on both JET and ITER. This paper presents the most recent results obtained within the JET TF-FT programme. The Tritium (T) retention measurements have confirmed high surface but little bulk T concentrations on the MKII-SRP divertor tiles and T thermal desorption tests confirmed the necessity to reach at least 600 deg. C. From the 2007 shutdown the MKII-HD (more ITER like) divertor has revealed some slight changes in the nature of the erosion/deposition. In order to improve analysis, time resolution devices such as quartz micro-balances and rotating collectors have been located beneath the divertor for deposition and plasma physics correlations. Due to improvement of dedicated models and technologies, in situ laser techniques for detritiation and characterisation/removal have provided encouraging results on quantitative characteristics (composition, thickness, adherence, temperature) of deposited films on plasma facing components. A particular effort on temperature control of the new metallic ITER-like wall (ILW) that is presently being installed in JET has been pursued with active laser infrared thermography. JET TF-FT also contributes to the operator strategy to comply with the safety agency requirements for T management. Recent results on two major topics purification of tritiated water and development of the {sup 3}He method for the determination of the T concentration in waste drums are presented. Finally, this paper also presents some activities in preparation of the ILW for the pre-characterisation of marker tiles and the refurbishment of diagnostics for deposition characterisation.

  9. A molecular model for membrane fusion based on solution studies of an amphiphilic peptide from HIV gp41.

    Fujii, G; Horvath, S.; Woodward, S.; Eiserling, F.; Eisenberg, D.

    1992-01-01

    The mechanism of protein-mediated membrane fusion and lysis has been investigated by solution-state studies of the effects of peptides on liposomes. A peptide (SI) corresponding to a highly amphiphilic C-terminal segment from the envelope protein (gp41) of the human immunodeficiency virus (HIV) was synthesized and tested for its ability to cause lipid membranes to fuse together (fusion) or to break open (lysis). These effects were compared to those produced by the lytic and fusogenic peptide ...

  10. Two Coiled-Coil Domains of Chlamydia trachomatis IncA Affect Membrane Fusion Events during Infection

    Ronzone, Erik; Paumet, Fabienne

    2013-01-01

    Chlamydia trachomatis replicates in a parasitophorous membrane-bound compartment called an inclusion. The inclusions corrupt host vesicle trafficking networks to avoid the degradative endolysosomal pathway but promote fusion with each other in order to sustain higher bacterial loads in a process known as homotypic fusion. The Chlamydia protein IncA (Inclusion protein A) appears to play central roles in both these processes as it participates to homotypic fusion and inhibits endocytic SNARE-me...

  11. A GALA lipopeptide mediates pH- and membrane charge dependent fusion with stable giant unilamellar vesicles

    Etzerodt, Thomas P.; Trier, Sofie; Henriksen, Jonas R.; Andresen, Thomas L.

    2012-01-01

    Peptides capable of mediating fusion between lipid membranes are widely observed in nature, and have attracted considerable attention in the liposome drug delivery field. However, studies that are proving the benefit of small synthetic fusion peptides as components in drug delivery systems remain...

  12. Septins promote macropinosome maturation and traffic to the lysosome by facilitating membrane fusion.

    Dolat, Lee; Spiliotis, Elias T

    2016-08-29

    Macropinocytosis, the internalization of extracellular fluid and material by plasma membrane ruffles, is critical for antigen presentation, cell metabolism, and signaling. Macropinosomes mature through homotypic and heterotypic fusion with endosomes and ultimately merge with lysosomes. The molecular underpinnings of this clathrin-independent endocytic pathway are largely unknown. Here, we show that the filamentous septin GTPases associate preferentially with maturing macropinosomes in a phosphatidylinositol 3,5-bisphosphate-dependent manner and localize to their contact/fusion sites with macropinosomes/endosomes. Septin knockdown results in large clusters of docked macropinosomes, which persist longer and exhibit fewer fusion events. Septin depletion and overexpression down-regulates and enhances, respectively, the delivery of fluid-phase cargo to lysosomes, without affecting Rab5 and Rab7 recruitment to macropinosomes/endosomes. In vitro reconstitution assays show that fusion of macropinosomes/endosomes is abrogated by septin immunodepletion and function-blocking antibodies and is induced by recombinant septins in the absence of cytosol and polymerized actin. Thus, septins regulate fluid-phase cargo traffic to lysosomes by promoting macropinosome maturation and fusion with endosomes/lysosomes. PMID:27551056

  13. The role of fusion activity of influenza A viruses in their biological properties.

    Jakubcová, L; Hollý, J; Varečková, E

    2016-06-01

    Influenza A viruses (IAVs) cause acute respiratory infections of humans, which are repeated yearly. Human IAV infections are associated with significant morbidity and mortality and therefore they represent a serious health problem. All human IAV strains are originally derived from avian IAVs, which, after their adaptation to humans, can spread in the human population and cause pandemics with more or less severe course of the disease. Presently, however, the potential of avian IAV to infect humans and to cause the disease cannot be predicted. Many studies are therefore focused on factors influencing the virulence and pathogenicity of IAV viruses in a given host. The virus-host interaction starts by virus attachment via the envelope glycoprotein hemagglutinin (HA) to the receptors on the cell surface. In addition to receptor binding, HA mediates also the fusion of viral and endosomal membranes, which follows the virus endocytosis. The fusion potential of HA trimer, primed by proteolytic cleavage, is activated by low pH in endosomes, resulting in HA refolding into the fusion-active form. The HA conformation change is predetermined by its 3-D structure, is pH-dependent, irreversible and strain-specific. The process of fusion activation of IAV hemagglutinin is crucial for virus entry into the cell and for the ability of the virus to replicate in the host. Here we discuss the known data about the characteristics of fusion activation of HA in relation to IAV virulence and pathogenicity. PMID:27265461

  14. Inhibition of Nipah Virus Infectin In Vivo: Targeting an Early Stage of Paramyxovirus Fusion Activation during Viral Entry

    M Porotto; B Rockx; C Yokoyama; A Talekar; I DeVito; l Palermo; J Liu; R Cortese; M Lu; et al.

    2011-12-31

    In the paramyxovirus cell entry process, receptor binding triggers conformational changes in the fusion protein (F) leading to viral and cellular membrane fusion. Peptides derived from C-terminal heptad repeat (HRC) regions in F have been shown to inhibit fusion by preventing formation of the fusogenic six-helix bundle. We recently showed that the addition of a cholesterol group to HRC peptides active against Nipah virus targets these peptides to the membrane where fusion occurs, dramatically increasing their antiviral effect. In this work, we report that unlike the untagged HRC peptides, which bind to the postulated extended intermediate state bridging the viral and cell membranes, the cholesterol tagged HRC-derived peptides interact with F before the fusion peptide inserts into the target cell membrane, thus capturing an earlier stage in the F-activation process. Furthermore, we show that cholesterol tagging renders these peptides active in vivo: the cholesterol-tagged peptides cross the blood brain barrier, and effectively prevent and treat in an established animal model what would otherwise be fatal Nipah virus encephalitis. The in vivo efficacy of cholesterol-tagged peptides, and in particular their ability to penetrate the CNS, suggests that they are promising candidates for the prevention or therapy of infection by Nipah and other lethal paramyxoviruses.

  15. Revised graphs of activation data for fusion reactor applications

    Activation data are required for calculation of induced activity in a fusion reactor. This report gives in graphical form, the activation data which have been evaluated based on recent measurements and calculations, for use in the activation calculation code system THIDA-2. It shows transmutation and decay chain data, activation cross sections and decay gamma-ray emission data for 152 nuclides of interest in terms of fusion reactor design. This report is an updated and enlarged version of a similar report compiled in 1982 for the activation data of 116 nuclides, which had been shown to be extremely effective in referring the activation data and in locating and correcting inappropriate data. (author)

  16. Active membrane cholesterol as a physiological effector.

    Lange, Yvonne; Steck, Theodore L

    2016-09-01

    Sterols associate preferentially with plasma membrane sphingolipids and saturated phospholipids to form stoichiometric complexes. Cholesterol in molar excess of the capacity of these polar bilayer lipids has a high accessibility and fugacity; we call this fraction active cholesterol. This review first considers how active cholesterol serves as an upstream regulator of cellular sterol homeostasis. The mechanism appears to utilize the redistribution of active cholesterol down its diffusional gradient to the endoplasmic reticulum and mitochondria, where it binds multiple effectors and directs their feedback activity. We have also reviewed a broad literature in search of a role for active cholesterol (as opposed to bulk cholesterol or lipid domains such as rafts) in the activity of diverse membrane proteins. Several systems provide such evidence, implicating, in particular, caveolin-1, various kinds of ABC-type cholesterol transporters, solute transporters, receptors and ion channels. We suggest that this larger role for active cholesterol warrants close attention and can be tested easily. PMID:26874289

  17. Health physics aspects of activation products from fusion reactors

    A review of the activation products from fusion reactors and their attendant impacts is discussed. This includes a discussion on their production, expected inventories, and the status of metabolic data on these products

  18. Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

    Bárbara Hissa

    Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

  19. cBid, Bax and Bcl-xL exhibit opposite membrane remodeling activities.

    Bleicken, S; Hofhaus, G; Ugarte-Uribe, B; Schröder, R; García-Sáez, A J

    2016-01-01

    The proteins of the Bcl-2 family have a crucial role in mitochondrial outer membrane permeabilization during apoptosis and in the regulation of mitochondrial dynamics. Current models consider that Bax forms toroidal pores at mitochondria that are responsible for the release of cytochrome c, whereas Bcl-xL inhibits pore formation. However, how Bcl-2 proteins regulate mitochondrial fission and fusion remains poorly understood. By using a systematic analysis at the single vesicle level, we found that cBid, Bax and Bcl-xL are able to remodel membranes in different ways. cBid and Bax induced a reduction in vesicle size likely related to membrane tethering, budding and fission, besides membrane permeabilization. Moreover, they are preferentially located at highly curved membranes. In contrast, Bcl-xL not only counterbalanced pore formation but also membrane budding and fission. Our findings support a mechanism of action by which cBid and Bax induce or stabilize highly curved membranes including non-lamellar structures. This molecular activity reduces the energy for membrane remodeling, which is a necessary step in toroidal pore formation, as well as membrane fission and fusion, and provides a common mechanism that links the two main functions of Bcl-2 proteins. PMID:26913610

  20. Membrane Fusion Mediated by pH-Low-Insertion-Peptide (pHLIP)

    Daniels, Jennifer; Yao, Lan; Engelman, Donald; Andreev, Oleg; Reshetnyak, Yana

    2012-02-01

    Liposomes are traditionally used as drug delivery carriers. The major mechanism of liposome entry into cell is endocytotic. First, the endocytotic pathway of cellular entry is non-specific: the delivery of therapeutics occurs to cells in both diseased and healthy tissues. Second, liposomes are usually trapped in endosome/lysosome, which prevents delivery of therapeutics to cytoplasm. We proposed to use pHLIP (pH-Low-Insertion-Peptide) to promote selective delivery of the liposome content to cytoplasm of cancer cells. We showed that liposomes coated with PEG polymer and pHLIP peptide enhance membrane fusion in acidic environments. pHLIP promotes fusion between lipid bilayer of liposome and plasma membrane or membrane of endosome/lysosome, which results in intracellular delivery of payload. Liposomes composed of 5 % pHLIP and 5 % PEG were ideal for the delivery. Since cancer and other pathological states produce an acid extracellular environment, this allows the liposome to target diseased tissue while avoiding healthy tissue (with neutral pH in extracellular space). The work is supported by NIH grants CA133890 to OAA, DME, YRK.

  1. Comparison of activation in fission and fusion spectrum neutron beams

    The materials used in the construction of fusion reactors have to satisfy a number of criterions, one of the important being low activation due to neutron irradiation. Experimental analysis of the activation of candidate materials for the first wall is performed with the irradiation of samples in various neutron fields, frequently in the field of a fission reactor. In the present work a calculation is performed to compare the expected activation of candidate materials intended to be used for the first wall in fusion reactors with the activation of a sample of the same material in a fission reactor beam. The FISPACT code is used for activation calculations. An investigation, to what extent the results of activation in a fission spectrum neutron beam, where most neutrons have energies of less than 2 MeV, mimic the real situation in a fusion reactor with the peak neutron energy around 14 MeV, is performed. (author)

  2. Accelerator and Fusion Research Division: summary of activities, 1983

    The activities described in this summary of the Accelerator and Fusion Research Division are diverse, yet united by a common theme: it is our purpose to explore technologically advanced techniques for the production, acceleration, or transport of high-energy beams. These beams may be the heavy ions of interest in nuclear science, medical research, and heavy-ion inertial-confinement fusion; they may be beams of deuterium and hydrogen atoms, used to heat and confine plasmas in magnetic fusion experiments; they may be ultrahigh-energy protons for the next high-energy hadron collider; or they may be high-brilliance, highly coherent, picosecond pulses of synchrotron radiation

  3. Study on a multi-component palladium alloy membrane for the fusion fuel cleanup system

    Demonstration Tests with (D,T)2 gas to examine the reported hydrogen embrittlement and helium damage on Pd and Pd-Ag binary alloy are needed for a palladium alloy membrane for its application to a fusion fuel system. T2-gas circulating and T2-gas immersion tests with a multi-component palladium alloy, which had been selected for use of tritum purification, have been performed in the Tritium Systems Test Assembly(TSTA) at Los Alamos National Laboratory under the Japan/US Fusion Cooperation Program. Mechanical tensile tests and metallographic studies have been conducted in these durability tests. Similar tests had been performed on the same material under tritium-free atmospheres(H2, N2) to analyse the data obtained by the T2-gas tests. This report describes the results of the mechanical tensile tests and the test conditions. (author)

  4. Ebola virus glycoprotein needs an additional trigger, beyond proteolytic priming for membrane fusion.

    Shridhar Bale

    2011-11-01

    Full Text Available BACKGROUND: Ebolavirus belongs to the family filoviridae and causes severe hemorrhagic fever in humans with 50-90% lethality. Detailed understanding of how the viruses attach to and enter new host cells is critical to development of medical interventions. The virus displays a trimeric glycoprotein (GP(1,2 on its surface that is solely responsible for membrane attachment, virus internalization and fusion. GP(1,2 is expressed as a single peptide and is cleaved by furin in the host cells to yield two disulphide-linked fragments termed GP1 and GP2 that remain associated in a GP(1,2 trimeric, viral surface spike. After entry into host endosomes, GP(1,2 is enzymatically cleaved by endosomal cathepsins B and L, a necessary step in infection. However, the functional effects of the cleavage on the glycoprotein are unknown. PRINCIPAL FINDINGS: We demonstrate by antibody binding and Hydrogen-Deuterium Exchange Mass Spectrometry (DXMS of glycoproteins from two different ebolaviruses that although enzymatic priming of GP(1,2 is required for fusion, the priming itself does not initiate the required conformational changes in the ectodomain of GP(1,2. Further, ELISA binding data of primed GP(1,2 to conformational antibody KZ52 suggests that the low pH inside the endosomes also does not trigger dissociation of GP1 from GP2 to effect membrane fusion. SIGNIFICANCE: The results reveal that the ebolavirus GP(1,2 ectodomain remains in the prefusion conformation upon enzymatic cleavage in low pH and removal of the glycan cap. The results also suggest that an additional endosomal trigger is necessary to induce the conformational changes in GP(1,2 and effect fusion. Identification of this trigger will provide further mechanistic insights into ebolavirus infection.

  5. Chemical studies of viral entry mechanisms: I. Hydrophobic protein-lipid interactions during Sendai virus membrane fusion. II. Kinetics of bacteriophage λ DNA injection

    Sendai virus glycoprotein interactions with target membranes during the early stages of fusion were examined using time-resolved hydrophobic photoaffinity labeling with the lipid-soluble carbene generator 3-(trifluoromethyl)-3-(m[125I] iodophenyl)diazirine. During Sendai virus fusion with liposomes composed of cardiolipin or phosphatidylserine, the viral fusion (F) protein is preferentially labeled at early time points, supporting the hypothesis that hydrophobic interaction of the fusion peptide at the N-terminus of the F1 subunit with the target membrane is an initiating event in fusion. Correlation of hydrophobic interactions with independently monitored fusion kinetics further supports this conclusion. The F1 subunit, containing the putative hydrophobic fusion sequence, is exclusively labeled, and the F2 subunit does not participate in fusion. Labeling shows temperature and pH dependence consistent with a need for protein conformational mobility and fusion at neutral pH. Higher amounts of labeling during fusion with CL vesicles than during virus-PS vesicle fusion reflects membrane packing regulation of peptide insertion into target membranes. Labeling of the viral hemagglutinin/neuraminidase (HN) at low pH indicates that HN-mediated fusion is triggered by hydrophobic interactions. Controls for diffusional labeling exclude a major contribution from this source. Labeling during reconstituted Sendai virus envelope-liposome fusion shows that functional reconstitution involves protein retention of the ability to undergo hydrophobic interactions. Examination of Sendai virus fusion with erythrocyte membranes indicates that hydrophobic interactions also trigger fusion between biological membranes. The data show that hydrophobic fusion protein interaction with both artificial and biological membranes is a triggering event in fusion

  6. A role for sperm surface protein disulfide isomerase activity in gamete fusion: evidence for the participation of ERp57.

    Ellerman, Diego A; Myles, Diana G; Primakoff, Paul

    2006-06-01

    In mammals, sperm-egg interaction is based on molecular events either unique to gametes or also present in somatic cells. In gamete fusion, it is unknown which features are gamete specific and which are shared with other systems. Conformational changes mediated by thiol-disulfide exchange are involved in the activation of some virus membrane fusion proteins. Here we asked whether that mechanism is also operative in sperm-egg fusion. Different inhibitors of protein disulfide isomerase (PDI) activity were able to inhibit sperm-egg fusion in vitro. While pretreatment of oocytes had no effect, pretreatment of sperm reduced their fusion ability. Some members of the PDI family were detected on the sperm head, and use of specific antibodies and substrates suggested that the oxidoreductase ERp57 has a role in gamete fusion. The results support the idea that thiol-disulfide exchange is a mechanism that may act in gamete fusion to produce conformational changes in fusion-active proteins. PMID:16740484

  7. Fusion alpha loss diagnostic for ITER using activation technique

    Bonheure, G.; Hult, M.; González de Orduña, R.; Vermaercke, P.; Murari, A.; Popovichev, S.; Mlynář, Jan

    2011-01-01

    Roč. 86, 6-8 (2011), s. 1298-1301. ISSN 0920-3796. [Symposium on Fusion Technology (SOFT) /26th./. Porto, 27.09.2010-01.10.2010] Institutional research plan: CEZ:AV0Z20430508 Keywords : ITER * fusion product * burning plasma diagnostics * alpha losses * activation technique Subject RIV: BL - Plasma and Gas Discharge Physics Impact factor: 1.490, year: 2011 http://www.sciencedirect.com/science/article/pii/S0920379611002778

  8. SNARE Molecules in Marchantia polymorpha: Unique and Conserved Features of the Membrane Fusion Machinery.

    Kanazawa, Takehiko; Era, Atsuko; Minamino, Naoki; Shikano, Yu; Fujimoto, Masaru; Uemura, Tomohiro; Nishihama, Ryuichi; Yamato, Katsuyuki T; Ishizaki, Kimitsune; Nishiyama, Tomoaki; Kohchi, Takayuki; Nakano, Akihiko; Ueda, Takashi

    2016-02-01

    The membrane trafficking pathway has been diversified in a specific way for each eukaryotic lineage, probably to fulfill specific functions in the organisms. In green plants, comparative genomics has supported the possibility that terrestrialization and/or multicellularization could be associated with the elaboration and diversification of membrane trafficking pathways, which have been accomplished by an expansion of the numbers of genes required for machinery components of membrane trafficking, including soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins. However, information regarding membrane trafficking pathways in basal land plant lineages remains limited. In the present study, we conducted extensive analyses of SNARE molecules, which mediate membrane fusion between target membranes and transport vesicles or donor organelles, in the liverwort, Marchantia polymorpha. The M. polymorpha genome contained at least 34 genes for 36 SNARE proteins, comprising fundamental sets of SNARE proteins that are shared among land plant lineages with low degrees of redundancy. We examined the subcellular distribution of a major portion of these SNARE proteins by expressing Citrine-tagged SNARE proteins in M. polymorpha, and the results showed that some of the SNARE proteins were targeted to different compartments from their orthologous products in Arabidopsis thaliana. For example, MpSYP12B was localized to the surface of the oil body, which is a unique organelle in liverworts. Furthermore, we identified three VAMP72 members with distinctive structural characteristics, whose N-terminal extensions contain consensus sequences for N-myristoylation. These results suggest that M. polymorpha has acquired unique membrane trafficking pathways associated with newly acquired machinery components during evolution. PMID:26019268

  9. Preliminary design of fusion reactor fuel cleanup system by palladium alloy membrane method

    A design of palladium diffuser and Fuel Cleanup System (FCU) for D-T fusion reactor is proposed. Feasibility of palladium alloy membrane method is discussed based on the early studies by the authors. Operating conditions of the palladium diffuser are determined experimentally. Dimensions of the diffuser are estimated from computer simulation. FCU system is designed under the feed conditions of Tritium Systems Test Assembly (TSTA) at Los Alamos Scientific Laboratory. The system is composed of Pd-diffusers, catalytic oxidizer, freezer and zink beds, and has some advantages in system layout and operation. This design can readily be extended to other conditions of plasma exhaust gases. (author)

  10. Sensor Fusion for Nuclear Proliferation Activity Monitoring

    Adel Ghanem, Ph D

    2007-03-30

    The objective of Phase 1 of this STTR project is to demonstrate a Proof-of-Concept (PoC) of the Geo-Rad system that integrates a location-aware SmartTag (made by ZonTrak) and a radiation detector (developed by LLNL). It also includes the ability to transmit the collected radiation data and location information to the ZonTrak server (ZonService). The collected data is further transmitted to a central server at LLNL (the Fusion Server) to be processed in conjunction with overhead imagery to generate location estimates of nuclear proliferation and radiation sources.

  11. Rating criteria for activated waste from fusion reactors

    In the past, Shallow Land Burial (SLB) was considered one suitable solution for the management of fusion activated waste, the other being recycling. These concepts have influenced the development of low activation materials for fusion (LAMs), having reduced long-term radioactivity. Present European studies, however, do not consider SLB a viable option and concentrate more on Geological Disposal (GD). A classification of activated materials into High-, Medium-, and Low-Level Waste is proposed, taking into account contact dose rates and decay heat levels, in compliance with the GD and Recycling alternatives. This proposal is also consistent with integral criteria to classify LAMs, based on both the short- and long-term radioactive behavior. Applications of these rating criteria to activated components from various fusion designs are shown

  12. Small Mismatches in Fatty Acyl Tail Lengths Can Effect Non Steroidal Anti-Inflammatory Drug Induced Membrane Fusion.

    Majumdar, Anupa; Sarkar, Munna

    2016-06-01

    Biological membranes are made up of a variety of lipids with diverse physicochemical properties. The lipid composition modulates different lipidic parameters, such as hydration, dynamics, lipid packing, curvature strain, etc. Changes in these parameters affect various membrane-mediated processes, such as membrane fusion which is an integral step in many biological processes. Packing defects, which originate either from mismatch in the headgroup region or in the hydrophobic acyl tail region, play a major role in modulating membrane dynamics. In this study, we demonstrate how even a small mismatch in the fatty acyl chain length, achieved by incorporation of low concentrations (up to 30 mol %) of dipalmitoylphosphatidylcholine (DPPC) into dimyristoylphosphatidylcholine (DMPC) small unilamellar vesicles (SUVs), alters several lipidic parameters like packing, dynamics, and headgroup hydration. This in turn affects non steroidal anti-inflammatory drug (NSAID) induced membrane fusion. Dynamic light scattering, differential scanning calorimetry, second-derivative absorption spectrophotometry, and steady-state and time-resolved fluorescence have been used to elucidate the effect of small mismatch in the tails in DMPC/DPPC mixed vesicles and how it modulates membrane fusion induced by the oxicam NSAIDs, meloxicam (Mx), piroxicam (Px), and tenoxicam (Tx). Fusion kinetics was monitored using fluorescence based fusion assays. At low DPPC concentration of 10 mol %, additional fluidization promotes lipid mixing to some extent for Mx, but at higher mol % of DPPC, subsequent increase in rigidity of membrane interior along with increase in headgroup hydration, synergistically inhibits fusion to various extents for the three different drugs, Mx, Px, and Tx. PMID:27153337

  13. Automatically Identifying Fusion Events between GLUT4 Storage Vesicles and the Plasma Membrane in TIRF Microscopy Image Sequences

    Jian Wu

    2015-01-01

    Full Text Available Quantitative analysis of the dynamic behavior about membrane-bound secretory vesicles has proven to be important in biological research. This paper proposes a novel approach to automatically identify the elusive fusion events between VAMP2-pHluorin labeled GLUT4 storage vesicles (GSVs and the plasma membrane. The differentiation is implemented to detect the initiation of fusion events by modified forward subtraction of consecutive frames in the TIRFM image sequence. Spatially connected pixels in difference images brighter than a specified adaptive threshold are grouped into a distinct fusion spot. The vesicles are located at the intensity-weighted centroid of their fusion spots. To reveal the true in vivo nature of a fusion event, 2D Gaussian fitting for the fusion spot is used to derive the intensity-weighted centroid and the spot size during the fusion process. The fusion event and its termination can be determined according to the change of spot size. The method is evaluated on real experiment data with ground truth annotated by expert cell biologists. The evaluation results show that it can achieve relatively high accuracy comparing favorably to the manual analysis, yet at a small fraction of time.

  14. Membrane separation technologies: their application to the fusion reactor fuel cycle

    The future fusion reactor fuel will be a mixture of deuterium and tritium. Deuterium is produced using traditional separation technology. Tritium, on the contrary, must be produced by means of a nuclear reaction between neutrons and lithium atoms within the reactor breeder which could be a lithiated ceramic material or a liquid metal containing lithium. The tritium produced in the breeder needs a proper extraction process to reach the required purity level. A conceptual modified version of the tritium recovery plant for a solid ceramic breeder, working with two membranes reaction/separation units, is studied in this work. The first considered membrane unit is a catalytic ceramic membrane reactor to remove, via oxidation, the hydrogen isotopes from the purge gas (He); the second is a Pd/Ag membrane permeator to separate the hydrogen isotopes from the water shift reactor gaseous products. The modelling and the mass balances have been obtained either on the basis of data in the literature or on experimental results. (orig.)

  15. Effects of retroviral envelope-protein cleavage upon trafficking, incorporation, and membrane fusion

    Retroviral envelope glycoproteins undergo proteolytic processing by cellular subtilisin-like proprotein convertases at a polybasic amino-acid site in order to produce the two functional subunits, SU and TM. Most previous studies have indicated that envelope-protein cleavage is required for rendering the protein competent for promoting membrane fusion and for virus infectivity. We have investigated the role of proteolytic processing of the Moloney murine leukemia virus envelope-protein through site-directed mutagenesis of the residues near the SU-TM cleavage site and have established that uncleaved glycoprotein is unable either to be incorporated into virus particles efficiently or to induce membrane fusion. Additionally, the results suggest that cleavage of the envelope protein plays an important role in intracellular trafficking of protein via the cellular secretory pathway. Based on our results it was concluded that a positively charged residue located at either P2 or P4 along with the arginine at P1 is essential for cleavage.

  16. Implication des peptides de fusion des glycoprotéines de fusion virales de classe I dans la fusion membranaire

    Brasseur R.

    2007-01-01

    Full Text Available The implication of fusion peptides of class I viral fusion glycoproteins in the membrane fusion. Viral infection involves fusion between the viral envelope and the target cell plasmic membrane. The fusion is induced by a glycoprotein anchored in the viral envelope. After activation, the glycoprotein undergoes a conformational change inducing the exposure of a region named « fusion peptide » essential for the fusion process. Studies on glycoproteins and on isolated fusion peptides have allowed to better understand the mechanisms involved in membrane fusion. It was notably shown that fusion peptides induce fusion and leakage of membranes. These peptides are able to insert obliquely in a membrane when helical. This orientation induces lipid destabilisation, favouring membrane fusion. However, to date, none of these in vitro, in vivo or in silico studies has determined the minimal sequence required for membrane fusion. Using the obliquity-fusogenicity relationship, the latter was determined by molecular modelling for two viruses, the Human Immunodeficiency Virus and the Bovine Leukaemia Virus. These new results are of particular interest in the development of vaccines and antiviral drugs.

  17. Identification and characterization of LFD-2, a predicted fringe protein required for membrane integrity during cell fusion in neurospora crassa.

    Palma-Guerrero, Javier; Zhao, Jiuhai; Gonçalves, A Pedro; Starr, Trevor L; Glass, N Louise

    2015-03-01

    The molecular mechanisms of membrane merger during somatic cell fusion in eukaryotic species are poorly understood. In the filamentous fungus Neurospora crassa, somatic cell fusion occurs between genetically identical germinated asexual spores (germlings) and between hyphae to form the interconnected network characteristic of a filamentous fungal colony. In N. crassa, two proteins have been identified to function at the step of membrane fusion during somatic cell fusion: PRM1 and LFD-1. The absence of either one of these two proteins results in an increase of germling pairs arrested during cell fusion with tightly appressed plasma membranes and an increase in the frequency of cell lysis of adhered germlings. The level of cell lysis in ΔPrm1 or Δlfd-1 germlings is dependent on the extracellular calcium concentration. An available transcriptional profile data set was used to identify genes encoding predicted transmembrane proteins that showed reduced expression levels in germlings cultured in the absence of extracellular calcium. From these analyses, we identified a mutant (lfd-2, for late fusion defect-2) that showed a calcium-dependent cell lysis phenotype. lfd-2 encodes a protein with a Fringe domain and showed endoplasmic reticulum and Golgi membrane localization. The deletion of an additional gene predicted to encode a low-affinity calcium transporter, fig1, also resulted in a strain that showed a calcium-dependent cell lysis phenotype. Genetic analyses showed that LFD-2 and FIG1 likely function in separate pathways to regulate aspects of membrane merger and repair during cell fusion. PMID:25595444

  18. Fusion

    Mahaffey, James A

    2012-01-01

    As energy problems of the world grow, work toward fusion power continues at a greater pace than ever before. The topic of fusion is one that is often met with the most recognition and interest in the nuclear power arena. Written in clear and jargon-free prose, Fusion explores the big bang of creation to the blackout death of worn-out stars. A brief history of fusion research, beginning with the first tentative theories in the early 20th century, is also discussed, as well as the race for fusion power. This brand-new, full-color resource examines the various programs currently being funded or p

  19. Fusion blanket materials development and recent R and D activities

    Development of structural materials plays an important role in the feasibility of fusion power plant. The candidate structural materials for future fusion reactors are Reduced Activation Ferritic Martensitic (RAFM) steel, nano structured ODS Steel, vanadium alloys and SiC/SiCf composite etc. RAFM steel is presently considered as the structural material for Lead Lithium Ceramic Breeder (LLCB) Test Blanket Module (TBM) because of its high void swelling resistance and improved thermal properties compared to austenitic steel. Development of RAFM steel in India is being carried out in full swing in collaboration with various research laboratories and steel industries. This paper presents an overview of the Indian activities on fusion blanket materials and describes in brief the efforts made to develop IN-RAFM steel as structural material for the LLCB TBM. In future, due to enhanced properties of vanadium base alloy and nano structured materials like ODS RAFMS, RAFM steel may be replaced by these materials for its application in DEMO relevant fusion reactor. Future R and D activities will be specifically towards the development of these structural materials for fusion reactor

  20. Effect of Amphipathic HIV Fusion Inhibitor Peptides on POPC and POPC/Cholesterol Membrane Properties: A Molecular Simulation Study

    Luís M. S. Loura

    2013-07-01

    Full Text Available T-20 and T-1249 fusion inhibitor peptides were shown to interact with 1-palmitoyl-2-oleyl-phosphatidylcholine (POPC (liquid disordered, ld and POPC/cholesterol (1:1 (POPC/Chol (liquid ordered, lo bilayers, and they do so to different extents. Although they both possess a tryptophan-rich domain (TRD, T-20 lacks a pocket binding domain (PBD, which is present in T-1249. It has been postulated that the PBD domain enhances FI interaction with HIV gp41 protein and with model membranes. Interaction of these fusion inhibitor peptides with both the cell membrane and the viral envelope membrane is important for function, i.e., inhibition of the fusion process. We address this problem with a molecular dynamics approach focusing on lipid properties, trying to ascertain the consequences and the differences in the interaction of T-20 and T-1249 with ld and lo model membranes. T-20 and T-1249 interactions with model membranes are shown to have measurable and different effects on bilayer structural and dynamical parameters. T-1249’s adsorption to the membrane surface has generally a stronger influence in the measured parameters. The presence of both binding domains in T-1249 appears to be paramount to its stronger interaction, and is shown to have a definite importance in membrane properties upon peptide adsorption.

  1. Applications of superpermeable membranes in fusion: the flux density problem and experimental progress

    Livshits, A. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Ohyabu, N. [National Institute for Fusion Science, Furoh-cho, Chikusa-ku, Nagoya 464-01 (Japan); Notkin, M. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Alimov, V. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Suzuki, H. [National Institute for Fusion Science, Furoh-cho, Chikusa-ku, Nagoya 464-01 (Japan); Samartsev, A. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Solovyov, M. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Grigoriadi, I. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Glebovsky, A. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Busnyuk, A. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Doroshin, A. [Bonch-Bruyevich Univ., St. Petersburg (Russian Federation); Komatsu, K. [National Institute for Fusion Science, Furoh-cho, Chikusa-ku, Nagoya 464-01 (Japan)

    1997-02-01

    Superpermeable membranes whose permeability to energetic hydrogen approaches the permeability of an opening of the same area can be employed to separate D/T and He in fusion machine exhausts, to control the edge plasma and divertor conditions (by pumping and/or arranging of gas circulation through SOL or divertor) and to pump and recuperate D/T in auxiliary systems e.g. in pellet or neutral beam injection. One of the key points is the operation at permeation flux densities of up to 10{sup 16}-10{sup 19} cm{sup -2} s{sup -1}. Theory predicts that the highest flux densities may be reached with superpermeable membranes based on the V group metals: the limit conditioned by a maximum permissible hydrogen concentration in bulk metal is expected to be as high as {proportional_to}10{sup 19} cm{sup -2} s{sup -1}. The experimental membrane system comprised a cylindrical Nb membrane and an incandescent Ta/Nb atomizer placed inside. The hydrogen pumping speed by this system amounts to {proportional_to}10{sup 3} l/s, with a specific pumping speed of {proportional_to}1 l/s per cm{sup 2} membrane area and {proportional_to}8 l/s per cm{sup 2} atomizer area. Superpermeability was observed at record parameters referring both to the flux density of 3 x 10{sup 17} H/cm{sup 2}/s (by one order of magnitude larger than ever before) and to the operational pressure of 3 x 10{sup -2} Torr. A long-term reliable operation of this system proved being possible even in a vacuum far inferior to UHV conditions. (orig.).

  2. TRAIL-CM4 fusion protein shows in vitro antibacterial activity and a stronger antitumor activity than solo TRAIL protein.

    Sang, Ming; Zhang, Jiaxin; Li, Bin; Chen, Yuqing

    2016-06-01

    A TRAIL-CM4 fusion protein in soluble form with tumor selective apoptosis and antibacterial functions was expressed in the Escherichia coli expression system and isolated through dialysis refolding and histidine-tag Nickel-affinity purification. Fresh Jurkat cells were treated with the TRAIL-CM4 fusion protein. Trypan blue staining and MTT analyses showed that, similar to a TRAIL positive control, Jurkat cell proliferation was significantly inhibited. Flow cytometry analyses using Annexin V-fluorescein revealed that Jurkat cells treated with the TRAIL-CM4 fusion protein exhibited increased apoptosis. Laser confocal microscopy showed that APB-CM4 and the fusion protein TRAIL-CM4 can bind to Jurkat cell membranes and initiate their destruction. ABP-CM4 enhances the antitumor activity of TRAIL by targeting and damaging the tumor cell membrane. In antibacterial experiments, agar well diffusion and bacterial growth inhibition curve assays revealed concentration-dependent TRAIL-CM4 antibacterial activity against Escherichia coli K12D31. The expressed TRAIL-CM4 fusion protein exhibited enhanced antitumor and antibacterial activities. Fusion protein expression allowed the two different proteins to function in combination. PMID:26926590

  3. Line tension at lipid phase boundaries as driving force for HIV fusion peptide-mediated fusion.

    Yang, Sung-Tae; Kiessling, Volker; Tamm, Lukas K

    2016-01-01

    Lipids and proteins are organized in cellular membranes in clusters, often called 'lipid rafts'. Although raft-constituent ordered lipid domains are thought to be energetically unfavourable for membrane fusion, rafts have long been implicated in many biological fusion processes. For the case of HIV gp41-mediated membrane fusion, this apparent contradiction can be resolved by recognizing that the interfaces between ordered and disordered lipid domains are the predominant sites of fusion. Here we show that line tension at lipid domain boundaries contributes significant energy to drive gp41-fusion peptide-mediated fusion. This energy, which depends on the hydrophobic mismatch between ordered and disordered lipid domains, may contribute tens of kBT to fusion, that is, it is comparable to the energy required to form a lipid stalk intermediate. Line-active compounds such as vitamin E lower line tension in inhomogeneous membranes, thereby inhibit membrane fusion, and thus may be useful natural viral entry inhibitors. PMID:27113279

  4. Comparative study of somatostatin-human serum albumin fusion proteins and natural somatostatin on receptor binding, internalization and activation.

    Ying Peng

    Full Text Available Albumin fusion technology, the combination of small molecular proteins or peptides with human serum albumin (HSA, is an effective method for improving the medicinal values of natural small molecular proteins or peptides. However, comparative studies between HSA-fusion proteins or peptides and the parent small molecules in biological and molecular mechanisms are less reported. In this study, we examined the binding property of two novel somatostatin-HSA fusion proteins, (SST142-HSA and (SST282-HSA, to human SSTRs in stably expressing SSTR1-5 HEK 293 cells; observed the regulation of receptor internalization and internalized receptor recycling; and detected the receptors activation of HSA fusion proteins in stably expressing SSTR2- and SSTR3-EGFP cells. We showed that both somatostatin-HSA fusion proteins had high affinity to all five SSTRs, stimulated the ERK1/2 phosphorylation and persistently inhibited the accumulation of forskolin-stimulated cAMP in SSTR2- and SSTR3-expressing cells; but were less potent than the synthetic somatostatin-14 (SST-14. Our experiments also showed that somatostatin-HSA fusion proteins did not induce the receptors internalization; rather, they accelerated the recycling of the internalized receptors induced by SST-14 to the plasma membrane. Our results indicated that somatostatin-HSA fusion proteins, different from SST-14, exhibit some particular properties in binding, regulating, and activating somatostatin receptors.

  5. Charged fusion product loss measurements using nuclear activation

    In ITER, α particle loss measurements will be required in order to understand the alpha particle physics. Techniques capable of operating in a fusion reactor environment need further development. Recent experimental studies on JET demonstrated the potential of nuclear activation to measure the flux of escaping MeV ions. New results from MeV ion induced activation of metallic, ceramic, and crystal samples placed near the plasma edge are reported. Activation products were measured as function of orientation with respect to the magnetic field as well as function of the distance to the plasma. Sample activity was measured using ultralow-level gamma-ray spectrometry. Distribution of 14.68 MeV fusion proton induced activation products is strongly anisotropic in agreement with simulations and falls off sharply with increasing distance to the plasma. Prospects for using the technique in ITER are discussed.

  6. Minor differences in the molecular machinery mediating regulated membrane fusion has major impact on metabolic health.

    Valladolid-Acebes, Ismael; Daraio, Teresa; Brismar, Kerstin; Hökfelt, Tomas; Bark, Christina

    2016-01-01

    The exocytosis of signaling molecules from neuronal, neuroendocrine and endocrine cells is regulated by membrane fusion involving SNAP-25 and associated SNARE proteins. The importance of this process for metabolic control recently became evident by studies of mouse mutants genetically engineered to only express one of 2 closely related, alternatively-spliced variants of SNAP-25. The results showed that even minor differences in the function of proteins regulating exocytosis are sufficient to provoke metabolic disease, including hyperglycaemia, liver steatosis, adipocyte hypertrophy and obesity. Thus, an imbalance in the dynamics of hormonal and/or neurotransmitter release can cause obesity and type 2 diabetes. This recent discovery highlights the fact that metabolic health requires a perfectly operating interplay between the SNARE protein machinery in excitable cells and the organs responding to these messengers. PMID:27617177

  7. Early Events in Chikungunya Virus Infection-From Virus Cell Binding to Membrane Fusion.

    van Duijl-Richter, Mareike K S; Hoornweg, Tabitha E; Rodenhuis-Zybert, Izabela A; Smit, Jolanda M

    2015-07-01

    Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne alphavirus causing millions of infections in the tropical and subtropical regions of the world. CHIKV infection often leads to an acute self-limited febrile illness with debilitating myalgia and arthralgia. A potential long-term complication of CHIKV infection is severe joint pain, which can last for months to years. There are no vaccines or specific therapeutics available to prevent or treat infection. This review describes the critical steps in CHIKV cell entry. We summarize the latest studies on the virus-cell tropism, virus-receptor binding, internalization, membrane fusion and review the molecules and compounds that have been described to interfere with virus cell entry. The aim of the review is to give the reader a state-of-the-art overview on CHIKV cell entry and to provide an outlook on potential new avenues in CHIKV research. PMID:26198242

  8. Early Events in Chikungunya Virus Infection—From Virus CellBinding to Membrane Fusion

    Mareike K. S. van Duijl-Richter

    2015-07-01

    Full Text Available Chikungunya virus (CHIKV is a rapidly emerging mosquito-borne alphavirus causing millions of infections in the tropical and subtropical regions of the world. CHIKV infection often leads to an acute self-limited febrile illness with debilitating myalgia and arthralgia. A potential long-term complication of CHIKV infection is severe joint pain, which can last for months to years. There are no vaccines or specific therapeutics available to prevent or treat infection. This review describes the critical steps in CHIKV cell entry. We summarize the latest studies on the virus-cell tropism, virus-receptor binding, internalization, membrane fusion and review the molecules and compounds that have been described to interfere with virus cell entry. The aim of the review is to give the reader a state-of-the-art overview on CHIKV cell entry and to provide an outlook on potential new avenues in CHIKV research.

  9. Highly specific inhibition of leukaemia virus membrane fusion by interaction of peptide antagonists with a conserved region of the coiled coil of envelope

    van Aalten Daan MF

    2008-08-01

    Full Text Available Abstract Background Human T-cell leukaemia virus (HTLV-1 and bovine leukaemia virus (BLV entry into cells is mediated by envelope glycoprotein catalyzed membrane fusion and is achieved by folding of the transmembrane glycoprotein (TM from a rod-like pre-hairpin intermediate to a trimer-of-hairpins. For HTLV-1 and for several virus groups this process is sensitive to inhibition by peptides that mimic the C-terminal α-helical region of the trimer-of-hairpins. Results We now show that amino acids that are conserved between BLV and HTLV-1 TM tend to map to the hydrophobic groove of the central triple-stranded coiled coil and to the leash and C-terminal α-helical region (LHR of the trimer-of-hairpins. Remarkably, despite this conservation, BLV envelope was profoundly resistant to inhibition by HTLV-1-derived LHR-mimetics. Conversely, a BLV LHR-mimetic peptide antagonized BLV envelope-mediated membrane fusion but failed to inhibit HTLV-1-induced fusion. Notably, conserved leucine residues are critical to the inhibitory activity of the BLV LHR-based peptides. Homology modeling indicated that hydrophobic residues in the BLV LHR likely make direct contact with a pocket at the membrane-proximal end of the core coiled-coil and disruption of these interactions severely impaired the activity of the BLV inhibitor. Finally, the structural predictions assisted the design of a more potent antagonist of BLV membrane fusion. Conclusion A conserved region of the HTLV-1 and BLV coiled coil is a target for peptide inhibitors of envelope-mediated membrane fusion and HTLV-1 entry. Nevertheless, the LHR-based inhibitors are highly specific to the virus from which the peptide was derived. We provide a model structure for the BLV LHR and coiled coil, which will facilitate comparative analysis of leukaemia virus TM function and may provide information of value in the development of improved, therapeutically relevant, antagonists of HTLV-1 entry into cells.

  10. Identification and purification of a sperm surface protein with a potential role in sperm-egg membrane fusion.

    Primakoff, P; Hyatt, H; Tredick-Kline, J

    1987-01-01

    Sperm-egg plasma membrane fusion during fertilization was studied using guinea pig gametes and mAbs to sperm surface antigens. The mAb, PH-30, strongly inhibited sperm-egg fusion in a concentration-dependent fashion. When zona-free eggs were inseminated with acrosome-reacted sperm preincubated in saturating (140 micrograms/ml) PH-30 mAb, the percent of eggs showing fusion was reduced 75%. The average number of sperm fused per egg was also reduced by 75%. In contrast a control mAb, PH-1, preincubated with sperm at 400 micrograms/ml, caused no inhibition. The PH-30 and PH-1 mAbs apparently recognize the same antigen but bind to two different determinants. Both mAbs immunoprecipitated the same two 125I-labeled polypeptides with Mr 60,000 (60 kD) and Mr 44,000 (44 kD). Boiling a detergent extract of sperm severely reduced the binding of PH-30 but had essentially no effect on the binding of PH-1, indicating that the two mAbs recognize different epitopes. Immunoelectron microscopy revealed that PH-30 mAb binding was restricted to the sperm posterior head surface and was absent from the equatorial region. The PH-30 and PH-1 mAbs did not bind to sperm from the testis, the caput, or the corpus epididymis. PH-30 mAb binding was first detectable on sperm from the proximal cauda epididymis, i.e., sperm at the developmental stage where fertilization competence appears. After purification by mAb affinity chromatography, the PH-30 protein retained antigenic activity, binding both the PH-30 and PH-1 mAbs. The purified protein showed two polypeptide bands of 60 and 44 kD on reducing SDS PAGE. The two polypeptides migrated further (to approximately 49 kD and approximately 33 kD) on nonreducing SDS PAGE, showing that they do not contain interchain disulfide bonds, but probably have intrachain disulfides. 44 kD appears not to be a proteolytic fragment of 60 kD because V8 protease digestion patterns did not reveal related peptide patterns from the 44- and 60-kD bands. In the absence of

  11. Functional Implications of Plasma Membrane Condensation for T Cell Activation

    Rentero, Carles; Zech, Tobias; Quinn, Carmel M.; Engelhardt, Karin; Williamson, David; Grewal, Thomas; Jessup, Wendy; Harder, Thomas; Gaus, Katharina

    2008-01-01

    The T lymphocyte plasma membrane condenses at the site of activation but the functional significance of this receptor-mediated membrane reorganization is not yet known. Here we demonstrate that membrane condensation at the T cell activation sites can be inhibited by incorporation of the oxysterol 7-ketocholesterol (7KC), which is known to prevent the formation of raft-like liquid-ordered domains in model membranes. We enriched T cells with 7KC, or cholesterol as control, to assess the importa...

  12. Reassessment of the lineage fusion hypothesis for the origin of double membrane bacteria.

    Kristen S Swithers

    Full Text Available In 2009, James Lake introduced a new hypothesis in which reticulate phylogeny reconstruction is used to elucidate the origin of gram-negative bacteria (Nature 460: 967-971. The presented data supported the gram-negative bacteria originating from an ancient endosymbiosis between the Actinobacteria and Clostridia. His conclusion was based on a presence-absence analysis of protein families that divided all prokaryotes into five groups: Actinobacteria, Double Membrane bacteria (DM, Clostridia, Archaea and Bacilli. Of these five groups, the DM are by far the largest and most diverse group compared to the other groupings. While the fusion hypothesis for the origin of double membrane bacteria is enticing, we show that the signal supporting an ancient symbiosis is lost when the DM group is broken down into smaller subgroups. We conclude that the signal detected in James Lake's analysis in part results from a systematic artifact due to group size and diversity combined with low levels of horizontal gene transfer.

  13. Convenient synthesis and application of versatile nucleic acid lipid membrane anchors in the assembly and fusion of liposomes

    Ries, Oliver; Löffler, Philipp M. G.; Vogel, Stefan

    2015-01-01

    Hydrophobic moieties like lipid membrane anchors are highly demanded modifications for nucleic acid oligomers. Membrane-anchor modified oligonucleotides are applicable in biomedicine leading to new delivery strategies as well as in biophysical investigations towards assembly and fusion of liposomes...... or the construction of DNA origami structures. We herein present the synthesis and applications of versatile lipid membrane anchor building blocks suitable for solid phase oligonucleotide synthesis. These are readily synthesized in bulk in five to seven steps from commercially available precursors...... and can be incorporated at any position within an oligonucleotide without significantly altering duplex stability and structure as proven by thermal denaturation experiments and circular dichroism. Furthermore, applicability could be demonstrated by assembly and fusion of liposomes mediated by lipid...

  14. Overview of Fusion Nuclear Technology Activities at SWIP

    The fusion nuclear technology (FNT) activities at SWIP are overviewed, which are including the design and R and D of the helium-cooled solid breeder test blanket module (TBM), development of Liquid metal blanket technology, structure and function materials research for ITER shielding and test blanket, ITER shielding blanket material development and the joining technologies and the R and D for ITER gravity support.

  15. The role of blood cell membrane lipids on the mode of action of HIV-1 fusion inhibitor sifuvirtide

    Research highlights: → Sifuvirtide interacts with erythrocyte and lymphocyte membrane in a concentration dependent manner by decreasing its dipole potential. → Dipole potential variations in lipid vesicles show sifuvirtide's lipid selectivity towards saturated phosphatidylcholines. → This peptide-membrane interaction may direct the drug towards raft-like membrane domains where the receptors used by HIV are located, facilitating its inhibitory action. -- Abstract: Sifuvirtide is a gp41 based peptide that inhibits HIV-1 fusion with the host cells and is currently under clinical trials. Previous studies showed that sifuvirtide partitions preferably to saturated phosphatidylcholine lipid membranes, instead of fluid-phase lipid vesicles. We extended the study to the interaction of the peptide with circulating blood cells, by using the dipole potential sensitive probe di-8-ANEPPS. Sifuvirtide decreased the dipole potential of erythrocyte and lymphocyte membranes in a concentration dependent manner, demonstrating its interaction. Also, the lipid selectivity of the peptide towards more rigid phosphatidylcholines was confirmed based on the dipole potential variations. Overall, the interaction of the peptide with the cell membranes is a contribution of different lipid preferences that presumably directs the peptide towards raft-like domains where the receptors are located, facilitating the reach of the peptide to its molecular target, the gp41 in its pre-fusion conformation.

  16. Low pH-dependent Hepatitis C Virus Membrane Fusion Depends on E2 Integrity, Target Lipid Composition, and Density of Virus Particles*

    Haid, Sibylle; Pietschmann, Thomas; Pécheur, Eve-Isabelle

    2009-01-01

    Hepatitis C virus (HCV) is an enveloped, positive strand RNA virus of about 9.6 kb. Like all enveloped viruses, the HCV membrane fuses with the host cell membrane during the entry process and thereby releases the genome into the cytoplasm, initiating the viral replication cycle. To investigate the features of HCV membrane fusion, we developed an in vitro fusion assay using cell culture-produced HCV and fluorescently labeled liposomes. With this model we could show that HCV-mediated fusion can...

  17. A cell penetrating peptide-integrated and enediyne-energized fusion protein shows potent antitumor activity.

    Ru, Qin; Shang, Bo-Yang; Miao, Qing-Fang; Li, Liang; Wu, Shu-Ying; Gao, Rui-Juan; Zhen, Yong-Su

    2012-11-20

    Arginine-rich peptides belong to a subclass of cell penetrating peptides that are taken up by living cells and can be detected freely diffusing inside the cytoplasm and nucleoplasm. This phenomenon has been attributed to either an endocytotic mode of uptake and a subsequent release from vesicles or a direct membrane penetration. Lidamycin is an antitumor antibiotic, which consists of an active enediyne chromophore (AE) and a noncovalently bound apoprotein (LDP). In the present study, a fusion protein (Arg)(9)-LDP composed of cell penetrating peptide (Arg)(9) and LDP was prepared by DNA recombination, and the enediyne-energized fusion protein (Arg)(9)-LDP-AE was prepared by molecular reconstitution. The data in fixed cells demonstrated that (Arg)(9)-LDP could rapidly enter cells, and the results based on fluorescence activated cell sorting indicated that the major route for (Arg)(9)-mediated cellular uptake of protein molecules was endocytosis. (Arg)(9)-LDP-AE demonstrated more potent cytotoxicity against different carcinoma cell lines than lidamycin in vitro. In the mouse hepatoma 22 model, (Arg)(9)-LDP-AE (0.3mg/kg) suppressed the tumor growth by 89.2%, whereas lidamycin (0.05 mg/kg) by 74.6%. Furthermore, in the glioma U87 xenograft model in nude mice, (Arg)(9)-LDP-AE at 0.2mg/kg suppressed tumor growth by 88.8%, compared with that of lidamycin by 62.9% at 0.05 mg/kg. No obvious toxic effects were observed in all groups during treatments. The results showed that energized fusion protein (Arg)(9)-LDP-AE was more effective than lidamycin and would be a promising candidate for glioma therapy. In addition, this approach to manufacturing fusion proteins might serve as a technology platform for the development of new cell penetrating peptides-based drugs. PMID:22982402

  18. A common landscape for membrane-active peptides

    Last, Nicholas B.; Schlamadinger, Diana E.; Miranker, Andrew D.

    2013-01-01

    Three families of membrane-active peptides are commonly found in nature and are classified according to their initial apparent activity. Antimicrobial peptides are ancient components of the innate immune system and typically act by disruption of microbial membranes leading to cell death. Amyloid peptides contribute to the pathology of diverse diseases from Alzheimer's to type II diabetes. Preamyloid states of these peptides can act as toxins by binding to and permeabilizing cellular membranes...

  19. Probing plasma membrane microdomains in cowpea protoplasts using lipidated GFP-fusion proteins and multimode FRET microscopy.

    Vermeer, J.E.M.; Munster, van, B.C.; Vischer, N O; Gadella, Th.W.J.

    2004-01-01

    Multimode fluorescence resonance energy transfer (FRET) microscopy was applied to study the plasma membrane organization using different lipidated green fluorescent protein (GFP)-fusion proteins co-expressed in cowpea protoplasts. Cyan fluorescent protein (CFP) was fused to the hyper variable region of a small maize GTPase (ROP7) and yellow fluorescent protein (YFP) was fused to the N-myristoylation motif of the calcium-dependent protein kinase 1 (LeCPK1) of tomato. Upon co-expressing in cowp...

  20. delta-Opioid receptors exhibit high efficiency when activating trimeric G proteins in membrane domains.

    Bourova, Lenka; Kostrnova, Alexandra; Hejnova, Lucie; Moravcova, Zuzana; Moon, Hyo-Eun; Novotny, Jiri; Milligan, Graeme; Svoboda, Petr

    2003-04-01

    Low-density membrane fragments (domains) were separated from the bulk of plasma membranes of human embryonic kidney (HEK)293 cells expressing a delta-opioid (DOP) receptor-Gi1alpha fusion protein by drastic homogenization and flotation on equilibrium sucrose density gradients. The functional activity of trimeric G proteins and capacity of the DOP receptor to stimulate both the fusion protein-linked Gi1alpha and endogenous pertussis-toxin sensitive G proteins was measured as d-Ala2, d-Leu5-enkephalin stimulated high-affinity GTPase or guanosine-5'-[gamma-35S]triphosphate ([35S]GTPgammaS) binding. The maximum d-Ala2-d-Leu5 enkephalin (DADLE)-stimulated GTPase was two times higher in low-density membrane fragments than in bulk of plasma membranes; 58 and 27 pmol/mg/min, respectively. The same difference was obtained for [35S]GTPgammaS binding. Contrarily, the low-density domains contained no more than half the DOP receptor binding sites (Bmax = 6.6 pmol/mg versus 13.6 pmol/mg). Thus, when corrected for expression levels of the receptor, low-density domains exhibited four times higher agonist-stimulated GTPase and [35S]GTPgammaS binding than the bulk plasma membranes. The regulator of G protein signaling RGS1, enhanced further the G protein functional activity but did not remove the difference between domain-bound and plasma membrane pools of G protein. The potency of the agonist in functional studies and the affinity of specific [3H]DADLE binding to the receptor were, however, the same in both types of membranes - EC50 = 4.5 +/- 0.1 x 10(-8) and 3.2 +/- 1.4 x 10(-8) m for GTPase; Kd = 1.2 +/- 0.1 and 1.3 +/- 0.1 nm for [3H]DADLE radioligand binding assay. Similar results were obtained when sodium bicarbonate was used for alkaline isolation of membrane domains. By contrast, detergent-insensitive membrane domains isolated following treatment of cells with Triton X100 exhibited no DADLE-stimulated GTPase or GTPgammaS binding. Functional coupling between the DOP receptor

  1. Dolichol phosphate induces non-bilayer structures, vesicle fusion and transbilayer movements of lipids in model membranes

    de Kruijff, B.; Van Duijn, G.; Valtersson, C.; Chojnacki, T.; Verkleij, A.J.; Dallner, G.

    1987-05-01

    The effect of dolichols, polyprenols, dolichol esterified with fatty acids, and dolichol phosphate on the structure and fluidity of model membranes was studied using different biophysical techniques. These studies suggest that (1) dolichol and dolichol derivatives destabilize unsaturated PE-containing bilayers and promote hexagonal II phase formation; (2) high concentrations of dolichol induce lipid structures characterized by isotropic T P NMR and particulate fracture faces. The effect of dolichol and dolichyl phosphate on fusion between large unilamellar vesicles of DOPC and DOPE was studied using a fluroescence resonance energy transfer assay. The influence of dolichyl phosphate on the transbilary movement of DOPC in multilamellar vesicles (MLV) and large unilamellar vesicles (LUV) composed of DOPC and DOPE (1:2) was investigated by using the PC-specified transfer protein. The results indicate that: (1) both dolichol and dolichyl phosphate enhance vesicle fusion in a comparable and concentration-dependent way; (2) the amount of exchangeable PC from MLVs is increased by dolichyl phosphate probably as a result of fusion processes. It thus appears that these polyprenols are potent destabilizers of bilayer structure and that this process is accompanied by membrane fusion and transbilayer transport of phospholipids.

  2. Dolichol phosphate induces non-bilayer structures, vesicle fusion and transbilayer movements of lipids in model membranes

    The effect of dolichols, polyprenols, dolichol esterified with fatty acids, and dolichol phosphate on the structure and fluidity of model membranes was studied using different biophysical techniques. These studies suggest that (1) dolichol and dolichol derivatives destabilize unsaturated PE-containing bilayers and promote hexagonal II phase formation; (2) high concentrations of dolichol induce lipid structures characterized by isotropic 31P NMR and particulate fracture faces. The effect of dolichol and dolichyl phosphate on fusion between large unilamellar vesicles of DOPC and DOPE was studied using a fluroescence resonance energy transfer assay. The influence of dolichyl phosphate on the transbilary movement of DOPC in multilamellar vesicles (MLV) and large unilamellar vesicles (LUV) composed of DOPC and DOPE (1:2) was investigated by using the PC-specified transfer protein. The results indicate that: (1) both dolichol and dolichyl phosphate enhance vesicle fusion in a comparable and concentration-dependent way; (2) the amount of exchangeable PC from MLVs is increased by dolichyl phosphate probably as a result of fusion processes. It thus appears that these polyprenols are potent destabilizers of bilayer structure and that this process is accompanied by membrane fusion and transbilayer transport of phospholipids

  3. Active Nuclear Import of Membrane Proteins Revisited

    Laba, Justyna K; Steen, Anton; Popken, Petra; Chernova, Alina; Poolman, Bert; Veenhoff, Liesbeth M

    2015-01-01

    It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the m

  4. Activation Calculation for a Fusion Experimental Breeder FEB-E

    FENGKaiming

    2002-01-01

    A fusion breeder might be an essential intermediate application of fusion energy at earlier term, since it has the potential to provide plenty of commercial fissile fuel. Based on fusion physics and technologies available at present and in the near future, the realistic fusion experimental breeder, FEB-E was designed.

  5. Saccharomyces cerevisiae nuclear fusion requires prior activation by alpha factor.

    Rose, M D; Price, B R; Fink, G. R.

    1986-01-01

    We have developed a protocol for efficient fusion of spheroplasts of the same mating type. Nuclear fusion in this whole-cell system is also efficient and closely parallels nuclear fusion in heterosexual mating of intact cells. In the spheroplast fusion system, nuclear fusion is dependent on both the KAR1 gene and prior exposure to alpha factor. The major products of nuclear fusion in the spheroplast fusion assay were true diploids that were homozygous at the mating-type locus. An additional 1...

  6. Membrane-associated chromate reductase activity from Enterobacter cloacae.

    P. C. Wang; Mori, T.; Toda, K.; Ohtake, H

    1990-01-01

    Washed cells of Enterobacter cloacae HO1 reduced hexavalent chromium (chromate: CrO4(2-) anaerobically. Chromate reductase activity was preferentially associated with the membrane fraction of the cells. Right-side-out membrane vesicles prepared from E. cloacae cells showed high chromate reductase activities when ascorbate-reduced phenazine methosulfate was added as an electron donor.

  7. Fusion of liposomes with the plasma membrane of epithelial cells: Fate of incorporated lipids as followed by freeze fracture and autoradiography of plastic sections

    Knoll, G.; Burger, K.N.J.; Bron, R.; van Meer, G.; Verkleij, A. J.

    1988-01-01

    The fusion of liposomes with the plasma membrane of influenza virus- infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells (van Meer et al., 1985. Biochemistry. 24:3593-3602), has been analyzed by morphological techniques. The distribution of liposomal lipids over the apical and basolateral plasma membrane domains after fusion was assessed by autoradiography of liposomal [3H]dipalmitoylphosphatidylcholine after rapid freezing or chemical fixation and further processi...

  8. Fusion of liposomes with the plasma membrane of epithelial cells: Fate of incorporated lipids as followed by freeze fracture and autoradiography of plastic sections

    Knoll, G; Burger, K.N.J.; Bron, R.; van Meer, G.; Verkleij, A J

    1988-01-01

    The fusion of liposomes with the plasma membrane of influenza virus-infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells (van Meer et al., 1985. Biochemistry, 24: 3593-3602), has been analyzed by morphological techniques. The distribution of liposomal lipids over the apical and basolateral plasma membrane domains after fusion was assessed by autoradiography of liposomal [3H]dipalmitoylphosphatidylcholine after rapid freezing or chemical fixation and further processi...

  9. Fusion of liposomes with the plasma membrane of epithelial cells: fate of incorporated lipids as followed by freeze fracture and autoradiography of plastic sections

    1988-01-01

    The fusion of liposomes with the plasma membrane of influenza virus- infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells (van Meer et al., 1985. Biochemistry. 24:3593-3602), has been analyzed by morphological techniques. The distribution of liposomal lipids over the apical and basolateral plasma membrane domains after fusion was assessed by autoradiography of liposomal [3H]dipalmitoylphosphatidylcholine after rapid freezing or chemical fixation and further processi...

  10. Water permeation through Nafion membranes: the role of water activity.

    Majsztrik, Paul; Bocarsly, Andrew; Benziger, Jay

    2008-12-25

    The permeation of water through 1100 equivalent weight Nation membranes has been measured for film thicknesses of 51-254 microm, temperatures of 30-80 degrees C, and water activities (a(w)) from 0.3 to 1 (liquid water). Water permeation coefficients increased with water content in Nafion. For feed side water activity in the range 0 permeation coefficients increased linearly with water activity and scaled inversely with membrane thickness. The permeation coefficients were independent of membrane thickness when the feed side of the membrane was in contact with liquid water (a(w) = 1). The permeation coefficient for a 127 microm thick membrane increased by a factor of 10 between contacting the feed side of the membrane to water vapor (a(w) = 0.9) compared to liquid water (a(w) = 1). Water permeation couples interfacial transport across the fluid membrane interface with water transport through the hydrophilic phase of Nafion. At low water activity the hydrophilic volume fraction is small and permeation is limited by water diffusion. The volume fraction of the hydrophilic phase increases with water activity, increasing water transport. As a(w) --> 1, the effective transport rate increased by almost an order of magnitude, resulting in a change of the limiting transport resistance from water permeation across the membrane to interfacial mass transport at the gas/membrane interface. PMID:19053672

  11. The effect of acclimation temperature on the fusion kinetics of lipid vesicles derived from endoplasmic reticulum membranes of rainbow trout (Oncorhynchus mykiss) liver.

    Miranda, Estuardo J; Hazel, Jeffrey R

    2002-02-01

    Membrane fusion is an obligatory step in many vital cellular processes. The well-established enrichment of bilayer-destabilizing lipids in membranes of poikilotherms subjected to growth at low temperatures leads to the prediction that such membranes will possess a greater propensity to undergo fusion. This hypothesis was explicitly tested in the present study by determining the kinetics of fusion between small unilamellar vesicles (SUVs) prepared from endoplasmic reticulum (ER) membranes of thermally-acclimated (to 5 and 20 degrees C) rainbow trout (Oncorhynchus mykiss) liver and bovine brain phosphatidylserine (BBPS). At temperatures above 10 degrees C, ER vesicles from 5 degrees C-acclimated trout, fused more rapidly and to a greater extent with BBPS vesicles (by average factors of 1.25- and 1.45-fold, respectively) than ER vesicles of 20 degrees C-acclimated trout. At temperatures >35 degrees C, apparent fusion rates declined while the extent of fusion increased in both acclimation groups. Fusion kinetics were found to be well correlated with and limited by the physical properties and phase state of the BBPS vesicles. These results indicate that dynamic attributes of biological membranes, such as the propensity to undergo fusion, are of potential regulatory significance and are partially conserved when growth or environmental temperature changes. PMID:11818217

  12. Charged fusion product loss measurements using nuclear activation

    Bonheure, G.; Hult, M.; González de Orduña, R.; Arnold, D.; Dombrowski, H.; Laubenstein, M.; Wieslander, E.; Vermaercke, P.; Murari, A.; Popovichev, S.; Mlynář, Jan

    2010-01-01

    Roč. 81, č. 10 (2010), 10D331. ISSN 0034-6748. [TOPICAL CONFERENCE ON HIGH-TEMPERATURE PLASMA DIAGNOSTICS/18th./. Wildwood, New Jersey, 16.05.2010-20.05.2010] R&D Projects: GA ČR GAP205/10/2055; GA MŠk LA08048 Institutional research plan: CEZ:AV0Z20430508 Keywords : fusion * fast particle loss * diagnostics * activation Subject RIV: BL - Plasma and Gas Discharge Physics Impact factor: 1.598, year: 2010 http://rsi.aip.org/resource/1/rsinak/v81/i10/p10D331_s1

  13. Fusing simulation and experiment: The effect of mutations on the structure and activity of the influenza fusion peptide

    Lousa, Diana; Pinto, Antónia R. T.; Victor, Bruno L.; Laio, Alessandro; Veiga, Ana S.; Castanho, Miguel A. R. B.; Soares, Cláudio M.

    2016-01-01

    During the infection process, the influenza fusion peptide (FP) inserts into the host membrane, playing a crucial role in the fusion process between the viral and host membranes. In this work we used a combination of simulation and experimental techniques to analyse the molecular details of this process, which are largely unknown. Although the FP structure has been obtained by NMR in detergent micelles, there is no atomic structure information in membranes. To answer this question, we performed bias-exchange metadynamics (BE-META) simulations, which showed that the lowest energy states of the membrane-inserted FP correspond to helical-hairpin conformations similar to that observed in micelles. BE-META simulations of the G1V, W14A, G12A/G13A and G4A/G8A/G16A/G20A mutants revealed that all the mutations affect the peptide’s free energy landscape. A FRET-based analysis showed that all the mutants had a reduced fusogenic activity relative to the WT, in particular the mutants G12A/G13A and G4A/G8A/G16A/G20A. According to our results, one of the major causes of the lower activity of these mutants is their lower membrane affinity, which results in a lower concentration of peptide in the bilayer. These findings contribute to a better understanding of the influenza fusion process and open new routes for future studies. PMID:27302370

  14. Fusing simulation and experiment: The effect of mutations on the structure and activity of the influenza fusion peptide.

    Lousa, Diana; Pinto, Antónia R T; Victor, Bruno L; Laio, Alessandro; Veiga, Ana S; Castanho, Miguel A R B; Soares, Cláudio M

    2016-01-01

    During the infection process, the influenza fusion peptide (FP) inserts into the host membrane, playing a crucial role in the fusion process between the viral and host membranes. In this work we used a combination of simulation and experimental techniques to analyse the molecular details of this process, which are largely unknown. Although the FP structure has been obtained by NMR in detergent micelles, there is no atomic structure information in membranes. To answer this question, we performed bias-exchange metadynamics (BE-META) simulations, which showed that the lowest energy states of the membrane-inserted FP correspond to helical-hairpin conformations similar to that observed in micelles. BE-META simulations of the G1V, W14A, G12A/G13A and G4A/G8A/G16A/G20A mutants revealed that all the mutations affect the peptide's free energy landscape. A FRET-based analysis showed that all the mutants had a reduced fusogenic activity relative to the WT, in particular the mutants G12A/G13A and G4A/G8A/G16A/G20A. According to our results, one of the major causes of the lower activity of these mutants is their lower membrane affinity, which results in a lower concentration of peptide in the bilayer. These findings contribute to a better understanding of the influenza fusion process and open new routes for future studies. PMID:27302370

  15. Active membrane having uniform physico-chemically functionalized ion channels

    Gerald, II, Rex E; Ruscic, Katarina J; Sears, Devin N; Smith, Luis J; Klingler, Robert J; Rathke, Jerome W

    2012-09-24

    The present invention relates to a physicochemically-active porous membrane for electrochemical cells that purports dual functions: an electronic insulator (separator) and a unidirectional ion-transporter (electrolyte). The electrochemical cell membrane is activated for the transport of ions by contiguous ion coordination sites on the interior two-dimensional surfaces of the trans-membrane unidirectional pores. One dimension of the pore surface has a macroscopic length (1 nm-1000 .mu.m) and is directed parallel to the direction of an electric field, which is produced between the cathode and the anode electrodes of an electrochemical cell. The membrane material is designed to have physicochemical interaction with ions. Control of the extent of the interactions between the ions and the interior pore walls of the membrane and other materials, chemicals, or structures contained within the pores provides adjustability of the ionic conductivity of the membrane.

  16. Development of low activation vanadium steel for fusion applications

    Proposed fusion reactors may enjoy significant advantages regarding public safety and waste disposal over current fission reactors. Neutron activation to the structural materials can be minimized by the appropriate choice of alloys. Unfortunately, commercially developed alloys for high temperature applications become activated with neutron absorption leading to sometimes very long decay chains. The present paper discusses the results of a new ''low activation'' ferritic alloy (UCVS-1) developed at UCLA. This new alloy, which contains vanadium instead of molybdenum for high temperature strength, shows very promising combinations of strength, ductility and low long-term radioactive products. It is shown in this paper that the strength and ductility of UCVS-1 are comparable to 2 1/4 Cr-1 Mo up to 4000C, achieving significant advantages regarding safety and radioactive waste disposal

  17. Nonequilibrium fluctuations, traveling waves, and instabilities in active membranes.

    Ramaswamy, S; Toner, J; Prost, J

    2000-04-10

    The stability of a flexible fluid membrane containing a distribution of mobile, active proteins (e.g., proton pumps) is shown to depend on the structure and functional asymmetry of the proteins. A stable active membrane is in a nonequilibrium steady state with height fluctuations whose statistical properties are governed by the protein activity. Disturbances are predicted to travel as waves at sufficiently long wavelength, with speed set by the normal velocity of the pumps. The unstable case involves a spontaneous, pump-driven undulation of the membrane, with clumping of the proteins in regions of high activity. PMID:11019123

  18. Mechanics of nonplanar membranes with force-dipole activity

    Lomholt, Michael Andersen

    2006-01-01

    A study is made of how active membrane proteins can modify the long wavelength mechanics of fluid membranes. The activity of the proteins is modelled as disturbing the protein surroundings through nonlocal force distributions of which a force-dipole distribution is the simplest example. An analyt...... contributions to mechanical properties such as tension and bending moments become apparent. It is also explained how the activity can induce a hydrodynamic attraction between the active proteins in the membrane. © 2006 The American Physical Society....

  19. Chloroquine Increases Glucose Uptake via Enhancing GLUT4 Translocation and Fusion with the Plasma Membrane in L6 Cells

    Qi Zhou

    2016-05-01

    Full Text Available Background/Aims: Chloroquine can induce an increase in the cellular uptake of glucose; however, the underlying mechanism is unclear. Methods: In this study, translocation of GLUT4 and intracellular Ca2+ changes were simultaneously observed by confocal microscope in L6 cells stably over-expressing IRAP-mOrange. The GLUT4 fusion with the plasma membrane (PM was traced using HA-GLUT4-GFP. Glucose uptake was measured using a cell-based glucose uptake assay. GLUT4 protein was detected by Western blotting and mRNA level was detected by RT-PCR. Results: We found that chloroquine induced significant increases in glucose uptake, glucose transporter GLUT4 translocation to the plasma membrane (GTPM, GLUT4 fusion with the PM, and intracellular Ca2+ in L6 muscle cells. Chloroquine-induced increases of GTPM and intracellular Ca2+ were inhibited by Gallein (Gβγ inhibitor and U73122 (PLC inhibitor. However, 2-APB (IP3R blocker only blocked the increase in intracellular Ca2+ but did not inhibit GTPM increase. These results indicate that chloroquine, via the Gβγ-PLC-IP3-IP3R pathway, induces elevation of Ca2+, and this Ca2+ increase does not play a role in chloroqui-ne-evoked GTPM increase. However, GLUT4 fusion with the PM and glucose uptake were significantly inhibited with BAPTA-AM. This suggests that Ca2+ enhances GLUT4 fusion with the PM resulting in glucose uptake increase. Conclusion: Our data indicate that chloroquine via Gβγ-PLC-IP3-IP3R induces Ca2+ elevation, which in turn promotes GLUT4 fusion with the PM. Moreover, chloroquine can enhance GLUT4 trafficking to the PM. These mechanisms eventually result in glucose uptake increase in control and insulin-resistant L6 cells. These findings suggest that chloroquine might be a potential drug for improving insulin tolerance in diabetic patients.

  20. Effect of components in activated sludge liquor on membrane fouling in a submerged membrane bioreactor

    YU Shui-li; ZHAO Fang-bo; ZHANG Xiao-hui; JING Guo-lin; ZHEN Xiang-hua

    2006-01-01

    By a membrane bioreactor with a settle tank in long-term operation and batch experiments, the effects of flocs, soluble microorganism products (SMPs) and metal ions in activated sludge liquor on membrane fouling were investigated. The results showed that foulants absorbed each other and formed a fouling layer as a "second membrane" influencing the permeability of the membrane.The "gel layer" caused by SMPs and "cake layer" by flocs showed great differences in morphology by analysis of scanning electron microscope and atomic force microscope. The "gel layer" was more compact and of poor permeability. When the membrane flux was MPa/h). SMPs played very important roles on membrane fouling. In the bu1king sludge, with SMPs increasing, the rate of membrane fouling (0.0132 MPa/h) was faster. While after flocculation of the SMPs, the rate of fouling decreased to 0.0034 MPa/h. Flocs could keep holes in their overlaps. They could alleviate membrane fouling by preventing the SMPs directly attaching on membrane surface.

  1. Accelerator ampersand Fusion Research Division 1991 summary of activities

    This report discusses research projects in the following areas: Heavy-ion fusion accelerator research; magnetic fusion energy; advanced light source; center for x-ray optics; exploratory studies; superconducting magnets; and bevalac operations

  2. Accelerator Fusion Research Division 1991 summary of activities

    Berkner, Klaus H.

    1991-12-01

    This report discusses research projects in the following areas: Heavy-ion fusion accelerator research; magnetic fusion energy; advanced light source; center for x-ray optics; exploratory studies; superconducting magnets; and bevalac operations.

  3. Accelerator & Fusion Research Division 1991 summary of activities

    1991-12-01

    This report discusses research projects in the following areas: Heavy-ion fusion accelerator research; magnetic fusion energy; advanced light source; center for x-ray optics; exploratory studies; superconducting magnets; and bevalac operations.

  4. Accelerator and fusion research division. 1992 Summary of activities

    1992-12-01

    This report contains brief discussions on research topics in the following area: Heavy-Ion Fusion Accelerator Research; Magnetic Fusion Energy; Advanced Light Source; Center for Beam Physics; Superconducting Magnets; and Bevalac Operations.

  5. Nonequilibrium Fluctuations, Travelling Waves, and Instabilities in Active Membranes

    Ramaswamy, Sriram; Toner, John; Prost, Jacques

    1999-01-01

    The stability of a flexible fluid membrane containing a distribution of mobile, active proteins (e.g. proton pumps) is shown to depend on the structure and functional asymmetry of the proteins. A stable active membrane is in a nonequilibrium steady state with height fluctuations whose statistical properties are governed by the protein activity. Disturbances are predicted to travel as waves at sufficiently long wavelength, with speed set by the normal velocity of the pumps. The unstable case i...

  6. Elementary Active Membranes Have the Power of Counting

    Porreca, Antonio E.; Leporati, Alberto; Mauri, Giancarlo; Zandron, Claudio; Research Group on Natural Computing (Universidad de Sevilla) (Coordinador)

    2011-01-01

    We prove that uniform families of P systems with active membranes operat- ing in polynomial time can solve the whole class of PP decision problems, without using nonelementary membrane division or dissolution rules. This result also holds for families having a stricter uniformity condition than the usual one.

  7. Molecular dynamics analysis of conformational change of paramyxovirus F protein during the initial steps of membrane fusion

    Highlights: ► Initial conformational change of paramyxovirus F protein is caused only by mechanical forces. ► HRA region undergoes a structural change from a beta + alpha conformation to an extended coil and then to an all-alpha conformation. ► HRS domains of F protein form three single α-helices prior to generation of the coiled coil. -- Abstract: The fusion of paramyxovirus to the cell membrane is mediated by fusion protein (F protein) present in the virus envelope, which undergoes a dramatic conformational change during the process. Unlike hemagglutinin in orthomyxovirus, this change is not mediated by an alteration of environmental pH, and its cause remains unknown. Steered molecular dynamics analysis leads us to suggest that the conformational modification is mediated only by stretching mechanical forces once the transmembrane fusion peptide of the protein is anchored to the cell membrane. Such elongating forces will generate major secondary structure rearrangement in the heptad repeat A region of the F protein; from β-sheet conformation to an elongated coil and then spontaneously to an α-helix. In addition, it is proposed that the heptad repeat A region adopts a final three-helix coiled coil and that this structure appears after the formation of individual helices in each monomer.

  8. Development of new cloning vectors for the production of immunogenic outer membrane fusion proteins in Escherichia coli

    Cornelis, P.; Sierra, J.C.; Lim, A. Jr.; Malur, A. [Vrije Universiteit Brussel, Paardenstraat (Belgium)] [and others

    1996-02-01

    The Pseudomonas aeruginosa lipoprotein gene (oprI) was modified by cloning an in-frame polylinker in both orientations at the end of oprI. The resulting plasmids pVUBI and pVUB2 allow high lipoprotein production in E. coli after IPTG induction. The modified lipoproteins are present in the outer membrane and surface-exposed. Outer membrane-bound fusion proteins of different sizes were produced and used to generate antibodies without use of adjuvant. An 87 bp DNA fragment from the vp72 capsid protein gene of African Swine Fever virus (ASFV) and the entire Leishmania major glycoprotein gp63 gene were expressed in this system. Finally, a fusion lipoprotein containing a 16 amino acid epitope from the preS2b region of Hepatitis B virus (HBV) was presented by an antigen-presenting cell line to a T-cell hybridoma while the corresponding cross-linked S2b peptide was not. The results suggest that OprI-based fusion proteins can be used to generate both humoral and cellular immune responses. 44 refs., 7 figs.

  9. Molecular dynamics analysis of conformational change of paramyxovirus F protein during the initial steps of membrane fusion

    Martin-Garcia, Fernando; Mendieta-Moreno, Jesus Ignacio; Mendieta, Jesus [Centro de Biologia Molecular ' Severo Ochoa' (CSIC/UAM), C/ Nicolas Cabrera, 1, Cantoblanco, 28049 Madrid (Spain); Biomol-Informatics SL, Parque Cientifico de Madrid, C/ Faraday, 7, Cantoblanco, 28049 Madrid (Spain); Gomez-Puertas, Paulino, E-mail: pagomez@cbm.uam.es [Centro de Biologia Molecular ' Severo Ochoa' (CSIC/UAM), C/ Nicolas Cabrera, 1, Cantoblanco, 28049 Madrid (Spain)

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer Initial conformational change of paramyxovirus F protein is caused only by mechanical forces. Black-Right-Pointing-Pointer HRA region undergoes a structural change from a beta + alpha conformation to an extended coil and then to an all-alpha conformation. Black-Right-Pointing-Pointer HRS domains of F protein form three single {alpha}-helices prior to generation of the coiled coil. -- Abstract: The fusion of paramyxovirus to the cell membrane is mediated by fusion protein (F protein) present in the virus envelope, which undergoes a dramatic conformational change during the process. Unlike hemagglutinin in orthomyxovirus, this change is not mediated by an alteration of environmental pH, and its cause remains unknown. Steered molecular dynamics analysis leads us to suggest that the conformational modification is mediated only by stretching mechanical forces once the transmembrane fusion peptide of the protein is anchored to the cell membrane. Such elongating forces will generate major secondary structure rearrangement in the heptad repeat A region of the F protein; from {beta}-sheet conformation to an elongated coil and then spontaneously to an {alpha}-helix. In addition, it is proposed that the heptad repeat A region adopts a final three-helix coiled coil and that this structure appears after the formation of individual helices in each monomer.

  10. Accelerator and Fusion Research Division 1989 summary of activities

    This report discusses the research being conducted at Lawrence Berkeley Laboratory's Accelerator and Fusion Research Division. The main topics covered are: heavy-ion fusion accelerator research; magnetic fusion energy; advanced light source; center for x-ray optics; exploratory studies; high-energy physics technology; and bevalac operations

  11. Accelerator and Fusion Research Division 1989 summary of activities

    1990-06-01

    This report discusses the research being conducted at Lawrence Berkeley Laboratory's Accelerator and Fusion Research Division. The main topics covered are: heavy-ion fusion accelerator research; magnetic fusion energy; advanced light source; center for x-ray optics; exploratory studies; high-energy physics technology; and bevalac operations.

  12. A Histidine Switch in Hemagglutinin-Neuraminidase Triggers Paramyxovirus-Cell Membrane Fusion▿

    Krishnan, Anuja; Santosh K Verma; Mani, Prashant; Gupta, Rahul; Kundu, Suman; Sarkar, Debi P

    2008-01-01

    Most paramyxovirus fusion proteins require coexpression of and activation by a homotypic attachment protein, hemagglutinin-neuraminidase (HN), to promote membrane fusion. However, the molecular mechanism of the activation remains unknown. We previously showed that the incorporation of a monohistidylated lipid into F-virosome (Sendai viral envelope containing only fusion protein) enhanced its fusion to hepatocytes, suggesting that the histidine residue in the lipid accelerated membrane fusion....

  13. Solid-State Nuclear Magnetic Resonance Measurements of HIV Fusion Peptide 13CO to Lipid 31P Proximities Support Similar Partially Inserted Membrane Locations of the α Helical and β Sheet Peptide Structures

    Gabrys, Charles M.; Qiang, Wei; Sun, Yan; Xie, Li; Schmick, Scott D.; Weliky, David P.

    2013-10-01

    Fusion of the human immunodeficiency virus (HIV) membrane and the host cell membrane is an initial step of infection of the host cell. Fusion is catalyzed by gp41, which is an integral membrane protein of HIV. The fusion peptide (FP) is the -25 N-terminal residues of gp41 and is a domain of gp41 that plays a key role in fusion catalysis likely through interaction with the host cell membrane. Much of our understanding of the FP domain has been accomplished with studies of -HFP-, i.e., a -25-residue peptide composed of the FP sequence but lacking the rest of gp41. HFP catalyzes fusion between membrane vesicles and serves as a model system to understand fusion catalysis. HFP binds to membranes and the membrane location of HFP is likely a significant determinant of fusion catalysis perhaps because the consequent membrane perturbation reduces the fusion activation energy. In the present study, many HFPs were synthesized and differed in the residue position that was 13CO backbone labeled. Samples were then prepared that each contained a singly 13CO labeled HFP incorporated into membranes that lacked cholesterol. HFP had distinct molecular populations with either α helical or oligomeric - sheet structure. Proximity between the HFP 13CO nuclei and 31P nuclei in the membrane headgroups was probed by solid-state NMR (SSNMR) rotational-echo double-resonance (REDOR) measurements. For many samples, there were distinct 13CO shifts for the α helical and - sheet structures so that the proximities to 31P nuclei could be determined for each structure. Data from several differently labeled HFPs were then incorporated into a membrane location model for the particular structure. In addition to the 13CO labeled residue position, the HFPs also differed in sequence and/or chemical structure. -HFPmn- was a linear peptide that contained the 23 N-terminal residues of gp41. -HFPmn_V2E- contained the V2E mutation that for HIV leads to greatly reduced extent of fusion and infection. The

  14. Fusion reactor technology studies common development activities of CEA and KFK in the frame of the european community fusion programme

    In 1982, it was decided in the European Community to complement the fusion programme by a technology part. The initial content of this technology programme was defined by European experts. The laboratories associated to Euratom were invited to participate with proposals, which were then selected by the Fusion Technology Steering Committee. Coherent with these activities, steps were taken to define the next generation of fusion experiments by the newly founded Next European Torus (NET) Team. To contribute in the most efficient way, CEA and KfK have agreed to cooperate by complementary activities, common experiments and by forming common research groups. The particular agreements have been kept open for partners from other EC laboratories to join. The different cooperative activities in the field of nuclear technologies are described below together with first results and findings

  15. Silver-enhanced block copolymer membranes with biocidal activity

    Madhavan, Poornima

    2014-11-12

    Silver nanoparticles were deposited on the surface and pore walls of block copolymer membranes with highly ordered pore structure. Pyridine blocks constitute the pore surfaces, complexing silver ions and promoting a homogeneous distribution. Nanoparticles were then formed by reduction with sodium borohydride. The morphology varied with the preparation conditions (pH and silver ion concentration), as confirmed by field emission scanning and transmission electron microscopy. Silver has a strong biocide activity, which for membranes can bring the advantage of minimizing the growth of bacteria and formation of biofilm. The membranes with nanoparticles prepared under different pH values and ion concentrations were incubated with Pseudomonas aeruginosa and compared with the control. The strongest biocidal activity was achieved with membranes containing membranes prepared under pH 9. Under these conditions, the best distribution with small particle size was observed by microscopy.

  16. Targeting of a chimeric human histone fusion mRNA to membrane-bound polysomes in HeLa cells

    The subcellular location of histone mRNA-containing polysomes may play a key role in the posttranscriptional events that mediate histone mRNA turnover following inhibition of DNA synthesis. Previously, it has been shown that histone mRNA is found primarily on free polysomes that are associated with the cytoskeleton. The authors report here the construction of an Escherichia coli pBR322 β-lactamase signal peptide-human H3 histone fusion gene. The fusion transcript is targeted to membrane-bound polysomes and remains stable following interruption of DNA replication. Relocating mRNA within the cell may provide a procedure for studying the posttranscriptional regulation of gene expression

  17. Targeting of a chimeric human histone fusion mRNA to membrane-bound polysomes in HeLa cells

    Zambetti, G.; Stein, J.; Stein, G.

    1987-05-01

    The subcellular location of histone mRNA-containing polysomes may play a key role in the posttranscriptional events that mediate histone mRNA turnover following inhibition of DNA synthesis. Previously, it has been shown that histone mRNA is found primarily on free polysomes that are associated with the cytoskeleton. The authors report here the construction of an Escherichia coli pBR322 ..beta..-lactamase signal peptide-human H3 histone fusion gene. The fusion transcript is targeted to membrane-bound polysomes and remains stable following interruption of DNA replication. Relocating mRNA within the cell may provide a procedure for studying the posttranscriptional regulation of gene expression.

  18. Pandemic H1N1 influenza A directly induces a robust and acute inflammatory gene signature in primary human bronchial epithelial cells downstream of membrane fusion

    Pandemic H1N1 influenza A (H1N1pdm) elicits stronger pulmonary inflammation than previously circulating seasonal H1N1 influenza A (sH1N1), yet mechanisms of inflammatory activation in respiratory epithelial cells during H1N1pdm infection are unclear. We investigated host responses to H1N1pdm/sH1N1 infection and virus entry mechanisms in primary human bronchial epithelial cells in vitro. H1N1pdm infection rapidly initiated a robust inflammatory gene signature (3 h post-infection) not elicited by sH1N1 infection. Protein secretion inhibition had no effect on gene induction. Infection with membrane fusion deficient H1N1pdm failed to induce robust inflammatory gene expression which was rescued with restoration of fusion ability, suggesting H1N1pdm directly triggered the inflammatory signature downstream of membrane fusion. Investigation of intra-virion components revealed H1N1pdm viral RNA (vRNA) triggered a stronger inflammatory phenotype than sH1N1 vRNA. Thus, our study is first to report H1N1pdm induces greater inflammatory gene expression than sH1N1 in vitro due to direct virus–epithelial cell interaction. - Highlights: • We investigated H1N1pdm/sH1N1 infection in primary epithelial cells. • H1N1pdm directly initiated a robust inflammatory gene signature, sH1N1 did not. • H1N1pdm viral RNA triggered a stronger response than sH1N1. • H1N1pdm induces greater response due to direct virus–cell interaction. • These results have potential to impact vaccine and therapeutic development

  19. Pandemic H1N1 influenza A directly induces a robust and acute inflammatory gene signature in primary human bronchial epithelial cells downstream of membrane fusion

    Paquette, Stéphane G. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Institute of Medical Science, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Banner, David [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Chi, Le Thi Bao [Department of Microbiology, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Carlo Urbani Centre, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Leon, Alberto J. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); Xu, Luoling; Ran, Longsi [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Huang, Stephen S.H. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Department of Immunology, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Farooqui, Amber [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); and others

    2014-01-05

    Pandemic H1N1 influenza A (H1N1pdm) elicits stronger pulmonary inflammation than previously circulating seasonal H1N1 influenza A (sH1N1), yet mechanisms of inflammatory activation in respiratory epithelial cells during H1N1pdm infection are unclear. We investigated host responses to H1N1pdm/sH1N1 infection and virus entry mechanisms in primary human bronchial epithelial cells in vitro. H1N1pdm infection rapidly initiated a robust inflammatory gene signature (3 h post-infection) not elicited by sH1N1 infection. Protein secretion inhibition had no effect on gene induction. Infection with membrane fusion deficient H1N1pdm failed to induce robust inflammatory gene expression which was rescued with restoration of fusion ability, suggesting H1N1pdm directly triggered the inflammatory signature downstream of membrane fusion. Investigation of intra-virion components revealed H1N1pdm viral RNA (vRNA) triggered a stronger inflammatory phenotype than sH1N1 vRNA. Thus, our study is first to report H1N1pdm induces greater inflammatory gene expression than sH1N1 in vitro due to direct virus–epithelial cell interaction. - Highlights: • We investigated H1N1pdm/sH1N1 infection in primary epithelial cells. • H1N1pdm directly initiated a robust inflammatory gene signature, sH1N1 did not. • H1N1pdm viral RNA triggered a stronger response than sH1N1. • H1N1pdm induces greater response due to direct virus–cell interaction. • These results have potential to impact vaccine and therapeutic development.

  20. An Extended Membrane System with Active Membranes to Solve Automatic Fuzzy Clustering Problems.

    Peng, Hong; Wang, Jun; Shi, Peng; Pérez-Jiménez, Mario J; Riscos-Núñez, Agustín

    2016-05-01

    This paper focuses on automatic fuzzy clustering problem and proposes a novel automatic fuzzy clustering method that employs an extended membrane system with active membranes that has been designed as its computing framework. The extended membrane system has a dynamic membrane structure; since membranes can evolve, it is particularly suitable for processing the automatic fuzzy clustering problem. A modification of a differential evolution (DE) mechanism was developed as evolution rules for objects according to membrane structure and object communication mechanisms. Under the control of both the object's evolution-communication mechanism and the membrane evolution mechanism, the extended membrane system can effectively determine the most appropriate number of clusters as well as the corresponding optimal cluster centers. The proposed method was evaluated over 13 benchmark problems and was compared with four state-of-the-art automatic clustering methods, two recently developed clustering methods and six classification techniques. The comparison results demonstrate the superiority of the proposed method in terms of effectiveness and robustness. PMID:26790484

  1. Accelerator and Fusion Research Division: 1984 summary of activities

    1985-05-01

    During fiscal 1984, major programmatic activities in AFRD continued in each of five areas: accelerator operations, highlighted by the work of nuclear science users, who produced clear evidence for the formation of compressed nuclear matter during heavy-ion collisions; high-energy physics, increasingly dominated by our participation in the design of the Superconducting Super Collider; heavy-ion fusion accelerator research, which focused on the design of a four-beam experiment as a first step toward assessing the promise of heavy-ion inertial-confinement fusion; and research at the Center for X-Ray Optics, which completed its first year of broadly based activities aimed at the exploitation of x-ray and ultraviolet radiation. At the same time, exploratory studies were under way, aimed at investigating major new programs for the division. During the past year, for example, we took a preliminary look at how we could use the Bevatron as an injector for a pair of colliding-beam rings that might provide the first glimpse of a hitherto unobserved state of matter called the quark-gluon plasma. Together with Livermore scientists, we also conducted pioneering high-gain free-electron laser (FEL) experiments and proposed a new FEL-based scheme (called the two-beam accelerator) for accelerating electrons to very high energies. And we began work on the design of the Coherent XUV Facility (CXF), an advanced electron storage ring for the production of intense coherent radiation from either undulators or free-electron lasers.

  2. Accelerator and Fusion Research Division: 1984 summary of activities

    During fiscal 1984, major programmatic activities in AFRD continued in each of five areas: accelerator operations, highlighted by the work of nuclear science users, who produced clear evidence for the formation of compressed nuclear matter during heavy-ion collisions; high-energy physics, increasingly dominated by our participation in the design of the Superconducting Super Collider; heavy-ion fusion accelerator research, which focused on the design of a four-beam experiment as a first step toward assessing the promise of heavy-ion inertial-confinement fusion; and research at the Center for X-Ray Optics, which completed its first year of broadly based activities aimed at the exploitation of x-ray and ultraviolet radiation. At the same time, exploratory studies were under way, aimed at investigating major new programs for the division. During the past year, for example, we took a preliminary look at how we could use the Bevatron as an injector for a pair of colliding-beam rings that might provide the first glimpse of a hitherto unobserved state of matter called the quark-gluon plasma. Together with Livermore scientists, we also conducted pioneering high-gain free-electron laser (FEL) experiments and proposed a new FEL-based scheme (called the two-beam accelerator) for accelerating electrons to very high energies. And we began work on the design of the Coherent XUV Facility (CXF), an advanced electron storage ring for the production of intense coherent radiation from either undulators or free-electron lasers

  3. RACC-PULSE, Neutron Activation in Fusion Reactor System

    1 - Description of program or function: CCC-0388/RACC was specifically developed to compute the radioactivity and radioactivity-related parameters (e.g. afterheat, biological hazard potential, etc.) due to neutron activation within Inertial Fusion Energy and Magnetic Fusion energy reactor systems. It can also be utilized to compute the radioactivity in fission, accelerator or any other neutron generating and neutron source system. This new designated RACC-PULSE is based on CCC-0388 and has the capability to model irradiation histories of varying flux levels having varying pulse widths (on times) and dwell periods (off times) and varying maintenance periods. This provides the user with the flexibility of modeling most any complexity of irradiation history beginning with simple steady state operating systems to complex multi-flux level pulse/intermittent operating systems. 2 - Method of solution: The solution method implemented within the RACC-PULSE code is a matrix based method which relies on the evaluation of the Matrix Exponential for the pulse period (on period), dwell period (off time) and post shutdown periods. For the pulsed and dwell periods, the Matrix Exponential was evaluated using the squaring and scaling technique outlined in a review article by Molar and Van Loan entitled Nineteen Dubious Ways to Compute the Exponential of a Matrix. A balanced binary tree method utilized for parameter storage in information systems was employed to evaluate the linear chains constructed for the post shutdown period. The RACC-Pulse code retains the capability of modeling the standard slab, cylinder, sphere and torus geometries in multi-dimensions as well as the point or zero-dimension geometry for Monte Carlo code interfacing. It provides easy interfacing with many of the standard multigroup, multidimensional neutron/photon transport code systems currently employed by the fusion community and implemented on the UNICOS Cray 2 System at NERSC. An auxiliary code is provided

  4. Effects of a perfusion bioreactor activated novel bone substitute in spine fusion in sheep

    Sørensen, Jesper Roed; Koroma, Kariatta Ester; Ding, Ming; Wendt, David; Jespersen, Stig; Juhl, Maria Vinther; Theilgaard, Naseem; Martin, Ivan; Overgaard, Søren

    2012-01-01

    To evaluate the effect of a large perfusion-bioreactor cell-activated bone substitute, on a two-level large posterolateral spine fusion sheep model.......To evaluate the effect of a large perfusion-bioreactor cell-activated bone substitute, on a two-level large posterolateral spine fusion sheep model....

  5. Drainin required for membrane fusion of the contractile vacuole in Dictyostelium is the prototype of a protein family also represented in man.

    Becker, M; Matzner, M; Gerisch, G

    1999-01-01

    The contractile vacuole expels water by forming a channel with the plasma membrane and thus enables cells to survive in a hypo-osmotic environment. Here we characterize drainin, a Dictyostelium protein involved in this process, as the first member of a protein family represented in fission yeast, Caenorhabditis elegans and man. Gene replacement in Dictyostelium shows that drainin acts at a checkpoint of channel formation between the contractile vacuole and the plasma membrane. A green fluorescent protein fusion of drainin localizes specifically to the contractile vacuole and rescues its periodic discharge in drainin-null cells. Drainin is a peripheral membrane protein, requiring a short hydrophobic stretch in its C-terminal region for localization and function. We suggest that drainin acts in a signaling cascade that couples a volume-sensing device in the vacuolar membrane to the membrane fusion machinery. PMID:10369671

  6. Development of transcriptional fusions to assess Leptospira interrogans promoter activity.

    Gustavo M Cerqueira

    Full Text Available BACKGROUND: Leptospirosis is a zoonotic infectious disease that affects both humans and animals. The existing genetic tools for Leptospira spp. have improved our understanding of the biology of this spirochete as well as the interaction of pathogenic leptospires with the mammalian host. However, new tools are necessary to provide novel and useful information to the field. METHODOLOGY AND PRINCIPAL FINDINGS: A series of promoter-probe vectors carrying a reporter gene encoding green fluorescent protein (GFP were constructed for use in L. biflexa. They were tested by constructing transcriptional fusions between the lipL41, Leptospiral Immunoglobulin-like A (ligA and Sphingomyelinase 2 (sph2 promoters from L. interrogans and the reporter gene. ligA and sph2 promoters were the most active, in comparison to the lipL41 promoter and the non-induced controls. The results obtained are in agreement with LigA expression from the L. interrogans Fiocruz L1-130 strain. CONCLUSIONS: The novel vectors facilitated the in vitro evaluation of L. interrogans promoter activity under defined growth conditions which simulate the mammalian host environment. The fluorescence and rt-PCR data obtained closely reflected transcriptional regulation of the promoters, thus demonstrating the suitability of these vectors for assessing promoter activity in L. biflexa.

  7. Recruitment and SNARE-mediated fusion of vesicles in furrow membrane remodeling during cytokinesis in zebrafish embryos

    Cytokinesis is the final stage in cell division that serves to partition cytoplasm and daughter nuclei into separate cells. Membrane remodeling at the cleavage plane is a required feature of cytokinesis in many species. In animal cells, however, the precise mechanisms and molecular interactions that mediate this process are not yet fully understood. Using real-time imaging in live, early stage zebrafish embryos, we demonstrate that vesicles labeled with the v-SNARE, VAMP-2, are recruited to the cleavage furrow during deepening in a microtubule-dependent manner. These vesicles then fuse with, and transfer their VAMP-2 fluorescent label to, the plasma membrane during both furrow deepening and subsequent apposition. This observation indicates that new membrane is being inserted during these stages of cytokinesis. Inhibition of SNAP-25 (a cognate t-SNARE of VAMP-2), using a monoclonal antibody, blocked VAMP-2 vesicle fusion and furrow apposition. Transient expression of mutant forms of SNAP-25 also produced defects in furrow apposition. SNAP-25 inhibition by either method, however, did not have any significant effect on furrow deepening. Thus, our data clearly indicate that VAMP-2 and SNAP-25 play an essential role in daughter blastomere apposition, possibly via the delivery of components that promote the cell-to-cell adhesion required for the successful completion of cytokinesis. Our results also support the idea that new membrane addition, which occurs during late stage cytokinesis, is not required for furrow deepening that results from contractile band constriction

  8. History, progress, achievement and future prospect of research activities on fusion materials by Japanese university researchers

    Research activities on fusion materials by Japanese university researchers are reviewed. Organized research on fusion materials has been initiated around mid 1970s under auspices of Monbusho (Ministry of Education, Science and Culture). Particularly effective was the Special Research Project on Fusion for fiscal year 1980 - 1989. At the same time, Japan/U.S. collaboration on fusion materials (1982 - 2000) has been very successful, yielding numerous useful results. The highlights of the technical achievement of these projects are briefly summarized. Both of these projects may be characterized to be composed of two major tasks, namely, fundamental aspects of alloy development for fusion and high fluence irradiation effects under fusion reactor environment. The basic philosophy of the project is discussed. The recent trend is to organize the university research activities into a comprehensive research network. (orig.)

  9. Induction of petite yeast mutants by membrane-active agents.

    Jiménez, J.; Longo, E.; Benítez, T

    1988-01-01

    Ethanol proved to be a strong mutagenic agent of Saccharomyces mitochondrial DNA. Other active membrane solvents, such as tert-butanol, isopropanol, and sodium dodecyl sulfate, also turned out to be powerful petite mutation [rho-] inducers. Mutants defective in ergosterol synthesis (erg mutants) showed an extremely high frequency of spontaneous petite cells, suggesting that mitochondrial membrane alterations that were caused either by changes in its composition, as in the erg mutants, or by t...

  10. Constant-space P systems with active membranes

    A. Leporati; Manzoni, L; Mauri, G; Porreca, A; Zandron, C

    2014-01-01

    We show that a constant amount of space is sufficient to simulate a polynomial-space bounded Turing machine by P systems with active membranes. We thus obtain a new characterisation of PSPACE, which raises interesting questions about the definition of space complexity for P systems. We then propose an alternative definition, where the size of the alphabet and the number of membrane labels of each P system are also taken into account. Finally we prove that, when less than a logarithmic number ...

  11. Modeling and vibration control of an active membrane mirror

    Ruggiero, Eric J.; Inman, Daniel J.

    2009-09-01

    The future of space satellite technology lies in ultra-large mirrors and radar apertures for significant improvements in imaging and communication bandwidths. The availability of optical-quality membranes drives a parallel effort for structural models that can capture the dominant dynamics of large, ultra-flexible satellite payloads. Unfortunately, the inherent flexibility of membrane mirrors wreaks havoc with the payload's on-orbit stability and maneuverability. One possible means of controlling these undesirable dynamics is by embedding active piezoelectric ceramics near the boundary of the membrane mirror. In doing so, active feedback control can be used to eliminate detrimental vibration, perform static shape control, and evaluate the health of the structure. The overall motivation of the present work is to design a control system using distributed bimorph actuators to eliminate any detrimental vibration of the membrane mirror. As a basis for this study, a piezoceramic wafer was attached in a bimorph configuration near the boundary of a tensioned rectangular membrane sample. A finite element model of the system was developed to capture the relevant system dynamics from 0 to 300 Hz. The finite element model was compared against experimental results, and fair agreement found. Using the validated finite element models, structural control using linear quadratic regulator control techniques was then used to numerically demonstrate effective vibration control. Typical results show that less than 12 V of actuation voltage is required to eliminate detrimental vibration of the membrane samples in less than 15 ms. The functional gains of the active system are also derived and presented. These spatially descriptive control terms dictate favorable regions within the membrane domain for placing sensors and can be used as a design guideline for structural control applications. The results of the present work demonstrate that thin plate theory is an appropriate modeling

  12. Activation of methane using solid oxide membranes

    Elshof, ten, J.E.; Hassel, van, E Edwin; Bouwmeester, H.J.M.

    1995-01-01

    Dense membranes of mixed-conducting perovskite-type oxides La0.6Sr0.4,CO0.8Fe0.2O3 and La0.8Ba0.2Co0.8Fe0.2O3 were used for methane coupling by application of pressure-driven O2 permeation. High operating temperatures, typically above 800°C, were needed to obtain reasonable oxygen fluxes. Conversions were small (1–3%). Both compositions showed comparable C2 selectivities at low methane partial pressures. At higher pressures the selectivity to C2 hydrocarbons for La0.6Sr0.4CO0.8Fe0.2O3 increas...

  13. Management of a water leak on actively cooled fusion devices

    ITER will be the most important machine equipped with actively cooled plasma facing components (PFCs). In case of abnormal events during a discharge, the PFC will be submitted to localized transient phenomena (high power densities, run away electrons, etc.), leading, in the worst case, to the degradation of the PFC wall and possibly to a water leak. In any case, a leak will have important consequences for the PFCs and equipment located in the vacuum vessel or connected to the ports such as seals, pumping systems or diagnostics. Considerable experience of these events has been gained at Tore Supra over a period of more than 10 years [J.J. Cordier, Ten years of maintenance on Tore Supra actively cooled components, in: Proceedings of the 21th Symp. of Fusion Technology (SOFT), Madrid, Spain, September, 2000.], which will be useful for the next step machines. This paper describes for each leak size type the procedures for maintaining save conditions in the vacuum vessel. It also presents the methods used at Tore Supra to drain-off the primary loop circuits and to identify the leaky PFC

  14. Beryllium neutron activation detector for pulsed DD fusion sources

    A compact fast neutron detector based on beryllium activation has been developed to perform accurate neutron fluence measurements on pulsed DD fusion sources. It is especially well suited to moderate repetition-rate (9Be(n,α)6He cross-section, energy calibration of the proportional counters, and numerical simulations of neutron interactions and beta-particle paths using MCNP5. The response function R(En) is determined over the neutron energy range 2-4 MeV. The count rate capability of the detector has been studied and the corrections required for high neutron fluence measurements are discussed. For pulsed DD neutron fluencies >3×104 cm-2, the statistical uncertainty in the fluence measurement is better than 1%. A small plasma focus device has been employed as a pulsed neutron source to test two of these new detectors, and their responses are found to be practically identical. Also the level of interfering activation is found to be sufficiently low as to be negligible.

  15. Activation and waste management considerations of fusion materials

    Cheng, E. T.; Saji, G.

    1994-09-01

    Inconel-625 (Ni625), SS316, Ti-6Al-4V (Ti64), ferritic steel (FS), reduced activity ferritic steel (RAFS), manganese steel (Mn-steel), and V5Cr5Ti (V55), were examined for a near-term experimental D-T fueled fusion power reactor with respect to waste management. Activation calculations for these materials were performed assuming one year continuous operation at 1 MW/m 2 wall loading. The results show that the blanket components made of V55, Ti64, Mn-steel, and FS will be allowed for transfer to an on-site dry storage facility after 10 years of cooling after discharge. To transport the discharged blanket components to a permanent disposal site, the cooling time needed can be within 10 years for Ti64 and V55, provided that the impurities (mainly Ni, Nb and Mo) be controlled to an acceptable level. The RAFS and Mn-steel will need about 30 y cooling time because of its Fe and Mn contents. Ni625, 316SS, and FS, however, will require more than 50000 y cooling time because of their Nb and Mo contents. The RAFS, Mn-steel, Ti64 and V55 can be shallow-land wastes if the impurity level for Nb and Mo is dropped below 10 ppm.

  16. Fusion Machinery

    Sørensen, Jakob Balslev; Milosevic, Ira

    2015-01-01

    SNARE proteins constitute the minimal machinery needed for membrane fusion. SNAREs operate by forming a complex, which pulls the lipid bilayers into close contact and provides the mechanical force needed for lipid bilayer fusion. At the chemical synapse, SNARE-complex formation between the vesicu......SNARE proteins constitute the minimal machinery needed for membrane fusion. SNAREs operate by forming a complex, which pulls the lipid bilayers into close contact and provides the mechanical force needed for lipid bilayer fusion. At the chemical synapse, SNARE-complex formation between...

  17. Expression Screening of Integral Membrane Proteins by Fusion to Fluorescent Reporters.

    Bird, Louise E; Nettleship, Joanne E; Järvinen, Valtteri; Rada, Heather; Verma, Anil; Owens, Raymond J

    2016-01-01

    The production of recombinant integral membrane proteins for structural and functional studies remains technically challenging due to their relatively low levels of expression. To address this problem, screening strategies have been developed to identify the optimal membrane sequence and expression host for protein production. A common approach is to genetically fuse the membrane protein to a fluorescent reporter, typically Green Fluorescent Protein (GFP) enabling expression levels, localization and detergent solubilisation to be assessed. Initially developed for screening the heterologous expression of bacterial membrane proteins in Escherichia coli, the method has been extended to eukaryotic hosts, including insect and mammalian cells. Overall, GFP-based expression screening has made a major impact on the number of membrane protein structures that have been determined in the last few years. PMID:27553231

  18. DNA Duplexes with Hydrophobic Modifications Inhibit Fusion between HIV-1 and Cell Membranes

    Xu, Liang; Cai, Lifeng; Chen, Xueliang; Jiang, Xifeng; Chong, Huihui; Zheng, Baohua; Wang, Kun; He, Junlin; Chen, Wei; Zhang, Tao; Cheng, Maosheng; He, Yuxian; Liu, Keliang

    2013-01-01

    Discovery of new drugs for the treatment of AIDS typically possessing unique structures associated with novel mechanisms of action has been of great importance due to the quick drug-resistant mutations of HIV-1 strains. The work presented in this report describes a novel class of DNA duplex-based HIV-1 fusion inhibitors. Hydrophobic groups were introduced into a DNA duplex skeleton either at one end, at both ends, or in the middle. These modified DNA duplexes inhibited fusion between HIV-1 an...

  19. Lipid protrusions membrane softness, and enzymatic activity

    Jensen, Morten Østergaard; Høyrup, P.; Callisen, T.H.;

    2004-01-01

    protrusion modes and mechanical softness of phospholipid bilayers and on the other side the activity of enzymes acting on lipid bilayers composed of different unsaturated lipids. Specifically, our experiments show a correlation between the bilayer bending rigidity and the apparent Arrhenius activation energy...

  20. Confidence building in and through fission and fusion activities

    The peaceful uses of atomic energy are most suitable for achieving worldwide confidence building for the following reasons. (1) In spite of the need for peaceful uses of nuclear energy, the world is facing difficulties in the public perception and acceptance of nuclear works and facilities. (2) The above difficulties are due to many factors, such as the two sides of nuclear energy peaceful and military, the possibility of a large-scale reactor accident, the lack of understanding about radiation and radioactivity, and finally, emotion and egoism. Some of these factors are unique to nuclear-energy, but in other cases of public reactions, there are many facets similar to the above factors. (3) The public concern about safety is at its highest, broadest and severest point ever, coincident with the highest life expectancy in history. Over-precaution and over-protection about certain things may sometimes spoil one's health. Nuclear energy is most definitely suffering from such a trend. As a result, a severe nuclear accident in any country results in severe damage worldwide no manner in what form the real physical effects reach other countries. (4) The huge science and technology efforts required for fission and fusion activities cannot be fully achieved by one country. Explanations of some of the above factors are given. 2 refs

  1. Dysferlin Binds SNAREs (Soluble N-Ethylmaleimide-sensitive Factor (NSF) Attachment Protein Receptors) and Stimulates Membrane Fusion in a Calcium-sensitive Manner.

    Codding, Sara J; Marty, Naomi; Abdullah, Nazish; Johnson, Colin P

    2016-07-01

    Resealing of tears in the sarcolemma of myofibers is a necessary step in the repair of muscle tissue. Recent work suggests a critical role for dysferlin in the membrane repair process and that mutations in dysferlin are responsible for limb girdle muscular dystrophy 2B and Miyoshi myopathy. Beyond membrane repair, dysferlin has been linked to SNARE-mediated exocytotic events including cytokine release and acid sphingomyelinase secretion. However, it is unclear whether dysferlin regulates SNARE-mediated membrane fusion. In this study we demonstrate a direct interaction between dysferlin and the SNARE proteins syntaxin 4 and SNAP-23. In addition, analysis of FRET and in vitro reconstituted lipid mixing assays indicate that dysferlin accelerates syntaxin 4/SNAP-23 heterodimer formation and SNARE-mediated lipid mixing in a calcium-sensitive manner. These results support a function for dysferlin as a calcium-sensing SNARE effector for membrane fusion events. PMID:27226605

  2. Towards a reduced activation structural materials database for fusion reactors

    Full text: The development of First Wall, Blanket and Divertor materials which are capable of withstanding many years the high neutron and heat fluxes, is a critical path to fusion power. Therefore, the timely availability of a sound materials database has become an indispensable element in international fusion road maps. In order to provide a related materials database for design, construction, licensing and safe operation of the ITER Test Blanket Modules and of a DEMO reactor, a wealth of R and D results on the European reduced activation ferritic-martensitic steel EUROFER, and on oxide dispersion strengthened (ODS) variants have become available, mainly in the temperature window 250-700 deg. C. Industrial EUROFER-batches of 3.5 and 8.0 tons have been produced with a variety of semi-finished, quality-assured product forms. Extensive chipless shaping and joining experience taking into account different welding procedures and powder technology product forms have demonstrated that EUROFER type steel complies with a wide range of established manufacturing processes. EUROFER is also resistant to high temperature aging, and the existing creep-rupture properties (∼30000 h) indicated long term stability and predictability. To increase the thermal efficiency of blankets beyond 45%, high temperature resistant SiCf/SiC channel inserts for liquid metal coolant tubes are developed. Mechanical and thermal properties of various SiCf/SiC composits have been measured after neutron radiation. Regarding radiation damage resistance of blanket structural materials, a broad based reactor irradiation programme counts several steps from 2 needs to be removed, the design is presently based on tiles made of W (∼2000 deg. C), as well as on structural materials like W-alloy (∼700-1300 deg. C) and RAF(M)-ODS steel (∼650 deg. C). Severe plastic deformation of pure W and W alloys improves ductility, but does not prevent from re-crystallisation between 850 and 1200 deg. C. For the

  3. Status of fusion power reactor design activity in JAERI

    A number of fusion power reactor design studies conducted in JAERI over the past 22 years are reviewed and the present status of reactor studies is introduced. A helium gas cooled power reactor which has a blanket with solid lithium ceramics such as Li2O for the breeding material and incolloy-800 for the structural material was proposed in 1973. This is one of the first reactor design using a lithium ceramics blanket concept. Another power reactor design called the Swimming Pool-type Tokamak Reactor (SPTR-P) was completed in 1983. In SPTR-P, the tokamak reactor is submerged in a water pool to utilize water as the shield to reduce long term radioactive waste and to achieve easy repair and maintenance. In 1990, the Steady State Tokamak Reactor (SSTR) concept was proposed as a realistic fusion reactor to be built in the near future. The major feature of SSTR is the maximum utilization of a bootstrap current in order to reduce the power required to maintain steady state operation. It is cooled by pressurized water and uses low activation ferritic steel as the structural material. A study of SSTR-2 to improve the safety and economic aspects of SSTR by replacing the water coolant by a mixture of helium gas and fine solid particles was carried out in 1992. Light yet highly heat resistant material, titanium aluminide intermetallic compound (TiAl), is used as the structural material. The net thermal efficiency larger than 40 % can be achieved with the gas-particulate mixture at 5 MPa and the outlet coolant temperature of 700degC. A concept of a Drastically Easy Maintenance (DREAM) tokamak reactor has recently been proposed. For easy replacement of blanket and divertor, a plasma configuration with high aspect ratio around 6 and a small number of torus sectoring of 12 are selected. A 1/12 torus sector is horizontally pulled out between the TF coils with a straight radial motion. It is estimated that the availability of 85 % can be achieved. (author)

  4. Re-evaluation of the use of low activation materials in waste management strategies for fusion

    The world fusion programs have had a long goal that fusion power stations should produce only low level waste and thus not pose a burden for the future generations. However, the environmental impact of waste material is determined not only by the level of activation, but also the total volume of activated material. Since a tokamak power plant is large, the potential to generate a correspondingly large volume of activated material exists. The adoption of low activation materials, while important for reducing the radiotoxicity of the most active components, should be done as part of a strategy that also minimizes the volume of waste material that might be categorized as radioactive, even if lower in level. In this paper we examine different fusion blanket and shield designs in terms of their ability to limit the activation of the large vessel/ex-vessel components (e.g. vacuum vessel, magnets) and we identify the trends that allow improved in-vessel shielding to result in reduced vessel/ex-vessel activation. Recycling and clearance are options for reducing the volume of radioactive waste in a fusion power plant. Thus, the performance of typical fusion power plant designs with respect to recycling and clearance criteria are also assessed, to show the potential for improvement in waste volume reduction by careful selection of materials' combinations. We discuss the impact of these results on fusion waste strategies and on the development of fusion power in the future

  5. Assessment report of research and development activities. Activity: 'Fusion research and development' (Interim report)

    Japan Atomic Energy Agency (hereinafter referred to as 'JAEA') asked the assessment committee, 'Evaluation Committee of Research and Development Activities for Fusion' (hereinafter referred to as 'Committee') for interim assessment of 'Research and Development of the technical system for extraction of fusion energy,' in accordance with 'General Guideline for the Evaluation of Government R and D Activities' by Cabinet Office, Government of Japan, 'Guideline for Evaluation of R and D in Ministry of Education, Culture, Sports, Science and Technology' and 'Regulation on Conduct for Evaluation of R and D Activities' by JAEA. In response to the JAEA's request, the Committee assessed the research program of the Fusion Research and Development Directorate (hereinafter referred to as 'FRDD') during the period of four years from October 2005 to August 2009. The Committee evaluated the management and research activities of the FRDD based on the explanatory documents prepared by the FRDD, the oral presentations with questions-and-answers by the Director General and the Deputy Director Generals. (author)

  6. Role of lipid phase separations and membrane hydration in phospholipid vesicle fusion.

    Hoekstra, D.

    1982-01-01

    The relationship between lipid phase separation and fusion of small unilamellar phosphatidylserine-containing vesicles was investigated. The kinetics of phase separation were monitored by following the increase of self-quenching of the fluorescent phospholipid analogue N-(7-nitro-2,1,3-benzoxadiazol

  7. Crystal Structure of Dengue Virus Type 1 Envelope Protein in the Postfusion Conformation and Its Implications for Membrane Fusion

    Nayak, Vinod; Dessau, Moshe; Kucera, Kaury; Anthony, Karen; Ledizet, Michel; Modis, Yorgo; (Yale); (L2 Diagnostics)

    2009-07-31

    Dengue virus relies on a conformational change in its envelope protein, E, to fuse the viral lipid membrane with the endosomal membrane and thereby deliver the viral genome into the cytosol. We have determined the crystal structure of a soluble fragment E (sE) of dengue virus type 1 (DEN-1). The protein is in the postfusion conformation even though it was not exposed to a lipid membrane or detergent. At the domain I-domain III interface, 4 polar residues form a tight cluster that is absent in other flaviviral postfusion structures. Two of these residues, His-282 and His-317, are conserved in flaviviruses and are part of the 'pH sensor' that triggers the fusogenic conformational change in E, at the reduced pH of the endosome. In the fusion loop, Phe-108 adopts a distinct conformation, forming additional trimer contacts and filling the bowl-shaped concavity observed at the tip of the DEN-2 sE trimer.

  8. Micropipette manipulation technique for the monitoring of pH-dependent membrane lysis as induced by the fusion peptide of influenza virus.

    Soltesz, S A; Hammer, D A

    1995-01-01

    We have assembled a micropipette aspiration assay to measure membrane destabilization events in which large (20-30 microns diameter) unilamellar vesicles are manipulated and exposed to membrane destabilizing agents. Single events can be seen with a light microscope and are recorded using both a video camera and a photomultiplier tube. We have performed experiments with a wild-type fusion peptide from influenza virus (X31) and found that it induces pH-dependent, stochastic lysis of large unila...

  9. Effect of increase in orientational order of lipid chains and head group spacing on non steroidal anti-inflammatory drug induced membrane fusion.

    Roy, Sutapa Mondal; Bansode, Amol S; Sarkar, Munna

    2010-12-21

    Membrane fusion is a key event in many biological processes. The fusion process, both in vivo and in vitro, is induced by different agents which include mainly proteins and peptides. For protein- and peptide-mediated membrane fusion, conformational reorganization serves as a driving force. Small drug molecules do not share this advantage; hence, drug induced membrane fusion occurring in absence of any other fusogenic agent and at physiologically relevant concentration of the drugs is a very rare event. To date, only three drugs, namely, meloxicam (Mx), piroxicam (Px), and tenoxicam (Tx), belonging to the oxicam group of non steroidal anti-inflammatory drugs (NSAIDs), have been shown by us to induce fusion at very low drug to lipid ratio without the aid of any other fusogenic agent. In our continued effort to understand the interplay of different physical and chemical parameters of both the participating drugs and the membrane on the mechanism of this drug induced membrane fusion, we present here the effect of increase in orientational order of the lipid chains and increase in head group spacing. This is achieved by studying the effect of low concentration cholesterol (gel to fluid transition temperature, is mainly known to increase orientational order of the lipid chains and increase head group spacing. To isolate the effect of these parameters, small unilameller vesicles (SUVs) formed by dimyristoylphosphatidylcholine (DMPC) with an average diameter of 50-60 nm were used as simple model membranes. Fluorescence assays were used to probe the time dependence of lipid mixing, content mixing, and leakage and also used to determine the partitioning of the drugs in the membrane bilayer. Differential scanning calorimetry (DSC) was used to study the effect of drugs in the presence of cholesterol on the chain-melting temperature which reflects the fluidization effect of the hydrophobic tail region of the bilayer. Our results show contradictory effect of low concentration

  10. Comparison of fusion methods based on DST and DBN in human activity recognition

    2011-01-01

    Ambient assistive living environments require sophisticated information fusion and reasoning techniques to accurately identify activities of a person under care. In this paper, we explain, compare and discuss the application of two powerful fusion methods, namely dynamic Bayesian networks (DBN) and Dempster-Shafer theory (DST), for human activity recognition. Both methods are described, the implementation of activity recognition based on these methods is explained, and model acquisition and composition are ...

  11. Active inhibition of herpes simplex virus type 1-induced cell fusion

    Bzik, D.J.; Person, S.; Read, G.S.

    1982-01-01

    Previous studies have demonstrated that syn mutant-infected cells fuse less well with nonsyncytial virus-infected cells than with uninfected cells, a phenomenon defined as function inhibition. The present study characterizes the kinetics as well as the requirements for expression of fusion inhibition. Initially, the capacity of sparse syn mutant-infected cells to fuse with uninfected surrounding cells was determined throughout infection. Of seven syn mutants examined, including representatives with alterations in two different viral genes that affect cell fusion, all showed an increase in fusion capacity up to 12 hr after infection and a decrease at later times. Fusion inhibition was examined in experiments employing sparse syn20-infected cells which had been incubated to a maximum fusion capacity; it was shown that surrounding cells infected with KOS, the parent of syn20, began to inhibit fusion by the syn20-infected cells at about 4 hr after infection, and that the maximum ability to inhibit fusion was attained at about 6 hr after infection. The metabolic blocking agents actinomycin D (RNA), cycloheximide (protein), 2-deoxyglucose, and tunicamycin (glycoslyation of glycoproteins) all showed the ability to inhibit the expression of fusion inhibition by KOS-infected cells if added shortly after infection. It is concluded that fusion inhibition is an active process that requires the synthesis of RNA, proteins, and glycoproteins. 17 references, 3 figures, 2 tables.

  12. Magnetic Fusion Energy Technology Fellowship Program: Summary of program activities for calendar year 1985

    This report summarizes the activities of the US Department of Energy (DOE) Magnetic Fusion Energy Technology Fellowship program (MFETF) for the 1985 calendar year. The MFETF program has continued to support the mission of the Office of Fusion Energy (OFE) and its Division of Development and Technology (DDT) by ensuring the availability of appropriately trained engineering manpower needed to implement the OFE/DDT magnetic fusion energy agenda. This program provides training and research opportunities to highly qualified students at DOE-designated academic, private sector, and government magnetic fusion energy institutions. The objectives of the Magnetic Fusion Energy Technology Fellowship program are: (1) to provide support for graduate study, training, and research in magnetic fusion energy technology; (2) to ensure an adequate supply of appropriately trained manpower to implement the nation's magnetic fusion energy agenda; (3) to raise the visibility of careers in magnetic fusion energy technology and to encourage students to pursue such careers; and (4) to make national magnetic fusion energy facilities available for manpower training

  13. The International Atomic Energy Agency Activities on Plasma Physics and Nuclear Fusion Research

    As a global facilitator in the nuclear field, the International Atomic Energy Agency (IAEA) encourages and assists research on controlled nuclear fusion in its Member States by fostering the exchange of scientific and technical information and promoting the exchange of scientists and experts. Within the Division of Physical and Chemical Sciences the Physics Section and the Nuclear Data Section work specifically on topics related to controlled nuclear fusion and organize conferences, technical meetings and workshops that promote information dissemination, training and education. International research is supported within Coordinated Research Projects (CRPs) and Technical Cooperation Projects, all open to all laboratories in the Member States. The International Fusion Research Council is the body that provides advice to the IAEA on programmatic orientations and activities with the view of promoting international cooperation in plasma physics and controlled nuclear fusion research and its applications. The IAEA holds one of the world’s leading fusion meetings. The biannual Fusion Energy Conference gathers more than 1000 participants from more than thirty eight countries and accommodates almost 600 scientific contributions covering the newest topics of research. Publication of the results presented is done in cooperation with the Nuclear Fusion Journal jointly published by the IAEA and IOPP. The IAEA Technical Meetings (TMs) are organised by the Agency and partly hosted by Member States to provide an opportunity for discussion on major concepts of fusion such as magnetic, inertial and pinch, and such as, for instance, steady state operation and burning plasma physics. A particular effort is put in the activities accompanying magnetic confinement research where the IAEA TMs bring together specialists to address specific issues that have a major impact on the success of fusion. Emphasis is put on topics with direct relevance to the effective use of fusion as a future

  14. Preliminary Estimation of Activated Corrosion Products in the Coolant System of Fusion Demo Reactor

    The second phase of the national program for fusion energy development in Korea starts from 2012 for design and construction of the fusion DEMO reactor. Radiological assessment for the fusion reactor is one of the key tasks to assure its licensability and the starting point of the assessment is determination of the source terms. As the first effort, the activities of the coolant due to activated corrosion product (ACP) were estimated. Data and experiences from fission reactors were used, in part, in the calculations of the ACP concentrations because of lack of operating experience for fusion reactors. The MCNPX code was used to determine neutron spectra and intensities at the coolant locations and the FISPACT code was used to estimate the ACP activities in the coolant of the fusion DEMO reactor. The calculated specific activities of the most nuclides in the fusion DEMO reactor coolant were 2-15 times lower than those in the PWR coolant, but the specific activities of 57Co and 57Ni were expected to be much higher than in the PWR coolant. The preliminary results of this study can be used to figure out the approximate radiological conditions and to establish a tentative set of radiological design criteria for the systems carrying coolant in the design phase of the fusion DEMO reactor.

  15. Exploring the membrane fusion mechanism through force-induced disassembly of HIV-1 six-helix bundle.

    Gao, Kai; Zhang, Yong; Lou, Jizhong

    2016-05-13

    Enveloped virus, such as HIV-1, employs membrane fusion mechanism to invade into host cell. HIV-1 gp41 ectodomain uses six-helix bundle configuration to accomplish this process. Using molecular dynamic simulations, we confirmed the stability of this six-helix bundle by showing high occupancy of hydrogen bonds and hydrophobic interactions. Key residues and interactions important for the bundle integration were characterized by force-induced unfolding simulations of six-helix bundle, exhibiting the collapse order of these groups of interactions. Moreover, our results in some way concerted with a previous theory that the formation of coiled-coil choose a route which involved cooperative interactions between the N-terminal and C-terminal helix. PMID:27079239

  16. Fusion of liposomes with the plasma membrane of epithelial cells: Fate of incorporated lipids as followed by freeze fracture and autoradiography of plastic sections

    Knoll, G.; Burger, K.N.J.; Bron, R.; van Meer, G.; Verkleij, A.J.

    1988-01-01

    The fusion of liposomes with the plasma membrane of influenza virus-infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells (van Meer et al., 1985. Biochemistry, 24: 3593-3602), has been analyzed by morphological techniques. The distribution of liposomal lipids over the apica

  17. Fusion of multisensor passive and active 3D imagery

    Fay, David A.; Verly, Jacques G.; Braun, Michael I.; Frost, Carl E.; Racamato, Joseph P.; Waxman, Allen M.

    2001-08-01

    We have extended our previous capabilities for fusion of multiple passive imaging sensors to now include 3D imagery obtained from a prototype flash ladar. Real-time fusion of low-light visible + uncooled LWIR + 3D LADAR, and SWIR + LWIR + 3D LADAR is demonstrated. Fused visualization is achieved by opponent-color neural networks for passive image fusion, which is then textured upon segmented object surfaces derived from the 3D data. An interactive viewer, coded in Java3D, is used to examine the 3D fused scene in stereo. Interactive designation, learning, recognition and search for targets, based on fused passive + 3D signatures, is achieved using Fuzzy ARTMAP neural networks with a Java-coded GUI. A client-server web-based architecture enables remote users to interact with fused 3D imagery via a wireless palmtop computer.

  18. Safety and personnel access aspects of low activation fusion blankets

    The use of silicon carbide and carbon materials for structural applications in fusion reactor first wall and blanket regions has been proposed and a continuing effort spent on the development of the ceramics technology. The advantages identified are an extremely low induced radioactivity inventory, a high temperature operating capability, abundant raw material resource availability, and minimized plasma impurity effects. One of the unique features of the applications of these materials to fusion reactor blanket designs is that no alloying element is needed in order to assure the specified mechanical properties such as occurs in metal alloys. The major source of long term radioactivity in these materials is impurities. The impurity elements and their concentrations carried over to the blanket structure during fabrication can be minimized by proper fabrication procedures and techniques. The safety and personnel access aspects of such fusion blankets in conjunction with the impurity element concentration are the main subjects of this paper

  19. Proceedings of a specialists' meeting on neutron activation cross sections for fission and fusion energy applications

    These proceedings of a specialists' meeting on neutron activation cross sections for fission and fusion energy applications are divided into 4 sessions bearing on: - data needs: 4 conferences - experimental work: 11 conferences - theoretical work: 4 conferences - evaluation work: 5 conferences

  20. IFMIF : International Fusion Materials Irradiation Facility Conceptual Design Activity: Executive summary

    This report is a summary of the results of the Conceptual Design Activity (CDA) on the International Fusion Materials Irradiation Facility (IFMIF), conducted during 1995 and 1996. The activity is under the auspices of the International Energy Agency (IEA) Implementing Agreement for a Programme of Research and Development on Fusion Materials. An IEA Fusion Materials Executive Subcommittee was charged with overseeing the IFMIF-CDA work. Participants in the CDA are the European Union, Japan, and the United States, with the Russian Federation as an associate member

  1. IFMIF : International Fusion Materials Irradiation Facility Conceptual Design Activity: Final report

    Martone, M. [ENEA, Centro Ricerche Frascati, Rome (Italy)

    1997-01-01

    This report documents the results of the Conceptual Design Activity (CDA) on the International Fusion Materials Irradiation Facility (IFMIF), conducted during 1995 and 1996. The activity is under the auspices of the International Energy Agency (IEA) Implementing Agreement for a Programme of Research and Development on Fusion Materials. An IEA Fusion Materials Executive Subcommittee was charged with overseeing the IFMIF-CDA work. Participants in the CDA are the European Union, Japan, and the United States, with the Russian Federation as an associate member.

  2. IFMIF : International Fusion Materials Irradiation Facility Conceptual Design Activity: Final report

    This report documents the results of the Conceptual Design Activity (CDA) on the International Fusion Materials Irradiation Facility (IFMIF), conducted during 1995 and 1996. The activity is under the auspices of the International Energy Agency (IEA) Implementing Agreement for a Programme of Research and Development on Fusion Materials. An IEA Fusion Materials Executive Subcommittee was charged with overseeing the IFMIF-CDA work. Participants in the CDA are the European Union, Japan, and the United States, with the Russian Federation as an associate member

  3. Accelerator ampersand Fusion Research Division: 1993 Summary of activities

    The Accelerator and Fusion Research Division (AFRD) is not only one of the largest scientific divisions at LBL, but also the one of the most diverse. Major efforts include: (1) investigations in both inertial and magnetic fusion energy; (2) operation of the Advanced Light Source, a state-of-the-art synchrotron radiation facility; (3) exploratory investigations of novel radiation sources and colliders; (4) research and development in superconducting magnets for accelerators and other scientific and industrial applications; and (5) ion beam technology development for nuclear physics and for industrial and biomedical applications. Each of these topics is discussed in detail in this book

  4. Accelerator & Fusion Research Division: 1993 Summary of activities

    Chew, J.

    1994-04-01

    The Accelerator and Fusion Research Division (AFRD) is not only one of the largest scientific divisions at LBL, but also the one of the most diverse. Major efforts include: (1) investigations in both inertial and magnetic fusion energy; (2) operation of the Advanced Light Source, a state-of-the-art synchrotron radiation facility; (3) exploratory investigations of novel radiation sources and colliders; (4) research and development in superconducting magnets for accelerators and other scientific and industrial applications; and (5) ion beam technology development for nuclear physics and for industrial and biomedical applications. Each of these topics is discussed in detail in this book.

  5. Inhibitory effect of mTOR activator MHY1485 on autophagy: suppression of lysosomal fusion.

    Yeon Ja Choi

    Full Text Available Autophagy is a major degradative process responsible for the disposal of cytoplasmic proteins and dysfunctional organelles via the lysosomal pathway. During the autophagic process, cells form double-membraned vesicles called autophagosomes that sequester disposable materials in the cytoplasm and finally fuse with lysosomes. In the present study, we investigated the inhibition of autophagy by a synthesized compound, MHY1485, in a culture system by using Ac2F rat hepatocytes. Autophagic flux was measured to evaluate the autophagic activity. Autophagosomes were visualized in Ac2F cells transfected with AdGFP-LC3 by live-cell confocal microscopy. In addition, activity of mTOR, a major regulatory protein of autophagy, was assessed by western blot and docking simulation using AutoDock 4.2. In the result, treatment with MHY1485 suppressed the basal autophagic flux, and this inhibitory effect was clearly confirmed in cells under starvation, a strong physiological inducer of autophagy. The levels of p62 and beclin-1 did not show significant change after treatment with MHY1485. Decreased co-localization of autophagosomes and lysosomes in confocal microscopic images revealed the inhibitory effect of MHY1485 on lysosomal fusion during starvation-induced autophagy. These effects of MHY1485 led to the accumulation of LC3II and enlargement of the autophagosomes in a dose- and time-dependent manner. Furthermore, MHY1485 induced mTOR activation and correspondingly showed a higher docking score than PP242, a well-known ATP-competitive mTOR inhibitor, in docking simulation. In conclusion, MHY1485 has an inhibitory effect on the autophagic process by inhibition of fusion between autophagosomes and lysosomes leading to the accumulation of LC3II protein and enlarged autophagosomes. MHY1485 also induces mTOR activity, providing a possibility for another regulatory mechanism of autophagy by the MHY compound. The significance of this study is the finding of a novel

  6. Activation of macrophages by lymphokines: enhancement of phagosome-lysosome fusion and killing of Coccidioides immitis.

    Beaman, L; Benjamini, E; Pappagianis, D

    1983-01-01

    Previously, it was shown that arthroconidia of Coccidioides immitis appear to inhibit phagosome-lysosome fusion and survive within normal mouse peritoneal macrophages. However, when these macrophages are exposed to antigen-stimulated T lymphocytes from immune mice, activation occurs, leading to enhanced phagosome-lysosome fusion and killing of C. immitis. Results indicate that the activation of macrophages can be effected after incubation with soluble lymphocyte product(s) (lymphokines). The ...

  7. TFEB activation promotes the recruitment of lysosomal glycohydrolases β-hexosaminidase and β-galactosidase to the plasma membrane

    Highlights: •TFEB activation promotes the increase of Hex and Gal activities. •The increase of Hex and Gal activities is related to transcriptional regulation. •TFEB promotes the recruitment of mature Hex and Gal on cell surface. -- Abstract: Lysosomes are membrane-enclosed organelles containing acid hydrolases. They mediate a variety of physiological processes, such as cellular clearance, lipid homeostasis, energy metabolism and pathogen defence. Lysosomes can secrete their content through a process called lysosome exocytosis in which lysosomes fuse with the plasma membrane realising their content into the extracellular milieu. Lysosomal exocytosis is not only responsible for the secretion of lysosomal enzymes, but it also has a crucial role in the plasma membrane repair. Recently, it has been demonstrated that lysosome response to the physiologic signals is regulated by the transcription factor EB (TFEB). In particular, lysosomal secretion is transcriptionally regulated by TFEB which induces both the docking and fusion of lysosomes with the plasma membrane. In this work we demonstrated that TFEB nuclear translocation is accompanied by an increase of mature glycohydrolases β-hexosaminidase and β-galactosidase on cell surface. This evidence contributes to elucidate an unknown TFEB biological function leading the lysosomal glycohydrolases on plasma membrane

  8. TFEB activation promotes the recruitment of lysosomal glycohydrolases β-hexosaminidase and β-galactosidase to the plasma membrane

    Magini, Alessandro [Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia (Italy); Department of Medical and Biological Sciences (DSMB), University of Udine, Udine (Italy); Polchi, Alice; Urbanelli, Lorena [Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia (Italy); Cesselli, Daniela; Beltrami, Antonio [Department of Medical and Biological Sciences (DSMB), University of Udine, Udine (Italy); Tancini, Brunella [Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia (Italy); Emiliani, Carla, E-mail: carla.emiliani@unipg.it [Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia (Italy)

    2013-10-18

    Highlights: •TFEB activation promotes the increase of Hex and Gal activities. •The increase of Hex and Gal activities is related to transcriptional regulation. •TFEB promotes the recruitment of mature Hex and Gal on cell surface. -- Abstract: Lysosomes are membrane-enclosed organelles containing acid hydrolases. They mediate a variety of physiological processes, such as cellular clearance, lipid homeostasis, energy metabolism and pathogen defence. Lysosomes can secrete their content through a process called lysosome exocytosis in which lysosomes fuse with the plasma membrane realising their content into the extracellular milieu. Lysosomal exocytosis is not only responsible for the secretion of lysosomal enzymes, but it also has a crucial role in the plasma membrane repair. Recently, it has been demonstrated that lysosome response to the physiologic signals is regulated by the transcription factor EB (TFEB). In particular, lysosomal secretion is transcriptionally regulated by TFEB which induces both the docking and fusion of lysosomes with the plasma membrane. In this work we demonstrated that TFEB nuclear translocation is accompanied by an increase of mature glycohydrolases β-hexosaminidase and β-galactosidase on cell surface. This evidence contributes to elucidate an unknown TFEB biological function leading the lysosomal glycohydrolases on plasma membrane.

  9. Reassessment of the Lineage Fusion Hypothesis for the Origin of Double Membrane Bacteria

    Swithers, Kristen S.; Fournier, Gregory P.; Anna G Green; Gogarten, J. Peter; Lapierre, Pascal

    2011-01-01

    In 2009, James Lake introduced a new hypothesis in which reticulate phylogeny reconstruction is used to elucidate the origin of Gram-negative bacteria (Nature 460: 967–971). The presented data supported the Gram-negative bacteria originating from an ancient endosymbiosis between the Actinobacteria and Clostridia. His conclusion was based on a presence-absence analysis of protein families that divided all prokaryotes into five groups: Actinobacteria, Double Membrane bacteria (DM), Clostridia, ...

  10. A minimal phycobilisome: fusion and chromophorylation of the truncated core-membrane linker and phycocyanin.

    Tang, Kun; Zeng, Xiao-Li; Yang, Yi; Wang, Zhi-Bin; Wu, Xian-Jun; Zhou, Ming; Noy, Dror; Scheer, Hugo; Zhao, Kai-Hong

    2012-07-01

    Phycobilisomes, the light-harvesting antennas in cyanobacteria and red algae, consist of an allophycocyanin core that is attached to the membrane via a core-membrane linker, and rods comprised of phycocyanin and often also phycoerythrin or phycoerythrocyanin. Phycobiliproteins show excellent energy transfer among the chromophores that renders them biomarkers with large Stokes-shifts absorbing over most of the visible spectrum and into the near infrared. Their application is limited, however, due to covalent binding of the chromophores and by solubility problems. We report construction of a water-soluble minimal chromophore-binding unit of the red-absorbing and fluorescing core-membrane linker. This was fused to minimal chromophore-binding units of phycocyanin. After double chromophorylation with phycocyanobilin, in E. coli, the fused phycobiliproteins absorbed light in the range of 610-660nm, and fluoresced at ~670nm, similar to phycobilisomes devoid of phycoerythr(ocyan)in. The fused phycobiliprotein could also be doubly chromophorylated with phycoerythrobilin, resulting in a chromoprotein absorbing around 540-575nm, and fluorescing at ~585nm. The broad absorptions and the large Stokes shifts render these chromoproteins candidates for imaging; they may also be helpful in studying phycobilisome assembly. PMID:22465853

  11. Endocrine activity of extraembryonic membranes extends beyond placental amniotes.

    Lori C Albergotti

    Full Text Available BACKGROUND: During development, all amniotes (mammals, reptiles, and birds form extraembryonic membranes, which regulate gas and water exchange, remove metabolic wastes, provide shock absorption, and transfer maternally derived nutrients. In viviparous (live-bearing amniotes, both extraembryonic membranes and maternal uterine tissues contribute to the placenta, an endocrine organ that synthesizes, transports, and metabolizes hormones essential for development. Historically, endocrine properties of the placenta have been viewed as an innovation of placental amniotes. However, an endocrine role of extraembryonic membranes has not been investigated in oviparous (egg-laying amniotes despite similarities in their basic structure, function, and shared evolutionary ancestry. In this study, we ask whether the oviparous chorioallantoic membrane (CAM of chicken (Gallus gallus has the capability to synthesize and receive signaling of progesterone, a major placental steroid hormone. METHODOLOGY/PRINCIPAL FINDINGS: We quantified mRNA expression of key steroidogenic enzymes involved in progesterone synthesis and found that 3beta-hydroxysteroid dehydrogenase, which converts pregnenolone to progesterone exhibited a 464 fold increase in the CAM from day 8 to day 18 of embryonic development (F(5, 68 = 89.282, p<0.0001. To further investigate progesterone synthesis, we performed explant culture and found that the CAM synthesizes progesterone in vitro in the presence of a steroid precursor. Finally, we quantified mRNA expression and performed protein immunolocalization of the progesterone receptor in the CAM. CONCLUSIONS/SIGNIFICANCE: Collectively, our data indicate that the chick CAM is steroidogenic and has the capability to both synthesize progesterone and receive progesterone signaling. These findings represent a paradigm shift in evolutionary reproductive biology by suggesting that endocrine activity of extraembryonic membranes is not a novel characteristic of

  12. Hydrodynamic collective effects of active proteins in biological membranes

    Koyano, Yuki; Mikhailov, Alexander S

    2016-01-01

    Lipid bilayers forming biological membranes are known to behave as viscous 2D fluids on submicrometer scales; usually they contain a large number of active protein inclusions. Recently, it has been shown [Proc. Nat. Acad. Sci. USA 112, E3639 (2015)] that such active proteins should in- duce non-thermal fluctuating lipid flows leading to diffusion enhancement and chemotaxis-like drift for passive inclusions in biomembranes. Here, a detailed analytical and numerical investigation of such effects is performed. The attention is focused on the situations when proteins are concentrated within lipid rafts. We demonstrate that passive particles tend to become attracted by active rafts and are accumulated inside them.

  13. Accelerator and Fusion Research Division: 1987 summary of activities

    1988-04-01

    An overview of the design and the initial studies for the Advanced Light Source is given. The research efforts for the Center for X-Ray Optics include x-ray imaging, multilayer mirror technology, x-ray sources and detectors, spectroscopy and scattering, and synchrotron radiation projects. The Accelerator Operations highlights include the research by users in nuclear physics, biology and medicine. The upgrade of the Bevalac is also discussed. The High Energy Physics Technology review includes the development of superconducting magnets and superconducting cables. A review of the Heavy-Ion Fusion Accelerator Research is also presented. The Magnetic Fusion Energy research included the development of ion sources, accelerators for negative ions, diagnostics, and theoretical plasma physics. (WRF)

  14. Accelerator and Fusion Research Division: 1987 summary of activities

    An overview of the design and the initial studies for the Advanced Light Source is given. The research efforts for the Center for X-Ray Optics include x-ray imaging, multilayer mirror technology, x-ray sources and detectors, spectroscopy and scattering, and synchrotron radiation projects. The Accelerator Operations highlights include the research by users in nuclear physics, biology and medicine. The upgrade of the Bevalac is also discussed. The High Energy Physics Technology review includes the development of superconducting magnets and superconducting cables. A review of the Heavy-Ion Fusion Accelerator Research is also presented. The Magnetic Fusion Energy research included the development of ion sources, accelerators for negative ions, diagnostics, and theoretical plasma physics

  15. Activity report of the fusion neutronics source from April 1, 2001 to March 31, 2004

    The Fusion Neutronics Source (FNS) is an accelerator based 14 MeV neutron generator established in 1981. FNS is a powerful tool for neutronics research aiming the fusion reactor development such as neutron cross section measurements, integral experiments and blanket neutronics experiments. This report reviews the FNS activities in the period from April 1, 2001 to March 31, 2004, including collaboration with universities and other research institutes. The 35 papers are indexed individually. (J.P.N.)

  16. Novel phenolic glycolipids: antioxidant activity and effect on membrane models

    Sainvitu, Pauline; Nasir, Mehmet Nail; Draguet, Florian; Nott, Katherine; Lins, Laurence; Crowet, Jean-Marc; Willems, Luc; Cosse, Jean-Philippe; Jérôme, Christine; Deleu, Magali

    2013-01-01

    Aromatic glycolipids are of both medical as well as pharmaceutical interest. Antimicrobial, antiviraland antiinflammatory activities have been reported (Otto, 2000, Journal of Molecular Catalysis B: Enzymatic). Moreover, they are expected to have interesting antioxidant properties when they contain phenolic groups. The alkyl chain should enhance their ability to penetrate into the cellular membrane (Nicolosi, 2002, Journal of Molecular Catalysis B: Enzymatic). The presence of a sugar unit co...

  17. Activated sludge filterability and full-scale membrane bioreactor operation

    Krzeminski, P.

    2013-01-01

    Despite continuous developments in the field of MBR technology, membrane fouling together with the associated energy demand and related costs issues remain major challenges. The efficiency of the filtration process in an MBR is governed by the activated sludge filterability, which is still limitedly understood and is determined by the interactions between the biomass, the wastewater and the applied process conditions. The purpose of this thesis is to increase understanding of the factors impa...

  18. Membraner

    Bach, Finn

    2009-01-01

    Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner......Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner...

  19. Erythrocyte membrane stabilization effect and antioxidant activity of methyl methacrylate

    Methyl methacrylate (MMK) is a synthetic product with mild impact on human health that is not well studied on cellular basis. Here, human erythrocytes were used to investigate the effects MMK exerts on acid and heat-induced hemolysis. Biphasic effect of MMK was observed for acid-induced hemolysis; i.e., protection at low (0 - 0.05% v/v) and stimulation at higher (0.1- 0.4% v/v) concentrations. The maximal protective effect was produced at 0.03% (v/v). At this concentration MMK increased the temperatures of heat denaturation of erythrocyte membrane proteins, spectrin and integral proteins, by about 20C and inhibited the heat-induced hemolysis by 20 %. This membrane stabilization effect of MMK is similar to that produced by some anti-inflammatory and antirheumatic drugs. The increased acid resistance possibly indicated anti-oxidant properties of MMK. The nonenzymatic antioxidant activity test evidenced that MMK has no superoxide dismutase-like activity but demonstrates strong catalase-like activity (about 900 kU/mmol at 0.05-0.1 mmol/l concentration). The results indicate that at low concentration MMK exerts benign effect on cellular membrane that could find therapeutic usage. (author)

  20. Study of synthesis parameters and active layer morphology of interfacially polymerized polyamide-polysulfone membranes

    Hermans, Sanne; Bernstein, Roy; Volodin, Alexander; Vankelecom, Ivo

    2015-01-01

    Thin film composite (TFC) polyamide membranes were prepared on a polysulfone support membrane and the effect of various synthesis conditions on the active layer morphology, the physicochemical properties and the membrane performance was investigated. The support membrane porosity factor had a significant effect on the TFC membrane performance. A polyamide top layer was formed within 15 s of reaction. Prolonging the reaction time, although resulting in a thicker active layer, only had a minor ...

  1. The GTPase-Activating Enzyme Gyp1p Is Required for Recycling of Internalized Membrane Material by Inactivation of the Rab/Ypt GTPase Ypt1p

    Lafourcade, Céline; Galan, Jean-Marc; Gloor, Yvonne; Haguenauer-Tsapis, Rosine; Peter, Matthias

    2004-01-01

    Rab/Ypt GTPases are key regulators of membrane trafficking and together with SNARE proteins mediate selective fusion of vesicles with target compartments. A family of GTPase-activating enzymes (GAPs) specific for Rab/Ypt GTPases has been discovered, but little is known about their function and substrate specificity in vivo. Here we show that the GAP activity of Gyp1p, a yeast member of this family, is specifically required for recycling of the SNARE Snc1p and the membrane dye FM4-64, implying...

  2. Photosynthesis Activates Plasma Membrane H+-ATPase via Sugar Accumulation.

    Okumura, Masaki; Inoue, Shin-Ichiro; Kuwata, Keiko; Kinoshita, Toshinori

    2016-05-01

    Plant plasma membrane H(+)-ATPase acts as a primary transporter via proton pumping and regulates diverse physiological responses by controlling secondary solute transport, pH homeostasis, and membrane potential. Phosphorylation of the penultimate threonine and the subsequent binding of 14-3-3 proteins in the carboxyl terminus of the enzyme are required for H(+)-ATPase activation. We showed previously that photosynthesis induces phosphorylation of the penultimate threonine in the nonvascular bryophyte Marchantia polymorpha However, (1) whether this response is conserved in vascular plants and (2) the process by which photosynthesis regulates H(+)-ATPase phosphorylation at the plasma membrane remain unresolved issues. Here, we report that photosynthesis induced the phosphorylation and activation of H(+)-ATPase in Arabidopsis (Arabidopsis thaliana) leaves via sugar accumulation. Light reversibly phosphorylated leaf H(+)-ATPase, and this process was inhibited by pharmacological and genetic suppression of photosynthesis. Immunohistochemical and biochemical analyses indicated that light-induced phosphorylation of H(+)-ATPase occurred autonomously in mesophyll cells. We also show that the phosphorylation status of H(+)-ATPase and photosynthetic sugar accumulation in leaves were positively correlated and that sugar treatment promoted phosphorylation. Furthermore, light-induced phosphorylation of H(+)-ATPase was strongly suppressed in a double mutant defective in ADP-glucose pyrophosphorylase and triose phosphate/phosphate translocator (adg1-1 tpt-2); these mutations strongly inhibited endogenous sugar accumulation. Overall, we show that photosynthesis activated H(+)-ATPase via sugar production in the mesophyll cells of vascular plants. Our work provides new insight into signaling from chloroplasts to the plasma membrane ion transport mechanism. PMID:27016447

  3. Antiviral activity of squalamine: Role of electrostatic membrane binding

    Beckerman, Bernard; Qu, Wei; Mishra, Abhijit; Zasloff, Michael; Wong, Gerard; Luijten, Erik

    2012-02-01

    Recent workootnotetextM. Zasloff et al., Proc. Nat. Acad. Sci. (USA) 108, 15978 (2011). has demonstrated that squalamine, a molecule found in the liver of sharks, exhibits broad-spectrum antiviral properties. It has been proposed that this activity results from the charge-density matching of squalamine and phospholipid membranes, causing squalamine to bind to membranes and displace proteins such as Rac1 that are crucial for the viral replication cycle. Here we investigate this hypothesis by numerical simulation of a coarse-grained model for the competition between Rac1 and squalamine in binding affinity to a flat lipid bilayer. We perform free-energy calculations to test the ability of squalamine to condense stacked bilayer systems and thereby displace bulkier Rac1 molecules. We directly compare our findings to small-angle x-ray scattering results for the same setup.

  4. A preliminary study on the activation analysis for a conceptual K DEMO fusion reactor

    It is known that the radioactive materials of nuclear fusion reactor have a low radioactivity in comparison with fission reactors. However, in the case of a specific module which is directly facing the fusion plasma, its radioactivity is so high that the module is needed to control and decommission by the legal regulation. The typical modules which have such a characteristic are Blanket Module (BM) and Divertor (Dv). These modules are regularly substituted with new module when irradiated sufficiently, so an accurate assessment of neutron activation is mandatory for those components of the fusion reactors because of the environmental effect and safety advantages. In order to reduce the neutron activation effect, two primary considerations are required. One is the use of the materials which abate the radioactive effects, and the other is the development of effective designs for low neutron activation. Various research groups of the ITER member countries have been performing the researches to satisfy these requirements. As part of the international research trends on neutron activation fields, the Korean research group has been designing the demonstration fusion reactor, which is named K DEMO. In this study, the activation calculations of K DEMO were carried out, and then those calculation results were compared with the calculation results of ITER model. In addition to two main modules, the activation calculations for Vacuum Vessel (VV) were performed, because that component represents the containment integrity of the fusion reactor. Neutron flux distributions in the fusion reactor were provided by a MCNP calculation. The activation calculations were performed by FISPACT 2007 code. The calculated fluxes were employed to FISPACT for the activation calculation

  5. Preparation and functional characterization of human vascular endothelial growth factor-melittin fusion protein with analysis of the antitumor activity in vitro and in vivo.

    Wang, Dingding; Hu, Lili; Su, Manman; Wang, Ju; Xu, Tianmin

    2015-09-01

    Vascular endothelial growth factor and its tyrosine kinase receptors have been identified as key mediators of the regulation of pathologic blood vessel growth and maintenance in the promotion of angiogenesis and tumor growth. Therefore, an alternative approach to destroying tumor endothelium would be to make this tissue particularly sensitive to VEGF-mediated drug delivery. To verify this hypothesis, we generated a protein containing VEGF165 fused to melittin. Melittin is a small linear peptide composed of 26 amino acid residues that can exert toxic or inhibitory effects on many types of tumor cells. This protein is a cytolytic peptide that attacks lipid membranes, leading to significant toxicity. In the present study, the Pichia pastoris expression system was used to express the fusion protein. Under optimal conditions, stable VEGF165-melittin production was achieved using a series of purification steps. The activity of VEGF165-melittin fusion protein was compared with melittin for its ability to suppress the growth of tumor cell line in vitro. The fusion toxin selectively inhibited growth of human hepatocellular carcinoma HepG-2 cell line with high expression of VEGFR-2. We found that sensitivity of VEGFR-2 transfected 293 cells to VEGF165-melittin enhanced as the cellular VEGFR-2 density increased. In an in vivo initial experiment, the fusion protein inhibited tumor growth in xenografts assays. Furthermore, successful expression and characterization of the fusion protein demonstrated its efficacy for use as a novel treatment strategy for cancer. PMID:26166416

  6. Enhanced bactericidal potency of nanoliposomes by modification of the fusion activity between liposomes and bacterium

    Ma YF

    2013-06-01

    Full Text Available Yufan Ma,1 Zhao Wang,1,2 Wen Zhao,1 Tingli Lu,1 Rutao Wang,1,2 Qibing Mei,1 Tao Chen1–3 1Key Laboratory for Space Bioscience and Biotechnology, School of Life Sciences, Northwestern Polytechnical University, Xi'an, Shaanxi, People's Republic of China; 2Shaanxi Liposome Research Center, Xi'an, Shaanxi, People's Republic of China; 3Xi'an Libang Pharmaceuticals Co, Ltd, Xi'an, People's Republic of China Background: Pseudomonas aeruginosa represents a good model of antibiotic resistance. These organisms have an outer membrane with a low level of permeability to drugs that is often combined with multidrug efflux pumps, enzymatic inactivation of the drug, or alteration of its molecular target. The acute and growing problem of antibiotic resistance of Pseudomonas to conventional antibiotics made it imperative to develop new liposome formulations to overcome these mechanisms, and investigate the fusion between liposome and bacterium. Methods: The rigidity, stability and charge properties of phospholipid vesicles were modified by varying the cholesterol, 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE, and negatively charged lipids 1,2-dimyristoyl-sn-glycero-3-phosphoglycerol sodium salt (DMPG, 1,2-dimyristoyl-sn-glycero-3-phopho-L-serine sodium salt (DMPS, 1,2-dimyristoyl-sn-glycero-3-phosphate monosodium salt (DMPA, nature phosphatidylserine sodium salt from brain and nature phosphatidylinositol sodium salt from soybean concentrations in liposomes. Liposomal fusion with intact bacteria was monitored using a lipid-mixing assay. Results: It was discovered that the fluid liposomes-bacterium fusion is not dependent on liposomal size and lamellarity. A similar degree of fusion was observed for liposomes with a particle size from 100 to 800 nm. The fluidity of liposomes is an essential pre-request for liposomes fusion with bacteria. Fusion was almost completely inhibited by incorporation of cholesterol into fluid liposomes. The increase in the

  7. Palmitoylation of SARS-CoV S protein is necessary for partitioning into detergent-resistant membranes and cell-cell fusion but not interaction with M protein

    Coronaviruses are enveloped RNA viruses that generally cause mild disease in humans. However, the recently emerged coronavirus that caused severe acute respiratory syndrome (SARS-CoV) is the most pathogenic human coronavirus discovered to date. The SARS-CoV spike (S) protein mediates virus entry by binding cellular receptors and inducing fusion between the viral envelope and the host cell membrane. Coronavirus S proteins are palmitoylated, which may affect function. Here, we created a non-palmitoylated SARS-CoV S protein by mutating all nine cytoplasmic cysteine residues. Palmitoylation of SARS-CoV S was required for partitioning into detergent-resistant membranes and for cell-cell fusion. Surprisingly, however, palmitoylation of S was not required for interaction with SARS-CoV M protein. This contrasts with the requirement for palmitoylation of mouse hepatitis virus S protein for interaction with M protein and may point to important differences in assembly and infectivity of these two coronaviruses.

  8. Mitochondrial matrix delivery using MITO-Porter, a liposome-based carrier that specifies fusion with mitochondrial membranes

    Yasuzaki, Yukari; Yamada, Yuma [Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Kita-ku, Sapporo 060-0812 (Japan); Harashima, Hideyoshi, E-mail: harasima@pharm.hokudai.ac.jp [Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Kita-ku, Sapporo 060-0812 (Japan)

    2010-06-25

    Mitochondria are the principal producers of energy in cells of higher organisms. It was recently reported that mutations and defects in mitochondrial DNA (mtDNA) are associated with various mitochondrial diseases including a variety of neurodegenerative and neuromuscular diseases. Therefore, an effective mitochondrial gene therapy and diagnosis would be expected to have great medical benefits. To achieve this, therapeutic agents need to be delivered into the innermost mitochondrial space (mitochondrial matrix), which contains the mtDNA pool. We previously reported on the development of MITO-Porter, a liposome-based carrier that introduces macromolecular cargos into mitochondria via membrane fusion. In this study, we provide a demonstration of mitochondrial matrix delivery and the visualization of mitochondrial genes (mtDNA) in living cells using the MITO-Porter. We first prepared MITO-Porter containing encapsulated propidium iodide (PI), a fluorescent dye used to stain nucleic acids to detect mtDNA. We then confirmed the emission of red-fluorescence from PI by conjugation with mtDNA, when the carriers were incubated in the presence of isolated rat liver mitochondria. Finally, intracellular observation by confocal laser scanning microscopy clearly verified that the MITO-Porter delivered PI to the mitochondrial matrix.

  9. Mitochondrial matrix delivery using MITO-Porter, a liposome-based carrier that specifies fusion with mitochondrial membranes

    Mitochondria are the principal producers of energy in cells of higher organisms. It was recently reported that mutations and defects in mitochondrial DNA (mtDNA) are associated with various mitochondrial diseases including a variety of neurodegenerative and neuromuscular diseases. Therefore, an effective mitochondrial gene therapy and diagnosis would be expected to have great medical benefits. To achieve this, therapeutic agents need to be delivered into the innermost mitochondrial space (mitochondrial matrix), which contains the mtDNA pool. We previously reported on the development of MITO-Porter, a liposome-based carrier that introduces macromolecular cargos into mitochondria via membrane fusion. In this study, we provide a demonstration of mitochondrial matrix delivery and the visualization of mitochondrial genes (mtDNA) in living cells using the MITO-Porter. We first prepared MITO-Porter containing encapsulated propidium iodide (PI), a fluorescent dye used to stain nucleic acids to detect mtDNA. We then confirmed the emission of red-fluorescence from PI by conjugation with mtDNA, when the carriers were incubated in the presence of isolated rat liver mitochondria. Finally, intracellular observation by confocal laser scanning microscopy clearly verified that the MITO-Porter delivered PI to the mitochondrial matrix.

  10. Retinoic acid inhibits calmodulin binding to human erythrocyte membranes and reduces membrane Ca2(+)-adenosine triphosphatase activity.

    Davis, F B; Smith, T. J.; Deziel, M R; Davis, P J; Blas, S D

    1990-01-01

    Ca2(+)-ATPase activity in human red cell membranes is dependent on the presence of calmodulin. All trans-retinoic acid inhibited human red cell membrane Ca2(+)-ATPase activity in vitro in a concentration-dependent manner (10(-8) to 10(-4) M). In contrast, retinol, retinal, 13-cis-retinoic acid and the benzene ring analogue of retinoic acid did not alter enzyme activity. Purified calmodulin (up to 500 ng/ml, 3 X 10(-8) M) added to red cell membranes, in the presence of inhibitory concentration...

  11. Modulation of Erythrocyte Plasma Membrane Redox System Activity by Curcumin

    Prabhakar Singh

    2016-01-01

    Full Text Available Plasma membrane redox system (PMRS is an electron transport chain system ubiquitously present throughout all cell types. It transfers electron from intracellular substrates to extracellular acceptors for regulation of redox status. Curcumin, isolated from Curcuma longa, has modulatory effects on cellular physiology due to its membrane interaction ability and antioxidant potential. The present study investigates the effect of curcumin on PMRS activity of erythrocytes isolated from Wistar rats in vitro and in vivo and validated through an in silico docking simulation study using Molegro Virtual Docker (MVD. Effects of curcumin were also evaluated on level of glutathione (GSH and the oxidant potential of plasma measured in terms of plasma ferric equivalent oxidative potentials (PFEOP. Results show that curcumin significantly (p<0.01 downregulated the PMRS activity in a dose-dependent manner. Molecular docking results suggest that curcumin interacts with amino acids at the active site cavity of cytochrome b5 reductase, a key constituent of PMRS. Curcumin also increased the GSH level in erythrocytes and plasma while simultaneously decreasing the oxidant potential (PFEOP of plasma. Altered PMRS activity and redox status are associated with the pathophysiology of several health complications including aging and diabetes; hence, the above finding may explain part of the role of curcumin in health beneficial effects.

  12. Measurements of fusion neutron yields by neutron activation technique: Uncertainty due to the uncertainty on activation cross-sections

    The neutron activation technique is routinely used in fusion experiments to measure the neutron yields. This paper investigates the uncertainty on these measurements as due to the uncertainties on dosimetry and activation reactions. For this purpose, activation cross-sections were taken from the International Reactor Dosimetry and Fusion File (IRDFF-v1.05) in 640 groups ENDF-6 format for several reactions of interest for both 2.5 and 14 MeV neutrons. Activation coefficients (reaction rates) have been calculated using the neutron flux spectra at JET vacuum vessel, both for DD and DT plasmas, calculated by MCNP in the required 640-energy group format. The related uncertainties for the JET neutron spectra are evaluated as well using the covariance data available in the library. These uncertainties are in general small, but not negligible when high accuracy is required in the determination of the fusion neutron yields

  13. LysoPC acyltransferase/PC transacylase activities in plant plasma membrane and plasma membrane-associated endoplasmic reticulum

    Tjellström Henrik

    2007-11-01

    Full Text Available Abstract Background The phospholipids of the plant plasma membrane are synthesized in the endoplasmic reticulum (ER. The majority of these lipids reach the plasma membrane independently of the secretory vesicular pathway. Phospholipid delivery to the mitochondria and chloroplasts of plant cells also bypasses the secretory pathway and here it has been proposed that lysophospholipids are transported at contact sites between specific regions of the ER and the respective organelle, followed by lysophospholipid acylation in the target organelle. To test the hypothesis that a corresponding mechanism operates to transport phospholipids to the plasma membrane outside the secretory pathway, we investigated whether lysolipid acylation occurs also in the plant plasma membrane and whether this membrane, like the chloroplasts and mitochondria, is in close contact with the ER. Results The plant plasma membrane readily incorporated the acyl chain of acyl-CoA into phospholipids. Oleic acid was preferred over palmitic acid as substrate and acyl incorporation occurred predominantly into phosphatidylcholine (PC. Phospholipase A2 stimulated the reaction, as did exogenous lysoPC when administered in above critical micellar concentrations. AgNO3 was inhibitory. The lysophospholipid acylation reaction was higher in a membrane fraction that could be washed off the isolated plasma membranes after repeated freezing and thawing cycles in a medium with lowered pH. This fraction exhibited several ER-like characteristics. When plasma membranes isolated from transgenic Arabidopsis expressing green fluorescent protein in the ER lumen were observed by confocal microscopy, membranes of ER origin were associated with the isolated plasma membranes. Conclusion We conclude that a lysoPC acylation activity is associated with plant plasma membranes and cannot exclude a PC transacylase activity. It is highly plausible that the enzyme(s resides in a fraction of the ER, closely

  14. Lateralized Difference in Tympanic Membrane Temperature: Emotion and Hemispheric Activity

    Ruth E Propper

    2013-03-01

    Full Text Available We review literature examining relationships between tympanic membrane temperature (TMT, affective/motivational orientation, and hemispheric activity. Lateralized differences in TMT might enable real-time monitoring of hemispheric activity in real-world conditions, and could serve as a corroborating marker of mental illnesses associated with specific affective dysregulation. We support the proposal that TMT holds potential for broadly indexing lateralized brain physiology during tasks demanding the processing and representation of emotional and/or motivational states, and for predicting trait-related affective/motivational orientations. The precise nature of the relationship between TMT and brain physiology, however, remains elusive. Indeed the limited extant research has sampled different participant populations and employed largely different procedures and measures, making for seemingly discrepant findings and implications. We propose, however, that many of these discrepancies can be resolved by considering how emotional states map onto motivational systems, and further examining how validated methods for inducing lateralized brain activity might affect TMT.

  15. v-SNARE transmembrane domains function as catalysts for vesicle fusion.

    Dhara, Madhurima; Yarzagaray, Antonio; Makke, Mazen; Schindeldecker, Barbara; Schwarz, Yvonne; Shaaban, Ahmed; Sharma, Satyan; Böckmann, Rainer A; Lindau, Manfred; Mohrmann, Ralf; Bruns, Dieter

    2016-01-01

    Vesicle fusion is mediated by an assembly of SNARE proteins between opposing membranes, but it is unknown whether transmembrane domains (TMDs) of SNARE proteins serve mechanistic functions that go beyond passive anchoring of the force-generating SNAREpin to the fusing membranes. Here, we show that conformational flexibility of synaptobrevin-2 TMD is essential for efficient Ca(2+)-triggered exocytosis and actively promotes membrane fusion as well as fusion pore expansion. Specifically, the introduction of helix-stabilizing leucine residues within the TMD region spanning the vesicle's outer leaflet strongly impairs exocytosis and decelerates fusion pore dilation. In contrast, increasing the number of helix-destabilizing, ß-branched valine or isoleucine residues within the TMD restores normal secretion but accelerates fusion pore expansion beyond the rate found for the wildtype protein. These observations provide evidence that the synaptobrevin-2 TMD catalyzes the fusion process by its structural flexibility, actively setting the pace of fusion pore expansion. PMID:27343350

  16. Activation Characteristics of Fuel Breeding Blanket Module in Fusion Driven Subcritical System

    HUANG Qun-Ying; LI Jian-Gang; CHEN Yi-Xue

    2004-01-01

    @@ Shortage of energy resources and production of long-lived radioactivity wastes from fission reactors are among the main problems which will be faced in the world in the near future. The conceptual design of a fusion driven subcritical system (FDS) is underway in Institute of Plasma Physics, Chinese Academy of Sciences. There are alternative designs for multi-functional blanket modules of the FDS, such as fuel breeding blanket module (FBB)to produce fuels for fission reactors, tritium breeding blanket module to produce the fuel, i.e. tritium, for fusion reactor and waste transmutation blanket module to try to permanently dispose of long-lived radioactivity wastes from fission reactors, etc. Activation of the fuel breeding blanket of the fusion driven subcritical system (FDS-FBB) by D-T fusion neutrons from the plasma and fission neutrons from the hybrid blanket are calculated and analysed under the neutron wall loading 0.5 MW/m2 and neutron fluence 15 MW. yr/m2. The neutron spectrum is calculated with the worldwide-used transport code MCNP/4C and activation calculations are carried out with the well known European inventory code FISPACT/99 with the latest released IAEA Fusion Evaluated Nuclear Data Library FENDL-2.0 and the ENDF/B-V uranium evaluated data. Induced radioactivities, dose rates and afterheats, etc, for different components of the FDS-FBB are compared and analysed.

  17. The impact of materials selection on long-term activation in fusion power plants

    Neutron-induced transmutation of materials in a D-T fusion power plant will give rise to the potential for long-term activation. To ensure that the attractive safety and environmental characteristics of fusion power are not degraded, careful design choices are necessary. An aim of optimising power plant design must be to minimise both the level of activation and the total volume of active material that might ultimately be categorised as waste requiring disposal. Materials selection is central to this optimisation. In this paper we assess the influence of materials choices for a power plant on the waste volume and the potential to clear (i.e. remove from regulatory control) and recycle material. Although the use of low activation materials in regions of high neutron flux is an important part of the strategy to minimise the level of activation, different choices may result from a strategy aimed at minimising the volume of active waste

  18. Metabolic and environmental aspects of fusion reactor activation products: niobium

    Easterly, C.E.; Shank, K.E.

    1977-11-01

    A summary of the metabolic and environmental aspects of niobium is presented. The toxicological symptoms from exposure to niobium are given, along with lethal concentration values for acute and chronic exposures. Existing human data are presented; animal uptake and retention data are analyzed for various routes of administration. Recommended metabolic values are also presented along with comments concerning their use and appropriateness. The natural distribution of niobium is given for freshwater, seawater, and the biosphere. Concentration factors and retention of /sup 95/Nb in the environment are discussed with reference to: plant retention via leaf absorption; plant retention via root uptake; uptake in terrestrial animals from plants; uptake in freshwater organisms; uptake in marine organisms; and movement in soil. Conclusions are drawn regarding needs for future work in these areas. This review was undertaken because niobium is expected to be a key metal in the development of commercial fusion reactors. It is recognized that niobium will likely not be used in the first generation reactors as a structural material but will appear as an alloy in such materials as superconducting wire.

  19. Putative BRAF activating fusion in a medullary thyroid cancer.

    Kasaian, Katayoon; Wiseman, Sam M; Walker, Blair A; Schein, Jacqueline E; Hirst, Martin; Moore, Richard A; Mungall, Andrew J; Marra, Marco A; Jones, Steven J M

    2016-03-01

    Medullary thyroid cancer (MTC) is a malignancy of the calcitonin-producing parafollicular cells of the thyroid gland. Surgery is the only curative treatment for this cancer. External beam radiation therapy is reserved for adjuvant treatment of MTC with aggressive features. Targeted therapeutics vandetanib and cabozantinib are approved for the treatment of aggressive and metastatic tumors that are not amenable to surgery. The use of these multikinase inhibitors are supported by the observed overactivation of the RET oncoprotein in a large subpopulation of MTCs. However, not all patients carry oncogenic alterations of this kinase. Hence, there is still a need for comprehensive molecular characterization of MTC utilizing whole-genome and transcriptome-sequencing methodologies with the aim of identifying targetable mutations. Here, we describe the genomic profiles of two medullary thyroid cancers and report the presence of a putative oncogenic BRAF fusion in one. Such alterations, previously observed in other malignancies and known targets of available drugs, can benefit patients who currently have no treatment options. PMID:27148585

  20. Action of erythropoietin in vitro on rabbit reticulocyte membrane Ca2+-ATPase activity.

    Lawrence, W D; Davis, P J; Blas, S D

    1987-01-01

    The mechanism of action of erythropoietin is thought to require specific interaction with the target cell surface and involve alteration of cellular calcium metabolism. Using the rabbit reticulocyte membrane as a model of the immature red cell membrane, we investigated the effects of human recombinant erythropoietin on membrane Ca2+-ATPase (calcium pump) activity in vitro. Erythropoietin in a concentration range of 0.025 to 3.0 U/ml progressively decreased membrane Ca2+-ATPase activity by up ...

  1. Flagellar membrane fusion and protein exchange in trypanosomes; a new form of cell-cell communication? [version 1; referees: 2 approved

    Simon Imhof

    2016-04-01

    Full Text Available Diverse structures facilitate direct exchange of proteins between cells, including plasmadesmata in plants and tunnelling nanotubes in bacteria and higher eukaryotes.  Here we describe a new mechanism of protein transfer, flagellar membrane fusion, in the unicellular parasite Trypanosoma brucei. When fluorescently tagged trypanosomes were co-cultured, a small proportion of double-positive cells were observed. The formation of double-positive cells was dependent on the presence of extracellular calcium and was enhanced by placing cells in medium supplemented with fresh bovine serum. Time-lapse microscopy revealed that double-positive cells arose by bidirectional protein exchange in the absence of nuclear transfer.  Furthermore, super-resolution microscopy showed that this process occurred in ≤1 minute, the limit of temporal resolution in these experiments. Both cytoplasmic and membrane proteins could be transferred provided they gained access to the flagellum. Intriguingly, a component of the RNAi machinery (Argonaute was able to move between cells, raising the possibility that small interfering RNAs are transported as cargo. Transmission electron microscopy showed that shared flagella contained two axonemes and two paraflagellar rods bounded by a single membrane. In some cases flagellar fusion was partial and interactions between cells were transient. In other cases fusion occurred along the entire length of the flagellum, was stable for several hours and might be irreversible. Fusion did not appear to be deleterious for cell function: paired cells were motile and could give rise to progeny while fused. The motile flagella of unicellular organisms are related to the sensory cilia of higher eukaryotes, raising the possibility that protein transfer between cells via cilia or flagella occurs more widely in nature.

  2. Activation of TRPA1 by membrane permeable local anesthetics

    Kronewald Sergej

    2011-08-01

    Full Text Available Abstract Background Low concentrations of local anesthetics (LAs suppress cellular excitability by inhibiting voltage-gated Na+ channels. In contrast, LAs at high concentrations can be excitatory and neurotoxic. We recently demonstrated that LA-evoked activation of sensory neurons is mediated by the capsaicin receptor TRPV1, and, to a lesser extent by the irritant receptor TRPA1. LA-induced activation and sensitization of TRPV1 involves a domain that is similar, but not identical to the vanilloid-binding domain. Additionally, activation of TRPV1 by LAs involves PLC and PI(4,5P2-signalling. In the present study we aimed to characterize essential structural determinants for LA-evoked activation of TRPA1. Results Recombinant rodent and human TRPA1 were expressed in HEK293t cells and investigated by means of whole-cell patch clamp recordings. The LA lidocaine activates TRPA1 in a concentration-dependent manner. The membrane impermeable lidocaine-derivative QX-314 is inactive when applied extracellularly. Lidocaine-activated TRPA1-currents are blocked by the TRPA1-antagonist HC-030031. Lidocaine is also an inhibitor of TRPA1, an effect that is more obvious in rodent than in human TRPA1. This species-specific difference is linked to the pore region (transmembrane domain 5 and 6 as described for activation of TRPA1 by menthol. Unlike menthol-sensitivity however, lidocaine-sensitivity is not similarly determined by serine- and threonine-residues within TM5. Instead, intracellular cysteine residues known to be covalently bound by reactive TRPA1-agonists seem to mediate activation of TRPA1 by LAs. Conclusions The structural determinants involved in activation of TRPA1 by LAs are disparate from those involved in activation by menthol or those involved in activation of TRPV1 by LAs.

  3. Activation analysis on HT-7U fusion experimental device

    Activation, dose rate and self-shielding effect of HT-7U device were calculated and analyzed by using one-dimensional discrete coordinate transport calculation code ANISN and the activation calculation code AFDKR with their data libraries based on one-dimensional model. Neutron spectrum, induced γ spectrum and their dose rate to the neighbour inhabitants and activity level were given and estimated

  4. Fusion material transmutation and activation analysis induced by fast neutrons: Anita-IEAF Activation Code Package

    This paper presents the Anita-IEAF code package for the activation characterization of materials exposed to neutrons with energies above 20 MeV. Its origins trace back to the Anita-2000 code (NEA-1638, RSICC CCC-606). Anita-IEAF is able to manage the many reaction channels for neutron energies up to 150 MeV. It computes the radioactive inventories of materials exposed to neutron irradiation, continuously or stepwise. It provides activity, isotopic nuclide density, decay heat, biological hazard, clearance index and gamma ray source spectra at shutdown and at different cooling times. The code package is provided with a complete database that includes neutron activation data library, decay, hazard and clearance data library, and gamma library. The Anita-IEAF neutron activation library was produced by processing the IEAF-2001 data activation files that have been recently released by FZK. It contains the neutron activation cross-sections for 679 nuclides in the 256 neutron energy group structure up to 150 MeV, in EAF format. That group structure includes the standard Vitamin-J 175 groups for energies below 20 MeV and 81 groups for the highest energies. The paper presents also an application of the Anita-IEAF code package to the neutron exposure characterization for the SS-316 liner and heat shield of the Test Cell area of the International Fusion Materials Irradiation Facility (IFMIF). The decay gamma source evaluation for SS-316, needed for dose rate calculations at beam-off IFMIF phase for shielding analysis, is discussed too. (authors)

  5. The Activities of the European Consortium on Nuclear Data Development and Analysis for Fusion

    This paper presents an overview of the activities of the European Consortium on Nuclear Data Development and Analysis for Fusion. The Consortium combines available European expertise to provide services for the generation, maintenance, and validation of nuclear data evaluations and data files relevant for ITER, IFMIF and DEMO, as well as codes and software tools required for related nuclear calculations

  6. The Activities of the European Consortium on Nuclear Data Development and Analysis for Fusion

    Fischer, U., E-mail: ulrich.fischer@kit.edu [Karlsruhe Institute of Technology, Institute for Neutron Physic and Reactor Technology, 76344 Eggenstein-Leopoldshafen (Germany); Avrigeanu, M.; Avrigeanu, V. [Horia Hulubei National Institute of Physics and Nuclear Engineering (IFIN-HH), RO-077125 Magurele (Romania); Cabellos, O. [Departamento de Ingenieria Nuclear, Universidad Politecnica de Madrid, 28006 Madrid (Spain); Kodeli, I. [Jozef Stefan Institute (JSI), Jamova 39, 1000 Ljubljana (Slovenia); Koning, A. [Nuclear Research and Consultancy Group (NRG), Westerduinweg 3, 1755 LE Petten (Netherlands); Konobeyev, A.Yu. [Karlsruhe Institute of Technology, Institute for Neutron Physic and Reactor Technology, 76344 Eggenstein-Leopoldshafen (Germany); Leeb, H. [Technische Universitaet Wien, Atominstitut, Wiedner Hauptstrasse 8–10, 1040 Wien (Austria); Rochman, D. [Nuclear Research and Consultancy Group (NRG), Westerduinweg 3, 1755 LE Petten (Netherlands); Pereslavtsev, P. [Karlsruhe Institute of Technology, Institute for Neutron Physic and Reactor Technology, 76344 Eggenstein-Leopoldshafen (Germany); Sauvan, P. [Universidad Nacional de Educacion a Distancia, C. Juan del Rosal, 12, 28040 Madrid (Spain); Sublet, J.-C. [Euratom/CCFE Fusion Association, Culham Science Centre, OX14 3DB (United Kingdom); Trkov, A. [Jozef Stefan Institute (JSI), Jamova 39, 1000 Ljubljana (Slovenia); Dupont, E. [OECD Nuclear Energy Agency, Paris (France); Leichtle, D.; Izquierdo, J. [Fusion for Energy, Barcelona (Spain)

    2014-06-15

    This paper presents an overview of the activities of the European Consortium on Nuclear Data Development and Analysis for Fusion. The Consortium combines available European expertise to provide services for the generation, maintenance, and validation of nuclear data evaluations and data files relevant for ITER, IFMIF and DEMO, as well as codes and software tools required for related nuclear calculations.

  7. Historical evolution of nuclear energy systems development and related activities in JAERI. Fission, fusion, accelerator utilization

    Tone, Tatsuzo [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment

    2001-03-01

    Overview of the historical evolution of nuclear energy systems development and related activities in JAERI is given in the report. This report reviews the research and development for light water reactor, fast breeder reactor, high temperature gas reactor, fusion reactor and utilization of accelerator-based neutron source. (author)

  8. Historical evolution of nuclear energy systems development and related activities in JAERI. Fission, fusion, accelerator utilization

    Overview of the historical evolution of nuclear energy systems development and related activities in JAERI is given in the report. This report reviews the research and development for light water reactor, fast breeder reactor, high temperature gas reactor, fusion reactor and utilization of accelerator-based neutron source. (author)

  9. Rab3A is a new interacting partner of synaptotagmin I and may modulate synaptic membrane fusion through a competitive mechanism

    Highlights: • Rab3A has been found to be a novel interacting protein of synaptotagmin I. • Rab3A binds to synaptotagmin I in a Ca2+-independent manner. • KKKK motif in C2B domain of synaptotagmin I is a key site for Rab3A binding. • Rab3A competitively inhibits the binding of C2B in synaptotagmin I to syntaxin 1B. • Rab3A may regulate synaptic membrane fusion and exocytosis in a competitive manner. - Abstract: Rab3 and synaptotagmin have been reported to be the key proteins that have opposite actions but cooperatively play critical regulatory roles in selecting and limiting the number of vesicles released at central synapses. However, the exact mechanism has not been fully understood. In this study, Rab3A and synaptotagmin I, the most abundant isoforms of Rab3 and synaptotagmin, respectively, in brain were for the first time demonstrated to directly interact with each other in a Ca2+-independent manner, and the KKKK motif in the C2B domain of synaptotagmin I was a key site for the Rab3A binding, which was further confirmed by the competitive inhibition of inositol hexakisphosphate. Further studies demonstrated that Rab3A competitively affected the synaptotagmin I interaction with syntaxin 1B that was involved in membrane fusion during the synaptic vesicle exocytosis. These data indicate that Rab3A is a new synaptotagmin I interacting partner and may participate in the regulation of synaptic membrane fusion and thus the vesicle exocytosis by competitively modulating the interaction of synaptotagmin with syntaxin of the t-SNARE complex in presynaptic membranes

  10. V-ATPase, ScNhxlp and Yeast Vacuole Fusion

    Quan-Sheng Qiu

    2012-01-01

    Membrane fusion is the last step in trafficking pathways during which membrane vesicles fuse with target organelles to deliver cargos.It is a central cellular reaction that plays important roles in signal transduction,protein sorting and subcellular compartmentation.Recent progress in understanding the roles of ion transporters in vacuole fusion in yeast is summanzed in this article.It is becoming increasingly evident that the vacuolar proton pump V-ATPase and vacuolar Na+/H+ antiporter ScNhxlp are key components of the vacuole fusion machinery in yeast.Yeast ScNhxlp regulates vacuole fusion by controlling the luminal pH.V-ATPases serve a dual role in vacuolar integrity in which they regulate both vacuole fusion and fission reactions in yeast.Fission defects are epistatic to fusion defects.Vacuole fission depends on the proton translocation activity of the V-ATPase; by contrast,the fusion reaction does not need the transport activity but requires the physical presence of the proton pump.Vo,the membrane-integral sector of the V-ATPase,forms trans-complexes between the opposing vacuoles in the terminal phase of vacuole fusion where the Vo trans-complexes build a continuous proteolipid channel at the fusion site to mediate the bilayer fusion.

  11. The feasibility of recycling and clearance of active materials from fusion power plants

    Zucchetti, M. [EURATOM/ENEA Fusion Association, Politecnico di Torino (Italy)]. E-mail: massimo.zucchetti@polito.it; El-Guebaly, L.A. [University of Wisconsin-Madison, Madison, WI (United States); Forrest, R.A. [EURATOM/UKAEA Fusion Association, Culham Science Centre, Abingdon (United Kingdom); Marshall, T.D. [Idaho National Laboratory, Idaho Falls (United States); Taylor, N.P. [EURATOM/UKAEA Fusion Association, Culham Science Centre, Abingdon (United Kingdom); Tobita, K. [Japan Atomic Energy Agency, Ibaraki (Japan)

    2007-08-01

    In order to minimize the quantity of active materials that require long-term storage, arising during operation and after fusion power plant decommissioning, maximum use should be made of both recycling within the nuclear industry and clearance. For the latter, revised limits have been recently issued at the international level and in the US and Europe. In this paper the implications for fusion materials of these new levels are considered. Concerning recycling, power plant studies have employed criteria based solely on radiological parameters. Reviews of remote procedures currently used within the nuclear industry suggest that these criteria have been unduly conservative and should be revised.

  12. Measurement and analysis of activation induced in titanium with fusion peak neutrons

    Klix, A., E-mail: axel.klix@kit.edu [Association FZK-Euratom, Karlsruhe Institute of Technology, Institute for Neutron Physics and Reactor Technology, D-76344 Eggenstein-Leopoldshafen (Germany); Domula, A. [Technische Universitaet Dresden, Institute for Nuclear and Particle Physics, D-01062 Dresden (Germany); Forrest, R. [Euratom/UKAEA Fusion Association, Culham Science Centre, Abingdon OX143DB (United Kingdom); Zuber, K. [Technische Universitaet Dresden, Institute for Nuclear and Particle Physics, D-01062 Dresden (Germany)

    2011-10-01

    The intense neutron flux densities in fusion reactor blankets produce activation in the blanket materials relevant to operational safety, decommissioning, etc. The aim of the present work is to check the European Activation System EASY-2007 for its capability to predict important gamma activities induced in titanium in a fusion neutron field. Many advanced low-activation materials for fusion applications contain titanium, most notably in the breeder material Li{sub 2}TiO{sub 3}. In the present work, a small sample of Ti was irradiated with the intense DT neutron generator of Technical University of Dresden. The gamma-radioactivity following irradiation was measured and nuclide activities were derived. For each of the measured gamma activities, the corresponding value was calculated with EASY, and calculation-to-experiment ratios (C/E) were determined. EASY predicted the induced gamma activities, isotopes of scandium, well with some overestimation for {sup 47}Sc. The results of this measurement together with available EXFOR and validated state-of-the-art activation libraries are discussed.

  13. Detection of closed influenza virus hemagglutinin fusion peptide structures in membranes by backbone {sup 13}CO-{sup 15}N rotational-echo double-resonance solid-state NMR

    Ghosh, Ujjayini; Xie Li; Weliky, David P., E-mail: weliky@chemistry.msu.edu [Michigan State University, Department of Chemistry (United States)

    2013-02-15

    The influenza virus fusion peptide is the N-terminal {approx}20 residues of the HA2 subunit of the hemagglutinin protein and this peptide plays a key role in the fusion of the viral and endosomal membranes during initial infection of a cell. The fusion peptide adopts N-helix/turn/C-helix structure in both detergent and membranes with reports of both open and closed interhelical topologies. In the present study, backbone {sup 13}CO-{sup 15}N REDOR solid-state NMR was applied to the membrane-associated fusion peptide to detect the distribution of interhelical distances. The data clearly showed a large fraction of closed and semi-closed topologies and were best-fitted to a mixture of two structures that do not exchange. One of the earlier open structural models may have incorrect G13 dihedral angles derived from TALOS analysis of experimentally correct {sup 13}C shifts.

  14. Potential low-level waste disposal limits for activation products from fusion

    Hanford Engineering Development Laboratory (HEDL) scientists are involved in studies considering alternative construction materials for the first wall of commercial fusion reactors. To permit a comparison of radioactivity levels, both the level of activation and an acceptable limit for the radionuclides present must be known. Generic material composition guidelines can be developed using the US Nuclear Regulatory Commission (NRC) regulations governing the near-surface disposal of low-level radioactive wastes. These regulations consider wastes defined as containing source, special nuclear, or by-product materials arising from research, industrial, medical, and nuclear fuel-cycle activities. However, not all of the activation products produced in low-level wastes from fusion reactors are considered by the NRC in their regulations. The purpose of this report is to present potential low-level waste-disposal limits for ten radionuclides resulting from fusion reactor operations that are not considered in the NRC low-level waste regulations. These potential limits will be used by HEDL scientists to complete their generic material composition guidelines for the first wall of commercial fusion reactors

  15. GS-5806 Inhibits Pre- to Postfusion Conformational Changes of the Respiratory Syncytial Virus Fusion Protein

    Samuel, Dharmaraj; Xing, Weimei; Niedziela-Majka, Anita; Wong, Jinny S; Hung, Magdeleine; Brendza, Katherine M.; Perron, Michel; Jordan, Robert; Sperandio, David; Liu, Xiaohong; Mackman, Richard; Sakowicz, Roman

    2015-01-01

    GS-5806 is a small-molecule inhibitor of human respiratory syncytial virus fusion protein-mediated viral entry. During viral entry, the fusion protein undergoes major conformational changes, resulting in fusion of the viral envelope with the host cell membrane. This process is reproduced in vitro using a purified, truncated respiratory syncytial virus (RSV) fusion protein. GS-5806 blocked these conformational changes, suggesting a possible mechanism for antiviral activity.

  16. DNA Triplex-Based Complexes Display Anti-HIV-1-Cell Fusion Activity.

    Xu, Liang; Zhang, Tao; Xu, Xiaoyu; Chong, Huihui; Lai, Wenqing; Jiang, Xifeng; Wang, Chao; He, Yuxian; Liu, Keliang

    2015-08-01

    DNA triplexes with hydrophobic modifications were designed and evaluated for their activity as inhibitors of the cell fusion of human immunodeficiency virus type 1 (HIV-1). Triplex inhibitors displayed low micromolar activities in the cell-cell fusion assay and nanomolar activities in the anti-HIV-1 pseudovirus test. Helix structure and the presence of sufficient numbers of hydrophobic regions were essential for the antifusion activity. Results from native polyacrylamide gel electrophoresis and a fluorescent resonance energy transfer-based inhibitory assay indicated that these triplexes may interact with the primary pocket at the glycoprotein 41 (gp41) N-heptad repeat, thereby inhibiting formation of the HIV-1 gp41 6-helical bundle. Triplex-based complexes may represent a novel category of HIV-1 inhibitors in anti-HIV-1 drug discovery. PMID:26192705

  17. Binding-Site Interactions between Epstein-Barr Virus Fusion Proteins gp42 and gH/gL Reveal a Peptide That Inhibits both Epithelial and B-Cell Membrane Fusion▿

    Kirschner, Austin N.; Lowrey, Amanda S.; Longnecker, Richard; Theodore S Jardetzky

    2007-01-01

    Herpesviruses require membrane-associated glycoproteins gB, gH, and gL for entry into host cells. Epstein-Barr virus (EBV) gp42 is a unique protein also required for viral entry into B cells. Key interactions between EBV gp42 and the EBV gH/gL complex were investigated to further elucidate their roles in membrane fusion. Deletion and point mutants within the N-terminal region of gp42 revealed residues important for gH/gL binding and membrane fusion. Many five-residue deletion mutants in the N...

  18. Candida albicans actively modulates intracellular membrane trafficking in mouse macrophage phagosomes.

    Fernández-Arenas, Elena; Bleck, Christopher K E; Nombela, César; Gil, Concha; Griffiths, Gareth; Diez-Orejas, Rosalía

    2009-04-01

    The intracellular trafficking/survival strategies of the opportunistic human pathogen Candida albicans are poorly understood. Here we investigated the infection of RAW264.7 macrophages with a virulent wild-type (WT) filamentous C. albicans strain and a hyphal signalling-defective mutant (efg1Delta/cph1Delta). A comparative analysis of the acquisition by phagosomes of actin, and of early/late endocytic organelles markers of the different fungal strains was performed and related to Candida's survival inside macrophages. Our results show that both fungal strains have evolved a similar mechanism to subvert the 'lysosomal' system, as seen by the inhibition of the phagosome fusion with compartments enriched in the lysobisphosphatidic acid and the vATPase, and thereby the acquisition of a low pH from the outset of infection. Besides, the virulent WT strain displayed additional specific survival strategies to prevent its targeting to compartmentsdisplaying late endosomal/lysosomal features, such as induction of active recycling out of phagosomes of the lysosomal membrane protein LAMP-1, the lysosomal protease cathepsin D and preinternalized colloidal gold. Finally, both virulent and efg1Delta/cph1Delta mutant fungal strains actively suppressed the production of macrophage nitric oxide (NO), although their cell wall extracts were potent inducers of NO. PMID:19134116

  19. Blood-group-Ii-active gangliosides of human erythrocyte membranes

    More than ten new types of gangliosides, in addition to haematoside and sialosylparagloboside, were isolated from human erythrocyte membranes. These were separated by successive chromatographies on DAEA-Sephadex, on porous silica-gel columns and on thin-layer silica gel as acetylated compounds. Highly potent blood-group-Ii and moderate blood-group-H activities were demonstrated in some of the ganglioside fractions. The gangliosides incorporated into chlolesterol/phosphatidylcholine liposomes stoicheiometrically inhibited binding of anti-(blood-group-I and i) antibodies to a radioiodinated blood-group-Ii-active glycoprotein. The fraction with the highest blood-group-I activity, I(g) fraction, behaved like sialosyl-deca- to dodeca-glycosylceramides on t.l.c. Certain blood-group-I and most of the i-determinants were in partially or completely cryptic form and could be unmasked by sialidase treatment. Thus the I and i antigens, which are known to occur on internal structures of blood-group-ABH-active glycoproteins in secretions, also occur in the interior of the carbohydrate chains of erythrocyte gangliosides. (author)

  20. A theory of Plasma Membrane Calcium Pump stimulation and activity

    Graupner, M; Meyer-Hermann, M; Erler, Frido; Graupner, Michael; Meyer-Hermann, Michael

    2003-01-01

    The ATP-driven Plasma Membrane Calcium (PMCA) pump is characterized by a high affinity to calcium and a low transport rate compared to other transmembrane calcium transport proteins. It plays a crucial role for calcium extrusion from cells. Calmodulin is an intracellular calcium buffering protein which is capable in its Calcium-liganded form to stimulate the PMCA pump by increasing both, the affinity to calcium and the maximum calcium transport rate. We introduce a new model of this stimulation process and deduce analytical expressions for experimental observables in order to determine the model parameter on the basis of specific experiments. Furthermore a model for the pumping activity is developed. In contrast to the biological process we have to describe the pumping rate behavior by assuming a ATP:Calcium stoichiometry of 2 in order to reproduce experimental data. The conjunction of the description of calcium pumping and the stimulation model fully and correctly simulates PMCA pump function. Therewith the ...

  1. Report of the Integrated Program Planning Activity for the DOE Fusion Energy Sciences Program

    This report of the Integrated Program Planning Activity (IPPA) has been prepared in response to a recommendation by the Secretary of Energy Advisory Board that, ''Given the complex nature of the fusion effort, an integrated program planning process is an absolute necessity.'' We, therefore, undertook this activity in order to integrate the various elements of the program, to improve communication and performance accountability across the program, and to show the inter-connectedness and inter-dependency of the diverse parts of the national fusion energy sciences program. This report is based on the September 1999 Fusion Energy Sciences Advisory Committee's (FESAC) report ''Priorities and Balance within the Fusion Energy Sciences Program''. In its December 5,2000, letter to the Director of the Office of Science, the FESAC has reaffirmed the validity of the September 1999 report and stated that the IPPA presents a framework and process to guide the achievement of the 5-year goals listed in the 1999 report. The National Research Council's (NRC) Fusion Assessment Committee draft final report ''An Assessment of the Department of Energy's Office of Fusion Energy Sciences Program'', reviewing the quality of the science in the program, was made available after the IPPA report had been completed. The IPPA report is, nevertheless, consistent with the recommendations in the NRC report. In addition to program goals and the related 5-year, 10-year, and 15-year objectives, this report elaborates on the scientific issues associated with each of these objectives. The report also makes clear the relationships among the various program elements, and cites these relationships as the reason why integrated program planning is essential. In particular, while focusing on the science conducted by the program, the report addresses the important balances between the science and energy goals of the program, between the MFE and IFE approaches, and between the domestic and international aspects

  2. Report of the Integrated Program Planning Activity for the DOE Fusion Energy Sciences Program

    None

    2000-12-01

    This report of the Integrated Program Planning Activity (IPPA) has been prepared in response to a recommendation by the Secretary of Energy Advisory Board that, ''Given the complex nature of the fusion effort, an integrated program planning process is an absolute necessity.'' We, therefore, undertook this activity in order to integrate the various elements of the program, to improve communication and performance accountability across the program, and to show the inter-connectedness and inter-dependency of the diverse parts of the national fusion energy sciences program. This report is based on the September 1999 Fusion Energy Sciences Advisory Committee's (FESAC) report ''Priorities and Balance within the Fusion Energy Sciences Program''. In its December 5,2000, letter to the Director of the Office of Science, the FESAC has reaffirmed the validity of the September 1999 report and stated that the IPPA presents a framework and process to guide the achievement of the 5-year goals listed in the 1999 report. The National Research Council's (NRC) Fusion Assessment Committee draft final report ''An Assessment of the Department of Energy's Office of Fusion Energy Sciences Program'', reviewing the quality of the science in the program, was made available after the IPPA report had been completed. The IPPA report is, nevertheless, consistent with the recommendations in the NRC report. In addition to program goals and the related 5-year, 10-year, and 15-year objectives, this report elaborates on the scientific issues associated with each of these objectives. The report also makes clear the relationships among the various program elements, and cites these relationships as the reason why integrated program planning is essential. In particular, while focusing on the science conducted by the program, the report addresses the important balances between the science and energy goals of the program, between the

  3. Conformational changes in Sindbis virions resulting from exposure to low pH and interactions with cells suggest that cell penetration may occur at the cell surface in the absence of membrane fusion

    Alphaviruses have the ability to induce cell-cell fusion after exposure to acid pH. This observation has served as an article of proof that these membrane-containing viruses infect cells by fusion of the virus membrane with a host cell membrane upon exposure to acid pH after incorporation into a cell endosome. We have investigated the requirements for the induction of virus-mediated, low pH-induced cell-cell fusion and cell-virus fusion. We have correlated the pH requirements for this process to structural changes they produce in the virus by electron cryo-microscopy. We found that exposure to acid pH was required to establish conditions for membrane fusion but that membrane fusion did not occur until return to neutral pH. Electron cryo-microscopy revealed dramatic changes in the structure of the virion as it was moved to acid pH and then returned to neutral pH. None of these treatments resulted in the disassembly of the virus protein icosahedral shell that is a requisite for the process of virus membrane-cell membrane fusion. The appearance of a prominent protruding structure upon exposure to acid pH and its disappearance upon return to neutral pH suggested that the production of a 'pore'-like structure at the fivefold axis may facilitate cell penetration as has been proposed for polio (J. Virol. 74 (2000) 1342) and human rhino virus (Mol. Cell 10 (2002) 317). This transient structural change also provided an explanation for how membrane fusion occurs after return to neutral pH. Examination of virus-cell complexes at neutral pH supported the contention that infection occurs at the cell surface at neutral pH by the production of a virus structure that breaches the plasma membrane bilayer. These data suggest an alternative route of infection for Sindbis virus that occurs by a process that does not involve membrane fusion and does not require disassembly of the virus protein shell

  4. Effect of Powdered Activated Carbon to Reduce Fouling in Membrane Bioreactors: A Sustainable Solution. Case Study

    Giuseppe Mancini; Antonella Luciano; Paolo Viotti; Sabrina Copelli; Massimo Raboni; Giordano Urbini; Vincenzo Torretta

    2013-01-01

    Membrane Bio Reactors (MBRs) are mainly used for industrial wastewaters applications where their costs can be more easily afforded. High costs are basically due to energy consumption and membrane cleaning or replacement. Membrane fouling is responsible for reducing treated water production and increasing maintenance as well as operation costs. According to previous researches, the addition of Powdered Activated Carbon (PAC) in high dosages could reduce membrane fouling; but such concentration...

  5. Global epigenomic analysis indicates protocadherin-7 activates osteoclastogenesis by promoting cell–cell fusion

    Highlights: • Identification of epigenetically regulated genes during osteoclastogenesis. • Pcdh7 is regulated by H3K4me3 and H3K27me3 during osteoclastogenesis. • Pcdh7 expression is increased by RANKL during osteoclastogenesis. • Establishment of novel cell fusion analysis for osteoclasts by imaging cytometer. • Pcdh7 regulates osteoclastogenesis by promoting cell fusion related gene expressions. - Abstract: Gene expression is dependent not only on genomic sequences, but also epigenetic control, in which the regulation of chromatin by histone modification plays a crucial role. Histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 27 trimethylation (H3K27me3) are related to transcriptionally activated and silenced sequences, respectively. Osteoclasts, the multinucleated cells that resorb bone, are generated by the fusion of precursor cells of monocyte/macrophage lineage. To elucidate the molecular and epigenetic regulation of osteoclast differentiation, we performed a chromatin immunoprecipitation sequencing (ChIP-seq) analysis for H3K4me3 and H3K27me3 in combination with RNA sequencing. We focused on the histone modification change from H3K4me3(+)H3K27me3(+) to H3K4me3(+)H3K27me3(–) and identified the protocadherin-7 gene (Pcdh7) to be among the genes epigenetically regulated during osteoclastogenesis. Pcdh7 was induced by RANKL stimulation in an NFAT-dependent manner. The knockdown of Pcdh7 inhibited RANKL-induced osteoclast differentiation due to the impairment of cell–cell fusion, accompanied by a decreased expression of the fusion-related genes Dcstamp, Ocstamp and Atp6v0d2. This study demonstrates that Pcdh7 plays a key role in osteoclastogenesis by promoting cell–cell fusion

  6. Global epigenomic analysis indicates protocadherin-7 activates osteoclastogenesis by promoting cell–cell fusion

    Nakamura, Haruhiko [Department of Orthopaedic Surgery, Faculty of Medicine, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-0033 (Japan); Department of Cell Signaling, Graduate School of Medical and Dental Science, Tokyo Medical and Dental University, Yushima 1-5-45, Bunkyo-ku, Tokyo 113-8549 (Japan); Nakashima, Tomoki [Department of Cell Signaling, Graduate School of Medical and Dental Science, Tokyo Medical and Dental University, Yushima 1-5-45, Bunkyo-ku, Tokyo 113-8549 (Japan); Japan Science and Technology Agency, PRESTO, Yushima 1-5-45, Bunkyo-ku, Tokyo 113-8549 (Japan); Hayashi, Mikihito [Department of Cell Signaling, Graduate School of Medical and Dental Science, Tokyo Medical and Dental University, Yushima 1-5-45, Bunkyo-ku, Tokyo 113-8549 (Japan); Japan Science and Technology Agency, ERATO, Takayanagi Osteonetwork Project, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Izawa, Naohiro; Yasui, Tetsuro [Department of Orthopaedic Surgery, Faculty of Medicine, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-0033 (Japan); Aburatani, Hiroyuki [Genome Science Division, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8904 (Japan); Tanaka, Sakae [Department of Orthopaedic Surgery, Faculty of Medicine, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-0033 (Japan); Takayanagi, Hiroshi, E-mail: takayana@m.u-tokyo.ac.jp [Japan Science and Technology Agency, ERATO, Takayanagi Osteonetwork Project, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Department of Immunology, Graduate School of Medicine and Faculty of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan)

    2014-12-12

    Highlights: • Identification of epigenetically regulated genes during osteoclastogenesis. • Pcdh7 is regulated by H3K4me3 and H3K27me3 during osteoclastogenesis. • Pcdh7 expression is increased by RANKL during osteoclastogenesis. • Establishment of novel cell fusion analysis for osteoclasts by imaging cytometer. • Pcdh7 regulates osteoclastogenesis by promoting cell fusion related gene expressions. - Abstract: Gene expression is dependent not only on genomic sequences, but also epigenetic control, in which the regulation of chromatin by histone modification plays a crucial role. Histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 27 trimethylation (H3K27me3) are related to transcriptionally activated and silenced sequences, respectively. Osteoclasts, the multinucleated cells that resorb bone, are generated by the fusion of precursor cells of monocyte/macrophage lineage. To elucidate the molecular and epigenetic regulation of osteoclast differentiation, we performed a chromatin immunoprecipitation sequencing (ChIP-seq) analysis for H3K4me3 and H3K27me3 in combination with RNA sequencing. We focused on the histone modification change from H3K4me3(+)H3K27me3(+) to H3K4me3(+)H3K27me3(–) and identified the protocadherin-7 gene (Pcdh7) to be among the genes epigenetically regulated during osteoclastogenesis. Pcdh7 was induced by RANKL stimulation in an NFAT-dependent manner. The knockdown of Pcdh7 inhibited RANKL-induced osteoclast differentiation due to the impairment of cell–cell fusion, accompanied by a decreased expression of the fusion-related genes Dcstamp, Ocstamp and Atp6v0d2. This study demonstrates that Pcdh7 plays a key role in osteoclastogenesis by promoting cell–cell fusion.

  7. Fusion cross sections of carbon isotopes obtained with an ionization chamber in active target mode

    Carbon fusion has provided questions to both physicists and astronomers for at least the last 50 years. From fundamental nuclear structure to recent discoveries in stellar phenomena there are still open topics. Fusion in the 12C + 12C system show oscillations that are not present in neighboring systems and are yet not completely understood. Unexplained behavior in the threshold between 1p and 2s1d shells is seen as fusion cross sections show significant changes in systems which differ by only a nucleon. A new type of stellar explosions, called super bursts, in X-ray binaries were recently observed and are thought to require fusion of radioactive carbon isotopes for an explanation, opening new paths for stellar nucleosynthesis. These are a few interesting examples that motivated the development of a new measurement technique, which comprises a Multi Sampling Ionization Chamber (Music) operated in active target mode, with methane gas (C H4) as both counting gas and reaction target. This offers a high efficiency detection method where excitation functions can be sampled, using a single beam energy, in a range determined by the ionization gas pressure. This is a great advantage since it drastically reduces the measurement time and the data are automatically normalized. The high efficiency of the detector makes it ideal for experiments where the reaction cross section and/or the beam intensity are low, i.e. for processes involving radioactive nuclei. Using the Music, fusion cross sections in systems with carbon isotopes of mass numbers A = 10, 12, 13, 14, 15 impinging on a carbon-12 target have been measured. Beam energies of about 3 MeV/A were used for obtaining fusion excitation functions in the center of mass energy range between 10 and 20 MeV. In this contribution, the operation principle of the Music is discussed. Then, the experimental excitation functions are presented and compared with previous data (3when available) and different theoretical models

  8. Activation analysis of the PULSAR-II fusion power reactor

    The PULSAR-II pulsed tokamak power plant design utilizes a blanket made of the vanadium alloy, V-5Cr-5Ti, and cooled with liquid lithium. The shield is made of a mixture of the low activation austenitic steel (Tenelon) and vanadium. The blanket is assumed to be replaced every 5.6 full power years (FPY) and the shield is assumed to stay in place for 30 FPY. The activity induced in the blanket at the end of its lifetime is higher than the activity induced in the shield after 30 FPY. At shutdown, the blanket and shield activities are 2678 MCi and 1747 MCi, respectively. One year after shutdown the shield activity drops to 18 MCi compared to 84 MCi for the blanket. The total decay heat generated in the blanket at the end of its lifetime is 34.7 MW and drops to 17.6 MW within an hour. At shutdown, 25.3 MW of decay heat are generated in the shield, dropping to only 0.1 MW within the first year. One week after shutdown, the values of the integrated decay heat are 1770 GJ for the blanket and 469 GJ for the shield. The radwaste classification of the reactor structure is evaluated according to both the NRC 10CFR61 and Fetter waste disposal concentration limits. After 5.6 years of irradiation, the blanket will only qualify for Class C low level waste. After 30 years of operation, the shield will also qualify for disposal as Class C waste. Only remote maintenance will be allowed inside the containment building

  9. Biophysical Characterization of a Vaccine Candidate against HIV-1: The Transmembrane and Membrane Proximal Domains of HIV-1 gp41 as a Maltose Binding Protein Fusion.

    Zhen Gong

    Full Text Available The membrane proximal region (MPR, residues 649-683 and transmembrane domain (TMD, residues 684-705 of the gp41 subunit of HIV-1's envelope protein are highly conserved and are important in viral mucosal transmission, virus attachment and membrane fusion with target cells. Several structures of the trimeric membrane proximal external region (residues 662-683 of MPR have been reported at the atomic level; however, the atomic structure of the TMD still remains unknown. To elucidate the structure of both MPR and TMD, we expressed the region spanning both domains, MPR-TM (residues 649-705, in Escherichia coli as a fusion protein with maltose binding protein (MBP. MPR-TM was initially fused to the C-terminus of MBP via a 42 aa-long linker containing a TEV protease recognition site (MBP-linker-MPR-TM. Biophysical characterization indicated that the purified MBP-linker-MPR-TM protein was a monodisperse and stable candidate for crystallization. However, crystals of the MBP-linker-MPR-TM protein could not be obtained in extensive crystallization screens. It is possible that the 42 residue-long linker between MBP and MPR-TM was interfering with crystal formation. To test this hypothesis, the 42 residue-long linker was replaced with three alanine residues. The fusion protein, MBP-AAA-MPR-TM, was similarly purified and characterized. Significantly, both the MBP-linker-MPR-TM and MBP-AAA-MPR-TM proteins strongly interacted with broadly neutralizing monoclonal antibodies 2F5 and 4E10. With epitopes accessible to the broadly neutralizing antibodies, these MBP/MPR-TM recombinant proteins may be in immunologically relevant conformations that mimic a pre-hairpin intermediate of gp41.

  10. Low-chromium reduced-activation ferritic steels for fusion

    Klueh, R.L.; Alexander, D.J.; Kenik, E.A. [Oak Ridge National Laboratory, TN (United States)

    1996-04-01

    Development of reduced-activation ferritic steels has concentrated on high-chromium (8-10 wt% Cr) steels. However, there are advantages for a low-chromium steel, and initial ORNL studies on reduced-activation steels were on compositions with 2.25 to 12% Cr. Those studies showed an Fe-2.25Cr-2W-0.25V-0.1C (2 1/4Cr-2WV) steel to have the highest strenglth of the steels studied. Although this steel had the best strength, Charpy impact properties were inferior to those of an Fe-9Cr-2W-0.25V-0.07Ta-0.1C (9Cr-2WVTa) and an Fe-2.25Cr-2W-0.1C (2 1/4Cr-2W) steel. Therefore, further development of the low-chromium Cr-W steels was required. These results indicate that it is possible to develop low-chromium reduced-activation ferritic steels that have tensile and impact properties as good or better than those of high-chromium (7-9% Cr) steels. Further improvement of properties should be possible by optimizing the composition.

  11. LegC3, an effector protein from Legionella pneumophila, inhibits homotypic yeast vacuole fusion in vivo and in vitro.

    Terry L Bennett

    Full Text Available During infection, the intracellular pathogenic bacterium Legionella pneumophila causes an extensive remodeling of host membrane trafficking pathways, both in the construction of a replication-competent vacuole comprised of ER-derived vesicles and plasma membrane components, and in the inhibition of normal phagosome:endosome/lysosome fusion pathways. Here, we identify the LegC3 secreted effector protein from L. pneumophila as able to inhibit a SNARE- and Rab GTPase-dependent membrane fusion pathway in vitro, the homotypic fusion of yeast vacuoles (lysosomes. This vacuole fusion inhibition appeared to be specific, as similar secreted coiled-coiled domain containing proteins from L. pneumophila, LegC7/YlfA and LegC2/YlfB, did not inhibit vacuole fusion. The LegC3-mediated fusion inhibition was reversible by a yeast cytosolic extract, as well as by a purified soluble SNARE, Vam7p. LegC3 blocked the formation of trans-SNARE complexes during vacuole fusion, although we did not detect a direct interaction of LegC3 with the vacuolar SNARE protein complexes required for fusion. Additionally, LegC3 was incapable of inhibiting a defined synthetic model of vacuolar SNARE-driven membrane fusion, further suggesting that LegC3 does not directly inhibit the activity of vacuolar SNAREs, HOPS complex, or Sec17p/18p during membrane fusion. LegC3 is likely utilized by Legionella to modulate eukaryotic membrane fusion events during pathogenesis.

  12. An Rh1–GFP Fusion Protein Is in the Cytoplasmic Membrane of a White Mutant Strain of Chlamydomonas reinhardtii

    Yoshihara, Corinne; Inoue, Kentaro; Schichnes, Denise; Ruzin, Steven; Inwood, William; Kustu, Sydney

    2008-01-01

    The major Rhesus (Rh) protein of the green alga Chlamydomonas reinhardtii, Rh1, is homologous to Rh proteins of humans. It is an integral membrane protein involved in transport of carbon dioxide. To localize a fusion of intact Rh1 to the green fluorescent protein (GFP), we used as host a white (lts1) mutant strain of C. reinhardtii, which is blocked at the first step of carotenoid biosynthesis. The lts1 mutant strain accumulated normal amounts of Rh1 heterotrophically in the dark and Rh1–GFP ...

  13. Efficient multiple object tracking using mutually repulsive active membranes.

    Yi Deng

    Full Text Available Studies of social and group behavior in interacting organisms require high-throughput analysis of the motion of a large number of individual subjects. Computer vision techniques offer solutions to specific tracking problems, and allow automated and efficient tracking with minimal human intervention. In this work, we adopt the open active contour model to track the trajectories of moving objects at high density. We add repulsive interactions between open contours to the original model, treat the trajectories as an extrusion in the temporal dimension, and show applications to two tracking problems. The walking behavior of Drosophila is studied at different population density and gender composition. We demonstrate that individual male flies have distinct walking signatures, and that the social interaction between flies in a mixed gender arena is gender specific. We also apply our model to studies of trajectories of gliding Myxococcus xanthus bacteria at high density. We examine the individual gliding behavioral statistics in terms of the gliding speed distribution. Using these two examples at very distinctive spatial scales, we illustrate the use of our algorithm on tracking both short rigid bodies (Drosophila and long flexible objects (Myxococcus xanthus. Our repulsive active membrane model reaches error rates better than 5 x 10(-6 per fly per second for Drosophila tracking and comparable results for Myxococcus xanthus.

  14. Enzymatically active high-flux selectively gas-permeable membranes

    Jiang, Ying-Bing; Cecchi, Joseph L.; Rempe, Susan; FU, Yaqin; Brinker, C. Jeffrey

    2016-01-26

    An ultra-thin, catalyzed liquid transport medium-based membrane structure fabricated with a porous supporting substrate may be used for separating an object species such as a carbon dioxide object species. Carbon dioxide flux through this membrane structures may be several orders of magnitude higher than traditional polymer membranes with a high selectivity to carbon dioxide. Other gases such as molecular oxygen, molecular hydrogen, and other species including non-gaseous species, for example ionic materials, may be separated using variations to the membrane discussed.

  15. The lipidic particle as an intermediate structure in membrane fusion processes and bilayer to hexagonal HII transitions

    Verkleij, A.J.; Echteld, C.J.A. van; Gerritsen, W.J.; Cullis, P.R.; de Kruijff, B.

    1980-01-01

    Small unilamellar vesicles comprised of a mixture of phosphatidylethanolamine/phosphatidylcholine/cholesterol (3 : 1 : 2) fuse to form large multilamellar vesicles on increasing the temperature from 0 to 50°C. This event is associated with the appearance of lipidic particles at the fusion sites, consistent with a role as intermediary structures during the fusion process. Further, for phosphatidylcholine/cardiolipin (1 : 1) liposomes in the presence of Mn2+ a direct relationship between lipidi...

  16. Low-Activation structural ceramic composites for fusion power reactors: materials development and main design issues

    This paper is devoted to the development of advanced Low-Activation Materials (LAMs) with favourable short-term activation characteristics for the use as structural materials in a fusion power reactor (in order to reduce the risk associated with a major accident, in particular those related with radio-isotopes release in the environment), and to try to approach the concept of an inherently safe reactor. LA Ceramics Composites (LACCs) are the most promising LAMs because of their relatively good thermo-mechanical properties. At present, SiC/SiC composite is the only LACC considered by the fusion community, and therefore is the one having the most complete data base. The preliminary design of a breeding blanket using SiC/SiC as structural material indicated that significant improvement of its thermal conductivity is required. (orig.)

  17. Low-activation structural ceramic composites for fusion power reactors: materials development and main design issues

    Development of advanced Low-Activation Materials (LAMs) with favourable short-term activation characteristics is discussed, for the use as structural materials in a fusion power reactor (in order to reduce the risk associated with a major accident, in particular those related with radio-isotopes release in the environment), and to try to approach the concept of an inherently safe reactor. LA Ceramics Composites (LACCs) are the most promising LAMs because of their relatively good thermo-mechanical properties. At present, SiC/SiC composite is the only LACC considered by the fusion community, and therefore is the one having the most complete data base. The preliminary design of a breeding blanket using SiC/SiC as structural material indicated that significant improvement of its thermal conductivity is required. (author) 11 refs.; 3 figs

  18. Fusion of a Short HA2-Derived Peptide Sequence to Cell-Penetrating Peptides Improves Cytosolic Uptake, but Enhances Cytotoxic Activity

    Igor Kitanovic

    2009-09-01

    Full Text Available Cell-penetrating peptides (CPP have become a widely used tool for efficient cargo delivery into cells. However, one limiting fact is their uptake by endocytosis causing the enclosure of the CPP-cargo construct within endosomes. One often used method to enhance the outflow into the cytosol is the fusion of endosome-disruptive peptide or protein sequences to CPP. But, until now, no studies exist investigating the effects of the fusion peptide to the cellular distribution, structural arrangements and cytotoxic behaviour of the CPP. In this study, we attached a short modified sequence of hemagglutinin subunit HA2 to different CPP and analysed the biologic activity of the new designed peptides. Interestingly, we observed an increased cytosolic distribution but also highly toxic activities in the micromolar range against several cell lines. Structural analysis revealed that attachment of the fusion peptide had profound implications on the whole conformation of the peptide, which might be responsible for membrane interaction and endosome disruption.

  19. IFMIF - International Fusion Materials Irradiation Facility Conceptual Design Activity/Interim Report

    Environmental acceptability, safety, and economic viability win ultimately be the keys to the widespread introduction of fusion power. This will entail the development of radiation- resistant and low- activation materials. These low-activation materials must also survive exposure to damage from neutrons having an energy spectrum peaked near 14 MeV with annual radiation doses in the range of 20 displacements per atom (dpa). Testing of candidate materials, therefore, requires a high-flux source of high energy neutrons. The problem is that there is currently no high-flux source of neutrons in the energy range above a few MeV. The goal, is therefore, to provide an irradiation facility for use by fusion material scientists in the search for low-activation and damage-resistant materials. An accellerator-based neutron source has been established through a number of international studies and workshops' as an essential step for materials development and testing. The mission of the International Fusion Materials Irradiation Facility (IFMIF) is to provide an accelerator-based, deuterium-lithium (D-Li) neutron source to produce high energy neutrons at sufficient intensity and irradiation volume to test samples of candidate materials up to about a full lifetime of anticipated use in fusion energy reactors. would also provide calibration and validation of data from fission reactor and other accelerator-based irradiation tests. It would generate material- specific activation and radiological properties data, and support the analysis of materials for use in safety, maintenance, recycling, decommissioning, and waste disposal systems

  20. IFMIF - International Fusion Materials Irradiation Facility Conceptual Design Activity/Interim Report

    Rennich, M.J.

    1995-12-01

    Environmental acceptability, safety, and economic viability win ultimately be the keys to the widespread introduction of fusion power. This will entail the development of radiation- resistant and low- activation materials. These low-activation materials must also survive exposure to damage from neutrons having an energy spectrum peaked near 14 MeV with annual radiation doses in the range of 20 displacements per atom (dpa). Testing of candidate materials, therefore, requires a high-flux source of high energy neutrons. The problem is that there is currently no high-flux source of neutrons in the energy range above a few MeV. The goal, is therefore, to provide an irradiation facility for use by fusion material scientists in the search for low-activation and damage-resistant materials. An accellerator-based neutron source has been established through a number of international studies and workshops` as an essential step for materials development and testing. The mission of the International Fusion Materials Irradiation Facility (IFMIF) is to provide an accelerator-based, deuterium-lithium (D-Li) neutron source to produce high energy neutrons at sufficient intensity and irradiation volume to test samples of candidate materials up to about a full lifetime of anticipated use in fusion energy reactors. would also provide calibration and validation of data from fission reactor and other accelerator-based irradiation tests. It would generate material- specific activation and radiological properties data, and support the analysis of materials for use in safety, maintenance, recycling, decommissioning, and waste disposal systems.

  1. Detergent disruption of bacterial inner membranes and recovery of protein translocation activity

    Isolation of the integral membrane components of protein translocation requires methods for fractionation and functional reconstitution. The authors treated inner-membrane vesicles of Escherichia coli with mixtures of octyl β-D-glucoside, phospholipids, and an integral membrane carrier protein under conditions that extract most of the membrane proteins into micellar solution. Upon dialysis, proteoliposomes were reconstituted that supported translocation of radiochemically pure [35S]pro-OmpA (the precursor of outer membrane protein A). Translocation into these proteoliposomes required ATP hydrolysis and membrane proteins, indicating that the reaction is that of the inner membrane. The suspension of membranes in detergent was separated into supernatant and pellet fractions by ultracentrifugation. After reconstitution, translocation activity was observed in both fractions, but processing by leader peptidase of translocated pro-OmpA to OmpA was not detectable in the reconstituted pellet fraction. Processing activity was restored by addition of pure leader peptidase as long as this enzyme was added before detergent removal, indicating that the translocation activity is not associated with detergent-resistant membrane vesicles. These results show that protein translocation activity can be recovered from detergent-disrupted membrane vesicles, providing a first step towards the goal of isolating the solubilized components

  2. Relationship between SU Subdomains That Regulate the Receptor-Mediated Transition from the Native (Fusion-Inhibited) to the Fusion-Active Conformation of the Murine Leukemia Virus Glycoprotein

    Lavillette, Dimitri; Ruggieri, Alessia; Boson, Bertrand; Maurice, Marielle; Cosset, François-Loïc

    2002-01-01

    Envelope glycoproteins (Env) of retroviruses are trimers of SU (surface) and TM (transmembrane) heterodimers and are expressed on virions in fusion-competent forms that are likely to be metastable. Activation of the viral receptor-binding domain (RBD) via its interaction with a cell surface receptor is thought to initiate a cascade of events that lead to refolding of the Env glycoprotein into its stable fusion-active conformation. While the fusion-active conformation of the TM subunit has bee...

  3. Salt stress in a membrane bioreactor: dynamics of sludge properties, membrane fouling and remediation through powdered activated carbon dosing.

    De Temmerman, L; Maere, T; Temmink, H; Zwijnenburg, A; Nopens, I

    2014-10-15

    Membrane bioreactors are a well-established technology for wastewater treatment. However, their efficiency is adversely impacted by membrane fouling, primarily inciting very conservative operations of installations that makes them less appealing from an economic perspective. This fouling propensity of the activated sludge is closely related to system disturbances. Therefore, improved insight into the impact of fouling is crucial towards increased membrane performance. In this work, the disturbance of a salt shock was investigated with respect to sludge composition and filterability in two parallel lab-scale membrane bioreactors. Several key sludge parameters (soluble microbial products, sludge-bound extracellular polymeric substances, supramicron particle size distributions (PSD), submicron particle concentrations) were intensively monitored prior to, during, and after a disturbance to investigate its impact as well as the potential governing mechanism. Upon salt addition, the supramicron PSD immediately shifted to smaller floc sizes, and the total fouling rate increased. Following a certain delay, an increase in submicron particles, supernatant proteins, and polysaccharides was observed as well as an increase in the irreversible membrane fouling rate. Recovery from the disturbance was evidenced with a simultaneous decrease in the above mentioned quantities. A similar experiment introducing powdered activated carbon (PAC) addition used for remediation resulted in either no or less significant changes in the above mentioned quantities, signifying its potential as a mitigation strategy. PMID:24999116

  4. Specific detection of Xanthomonas oryzae pv. oryzicola in infected rice plant by use of PCR assay targeting a membrane fusion protein gene.

    Kang, Man Jung; Shim, Jae Kyung; Cho, Min Seok; Seol, Young Joo; Hahn, Jang Ho; Hwang, Duk Ju; Park, Dong Suk

    2008-09-01

    Successful control of Xanthomonas oryzae pv. oryzicola, the causal agent of bacterial leaf streak, requires a specific and reliable diagnostic tool. A pathovar-specific PCR assay was developed for the rapid and accurate detection of the plant pathogenic bacterium Xanthomonas oryzae pv. oryzicola in diseased plant. Based on differences in a membrane fusion protein gene of Xanthomonas oryzae pv. oryzicola and other microorganisms, which was generated from NCBI (http://www.ncbi.nlm.nih.gov/) and CMR (http://cmr.tigr.org/) BLAST searches, one pair of pathovar-specific primers, XOCMF/XOCMR, was synthesized. Primers XOCMF and XOCMR from a membrane fusion protein gene were used to amplify a 488-bp DNA fragment. The PCR product was only produced from 4 isolates of Xanthomonas oryzae pv. oryzicola among 37 isolates of other pathovars and species of Xanthomonas, Pectobacterium, Pseudomonas, Burkholderia, Escherichia coli, and Fusarium oxysporum f.sp. dianthi. The results suggested that the assay detected the pathogen more rapidly and accurately than standard isolation methods. PMID:18852502

  5. ACTIVE CALCIUM TRANSPORT IN PLASMA MEMBRANE VESICLES FROM DEVELOPING COTYLEDONS OF COMMON BEAN

    黄建中; 陈子元

    1995-01-01

    Plasma membrane vesicles were prepared from the developing cotyledons of common bean (Phaseolus vulgaris L cv Diyundou)by aqueous two-phase partitioning and characterized as to their purity by assaying marker enzymes for other membranes.The putative plasma membrane fraction was minimalyy contaminated by membranes other than plasma membrane and hence was of high purity,It exhibited a Ca2+-dependent ATPase activity,which was inhibited by 1umol/L EB and promoted by calcium ionophore A23187.Such an activity was responsible for the observed ATP dependent 45Ca2+ uptake into inside-out plasma membrane vesicles.This process was stimulated by 0.5μmol/L CaM and 20μmol/L IAA but inhibited by 2μmol/L ABA and abolished by A23187,Possible role of cytoplasmic Ca2+ in mediating phytohormones activity is discussed.

  6. Active calcium transport in plasma membrane vesicles from developing cotyledons of common bean

    Plasma membrane vesicles were prepared from the developing cotyledons of common bean (Phaseolus vulgaris L cv Diyundou) by aqueous two-phase partitioning and characterized as to their purity by assaying marker enzymes for other membranes. The putative plasma membrane fraction was minimally contaminated by membranes other than plasma membrane and hence was of high purity. It exhibited a Ca2+-dependent ATPase activity, which was inhibited by 1 μmol/L EB and promoted by calcium ionophore A23187. Such an activity was responsible for the observed ATP-dependent 45Ca2+ uptake into inside-out plasma membrane vesicles. This process was stimulated by 0.6 μmol/L CaM and 20 μmol/L IAA but inhibited by 2 μmol/L ABA and abolished by A23187. Possible role of cytoplasmic Ca2+ in mediating phytohormones activity is discussed

  7. Joining techniques for a reduced activation 12Cr steel for inertial fusion energy

    Highlights: • We characterized joining techniques for a candidate material for inertial confinement fusion: reduced activation ferritic martensitic 12% chromium steel. • E-beam, TIG, and laser welds were completed with good quality without significant cracking or porosity. • A heat treatment of 950 °C for 1 h normalized the weld fusion zone and heat-affected zone to the base metal microstructure. • Diffusion bonding at 950 °C or greater for 2 h produced an interface with over 600 MPa tensile strength. - Abstract: At Lawrence Livermore National Laboratory, we are developing a reduced activation ferritic martensitic steel that is based on the ferritic martensitic steel HT-9. As a part of the development of this steel, we tested a series of welding processes for characterization, including conventional welds (electron beam, tungsten inert gas, and laser) as well as solid-state welds (hot isostatic pressing). We also heat treated the joints at various temperatures between 750 °C and 1050 °C to find a suitable normalization scheme. The modified HT-9 reduced activation ferritic martensitic steel appears highly suitable to welding and diffusion bonding. All welds showed good quality fusion zones with insignificant cracking or porosity. Additionally, a heat treatment schedule of 950 °C for one hour caused minimal grain growth while still converging the hardness of the base metal with that of the fusion and heat-affected zones. Also, modified HT-9 diffusion bonds that were created at temperatures of at least 950 °C for two hours at 103 MPa had interface tensile strengths of greater than 600 MPa. The diffusion bonds showed no evidence of increased hardness nor void formation at the diffusion bonded interface

  8. Circulating polymerase chain reaction chips utilizing multiple-membrane activation

    Wang, Chih-Hao; Chen, Yi-Yu; Liao, Chia-Sheng; Hsieh, Tsung-Min; Luo, Ching-Hsing; Wu, Jiunn-Jong; Lee, Huei-Huang; Lee, Gwo-Bin

    2007-02-01

    This paper reports a new micromachined, circulating, polymerase chain reaction (PCR) chip for nucleic acid amplification. The PCR chip is comprised of a microthermal control module and a polydimethylsiloxane (PDMS)-based microfluidic control module. The microthermal control modules are formed with three individual heating and temperature-sensing sections, each modulating a specific set temperature for denaturation, annealing and extension processes, respectively. Micro-pneumatic valves and multiple-membrane activations are used to form the microfluidic control module to transport sample fluids through three reaction regions. Compared with other PCR chips, the new chip is more compact in size, requires less time for heating and cooling processes, and has the capability to randomly adjust time ratios and cycle numbers depending on the PCR process. Experimental results showed that detection genes for two pathogens, Streptococcus pyogenes (S. pyogenes, 777 bps) and Streptococcus pneumoniae (S. pneumoniae, 273 bps), can be successfully amplified using the new circulating PCR chip. The minimum number of thermal cycles to amplify the DNA-based S. pyogenes for slab gel electrophoresis is 20 cycles with an initial concentration of 42.5 pg µl-1. Experimental data also revealed that a high reproducibility up to 98% could be achieved if the initial template concentration of the S. pyogenes was higher than 4 pg µl-1. The preliminary results of the current paper were presented at the 19th IEEE International Conference on Micro Electro Mechanical Systems (IEEE MEMS 2006), Istanbul, Turkey, 22-26 January, 2006.

  9. [3H]PN200-110 and [3H]ryanodine binding and reconstitution of ion channel activity with skeletal muscle membranes

    Skeletal muscle membranes derived either from the tubular (T) network or from the sarcoplasmic reticulum (SR) were characterized with respect to the binding of the dihydropyridine, [3H]PN200-110, and the alkaloid, [3H]ryanodine; polypeptide composition; and ion channel activity. Conditions for optimizing the binding of these radioligands are discussed. A bilayer pulsing technique is described and is used to examine the channels present in these membranes. Fusion of T-tubule membranes into bilayers revealed the presence of chloride channels and dihydropyridine-sensitive calcium channels with three distinct conductances. The dihydropyridine-sensitive channels were further characterized with respect to their voltage dependence. Pulsing experiments indicated that two different populations of dihydropyridine-sensitive channels existed. Fusion of heavy SR vesicles revealed three different ion channels; the putative calcium release channel, a potassium channel, and a chloride channel. Thus, this fractionation procedure provides T-tubules and SR membranes which, with radioligand binding and single channel recording techniques, provide a useful tool to study the characteristics of skeletal muscle ion channels and their possible role in excitation-contraction coupling

  10. Interaction of peptides with cell membranes: insights from molecular modeling

    The investigation of the interaction of peptides with cell membranes is the focus of active research. It can enhance the understanding of basic membrane functions such as membrane transport, fusion, and signaling processes, and it may shed light on potential applications of peptides in biomedicine. In this review, we will present current advances in computational studies on the interaction of different types of peptides with the cell membrane. Depending on the properties of the peptide, membrane, and external environment, the peptide–membrane interaction shows a variety of different forms. Here, on the basis of recent computational progress, we will discuss how different peptides could initiate membrane pores, translocate across the membrane, induce membrane endocytosis, produce membrane curvature, form fibrils on the membrane surface, as well as interact with functional membrane proteins. Finally, we will present a conclusion summarizing recent progress and providing some specific insights into future developments in this field. (topical review)

  11. Adjoint Monte Carlo Simulation of Fusion Product Activation Probe Experiment in ASDEX Upgrade tokamak

    Äkäslompolo, Simppa; Tardini, Giovanni; Kurki-Suonio, Taina

    2015-01-01

    The activation probe is a robust tool to measure flux of fusion products from a magnetically confined plasma. A carefully chosen solid sample is exposed to the flux, and the impinging ions transmute the material makig it radioactive. Ultra-low level gamma-ray spectroscopy is used post mortem to measure the activity and, thus, the number of fusion products. This contribution presents the numerical analysis of the first measurement in the ASDEX Upgrade tokamak, which was also the first experiment to measure a single discharge. The ASCOT suite of codes was used to perform adjoint/reverse Monte-Carlo calculations of the fusion products. The analysis facilitated, for the first time, a comparison of numerical and experimental values for absolutely calibrated flux. The results agree to within 40%, which can be considered remarkable considering the fact that all features of the plasma cannot be accounted in the simulations. Also an alternative probe orientation was studied. The results suggest that a better optimized...

  12. ZZ MONTAGE-400, Neutron Activation 100-Group Cross-Section Library of Fusion Reactor Materials

    1 - Description of problem or function: Format: GAM-II group structure and ANISN; Number of groups: 100-group cross sections. Nuclides: H, He, Li, Be, B, N, O, F, Na, Al, Si, P, S, Cl, K, Ca, Sc, Ti, V, Cr, Mn, Fe, Co, Ni, Cu, Y, Zr, Nb, Mo, Tc, Ru, Ag, Sn, Cs, Hf, Ta, W, Re, Au, Pb. Origin: derived from ENDF/B, or calculated at Brookhaven National Laboratory. Weighting spectrum: 1/E except near 14 MeV where a thermally broadened fusion peak, assuming a temperature of 20 MeV, is employed. This data library contains 100- group cross sections, with GAM-II group structure, for 421 neutron activation reactions with fusion reactor structural and coolant materials. The weighting function is 1/E except near 14 MeV where a thermally broadened fusion peak, assuming a temperature of 20 MeV, is employed. The library also contains half life information for the activated nuclei. 2 - Method of solution: The thermal group cross sections were calculated from the 2200 m/s value, when available, otherwise from the group 99 value. The majority of the non-thermal cross sections were derived from pointwise data derived from ENDF/B, or calculated at Brookhaven National Laboratory using the nuclear systematics code THRESH. These were converted to multigroup from using the codes ETOG and NJOY

  13. Biological effects of activation products and other chemicals released from fusion power plants

    Literature reviews indicate that existing information is incomplete, often contradictory, and of questionable value for the prediction and assessment of ultimate impact from fusion-associated activation products and other chemical releases. It is still uncertain which structural materials will be used in the blanket and first wall of fusion power plants. However, niobium, vanadium, vanadium-chromium alloy, vanadium-titanium alloy, sintered aluminum product, and stainless steel have been suggested. The activation products of principal concern will be the longer-lived isotopes of 26Al, 49V, 51Cr, 54Mn, 55Fe, 58Co, 60Co, 93Nb, and 94Nb. Lithium released to the environment either during the mining cycle, from power plant operation or accident, may be in the form of a number of compound types varying in solubility and affinity for biological organisms. The effects of a severe liquid metal fire or explosion involving Na or K will vary according to inherent abiotic and biotic features of the affected site. Saline, saline-alkaline, and sodic soils of arid lands would be particularly susceptible to alkaline stress. Beryllium released to the environment during the mining cycle or reactor accident situation could be in the form of a number of compound types. Adverse effects to aquatic species from routine chemical releases (biocides, corrosion inhibitors, dissolution products) may occur in the discharge of both fission and fusion power plant designs

  14. Nuclear model calculations of activation cross sections for fusion reactor technology

    Intense neutron fluxes within fusion reactors that are currently being designed will lead to the activation of structural components. To assess and minimize this radioactivity, nuclear cross sections are needed for neutrons with energies up to 20 MeV. We describe research performed for the International Atomic Energy Agency (IAEA) Coordinated Research Programme on activation cross sections for fusion reactor technology, which has selected certain high-priority reactions for both experimental and theoretical study. Using statistical model codes, we have investigated: (1) excitation function cross sections for radionuclide production in the reactions 94Mo(n,p)94Nb, 109Ag(n,2n)108mAg, 151Eu(n,2n)150mEu, 153Eu(n,2n)152g+m2Eu, 159Tb(n,2n)158Tb, 187Re(n,2n)186mRe, 179Hf(n,2n)178m2Hf, 193Ir(n,2n)192m2Ir; and (2) the systematical dependence of isomeric ratios on isomer spin and incident-energy. Using our calculated results for the excitation functions, along with calculations by other groups, the theoretical excitation functions have been normalized to experimental values at 14.5 MeV to produce evaluated excitation functions. These evaluations can be used within radiation transport and nuclide inventory codes to design and assess the environmental impact of fusion reactors. (orig.)

  15. Adjoint Monte Carlo simulation of fusion product activation probe experiment in ASDEX Upgrade tokamak

    The activation probe is a robust tool to measure flux of fusion products from a magnetically confined plasma. A carefully chosen solid sample is exposed to the flux, and the impinging ions transmute the material making it radioactive. Ultra-low level gamma-ray spectroscopy is used post mortem to measure the activity and, thus, the number of fusion products. This contribution presents the numerical analysis of the first measurement in the ASDEX Upgrade tokamak, which was also the first experiment to measure a single discharge. The ASCOT suite of codes was used to perform adjoint/reverse Monte Carlo calculations of the fusion products. The analysis facilitates, for the first time, a comparison of numerical and experimental values for absolutely calibrated flux. The results agree to within a factor of about two, which can be considered a quite good result considering the fact that all features of the plasma cannot be accounted in the simulations.Also an alternative to the present probe orientation was studied. The results suggest that a better optimized orientation could measure the flux from a significantly larger part of the plasma. A shorter version of this contribution is due to be published in PoS at: 1st EPS conference on Plasma Diagnostics

  16. In vitro Activity and Function of B7-H4-Ig Fusion Protein

    Rasmussen, Susanne B; Kosicki, Michael; Svendsen, Signe Goul;

    2013-01-01

    -Ig fusion protein has been documented to assuage the symptoms in mouse models of RA, T1D, and multiple sclerosis in vivo. In the present study, B7-H4-Ig bound to the majority of human peripheral blood monocytes and NK cells, but not to either normal or activated T cells. B7-H4-Ig fusion protein was......B7-H4 has been shown to inhibit T cell proliferation, cytokine production and cell cycle in vitro. B7-H4 deficient mice develop exacerbated disease in the mouse models of Rheumatoid Arthritis (RA), Type 1 Diabetes (T1D) and Experimental Autoimmune Encephalomyelitis (EAE). On the other hand, B7-H4...

  17. Analysis of Induced Gamma Activation by D-T Neutrons in Selected Fusion Reactor Relevant Materials with EAF-2010

    Klix, Axel; Fischer, Ulrich; Gehre, Daniel

    2016-02-01

    Samples of lanthanum, erbium and titanium which are constituents of structural materials, insulating coatings and tritium breeder for blankets of fusion reactor designs have been irradiated in a fusion peak neutron field. The induced gamma activities were measured and the results were used to check calculations with the European activation system EASY-2010. Good agreement for the prediction of major contributors to the contact dose rate of the materials was found, but for minor contributors the calculation deviated up to 50%.

  18. Analysis of induced gamma activation by D-T neutrons in selected fusion reactor relevant materials with EAF-2010

    Klix Axel; Fischer Ulrich; Gehre Daniel

    2016-01-01

    Samples of lanthanum, erbium and titanium which are constituents of structural materials, insulating coatings and tritium breeder for blankets of fusion reactor designs have been irradiated in a fusion peak neutron field. The induced gamma activities were measured and the results were used to check calculations with the European activation system EASY-2010. Good agreement for the prediction of major contributors to the contact dose rate of the materials was found, but for minor contributors t...

  19. VAMP7 regulates constitutive membrane incorporation of the cold-activated channel TRPM8.

    Ghosh, Debapriya; Pinto, Silvia; Danglot, Lydia; Vandewauw, Ine; Segal, Andrei; Van Ranst, Nele; Benoit, Melissa; Janssens, Annelies; Vennekens, Rudi; Vanden Berghe, Pieter; Galli, Thierry; Vriens, Joris; Voets, Thomas

    2016-01-01

    The cation channel TRPM8 plays a central role in the somatosensory system, as a key sensor of innocuously cold temperatures and cooling agents. Although increased functional expression of TRPM8 has been implicated in various forms of pathological cold hypersensitivity, little is known about the cellular and molecular mechanisms that determine TRPM8 abundance at the plasma membrane. Here we demonstrate constitutive transport of TRPM8 towards the plasma membrane in atypical, non-acidic transport vesicles that contain lysosomal-associated membrane protein 1 (LAMP1), and provide evidence that vesicle-associated membrane protein 7 (VAMP7) mediates fusion of these vesicles with the plasma membrane. In line herewith, VAMP7-deficient mice exhibit reduced functional expression of TRPM8 in sensory neurons and concomitant deficits in cold avoidance and icilin-induced cold hypersensitivity. Our results uncover a cellular pathway that controls functional plasma membrane incorporation of a temperature-sensitive TRP channel, and thus regulates thermosensitivity in vivo. PMID:26843440

  20. Progress in the US program to develop low-activation structural materials for fusion

    It has long been recognized that attainment of the safety and environmental potential of fusion energy requires the successful development of low-activation materials for the first wall, blanket and other high heat flux structural components. Only a limited number of materials potentially possess the physical, mechanical and low-activation characteristics required for this application. The current US structural materials research effort is focused on three candidate materials: advanced ferritic steels, vanadium alloys, and silicon carbide composites. Recent progress has been made in understanding the response of these materials to neutron irradiation. (author)

  1. Progress in the U.S. program to develop low-activation structural materials for fusion

    It has long been recognized that attainment of the safety and environmental potential of fusion energy requires the successful development of low-activation materials for the first wall, blanket and other high heat flux structural components. Only a limited number of materials potentially possess the physical, mechanical and low-activation characteristics required for this application. The current U.S. structural materials research effort is focused on three candidate materials: advanced ferritic steels, vanadium alloys, and silicon carbide composites. Recent progress has been made in understanding the response of these materials to neutron irradiation. (author)

  2. Membrane topology analysis of HIV-1 envelope glycoprotein gp41

    Xiao Dan

    2010-11-01

    Full Text Available Abstract Background The gp41 subunit of the HIV-1 envelope glycoprotein (Env has been widely regarded as a type I transmembrane protein with a single membrane-spanning domain (MSD. An alternative topology model suggested multiple MSDs. The major discrepancy between the two models is that the cytoplasmic Kennedy sequence in the single MSD model is assigned as the extracellular loop accessible to neutralizing antibodies in the other model. We examined the membrane topology of the gp41 subunit in both prokaryotic and mammalian systems. We attached topological markers to the C-termini of serially truncated gp41. In the prokaryotic system, we utilized a green fluorescent protein (GFP that is only active in the cytoplasm. The tag protein (HaloTag and a membrane-impermeable ligand specific to HaloTag was used in the mammalian system. Results In the absence of membrane fusion, both the prokaryotic and mammalian systems (293FT cells supported the single MSD model. In the presence of membrane fusion in mammalian cells (293CD4 cells, the data obtained seem to support the multiple MSD model. However, the region predicted to be a potential MSD is the highly hydrophilic Kennedy sequence and is least likely to become a MSD based on several algorithms. Further analysis revealed the induction of membrane permeability during membrane fusion, allowing the membrane-impermeable ligand and antibodies to cross the membrane. Therefore, we cannot completely rule out the possible artifacts. Addition of membrane fusion inhibitors or alterations of the MSD sequence decreased the induction of membrane permeability. Conclusions It is likely that a single MSD model for HIV-1 gp41 holds true even in the presence of membrane fusion. The degree of the augmentation of membrane permeability we observed was dependent on the membrane fusion and sequence of the MSD.

  3. Electrospun nanofiber membranes for electrically activated shape memory nanocomposites

    A novel shape memory nanocomposite system, consisting of a thermoplastic Nafion polymer and ultrathin electrospun polyacrylonitrile (PAN)-based carbonization nanofiber membranes, is successfully synthesized. PAN-based carbonization nanofiber networks that offer responses to deformations are considered to be an excellent actuation source. Significant improvement in the electrical conductivity of carbon nanofiber membranes is found by adjusting the applied voltage power in the electrospinning PAN process varying from 7.85 to 12.30 S cm−1. The porous structure of the carbon nanofiber membranes provides a large specific surface area and interfacial contact area when combined with the polymer matrix. Shape memory Nafion nanocomposites filled with interpenetrating non-woven electrospun PAN carbonization membranes can be actuated by applying 14 V electrical voltage within 5 s. The results, as demonstrated through morphology, electrical and thermal measurements and a shape recovery test, suggest a valuable route to producing soft nanocomposites. (papers)

  4. Alteration of membrane phospholipid methylation by adenosine analogs does not affect T lymphocyte activation

    Membrane phospholipid methylation has been described during activation of various immune cells. Moreover recent data indicated modulation of immune cells functions by adenosine. As S-adenosyl-methionine and S-adenosyl-homocysteine are adenosine analogs and modulators of transmethylation reactions, the effects of SAH and SAM were investigated on membrane phospholipid methylation and lymphocyte activation. SAM was shown to induce the membrane phospholipid methylation as assessed by the 3Hmethyl-incorporation in membrane extract. This effect was inhibited by SAH. In contrast SAM and SAH did not affect the phytohemagglutinin-induced proliferative response of peripheral blood mononuclear cells. SAH neither modified the early internalization of membrane CD3 antigens nor did it prevent the late expression of HLA-DR antigens on lymphocytes activated by phytohemagglutinin. These results indicate that in vitro alteration of phospholipid methylation does not affect subsequent steps of human T lymphocyte activation and proliferation

  5. Perspective on fusion research in China (2) fusion activities in China with special intonation on hybrid reactor program

    Lijian, Qiu

    2001-09-01

    Chinese fusion research was started from 1958, but with more clear problem definition it has been set up as the national program for development of the hybrid reactor in 1986. In this paper, it will be described how the organized program is going on.

  6. Modification of trout sperm membranes associated with activation and cryopreservation. Implications for fertilizing potential.

    Purdy, P H; Barbosa, E A; Praamsma, C J; Schisler, G J

    2016-08-01

    We investigated the effects of two trout sperm activation solutions on sperm physiology and membrane organization prior to and following cryopreservation using flow cytometry and investigated their impact on in vitro fertility. Overall, frozen-thawed samples had greater phospholipid disorder when compared with fresh samples (high plasma membrane fluidity; P < 0.0001) and sperm activated with water also had high plasma membrane fluidity when compared to sperm activated with Lahnsteiner solution (LAS; P < 0.0001). Following cryopreservation water activated samples had membranes with greater membrane protein disorganization compared with LAS but the membrane protein organization of LAS samples was similar to samples prior to freezing (P < 0.0001). Post-thaw water activation resulted in significant increases in intracellular calcium compared to LAS (P < 0.002). In vitro fertility trials with frozen-thawed milt and LAS activation resulted in greater fertility (45%) compared to water activated samples (10%; P < 0.0001). Higher fertility rates correlated with lower intracellular calcium with water (R(2) = -0.9; P = 0.01) and LAS (R(2) = -0.85; P = 0.03) activation. Greater plasma membrane phospholipid (R(2) = -0.89; P = 0.02) and protein (R(2) = -0.84; P = 0.04) disorder correlated with lower water activation fertility rates. These membrane organization characteristics only approached significance with LAS activation in vitro fertility (P = 0.09, P = 0.06, respectively). Potentially the understanding of sperm membrane reorganizations and the physiology associated with activation following cryopreservation may enable users in a repository or hatchery setting to estimate the fertilizing potential of a sample and determine its value. PMID:27234987

  7. Evidence that Rabies Virus Forms Different Kinds of Fusion Machines with Different pH Thresholds for Fusion

    Roche, Stéphane; Gaudin, Yves

    2004-01-01

    Fusion of rabies virus with membranes is triggered at a low pH and is mediated by a viral glycoprotein (G). Fusion of rabies virus with liposomes was monitored by using a lipid mixing assay based on fluorescence resonance energy transfer. Fusion was detected below pH 6.4, and its extent increased with H+ concentrations to be maximal around pH 6.15. The origin of the partial fusion activity of rabies virus under suboptimal pH conditions (i.e., between pH 6.15 and 6.4) was investigated. We demo...

  8. Actomyosin dynamics drive local membrane component organization in an in vitro active composite layer.

    Köster, Darius Vasco; Husain, Kabir; Iljazi, Elda; Bhat, Abrar; Bieling, Peter; Mullins, R Dyche; Rao, Madan; Mayor, Satyajit

    2016-03-22

    The surface of a living cell provides a platform for receptor signaling, protein sorting, transport, and endocytosis, whose regulation requires the local control of membrane organization. Previous work has revealed a role for dynamic actomyosin in membrane protein and lipid organization, suggesting that the cell surface behaves as an active composite composed of a fluid bilayer and a thin film of active actomyosin. We reconstitute an analogous system in vitro that consists of a fluid lipid bilayer coupled via membrane-associated actin-binding proteins to dynamic actin filaments and myosin motors. Upon complete consumption of ATP, this system settles into distinct phases of actin organization, namely bundled filaments, linked apolar asters, and a lattice of polar asters. These depend on actin concentration, filament length, and actin/myosin ratio. During formation of the polar aster phase, advection of the self-organizing actomyosin network drives transient clustering of actin-associated membrane components. Regeneration of ATP supports a constitutively remodeling actomyosin state, which in turn drives active fluctuations of coupled membrane components, resembling those observed at the cell surface. In a multicomponent membrane bilayer, this remodeling actomyosin layer contributes to changes in the extent and dynamics of phase-segregating domains. These results show how local membrane composition can be driven by active processes arising from actomyosin, highlighting the fundamental basis of the active composite model of the cell surface, and indicate its relevance to the study of membrane organization. PMID:26929326

  9. Crystal Structure of a Soluble Fragment of the Membrane Fusion Protein HlyD in a Type I Secretion System of Gram-Negative Bacteria.

    Kim, Jin-Sik; Song, Saemee; Lee, Minho; Lee, Seunghwa; Lee, Kangseok; Ha, Nam-Chul

    2016-03-01

    The protein toxin HlyA of Escherichia coli is exported without a periplasmic intermediate by the type I secretion system (T1SS). The T1SS is composed of an inner membrane ABC transporter HlyB, an outer-membrane channel protein TolC, and a membrane fusion protein HlyD. However, the assembly of the T1SS remains to be elucidated. In this study, we determine the crystal structure of a part of the C-terminal periplasmic domain of HlyD. The long α-helical domain consisting of three α helices and a lipoyl domain was identified in the crystal structure. Based on the HlyD structure, we modeled the hexameric assembly of HlyD with a long α-helical barrel, which formed a complex with TolC in an intermeshing cogwheel-to-cogwheel manner, as observed in tripartite RND-type drug efflux pumps. These observations provide a structural blueprint for understanding the type I secretion system in pathogenic Gram-negative bacteria. PMID:26833388

  10. Crystal Structure of Dengue Type 1 Envelope Protein in the Postfusion Conformation and its Implication for Receptor Binding, Membrane Fusion and Antibody Recognition

    Nayak, V.; Dessau, M; Kucera, K; Anthony, K; Ledizet, M; Modis, Y

    2009-01-01

    Dengue virus relies on a conformational change in its envelope protein, E, to fuse the viral lipid membrane with the endosomal membrane and thereby deliver the viral genome into the cytosol. We have determined the crystal structure of a soluble fragment E (sE) of dengue virus type 1 (DEN-1). The protein is in the postfusion conformation even though it was not exposed to a lipid membrane or detergent. At the domain I-domain III interface, 4 polar residues form a tight cluster that is absent in other flaviviral postfusion structures. Two of these residues, His-282 and His-317, are conserved in flaviviruses and are part of the pH sensor that triggers the fusogenic conformational change in E, at the reduced pH of the endosome. In the fusion loop, Phe-108 adopts a distinct conformation, forming additional trimer contacts and filling the bowl-shaped concavity observed at the tip of the DEN-2 sE trimer.

  11. Towards a reduced activation structural materials database for fusion DEMO reactors

    The development of First Wall, Blanket and Divertor materials which are capable of withstanding many years the high neutron and heat fluxes, is a critical path to fusion power. Therefore, the timely availability of a sound materials database has become an indispensable element in international fusion road maps. In order to provide materials design data for short term needs of ITER Test Blanket Modules and for a DEMOnstration fusion reactor, a wealth of R and D results on the European reduced activation ferritic-martensitic steel EUROFER, and on oxide dispersion strengthened variants are being characterized, mainly in the temperature window 250-650 deg. C. The characterisation includes irradiations up to 15 dpa in the mixed spectrum reactor HFR and up to 75 dpa in the fast breeder reactor BOR60. Industrial EUROFER-batches of 3.5 and 7.5 tons have been produced with a variety of semi-finished, quality-assured product forms. To increase thermal efficiency of blankets, high temperature resistant SiCf/SiC channel inserts for liquid metal coolant tubes are also developed. Regarding radiation damage resistance, a broad based reactor irradiation programs counts several steps from ≤5dpa (ITER TBMs) up to 75 dpa (DEMO). For the European divertor designers, a materials data base is presently being set up for pure W and W alloys, and related reactor irradiations are foreseen with temperatures from 650-1000 deg. C. (author)

  12. Position control of active magnetic levitation using sphere-shaped HTS bulk for inertial nuclear fusion

    We have developed an active magnetic levitation system that comprises a field-cooled disk-shaped or sphere-shaped HTS bulk and multiple ring-shaped electromagnets. In this system, the levitation height of HTS bulk can be controlled by adjusting the operating current of each electromagnet individually. Further, the application of the vertical noncontact levitation system is expected due to its levitation stability without mechanical supports. We assume that this system is applied to inertial nuclear fusion. However, one of the important issues is to achieve position control with high accuracy of the fusion fuel in order to illuminate the target evenly over the entire surface. Therefore, this system is applied to the levitation and position control of a sphere-shaped superconducting capsule containing nuclear fusion fuel. In this study, we designed and constructed a position control system for the sphere-shaped HTS bulk with a diameter of 5 mm by using numerical simulation based on hybrid finite element and boundary element analysis. We then carried out the experiment of levitation height and position control characteristics of the HTS bulk in this system. With regard to position control, accuracies within 59 μm are obtained

  13. Spatial distribution and activity of Na(+)/K(+)-ATPase in lipid bilayer membranes with phase boundaries.

    Bhatia, Tripta; Cornelius, Flemming; Brewer, Jonathan; Bagatolli, Luis A; Simonsen, Adam C; Ipsen, John H; Mouritsen, Ole G

    2016-06-01

    We have reconstituted functional Na(+)/K(+)-ATPase (NKA) into giant unilamellar vesicles (GUVs) of well-defined binary and ternary lipid composition including cholesterol. The activity of the membrane system can be turned on and off by ATP. The hydrolytic activity of NKA is found to depend on membrane phase, and the water relaxation in the membrane on the presence of NKA. By collapsing and fixating the GUVs onto a solid support and using high-resolution atomic-force microscopy (AFM) imaging we determine the protein orientation and spatial distribution at the single-molecule level and find that NKA is preferentially located at lo/ld interfaces in two-phase GUVs and homogeneously distributed in single-phase GUVs. When turned active, the membrane is found to unbind from the support suggesting that the protein function leads to softening of the membrane. PMID:26994932

  14. Comparison of membrane fouling during short-term filtration of aerobic granular sludge and activated sludge

    2007-01-01

    Aerobic granular sludge was cultivated adopting internal-circulate sequencing batch airlift reactor. The contradistinctive experiment about short-term membrane fouling between aerobic granular sludge system and activated sludge system were investigated. The membrane foulants was also characterized by Fourier Transform Infrared (FTIR) spectroscopy technique. The results showed that the aerobic granular sludge had excellent denitrification ability; the removal efficiency of TN could reach 90%. The aerobic granular sludge could alleviate membrane fouling effectively. The steady membrane flux of aerobic granular sludge was twice as much as that of activated sludge system. In addition, it was found that the aerobic granular sludge could result in severe membrane pore-blocking, however, the activated sludge could cause severe cake fouling. The major components of the foulants were identified as comprising of proteins and polysaccharide materials.

  15. The advanced 3D method for activation analysis of fusion reactor materials

    The method allows analyzing the complex objects activated by neutrons (e.g. fusion reactors) combining advantages of the 3D radiation transport by MCNP program with calculations of multiple activation and radioactive decay chains by FISPACT program. The problem of preparing the gamma-ray sources in cells of 3D geometry was solved by creation of an interface between the MCNP and FISPACT programs. The interface allows optimizing the process of activation analysis by revealing dominant sources of radiation. The developed interface essentially reduces the time needed for calculations. The main advantage of the method is realization of so-called 'multibox' procedure for decay gamma source sampling during decay gamma transport in very large and complex fusion reactor models. Shutdown dose rate calculations are faster (up to 600 times in ITER cryostat) in comparison with applied MCNP standard source definition by using an external user-supplied source subroutine of the 'multibox' procedure. The offered method is intended for solution of the activation tasks with deep penetration of radiation. The method was used in the engineering design of ITER-FEAT and RF DEMO-S

  16. Fusion-neutron-yield, activation measurements at the Z accelerator: Design, analysis, and sensitivity

    Hahn, K. D., E-mail: kdhahn@sandia.gov; Ruiz, C. L.; Fehl, D. L.; Chandler, G. A.; Knapp, P. F.; Smelser, R. M.; Torres, J. A. [Sandia National Laboratories, Diagnostics and Target Physics, Albuquerque, New Mexico 87123 (United States); Cooper, G. W.; Nelson, A. J. [Department of Chemical and Nuclear Engineering, University of New Mexico, Albuquerque, New Mexico 87131 (United States); Leeper, R. J. [Los Alamos National Laboratories, Plasma Physics Group, Los Alamos, New Mexico 87545 (United States)

    2014-04-15

    We present a general methodology to determine the diagnostic sensitivity that is directly applicable to neutron-activation diagnostics fielded on a wide variety of neutron-producing experiments, which include inertial-confinement fusion (ICF), dense plasma focus, and ion beam-driven concepts. This approach includes a combination of several effects: (1) non-isotropic neutron emission; (2) the 1/r{sup 2} decrease in neutron fluence in the activation material; (3) the spatially distributed neutron scattering, attenuation, and energy losses due to the fielding environment and activation material itself; and (4) temporally varying neutron emission. As an example, we describe the copper-activation diagnostic used to measure secondary deuterium-tritium fusion-neutron yields on ICF experiments conducted on the pulsed-power Z Accelerator at Sandia National Laboratories. Using this methodology along with results from absolute calibrations and Monte Carlo simulations, we find that for the diagnostic configuration on Z, the diagnostic sensitivity is 0.037% ± 17% counts/neutron per cm{sup 2} and is ∼ 40% less sensitive than it would be in an ideal geometry due to neutron attenuation, scattering, and energy-loss effects.

  17. Quercetin modulates activities of Taiwan cobra phospholipase A2 via its effects on membrane structure and membrane-bound mode of phospholipase A2

    Yi-Ling Chiou; Shinne-Ren Lin; Wan-Ping Hu; Long-Sen Chang

    2012-06-01

    The goal of the present study is to elucidate the mechanism of quercetin on modulating Naja naja atra phospholipase A2 (PLA2) activities. Sphingomyelin inhibited PLA2 enzymatic activity and membrane-damaging activity against egg yolk phosphatidylcholine (EYPC), while cholesterol and quercetin abrogated the sphingomeyelin inhibitory effect. Quercetin incorporation led to a reduction in PLA2 enzymatic activity and membrane-damaging activity toward EYPC/sphingomyelin/cholesterol vesicles. Both cholesterol and quercetin increased detergent resistance and reduced membrane fluidity of EYPC/sphingomyelin vesicles. Quercetin reduced detergent insolubility but increased ordered lipid packing of EYPC/sphingomyelin/cholesterol vesicles. Acrylamide quenching studies and trinitrophenylation of Lys residues revealed that quercetin altered the membrane-bound mode of PLA2 differently upon absorption onto the membrane bilayers of different lipid compositions. However, 8-anilinonaphthalene sulphonate-binding assay revealed that quercetin marginally affected the interaction between active site of PLA2 with phospholipid vesicles. Collectively, our data indicate that membrane-inserted quercetin modulates PLA2 interfacial activity and membrane-damaging activity via its effects on membrane structure and membrane-bound mode of PLA2.

  18. New insights into membrane-active action in plasma membrane of fungal hyphae by the lipopeptide antibiotic bacillomycin L.

    Zhang, Bao; Dong, Chunjuan; Shang, Qingmao; Han, Yuzhu; Li, Pinglan

    2013-09-01

    Bacillomycin L, a natural iturinic lipopeptide produced by Bacillus amyloliquefaciens, is characterized by strong antifungal activities against a variety of agronomically important filamentous fungi including Rhizoctonia solani Kühn. Prior to this study, the role of membrane permeabilization in the antimicrobial activity of bacillomycin L against plant pathogenic fungi had not been investigated. To shed light on the mechanism of this antifungal activity, the permeabilization of R. solani hyphae by bacillomycin L was investigated and compared with that by amphotericin B, a polyene antibiotic which is thought to act primarily through membrane disruption. Our results derived from electron microscopy, various fluorescent techniques and gel retardation experiments revealed that the antifungal activity of bacillomycin L may be not solely a consequence of fungal membrane permeabilization, but related to the interaction of it with intracellular targets. Our findings provide more insights into the mode of action of bacillomycin L and other iturins, which could in turn help to develop new or improved antifungal formulations or result in novel strategies to prevent fungal spoilage. PMID:23756779

  19. Influence of activating hormones on human platelet membrane Ca/sup 2 +/-ATPase activity

    Resink, T.J.; Dimitrov, D.; Stucki, S.; Buehler, F.R.

    1986-07-16

    Intact platelets were pretreated with hormones and thereafter membranes were prepared and Ca/sup 2 +/-ATPase activity determined. Thrombin decreased the V/sub max/ of Ca/sup 2 +/-ATPase after pretreatment of intact platelets. Platelet activating factor, vasopressin and ADP also decreased Ca/sup 2 +/-ATPase activity. 12-O-tetradecanoylphorbol-13-acetate (TPA) or A23187 or ionomycin alone had no effect, while the simultaneous pretreatment with TPA and Ca/sup 2 +/-ionophore decreased Ca/sup 2 +/-ATPase activity. cAMP elevating agents prostaglandin E/sub 1/ (PGE/sub 1/) and forskolin had no influence per se on Ca/sup 2 +/-ATPase, but antagonized the inhibitory effect of thrombin. The data suggest a close connection between phosphoinositide metabolism and the Ca/sup 2 +/-ATPase system.

  20. The influence of activation of heterogeneous ion-exchange membranes on their electrochemical properties

    Brožová, Libuše; Křivčík, J.; Neděla, D.; Kysela, V.; Žitka, Jan

    2015-01-01

    Roč. 56, č. 12 (2015), s. 3228-3232. ISSN 1944-3994. [International Conference on Membrane and Electromembrane Processes - MELPRO 2014. Prague, 18.05.2014-21.05.2014] Institutional support: RVO:61389013 Keywords : heterogeneous ion-exchange membranes * electrochemical properties * activation Subject RIV: JP - Industrial Processing Impact factor: 1.173, year: 2014

  1. Effect of low dosages of powdered activated carbon on membrane bioreactor performance

    Remy, M.J.J.; Temmink, H.; Rulkens, W.H.

    2012-01-01

    Previous research has demonstrated that powdered activated carbon (PAC), when applied at very low dosages and long SRTs, reduces membrane fouling in membrane bioreactors (MBRs). This effect was related to the formation of stronger sludge flocs, which are less sensitive to shear. In this contribution

  2. Why low powdered activated carbon addition reduces membrane fouling in MBRs

    Remy, M.J.J.; Potier, V.; Temmink, B.G.; Rulkens, W.H.

    2010-01-01

    Previous research had demonstrated that powdered activated carbon (PAC), when applied at very low dosages and long SRTs, reduces membrane fouling in membrane bioreactor (MBRs). In this contribution several mechanisms to explain this beneficial effect of PAC were investigated, including enhanced scou

  3. One-way membrane trafficking of SOS in receptor-triggered Ras activation.

    Christensen, Sune M; Tu, Hsiung-Lin; Jun, Jesse E; Alvarez, Steven; Triplet, Meredith G; Iwig, Jeffrey S; Yadav, Kamlesh K; Bar-Sagi, Dafna; Roose, Jeroen P; Groves, Jay T

    2016-09-01

    SOS is a key activator of the small GTPase Ras. In cells, SOS-Ras signaling is thought to be initiated predominantly by membrane recruitment of SOS via the adaptor Grb2 and balanced by rapidly reversible Grb2-SOS binding kinetics. However, SOS has multiple protein and lipid interactions that provide linkage to the membrane. In reconstituted-membrane experiments, these Grb2-independent interactions were sufficient to retain human SOS on the membrane for many minutes, during which a single SOS molecule could processively activate thousands of Ras molecules. These observations raised questions concerning how receptors maintain control of SOS in cells and how membrane-recruited SOS is ultimately released. We addressed these questions in quantitative assays of reconstituted SOS-deficient chicken B-cell signaling systems combined with single-molecule measurements in supported membranes. These studies revealed an essentially one-way trafficking process in which membrane-recruited SOS remains trapped on the membrane and continuously activates Ras until being actively removed via endocytosis. PMID:27501536

  4. Structural investigations of calcium binding and its role in activity and activation of outer membrane phospholipase A from Escherichia coli

    Snijder, H.J.; Kingma, R.L.; Kalk, K.H.; Egmond, M.R.; Dijkstra, B.W.

    2001-01-01

    Outer membrane phospholipase A (OMPLA) is an integral membrane enzyme that catalyses the hydrolysis of phospholipids. Enzymatic activity is regulated by reversible dimerisation and calcium-binding. We have investigated the role of calcium by X-ray crystallography. In monomeric OMPLA, one calcium ion

  5. Methodologies in the study of cell-cell fusion.

    Cohen, F S; Melikyan, G B

    1998-10-01

    The process of membrane fusion has been profitably studied by fusing cells that express fusion proteins on their surfaces to the membranes of target cells. Primary methods for monitoring the occurrence of fusion between cells are measurement of formation of heterokaryons, measurement of activation of reporter genes, measurement of transfer of lipidic and aqueous fluorescent dyes, and electrophysiological recording of fusion pores. Fluorescence and electrical methods have been well developed for fusion of a nucleated cell expressing viral fusion proteins to red blood cell targets. These techniques are now being extended to the study of fusion between two nucleated cells. Microscopic observation of spread of fluorescent dyes from one cell to another is a sensitive and convenient means of detecting fusion on the level of single events. In such studies, both the membrane and the aqueous continuities that occur as a result of fusion can be measured in the same experiment. By following spread of aqueous dyes of different sizes from one cell to another, the growth of a fusion pore can also be followed. By labeling cells with fluorescent probes, a state of hemifusion can be identified if probes in outer membrane leaflets transfer but probes in inner leaflets or aqueous spaces do not. Electrical measurements-both capacitance and double-whole-cell voltage-clamp techniques-are the most sensitive methods yet developed for detecting the formation of pores and for quantifying their growth. These powerful single-event methodologies should be directly applicable to further advances in expressing nonviral fusion proteins on cell surfaces. PMID:9790869

  6. The ORNL Controlled Fusion Atomic Data Center: Overview of Activities 2011

    The Controlled Fusion Atomic Data Center (CFADC) of the Oak Ridge National Laboratory continued operation aimed at collecting, evaluating, and disseminating atomic, molecular, and particle-surface interaction (AM and PSI) data needed by both the U.S. and international plasma science communities. This work has been carried out within an overarching atomic physics research group which produces much of the required data through an active experimental and theoretical science program. The production of an annotated bibliography of AM and PSI literature relevant to plasma science continues to be among the most important activities of the data center, forming the basis for the CFADC on-line bibliographic search engine and a significant part of the IAEA A+M Data Unit's 'International Bulletin on Atomic and Molecular Data for Fusion.' Also chief among the data center's activities are responses to specific data requests from the plasma science community, leading to either rapid feedback using existing data resources or long term data production projects, as well as participation in IAEA Coordinated Research Programs including recently 'Data for Surface Composition Dynamics Relevant to Erosion Processes' and 'Atomic and Molecular Data for Plasma Modeling.' Highlights of recent data production projects include the following: Experimental and theoretical data for inelastic electron-hydrocarbon reactions, large scale computational results for particle reflection from surfaces, measurements of chemical sputtering from carbon, inaugural experiments considering molecular ion collisions with neutral hydrogen, and expansion of the database of elastic and related transport cross sections calculated for intrinsic and extrinsic impurities in hydrogen plasmas. Progress is being hampered owing to news from the US Department of Energy that it plans to close out the program after a ramp down of funding in 2012, following a distinguished 52 year history of contributions to the US and

  7. Neutronics and activation analysis of a SS316 based experimental D-T fusion power reactor

    Neutronics and activation analysis for a SS316 based experimental tokamak fusion power reactor was completed using a two-dimensional neutronics model because of its practical features. Operating and decay power densities in all reactor components were obtained. The significant increase of neutron fluxes in the vacuum duct region was revealed. Maintenance and waste management aspects of the reactor components were investigated. Impacts on these aspects due to important minor alloying elements and impurities such as Mn, Co, Mo, and Nb were addressed

  8. FENDL/A-2.0. Neutron activation cross section data library for fusion applications

    This document describes the contents of a comprehensive neutron cross section data library for 13,006 neutron activation reactions with 739 target nuclides from H (A=1,Z=1) to Cm (A=248,Z=96), in the incident energy range up to 20 MeV. FENDL/A-2 is a sublibrary of FENDL-2, the second revision of the evaluated nuclear data library for fusion applications. It is supplemented by a decay data library FENDL/D-2 in ENDF-6 format for 1867 nuclides. The data are available from the IAEA Nuclear Data Section online via INTERNET by FTP command, or on magnetic tape upon request. (author)

  9. Membrane lipids regulate ganglioside GM2 catabolism and GM2 activator protein activity.

    Anheuser, Susi; Breiden, Bernadette; Schwarzmann, Günter; Sandhoff, Konrad

    2015-09-01

    Ganglioside GM2 is the major lysosomal storage compound of Tay-Sachs disease. It also accumulates in Niemann-Pick disease types A and B with primary storage of SM and with cholesterol in type C. Reconstitution of GM2 catabolism with β-hexosaminidase A and GM2 activator protein (GM2AP) at uncharged liposomal surfaces carrying GM2 as substrate generated only a physiologically irrelevant catabolic rate, even at pH 4.2. However, incorporation of anionic phospholipids into the GM2 carrying liposomes stimulated GM2 hydrolysis more than 10-fold, while the incorporation of plasma membrane stabilizing lipids (SM and cholesterol) generated a strong inhibition of GM2 hydrolysis, even in the presence of anionic phospholipids. Mobilization of membrane lipids by GM2AP was also inhibited in the presence of cholesterol or SM, as revealed by surface plasmon resonance studies. These lipids also reduced the interliposomal transfer rate of 2-NBD-GM1 by GM2AP, as observed in assays using Förster resonance energy transfer. Our data raise major concerns about the usage of recombinant His-tagged GM2AP compared with untagged protein. The former binds more strongly to anionic GM2-carrying liposomal surfaces, increases GM2 hydrolysis, and accelerates intermembrane transfer of 2-NBD-GM1, but does not mobilize membrane lipids. PMID:26175473

  10. Luteinizing hormone-releasing hormone fusion protein vaccines block estrous cycle activity in beef heifers.

    Stevens, J D; Sosa, J M; deAvila, D M; Oatley, J M; Bertrand, K P; Gaskins, C T; Reeves, J J

    2005-01-01

    Two LHRH fusion proteins, thioredoxin and ovalbumin, each containing seven LHRH inserts were tested for their ability to inhibit estrous cycle activity. The objective was to evaluate immune and biological responses from alternating the two fusion proteins in an immunization schedule. One hundred ten heifers were divided equally into 11 groups. Two control groups consisted of either spayed or intact, untreated heifers. Heifers in the other nine groups were immunized on wk 0, 4, and 9. Treatments were immunizations of the same protein throughout or alternating the proteins in different booster sequences. Blood was collected weekly for 22 wk, and serum was assayed for concentrations of progesterone and titers of anti-LHRH. At slaughter, reproductive tracts were removed from each heifer and weighed. Heifers with >or=1 ng/mL of progesterone were considered to have a functional corpus luteum and thus to have estrous cycle activity. All LHRH-immunized groups of heifers had a smaller (P spayed heifers during wk 9 to 22. Anti-LHRH did not differ among immunized groups during wk 1 to 9. Starting at wk 10 and continuing through the conclusion of the study, there was an overall difference among treatment groups for anti-LHRH (P spaying in suppression of estrous cycle activity, but alternating the two proteins in an immunization schedule did not enhance the immunological or biological effectiveness of the vaccine. PMID:15583055

  11. Conformational activation of visual rhodopsin in native disc membranes

    Malmerberg, E.; Bovee-Geurts, P.H.M.; Katona, G.; Deupi, X.; Arnlund, D.; Wickstrand, C.; Johansson, L.C.; Westenhoff, S.; Nazarenko, E.; GF, X.S.; Menzel, A.; Grip, W.J. de; Neutze, R.

    2015-01-01

    Rhodopsin is the G protein-coupled receptor (GPCR) that serves as a dim-light receptor for vision in vertebrates. We probed light-induced conformational changes in rhodopsin in its native membrane environment at room temperature using time-resolved wide-angle x-ray scattering. We observed a rapid co

  12. Cellulose microfibril deposition: coordinated activity at the plant plasma membrane

    Lindeboom, J.J.; Mulder, B.; Vos, J.W.; Ketelaar, M.J.; Emons, A.M.C.

    2008-01-01

    Plant cell wall production is a membrane-bound process. Cell walls are composed of cellulose microfibrils, embedded inside a matrix of other polysaccharides and glycoproteins. The cell wall matrix is extruded into the existing cell wall by exocytosis. This same process also inserts the cellulose syn

  13. G-protein activity in Percoll-purified plasma membranes, bulk plasma membranes, and low-density plasma membranes isolated from rat cerebral cortex

    Bouřová, Lenka; Stöhr, Jiří; Lisý, Václav; Rudajev, Vladimír; Novotný, Jiří; Svoboda, Petr

    2009-01-01

    Roč. 15, č. 4 (2009), BR111-BR122. ISSN 1234-1010 R&D Projects: GA MŠk(CZ) LC554; GA MŠk(CZ) LC06063; GA ČR(CZ) GA309/06/0121; GA AV ČR(CZ) IAA500110606 Institutional research plan: CEZ:AV0Z50110509 Keywords : rat cerebral cortex * plasma membrane * G-protein activity Subject RIV: CE - Biochemistry Impact factor: 1.543, year: 2009

  14. A fusion-inhibiting peptide against Rift Valley fever virus inhibits multiple, diverse viruses.

    Jeffrey W Koehler

    Full Text Available For enveloped viruses, fusion of the viral envelope with a cellular membrane is critical for a productive infection to occur. This fusion process is mediated by at least three classes of fusion proteins (Class I, II, and III based on the protein sequence and structure. For Rift Valley fever virus (RVFV, the glycoprotein Gc (Class II fusion protein mediates this fusion event following entry into the endocytic pathway, allowing the viral genome access to the cell cytoplasm. Here, we show that peptides analogous to the RVFV Gc stem region inhibited RVFV infectivity in cell culture by inhibiting the fusion process. Further, we show that infectivity can be inhibited for diverse, unrelated RNA viruses that have Class I (Ebola virus, Class II (Andes virus, or Class III (vesicular stomatitis virus fusion proteins using this single peptide. Our findings are consistent with an inhibition mechanism similar to that proposed for stem peptide fusion inhibitors of dengue virus in which the RVFV inhibitory peptide first binds to both the virion and cell membranes, allowing it to traffic with the virus into the endocytic pathway. Upon acidification and rearrangement of Gc, the peptide is then able to specifically bind to Gc and prevent fusion of the viral and endocytic membranes, thus inhibiting viral infection. These results could provide novel insights into conserved features among the three classes of viral fusion proteins and offer direction for the future development of broadly active fusion inhibitors.

  15. Semiallogenic fusions of MSI+ tumor cells and activated B cells induce MSI-specific T cell responses

    Klier Ulrike

    2011-09-01

    Full Text Available Abstract Background Various strategies have been developed to transfer tumor-specific antigens into antigen presenting cells in order to induce cytotoxic T cell responses against tumor cells. One approach uses cellular vaccines based on fusions of autologous antigen presenting cells and allogeneic tumor cells. The fusion cells combine antigenicity of the tumor cell with optimal immunostimulatory capacity of the antigen presenting cells. Microsatellite instability caused by mutational inactivation of DNA mismatch repair genes results in translational frameshifts when affecting coding regions. It has been shown by us and others that these mutant proteins lead to the presentation of immunogenic frameshift peptides that are - in principle - recognized by a multiplicity of effector T cells. Methods We chose microsatellite instability-induced frameshift antigens as ideal to test for induction of tumor specific T cell responses by semiallogenic fusions of microsatellite instable carcinoma cells with CD40-activated B cells. Two fusion clones of HCT116 with activated B cells were selected for stimulation of T cells autologous to the B cell fusion partner. Outgrowing T cells were phenotyped and tested in functional assays. Results The fusion clones expressed frameshift antigens as well as high amounts of MHC and costimulatory molecules. Autologous T cells stimulated with these fusions were predominantly CD4+, activated, and reacted specifically against the fusion clones and also against the tumor cell fusion partner. Interestingly, a response toward 6 frameshift-derived peptides (of 14 tested could be observed. Conclusion Cellular fusions of MSI+ carcinoma cells and activated B cells combine the antigen-presenting capacity of the B cell with the antigenic repertoire of the carcinoma cell. They present frameshift-derived peptides and can induce specific and fully functional T cells recognizing not only fusion cells but also the carcinoma cells. These

  16. Semiallogenic fusions of MSI+ tumor cells and activated B cells induce MSI-specific T cell responses

    Various strategies have been developed to transfer tumor-specific antigens into antigen presenting cells in order to induce cytotoxic T cell responses against tumor cells. One approach uses cellular vaccines based on fusions of autologous antigen presenting cells and allogeneic tumor cells. The fusion cells combine antigenicity of the tumor cell with optimal immunostimulatory capacity of the antigen presenting cells. Microsatellite instability caused by mutational inactivation of DNA mismatch repair genes results in translational frameshifts when affecting coding regions. It has been shown by us and others that these mutant proteins lead to the presentation of immunogenic frameshift peptides that are - in principle - recognized by a multiplicity of effector T cells. We chose microsatellite instability-induced frameshift antigens as ideal to test for induction of tumor specific T cell responses by semiallogenic fusions of microsatellite instable carcinoma cells with CD40-activated B cells. Two fusion clones of HCT116 with activated B cells were selected for stimulation of T cells autologous to the B cell fusion partner. Outgrowing T cells were phenotyped and tested in functional assays. The fusion clones expressed frameshift antigens as well as high amounts of MHC and costimulatory molecules. Autologous T cells stimulated with these fusions were predominantly CD4+, activated, and reacted specifically against the fusion clones and also against the tumor cell fusion partner. Interestingly, a response toward 6 frameshift-derived peptides (of 14 tested) could be observed. Cellular fusions of MSI+ carcinoma cells and activated B cells combine the antigen-presenting capacity of the B cell with the antigenic repertoire of the carcinoma cell. They present frameshift-derived peptides and can induce specific and fully functional T cells recognizing not only fusion cells but also the carcinoma cells. These hybrid cells may have great potential for cellular immunotherapy and

  17. Preparation of polysaccharide loaded collagen membrane with anti-oxidative activity.

    Shu, Zibin; Ding, Shengli; He, Xiaohong; Dai, Xuemei; Xiao, Qian; Yang, Min; Leng, Xue; Ma, Yanshun; Yang, Hua

    2015-01-01

    The scavenging activity of polysaccharides from Lycium barbarum, Lentinus edodes and Ganoderma Lucidum Karst to DPPH free radicals was investigated. It was found that among the three polysaccharides, Lycium barbarum polysaccharides (LBP) exhibits the best scavenging activity. Polysaccharide loaded collagen membranes were prepared by mixing LBP with collagen, starch, glycerol, sodium carboxymethyl cellulose and glutaraldehyde. In vitro drug release from membranes was evaluated. With increasing the immersion time, the release rate first increases and then slows down. Meanwhile, the scavenging activity to DPPH radicals exhibits similar variation, in agreement with a good release effect of the membrane. The optimal formulation of collagen membrane and preparation parameters were obtained considering the overall properties and the scavenging activity to radicals. PMID:26406078

  18. Extra amino group-containing gramicidin S analogs possessing outer membrane-permeabilizing activity

    Kawai, Masao; Tanaka, Ryoji; Yamamura, Hatsuo; Yasuda, Keiko; Narita, Shizuto; Umemoto, Hiroshi; Ando, Setsuko; Katsu, Takashi; ヤマムラ, ハツオ; 山村, 初雄

    2003-01-01

    Novel (2S,4R)- and (2S,4S)-4-aminoproline residue-containing analogs of the cyclic decapeptide antibiotic gramicidin S were synthesized, which exhibited marked permeabilizing activity on the outer membrane of gram-negative bacteria.

  19. Gβ1γ2 Activates Phospholipase A2-Dependent Golgi Membrane Tubule Formation

    WilliamJBrown

    2014-02-01

    Full Text Available Heterotrimeric G proteins transduce the ligand binding of transmembrane G protein coupled receptors into a variety of intracellular signaling pathways. Recently, heterotrimeric Gβγ subunit signaling at the Golgi complex has been shown to regulate the formation of vesicular transport carriers that deliver cargo from the Golgi to the plasma membrane. In addition to vesicles, membrane tubules have also been shown to mediate export from the Golgi complex, which requires the activity of cytoplasmic phospholipase A2 (PLA2 enzyme activity. Through the use of an in vitro reconstitution assay with isolated Golgi complexes, we provide evidence that Gβ1γ2 signaling also stimulates Golgi membrane tubule formation. In addition, we show that an inhibitor of Gβγ activation of PLA2 enzymes inhibits in vitro Golgi membrane tubule formation. Additionally, purified Gβγ protein stimulates membrane tubules in the presence of low (sub-threshold cytosol concentrations. Importantly, this Gβγ stimulation of Golgi membrane tubule formation was inhibited by treatment with the PLA2 antagonist ONO-RS-082. These studies indicate that Gβ1γ2 signaling activates PLA2 enzymes required for Golgi membrane tubule formation, thus establishing a new layer of regulation for this process.

  20. The Computational Power of Exponential-Space P Systems with Active Membranes

    Alhazov, Artiom; Leporati, Alberto; Mauri, Giancarlo; Porreca, Antonio E.; Zandron, Claudio; Research Group on Natural Computing (Universidad de Sevilla) (Coordinador)

    2012-01-01

    We show that exponential-space P systems with active membranes characterize the complexity class EXPSPACE. This result is proved by simulating Turing machines working in exponential space via uniform families of P systems with restricted elementary active membranes; the simulation is e cient, in the sense that the time and space required are at most polynomial with respect to the resources employed by the simulated Turing machine.

  1. Complete Problems for a Variant of P Systems with Active Membranes

    Porreca, Antonio E.; Leporati, Alberto; Mauri, Giancarlo; Zandron, Claudio; Research Group on Natural Computing (Universidad de Sevilla) (Coordinador)

    2010-01-01

    We identify a family of decision problems that are hard for some complexity classes defined in terms of P systems with active membranes working in polynomial time. Furthermore, we prove the completeness of these problems in the case where the systems are equipped with a form of priority that linearly orders their rules. Finally, we highlight some possible connections with open problems related to the computational complexity of P systems with active membranes.

  2. Activation calculation and environmental safety analysis for fusion experimental breeder (FEB)

    An activation calculation code FDKR and decay chain data library AFDCDLIB are used to calculate the radioactivity, decay heat, dose rate and biological hazard potential (BHP) form activation products, actinides and fission products in a Fusion Experiment Breeder (FEB). The code and library are introduced briefly, and calculation results and decay curves of related hazards after one year operation with 150 MW fusion power are given. The total radioactivity inventory, decay heat and BHP are 5.74 x 1020 Bq, 8.34 MW and 4.08 x 108 km3 of air, respectively, at shutdown. Results obtained show that the first wall of FEB can meet the nuclear waste disposal criteria for the NRC 10 CFR61 Class C after a few weeks from shutdown. The inventory of important actinides for the fuel reprocessing, such as 232U and 237Np were also calculated. It was shown that their concentrations do not excess the limit value of environmental safety required. (9 refs., 4 figs., 9 tabs.)

  3. Joining techniques for a reduced activation 12Cr steel for inertial fusion energy

    Hunt, R. M. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); El-Dasher, B. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Choi, B. W. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Torres, S. G. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2014-10-01

    At Lawrence Livermore National Laboratory, we are developing a reduced activation ferritic martensitic steel that is based on the ferritic martensitic steel HT-9. As a part of the development of this steel, we tested a series of welding processes for characterization, including conventional welds (electron beam, tungsten inert gas, and laser) as well as solid-state welds (hot isostatic pressing). We also heat treated the joints at various temperatures between 750 °C and 1050 °C to find a suitable normalization scheme. The modified HT-9 reduced activation ferritic martensitic steel appears highly suitable to welding and diffusion bonding. All welds showed good quality fusion zones with insignificant cracking or porosity. Additionally, a heat treatment schedule of 950 °C for one hour caused minimal grain growth while still converging the hardness of the base metal with that of the fusion and heat-affected zones. Also, modified HT-9 diffusion bonds that were created at temperatures of at least 950 °C for two hours at 103 MPa had interface tensile strengths of greater than 600 MPa. The diffusion bonds showed no evidence of increased hardness nor void formation at the diffusion bonded interface.

  4. Synthetic aperture microwave imaging with active probing for fusion plasma diagnostics

    A Synthetic Aperture Microwave Imaging (SAMI) system has been designed and built to obtain 2-D images at several frequencies from fusion plasmas. SAMI uses a phased array of linearly polarised antennas. The array configuration has been optimised to achieve maximum synthetic aperture beam efficiency. The signals received by antennas are down-converted to the intermediate frequency range and then recorded in a full vector form. Full vector signals allow beam focusing and image reconstruction in both real time and a post processing mode. SAMI can scan over 16 preprogrammed frequencies in the range of 10–35 GHz with a switching time of 300ns. The system operates in 2 different modes simultaneously: both a 'passive' imaging of plasma emission and also an 'active' imaging of the back-scattered signal of the radiation launched by one of the antennas from the same array. This second mode is similar to so-called Doppler backscattering (DBS) reflectometry with 2-D resolution of the propagation velocity of turbulent structures. Both modes of operation show good performance in a real fusion plasma experiments on Mega Amp Spherical Tokamak (MAST). We have obtained the first ever 2-D images of BXO mode conversion windows. With active probing, the first ever turbulence velocity maps have been obtained. In this article we present an overview of the diagnostic and discuss recent results.

  5. Constant-Space P Systems with Active Membranes

    Leporati, Alberto; Manzoni, Luca; Mauri, Giancarlo; Porreca, Antonio E.; Zandron, Claudio; Research Group on Natural Computing (Universidad de Sevilla) (Coordinador)

    2014-01-01

    We continue the investigation of the computational power of space- constrained P systems. We show that only a constant amount of space is needed in order to simulate a polynomial-space bounded Turing machine. Due to this result, we propose an alternative de nition of space complexity for P systems, where the amount of information contained in individual objects and membrane labels is also taken into ac- count. Finally, we prove that, when less than a logarithmic number of membr...

  6. A stimulus-activated conductance in isolated taste epithelial membranes.

    Teeter, J H; Brand, J. G.; Kumazawa, T.

    1990-01-01

    Membrane vesicles isolated from the cutaneous taste epithelium of the catfish were incorporated into phospholipid bilayers on the tips of patch pipettes. Voltage-dependent conductances were observed in approximately 50% of the bilayers and single-channel currents having conductances from 8 to greater than 250 pS were recorded. In 40% of the bilayers displaying no voltage-dependent conductances, micromolar concentrations of L-arginine, a potent stimulus for one class of catfish amino acid tast...

  7. High-pressure stainless steel active membrane microvalves

    Sharma, G.; Svensson, S.; Ogden, S.; Klintberg, L.; Hjort, K.

    2011-07-01

    In this work, high-pressure membrane microvalves have been designed, manufactured and evaluated. The valves were able to withstand back-pressures of 200 bar with a response time of less than 0.6 s. These stainless steel valves, manufactured with back-end batch production, utilize the large volume expansion coupled to the solid-liquid phase transition in paraffin wax. When membrane materials were evaluated, parylene coated stainless steel was found to be the best choice as compared to polydimethylsiloxane and polyimide. Also, the influence of the orifice placement and diameter is included in this work. If the orifice is placed too close to the rim of the membrane, the valve can stay sealed even after turning the power off, and the valve will not open until the pressure in the system is released. The developed steel valves, evaluated for both water and air, provide excellent properties in terms of mechanical stability, ease of fabrication, and low cost. Possible applications include sampling at high pressures, chemical microreactors, high performance liquid chromatography, pneumatics, and hydraulics.

  8. Myristoylated and non-myristoylated forms of the pH sensor protein hisactophilin II: intracellular shuttling to plasma membrane and nucleus monitored in real time by a fusion with green fluorescent protein.

    Hanakam, F; Albrecht, R; Eckerskorn, C; Matzner, M; Gerisch, G

    1996-01-01

    Hisactophilins are myristoylated proteins that are rich in histidine residues and known to exist in Dictyostelium cells in a plasma membrane-bound and a soluble cytoplasmic state. Intracellular translocation of these proteins in response to pH changes was monitored using hisactophilin fusions with green fluorescent protein (GFP) and confocal laser scanning microscopy. Both the normal and a mutated non-myristoylated fusion protein shuffled within the cells in a pH-dependent manner. After lowering the pH, these proteins translocated within minutes between the cytoplasm, the plasma membrane and the nucleus. The role of histidine clusters on the surface of hisactophilin molecules in binding of the proteins to the plasma membrane and in their transfer to the nucleus is discussed on the basis of a pH switch mechanism. Images PMID:8670794

  9. Management of water leaks on Tore Supra actively cooled fusion device

    Up to now, Tore Supra is the only fusion device fully equipped with actively cooled Plasma Facing Components (PFCs). In case of abnormal events during a plasma discharge, the PFCs could be submitted to a transient high power density (run away electrons) or to a continuous phenomena as local thermal flux induced by trapped suprathermal electrons or ions). It could lead to a degradation of the PFC integrity and in the worst case to a water leak occurrence. Such water leak has important consequence on the tokamak operation that concerns PFCs themselves, monitoring equipment located in the vacuum vessel or connected to the ports as RF antennas, diagnostics or pumping systems. Following successive water leak events (the most important water leak, that occurred in September 2002, is described in the paper), a large feedback experience has been gained on Tore supra since more than 15 years that could be useful to actively cooled next devices as W7X and ITER. (authors)

  10. Convergent mutations and kinase fusions lead to oncogenic STAT3 activation in anaplastic large cell lymphoma.

    Crescenzo, Ramona; Abate, Francesco; Lasorsa, Elena; Tabbo', Fabrizio; Gaudiano, Marcello; Chiesa, Nicoletta; Di Giacomo, Filomena; Spaccarotella, Elisa; Barbarossa, Luigi; Ercole, Elisabetta; Todaro, Maria; Boi, Michela; Acquaviva, Andrea; Ficarra, Elisa; Novero, Domenico; Rinaldi, Andrea; Tousseyn, Thomas; Rosenwald, Andreas; Kenner, Lukas; Cerroni, Lorenzo; Tzankov, Alexander; Ponzoni, Maurilio; Paulli, Marco; Weisenburger, Dennis; Chan, Wing C; Iqbal, Javeed; Piris, Miguel A; Zamo', Alberto; Ciardullo, Carmela; Rossi, Davide; Gaidano, Gianluca; Pileri, Stefano; Tiacci, Enrico; Falini, Brunangelo; Shultz, Leonard D; Mevellec, Laurence; Vialard, Jorge E; Piva, Roberto; Bertoni, Francesco; Rabadan, Raul; Inghirami, Giorgio

    2015-04-13

    A systematic characterization of the genetic alterations driving ALCLs has not been performed. By integrating massive sequencing strategies, we provide a comprehensive characterization of driver genetic alterations (somatic point mutations, copy number alterations, and gene fusions) in ALK(-) ALCLs. We identified activating mutations of JAK1 and/or STAT3 genes in ∼20% of 88 [corrected] ALK(-) ALCLs and demonstrated that 38% of systemic ALK(-) ALCLs displayed double lesions. Recurrent chimeras combining a transcription factor (NFkB2 or NCOR2) with a tyrosine kinase (ROS1 or TYK2) were also discovered in WT JAK1/STAT3 ALK(-) ALCL. All these aberrations lead to the constitutive activation of the JAK/STAT3 pathway, which was proved oncogenic. Consistently, JAK/STAT3 pathway inhibition impaired cell growth in vitro and in vivo. PMID:25873174

  11. Arf6 recruits the Rac GEF Kalirin to the plasma membrane facilitating Rac activation

    Donaldson Julie G

    2007-07-01

    Full Text Available Abstract Background Many studies implicate Arf6 activity in Rac-mediated membrane ruffling and cytoskeletal reorganization. Although Arf6 facilitates the trafficking of Rac1 to the plasma membrane and in many cases Arf6 activation leads to the activation of Rac1, the details of how Arf6 influences Rac function remain to be elucidated. Results We demonstrate in binding assays and by co-immunoprecipitation that GDP-bound Arf6 binds to Kalirin5, a Rho family guanine nucleotide exchange factor, through interaction with the spectrin repeat region. In cells, expression of wild type Arf6 recruits spectrin repeat 5 and Kalirin to the plasma membrane and leads to enhanced Kalirin5-induced ruffling. By contrast, expression of an Arf6 mutant that cannot become activated, Arf6 T27N, still recruits spectrin repeat 5 and Kalirin to membranes but inhibits Kalirin5-induced ruffling in HeLa cells. Kalirin5-induced Rac1 activation is increased by the expression of wild type Arf6 and decreased by Arf6T27N. Furthermore, expression of a catalytically-inactive mutant of Kalirin5 inhibits cytoskeletal changes observed in cells expressing EFA6, an Arf6 guanine nucleotide exchange factor that leads to activation of Rac. Conclusion We show here with over-expressed proteins that the GDP-bound form of Arf6 can bind to the spectrin repeat regions in Kalirin Rho family GEFs thereby recruiting Kalirin to membranes. Although Kalirin is recruited onto membranes by Arf6-GDP, subsequent Rac activation and membrane ruffling requires Arf6 activation. From these results, we suggest that Arf6 can regulate through its GTPase cycle the activation of Rac.

  12. Recognition of acidic phospholipase A2 activity in plasma membranes of resident peritoneal macrophages

    Phospholipase (PLase) activities in the plasma membrane of guinea pig peritoneal macrophages were studied, as these enzymes having such activity may be candidates for the release of arachidonic acid (AA) from phosphatidylcholine (PC). An AA release system operating at acidic pH was identified in the macrophage plasma membrane and characterized. This membrane-bound acidic PLase A2 had an optimum pH at 4.5, and enzyme activation was observed in Ca++-free medium; but the maximum activity was found at 0.5 mM Ca++ concentration. The Km value for PC of acidic PLase A2 was 4.2 μM, and a Michaelis-Menten relationship was evident. Calcium might act as a cofactor at some intermediate step during the activation of acidic PLase A2 in light of the uncompetitive manner of Ca++ action. Furthermore, the release of [3H]-AA from preradiolabelled macrophage plasma membranes occurred with the addition of Ca++ at pH 4.5. These data suggest that the acid PLase A2 is a component of the plasma membrane and is not due to lysosomal contamination since membrane-bound acidic PLase A2 properties are opposite to those found for lysosomal PLase A2

  13. Sodium pump molecular activity and membrane lipid composition in two disparate ectotherms, and comparison with endotherms.

    Turner, Nigel; Hulbert, A J; Else, Paul L

    2005-02-01

    Previous research has shown that the lower sodium pump molecular activity observed in tissues of ectotherms compared to endotherms, is largely related to the lower levels of polyunsaturates and higher levels of monounsaturates found in the cell membranes of ectotherms. Marine-based ectotherms, however, have very polyunsaturated membranes, and in the current study, we measured molecular activity and membrane lipid composition in tissues of two disparate ectothermic species, the octopus (Octopus vulgaris) and the bearded dragon lizard (Pogona vitticeps), to determine whether the high level of membrane polyunsaturation generally observed in marine-based ectotherms is associated with an increased sodium pump molecular activity relative to other ectotherms. Phospholipids from all tissues of the octopus were highly polyunsaturated and contained high concentrations of the omega-3 polyunsaturate, docosahexaenoic acid (22:6 (n-3)). In contrast, phospholipids from bearded dragon tissues contained higher proportions of monounsaturates and lower proportions of polyunsaturates. Sodium pump molecular activity was only moderately elevated in tissues of the octopus compared to the bearded dragon, despite the much greater level of polyunsaturation in octopus membranes. When the current data were combined with data for the ectothermic cane toad, a significant (P = 0.003) correlation was observed between sodium pump molecular activity and the content of 22:6 (n-3) in the surrounding membrane. These results are discussed in relation to recent work which shows a similar relationship in endotherms. PMID:15726386

  14. Nuclear envelope breakdown induced by herpes simplex virus type 1 involves the activity of viral fusion proteins

    Herpesvirus infection reorganizes components of the nuclear lamina usually without loss of integrity of the nuclear membranes. We report that wild-type HSV infection can cause dissolution of the nuclear envelope in transformed mouse embryonic fibroblasts that do not express torsinA. Nuclear envelope breakdown is accompanied by an eight-fold inhibition of virus replication. Breakdown of the membrane is much more limited during infection with viruses that lack the gB and gH genes, suggesting that breakdown involves factors that promote fusion at the nuclear membrane. Nuclear envelope breakdown is also inhibited during infection with virus that does not express UL34, but is enhanced when the US3 gene is deleted, suggesting that envelope breakdown may be enhanced by nuclear lamina disruption. Nuclear envelope breakdown cannot compensate for deletion of the UL34 gene suggesting that mixing of nuclear and cytoplasmic contents is insufficient to bypass loss of the normal nuclear egress pathway. - Highlights: • We show that wild-type HSV can induce breakdown of the nuclear envelope in a specific cell system. • The viral fusion proteins gB and gH are required for induction of nuclear envelope breakdown. • Nuclear envelope breakdown cannot compensate for deletion of the HSV UL34 gene

  15. Nuclear envelope breakdown induced by herpes simplex virus type 1 involves the activity of viral fusion proteins

    Maric, Martina; Haugo, Alison C. [Department of Microbiology, University of Iowa, Iowa City, IA 52242 (United States); Dauer, William [Department of Neurology, University of Michigan, Ann Arbor, MI 48109 (United States); Johnson, David [Department of Microbiology and Immunology, Oregon Health Sciences University, Portland, OR 97201 (United States); Roller, Richard J., E-mail: richard-roller@uiowa.edu [Department of Microbiology, University of Iowa, Iowa City, IA 52242 (United States)

    2014-07-15

    Herpesvirus infection reorganizes components of the nuclear lamina usually without loss of integrity of the nuclear membranes. We report that wild-type HSV infection can cause dissolution of the nuclear envelope in transformed mouse embryonic fibroblasts that do not express torsinA. Nuclear envelope breakdown is accompanied by an eight-fold inhibition of virus replication. Breakdown of the membrane is much more limited during infection with viruses that lack the gB and gH genes, suggesting that breakdown involves factors that promote fusion at the nuclear membrane. Nuclear envelope breakdown is also inhibited during infection with virus that does not express UL34, but is enhanced when the US3 gene is deleted, suggesting that envelope breakdown may be enhanced by nuclear lamina disruption. Nuclear envelope breakdown cannot compensate for deletion of the UL34 gene suggesting that mixing of nuclear and cytoplasmic contents is insufficient to bypass loss of the normal nuclear egress pathway. - Highlights: • We show that wild-type HSV can induce breakdown of the nuclear envelope in a specific cell system. • The viral fusion proteins gB and gH are required for induction of nuclear envelope breakdown. • Nuclear envelope breakdown cannot compensate for deletion of the HSV UL34 gene.

  16. The higher level of complexity of K-Ras4B activation at the membrane.

    Jang, Hyunbum; Banerjee, Avik; Chavan, Tanmay S; Lu, Shaoyong; Zhang, Jian; Gaponenko, Vadim; Nussinov, Ruth

    2016-04-01

    Is nucleotide exchange sufficient to activate K-Ras4B? To signal, oncogenic rat sarcoma (Ras) anchors in the membrane and recruits effectors by exposing its effector lobe. With the use of NMR and molecular dynamics (MD) simulations, we observed that in solution, farnesylated guanosine 5'-diphosphate (GDP)-bound K-Ras4B is predominantly autoinhibited by its hypervariable region (HVR), whereas the GTP-bound state favors an activated, HVR-released state. On the anionic membrane, the catalytic domain adopts multiple orientations, including parallel (∼180°) and perpendicular (∼90°) alignments of the allosteric helices, with respect to the membrane surface direction. In the autoinhibited state, the HVR is sandwiched between the effector lobe and the membrane; in the active state, with membrane-anchored farnesyl and unrestrained HVR, the catalytic domain fluctuates reinlessly, exposing its effector-binding site. Dimerization and clustering can reduce the fluctuations. This achieves preorganized, productive conformations. Notably, we also observe HVR-autoinhibited K-Ras4B-GTP states, with GDP-bound-like orientations of the helices. Thus, we propose that the GDP/GTP exchange may not be sufficient for activation; instead, our results suggest that the GDP/GTP exchange, HVR sequestration, farnesyl insertion, and orientation/localization of the catalytic domain at the membrane conjointly determine the active or inactive state of K-Ras4B. Importantly, K-Ras4B-GTP can exist in active and inactive states; on its own, GTP binding may not compel K-Ras4B activation.-Jang, H., Banerjee, A., Chavan, T. S, Lu, S., Zhang, J., Gaponenko, V., Nussinov, R. The higher level of complexity of K-Ras4B activation at the membrane. PMID:26718888

  17. Powder Activated Carbon Pretreatment of a Microfiltration Membrane for the Treatment of Surface Water

    Yali Song

    2015-09-01

    Full Text Available This study focused on the effect of powder activated carbon (PAC adsorption on microfiltration (MF membrane performance. The results showed that PAC pretreatment offered high organic matter removal rates for both dissolved organic carbon (DOC and ultraviolet absorbance at 254 nm (UV254 during 10–200 mg/L PAC dosage. The removal efficiencies of organic matter by MF membrane filtration decreased with the increase of organic matter removal rate by PAC adsorption. PAC mainly removed organic matter of about 3 kDa molecular weight (MW. MF membrane maintained more than 5 kDa MW organic matter on the membrane after PAC adsorption. The results of membrane filtration indicated that PAC pretreatment slightly promoted membrane flux, regardless of PAC dosage. It seems that the organic matter fouling membrane was concentrated in more than 3 kDa MW. PAC removed markedly less than 3 kDa MW organic matter and had less effect on more than 3 kDa organic matter. Thus, PAC cannot reduce membrane fouling.

  18. The antifungal activity and membrane-disruptive action of dioscin extracted from Dioscorea nipponica.

    Cho, Jaeyong; Choi, Hyemin; Lee, Juneyoung; Kim, Mi-Sun; Sohn, Ho-Yong; Lee, Dong Gun

    2013-03-01

    Dioscin is a kind of steroidal saponin isolated from the root bark of wild yam Dioscorea nipponica. We investigated the antifungal effect of dioscin against different fungal strains and its antifungal mechanism(s) in Candida albicans cells. Using the propidium iodide assay and calcein-leakage measurement, we confirmed that dioscin caused fungal membrane damage. Furthermore, we evaluated the ability of dioscin to disrupt the plasma membrane potential, using 3,3'-dipropylthiadicarbocyanine iodide [DiSC(3)(5)] and bis-(1,3-dibarbituric acid)-trimethine oxanol [DiBAC(4)(3)]. Cells stained with the dyes had a significant increase in fluorescent intensity after exposure to dioscin, indicating that dioscin has an effect on the membrane potential. To visualize the effect of dioscin on the cell membrane, we synthesized rhodamine-labeled giant unilamellar vesicles (GUVs) mimicking the outer leaflet of the plasma membrane of C. albicans. As seen in the result, the membrane disruptive action of dioscin caused morphological change and rhodamine leakage of the GUVs. In three-dimensional contour-plot analysis using flow cytometry, we observed a decrease in cell size, which is in agreement with our result from the GUV assay. These results suggest that dioscin exerts a considerable antifungal activity by disrupting the structure in membrane after invading into the fungal membrane, resulting in fungal cell death. PMID:23262192

  19. Powder Activated Carbon Pretreatment of a Microfiltration Membrane for the Treatment of Surface Water

    Song, Yali; Dong, Bingzhi; Gao, Naiyun; Ma, Xiaoyan

    2015-01-01

    This study focused on the effect of powder activated carbon (PAC) adsorption on microfiltration (MF) membrane performance. The results showed that PAC pretreatment offered high organic matter removal rates for both dissolved organic carbon (DOC) and ultraviolet absorbance at 254 nm (UV254) during 10–200 mg/L PAC dosage. The removal efficiencies of organic matter by MF membrane filtration decreased with the increase of organic matter removal rate by PAC adsorption. PAC mainly removed organic matter of about 3 kDa molecular weight (MW). MF membrane maintained more than 5 kDa MW organic matter on the membrane after PAC adsorption. The results of membrane filtration indicated that PAC pretreatment slightly promoted membrane flux, regardless of PAC dosage. It seems that the organic matter fouling membrane was concentrated in more than 3 kDa MW. PAC removed markedly less than 3 kDa MW organic matter and had less effect on more than 3 kDa organic matter. Thus, PAC cannot reduce membrane fouling. PMID:26378552

  20. Activation of Endothelial Nitric Oxide (eNOS Occurs through Different Membrane Domains in Endothelial Cells.

    Jason Tran

    Full Text Available Endothelial cells respond to a large range of stimuli including circulating lipoproteins, growth factors and changes in haemodynamic mechanical forces to regulate the activity of endothelial nitric oxide synthase (eNOS and maintain blood pressure. While many signalling pathways have been mapped, the identities of membrane domains through which these signals are transmitted are less well characterized. Here, we manipulated bovine aortic endothelial cells (BAEC with cholesterol and the oxysterol 7-ketocholesterol (7KC. Using a range of microscopy techniques including confocal, 2-photon, super-resolution and electron microscopy, we found that sterol enrichment had differential effects on eNOS and caveolin-1 (Cav1 colocalisation, membrane order of the plasma membrane, caveolae numbers and Cav1 clustering. We found a correlation between cholesterol-induced condensation of the plasma membrane and enhanced high density lipoprotein (HDL-induced eNOS activity and phosphorylation suggesting that cholesterol domains, but not individual caveolae, mediate HDL stimulation of eNOS. Vascular endothelial growth factor (VEGF-induced and shear stress-induced eNOS activity was relatively independent of membrane order and may be predominantly controlled by the number of caveolae on the cell surface. Taken together, our data suggest that signals that activate and phosphorylate eNOS are transmitted through distinct membrane domains in endothelial cells.

  1. Nonlinear Dielectric Spectroscopy as an Indirect Probe of Metabolic Activity in Thylakoid Membrane

    John H. Miller

    2011-01-01

    Full Text Available Nonlinear dielectric spectroscopy (NDS is a non-invasive probe of cellular metabolic activity with potential application in the development of whole-cell biosensors. However, the mechanism of NDS interaction with metabolic membrane proteins is poorly understood, partly due to the inherent complexity of single cell organisms. Here we use the light-activated electron transport chain of spinach thylakoid membrane as a model system to study how NDS interacts with metabolic activity. We find protein modification, as opposed to membrane pump activity, to be the dominant source of NDS signal change in this system. Potential mechanisms for such protein modifications include reactive oxygen species generation and light-activated phosphorylation.

  2. Analysis of Induced Gamma Activation by D-T Neutrons in Selected Fusion Reactor Relevant Materials with EAF-2010

    Klix Axel

    2016-01-01

    Full Text Available Samples of lanthanum, erbium and titanium which are constituents of structural materials, insulating coatings and tritium breeder for blankets of fusion reactor designs have been irradiated in a fusion peak neutron field. The induced gamma activities were measured and the results were used to check calculations with the European activation system EASY-2010. Good agreement for the prediction of major contributors to the contact dose rate of the materials was found, but for minor contributors the calculation deviated up to 50%.

  3. Towards fusion energy as a sustainable energy source: Activities at DTU Physics

    Rasmussen, Jesper; Christensen, Alexander Simon; Dam, Magnus;

    2014-01-01

    Nuclear fusion – the process from which the Sun derives its energy – holds the potential to become a clean,safe, highly efficient, and virtually inexhaustible energy source for the future. To mimic this process on earth, experimental fusion devices seek to heat gas to millions of degrees (creating...... a fusion plasma) and to confine it within magnetic fields. Learning how such plasmas behave and can be controlled is a crucial step towards realizing fusion as a sustainable energy source.At the Plasma Physics and Fusion Energy (PPFE) section at DTU Physics, we are exploring these issues,focusing on areas...... of high priority on the way towards a working fusion power plant. On the theoreticalfront, we are simulating plasma turbulence and transport of heat and particles in fusion plasmas (Fig. 1a). These issues play a key role in determining how the plasma behaves globally and how well it remains confined...

  4. Membrane processes for removal of pharmaceutically active compounds (PhACs) from water and wastewaters.

    Taheran, Mehrdad; Brar, Satinder K; Verma, M; Surampalli, R Y; Zhang, T C; Valero, J R

    2016-03-15

    Pharmaceutically active compounds (PhACs), which find their way easily into the water sources, are emerging as a major concern for drinking water quality and aquatic species. Therefore, their removal from water sources is a priority from environmental point of view. During the past decade, different methods including membrane separation, adsorption systems and chemical transformation have been evaluated for removal of these compounds. This paper reviews different aspects of PhAC removal by using membrane separation processes, as they have been conventionally known to show high potential in the production of superior quality drinking and industrial water. In brief, osmosis membranes can efficiently remove almost all PhACs though its operational cost is relatively high and nanofiltration (NF) membranes are highly influenced by electrostatic and hydrophobic interaction. Moreover, the efficiency of membrane bioreactors (MBRs) is difficult to predict due to the complex interaction of compounds with microorganisms. To improve the performance and robustness of membrane technology, it is suggested to combine membranes with other systems, such as activated carbon and enzymatic degradation. PMID:26789358

  5. A study on flow characteristics of fountain-pen nano-lithography with active membrane pumping

    In this study, the flow characteristics of a FPN (Fountain Pen Nano-Lithography) using active membrane pumping are investigated. The FPN has integrated chamber, micro channel, and high capacity reservoir for continuous ink feed. The most important aspect in this probe provided control of fluid injection using active membrane pumping in chamber. The flow rates in channel by capillary force are theoretically analyzed, including the control of the mass flow rates by the deflection of the membrane. The above results are compared with the numerical simulations that calculated by commercial code, FLUENT. The velocity of the fluid in micro channel shows linear behaviors. And the mass flows are proportional to the second order function of the pumping pressure that is imposed to the membrane

  6. Controlled release and antibacterial activity of tetracycline hydrochloride-loaded bacterial cellulose composite membranes.

    Shao, Wei; Liu, Hui; Wang, Shuxia; Wu, Jimin; Huang, Min; Min, Huihua; Liu, Xiufeng

    2016-07-10

    Bacterial cellulose (BC) is widely used in biomedical applications. In this study, we prepared an antibiotic drug tetracycline hydrochloride (TCH)-loaded bacterial cellulose (BC) composite membranes, and evaluated the drug release, antibacterial activity and biocompatibility. The structure and morphology of the fabricated BC-TCH composite membranes were characterized using scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). The TCH release results show that the incorporation of BC matrix to load TCH is able to control the release. In vitro antibacterial assay demonstrate that the developed BC-TCH composites displayed excellent antibacterial activity solely associated with the loaded TCH drug. More importantly, the BC-TCH composite membranes display good biocompatibility. These characteristics of BC-TCH composite membranes indicate that they may successfully serve as wound dressings and other medical biomaterials. PMID:27106158

  7. Activation and clearance of vanadium alloys and beryllium multipliers in fusion reactors

    Bartenev, S.A. [V.G. Khlopin Radium Institute, 2nd Murinskij Prospect 28, 194021 St. Petersburg (Russian Federation); Ciampichetti, A. [EURATOM/ENEA Fusion Association, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Turin (Italy); Firsin, N.G. [V.G. Khlopin Radium Institute, 2nd Murinskij Prospect 28, 194021 St. Petersburg (Russian Federation); Forrest, R. [EURATOM/UKAEA Fusion Association, Culham Science Centre, Abingdon OX14 3DB (United Kingdom); Kolbasov, B.N. [Nuclear Fusion Institute, Russian Research Centre ' Kurchatov Institute' , pl. Kurchatova 1, 123182 Moscow (Russian Federation); Romanov, P.V. [Federal Agency for Atomic Energy, Bolshaya Ordynka 24-26, 109017 Moscow (Russian Federation); Romanovskij, V.N. [V.G. Khlopin Radium Institute, 2nd Murinskij Prospect 28, 194021 St. Petersburg (Russian Federation); Zucchetti, M. [EURATOM/ENEA Fusion Association, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Turin (Italy)], E-mail: zucchetti@polito.it

    2007-10-15

    The possibility of clearance of vanadium-chromium-titanium (V-Cr-Ti) alloys is analysed. These alloys after their service in fusion power plants, have the potential to reach clearance if they are purified from activation products. The extraction part of the technological scheme for radiochemical separation of components of irradiated V-Cr-Ti alloy and their purification from metallic activation products, developed earlier, was tested for the first time in laboratory conditions using activated alloy specimens. The replacement of the acid reextraction of V with peroxide and of acid reextraction of Cr with alkaline improved characteristics of the extraction reprocessing. Duration of the V and Cr reextraction was shortened by about an order of magnitude, the output of these alloy components was increased, V purification from rare-earth metals became two times as great, and Cr decontamination from Co increased by two orders of magnitude. Activation of Be contaminated with trace quantities of uranium is an issue: estimation of Be activation in the blanket of the Power Plant Conceptual Study (PPCS) has suggested that traces of U impurity in Be should be removed - or substantially reduced - prior to use.

  8. Luminol electrochemiluminescence for the analysis of active cholesterol at the plasma membrane in single mammalian cells.

    Ma, Guangzhong; Zhou, Junyu; Tian, Chunxiu; Jiang, Dechen; Fang, Danjun; Chen, Hongyuan

    2013-04-16

    A luminol electrochemiluminescence assay was reported to analyze active cholesterol at the plasma membrane in single mammalian cells. The cellular membrane cholesterol was activated by the exposure of the cells to low ionic strength buffer or the inhibition of intracellular acyl-coA/cholesterol acyltransferase (ACAT). The active membrane cholesterol was reacted with cholesterol oxidase in the solution to generate a peak concentration of hydrogen peroxide on the electrode surface, which induced a measurable luminol electrochemiluminescence. Further treatment of the active cells with mevastatin decreased the active membrane cholesterol resulting in a drop in luminance. No change in the intracellular calcium was observed in the presence of luminol and voltage, which indicated that our analysis process might not interrupt the intracellular cholesterol trafficking. Single cell analysis was performed by placing a pinhole below the electrode so that only one cell was exposed to the photomultiplier tube (PMT). Twelve single cells were analyzed individually, and a large deviation on luminance ratio observed exhibited the cell heterogeneity on the active membrane cholesterol. The smaller deviation on ACAT/HMGCoA inhibited cells than ACAT inhibited cells suggested different inhibition efficiency for sandoz 58035 and mevastatin. The new information obtained from single cell analysis might provide a new insight on the study of intracellular cholesterol trafficking. PMID:23527944

  9. Fast serial analysis of active cholesterol at the plasma membrane in single cells.

    Tian, Chunxiu; Zhou, Junyu; Wu, Zeng-Qiang; Fang, Danjun; Jiang, Dechen

    2014-01-01

    Previously, our group has utilized the luminol electrochemiluminescence to analyze the active cholesterol at the plasma membrane in single cells by the exposure of one cell to a photomultiplier tube (PMT) through a pinhole. In this paper, fast analysis of active cholesterol at the plasma membrane in single cells was achieved by a multimicroelectrode array without the pinhole. Single cells were directly located on the microelectrodes using cell-sized microwell traps. A cycle of voltage was applied on the microelectrodes sequentially to induce a peak of luminescence from each microelectrode for the serial measurement of active membrane cholesterol. A minimal time of 1.60 s was determined for the analysis of one cell. The simulation and the experimental data exhibited a semisteady-state distribution of hydrogen peroxide on the microelectrode after the reaction of cholesterol oxidase with the membrane cholesterol, which supported the relative accuracy of the serial analysis. An eight-microelectrode array was demonstrated to analyze eight single cells in 22 s serially, including the channel switching time. The results from 64 single cells either activated by low ion strength buffer or the inhibition of intracellular acyl-coA/cholesterol acyltransferase (ACAT) revealed that most of the cells analyzed had the similar active membrane cholesterol, while few cells had more active cholesterol resulting in the cellular heterogeneity. The fast single-cell analysis platform developed will be potentially useful for the analysis of more molecules in single cells using proper oxidases. PMID:24328095

  10. Plasma membrane recruitment of dephosphorylated β-catenin upon activation of the Wnt pathway

    Hendriksen, Jolita; Jansen, Marnix; Brown, Carolyn; van der Velde, Hella; van Ham, Marco; Galjart, Niels; Offerhaus, Johan; Fagotto, Francois; Fornerod, Maarten

    2008-01-01

    textabstractThe standard model of Wnt signaling specifies that after receipt of a Wnt ligand at the membranous receptor complex, downstream mediators inhibit a cytoplasmic destruction complex, allowing β-catenin to accumulate in the cytosol and nucleus and co-activate Wnt target genes. Unexpectedly, shortly after Wnt treatment, we detected the dephosphorylated form of β-catenin at the plasma membrane, where it displayed a discontinuous punctate labeling. This pool of β-catenin could only be d...

  11. Trans-activity of Plasma Membrane-associated Ganglioside Sialyltransferase in Mammalian Cells*

    Vilcaes, Aldo A.; Demichelis, Vanina Torres; Daniotti, Jose L.

    2011-01-01

    Gangliosides are acidic glycosphingolipids that contain sialic acid residues and are expressed in nearly all vertebrate cells. They are synthesized at the Golgi complex by a combination of glycosyltransferase activities followed by vesicular delivery to the plasma membrane, where they participate in a variety of physiological as well as pathological processes. Recently, a number of enzymes of ganglioside anabolism and catabolism have been shown to be associated with the plasma membrane. In pa...

  12. Transport and deposition of activation products in a helium cooled fusion power plant

    The transport and deposition of neutron activation products in a helium cooled tokamak fusion power plant are investigated. Stainless steel is used as coolant channel material for a helium/steam system. The important gamma emitting nuclides 56Mn, 54Mn, 57Co, 58Co, 60Co, 51Cr, and 99Mo are considered. The dominant release mechanism identified is direct daughter recoil emission from (n,x) type reactions. Corrosion and evaporation are discussed. The radionuclide inventory released by these mechanisms is predicted to exceed 1 x 104 Ci for a reference reactor design after only several days of operation, and approach 3.5 x 104 Ci in equilibrium. A mass transport model is then used to predict the deposition pattern of this inventory in the reactor cooling system

  13. Improved antibacterial activity of nanofiltration polysulfone membranes modified with silver nanoparticles.

    Andrade, Patricia Fernanda; de Faria, Andreia Fonseca; Oliveira, Silvana Ruella; Arruda, Marco Aurélio Zezzi; Gonçalves, Maria do Carmo

    2015-09-15

    Polysulfone membranes (PSf) containing silver nanoparticles were prepared by the wet phase-inversion process. Silver nanoparticles (AgNP) were dispersed into the polymer matrix using two different methodologies. In the first one, the AgNP were synthesized and further dispersed into the polymer solution (ex situ process). In the second method, the formation of the AgNP was performed in situ. The AgNP crystalline structure in the PSf membranes was confirmed by X-ray diffraction. Field emission scanning electron microscopy images showed that the addition of AgNP in PSf membranes caused no significant changes to the finger-like morphology. When the ex situ methodology was applied, 45 nm average size AgNP were uniformly distributed in the internal pores of the membranes. However, when the AgNP were formed through the in situ process, the AgNP were uniformly and preferentially distributed on the top and bottom surfaces of the membrane. In the last case, the AgNP showed cubic morphology when present in the bottom and top surfaces, however, when inside the membrane their morphology was spherical. The cubic-like nanoparticles displayed a 38 nm average edge length. The silver ion released from the membrane during water filtration was measured using inductively coupled plasma mass spectrometry, which showed a silver leaching of approximately 2 μg L(-1). The nanocomposite membranes prepared by the in situ method exhibited a better antibacterial activity, in comparison to those prepared by ex situ, and also a decrease in 90% Escherichia coli adhered cells compared to the pristine PSf membranes. In conclusion, the in situ procedure can be considered a feasible, simple, and reproducible methodology to prepare anti-biofouling polysulfone membranes containing AgNP. PMID:26099831

  14. Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity

    Lim, Seng Koon; Sandén, Camilla; Selegård, Robert; Liedberg, Bo; Aili, Daniel

    2016-02-01

    Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

  15. Biologically Inspired Photocatalytically Active Membranes for Water Treatment

    Kinsinger, Nichola M.

    There is an alarming increase of a variety of new chemicals that are now being discharged into the wastewater system causing increased concern for public health and safety because many are not removed by typical wastewater treatment practices. Titanium Dioxide (TiO2) is a heterogeneous photocatalytic material that rapidly and completely mineralizing organics without harmful byproducts. TiO2 is synthesized by various methods, which lack the necessary control of crystal size, phase, and morphological features that yield optimized semiconductor materials. Mineralizing organisms demonstrate how nature can produce elegant structures at room temperature through controlled organic-mineral interactions. Here, we utilize biologically-inspired scaffolds to template the nucleation and growth of inorganic materials such as TiO2, which aid in controlling the size and phase of these particles and ultimately, their properties. Nanosized rutile and anatase particles were synthesized under solution conditions at relatively low temperatures and mild pH conditions. The effects of reaction conditions on phase and grain size were investigated and discussed from coordination chemistry and coarsening mechanisms. Photocatalytic characterization of TiO2 phase mixtures was performed to investigate their synergistic effect. The suspension conditions of these catalytic nanomaterials were modulated to optimize the degradation rate of organic analytes. Through the addition of an organic scaffold during the synthesis reaction, a mechanically robust (elastic) composite material containing TiO2 nanoparticles was produced. This composite was subsequently heat-treated to produce a porous, high surface area TiO2 nanoparticulate membrane. Processing conditions were investigated to characterize the growth and phase transformation of TiO2, which ultimately impacts photocatalytic performance. These bulk porous TiO2 structures can be fabricated and tailored to act as stand-alone photocatalytic membranes

  16. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants

  17. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    Roehrig, John T., E-mail: jtr1@cdc.gov [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Butrapet, Siritorn; Liss, Nathan M. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Bennett, Susan L. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Luy, Betty E.; Childers, Thomas; Boroughs, Karen L.; Stovall, Janae L.; Calvert, Amanda E. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Blair, Carol D. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Huang, Claire Y.-H. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States)

    2013-07-05

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants.

  18. Biological Activity of Blackcurrant Extracts (Ribes nigrum L. in Relation to Erythrocyte Membranes

    Dorota Bonarska-Kujawa

    2014-01-01

    Full Text Available Compounds contained in fruits and leaves of blackcurrant (Ribes nigrum L. are known as agents acting preventively and therapeutically on the organism. The HPLC analysis showed they are rich in polyphenol anthocyanins in fruits and flavonoids in leaves, that have antioxidant activity and are beneficial for health. The aim of the research was to determine the effect of blackcurrant fruit and leaf extracts on the physical properties of the erythrocyte membranes and assess their antioxidant properties. The effect of the extracts on osmotic resistance, shape of erythrocytes and hemolytic and antioxidant activity of the extracts were examined with spectrophotometric methods. The FTIR investigation showed that extracts modify the erythrocyte membrane and protect it against free radicals induced by UV radiation. The results show that the extracts do not induce hemolysis and even protect erythrocytes against the harmful action of UVC radiation, while slightly strengthening the membrane and inducing echinocytes. The compounds contained in the extracts do not penetrate into the hydrophobic region, but bind to the membrane surface inducing small changes in the packing arrangement of the polar head groups of membrane lipids. The extracts have a high antioxidant activity. Their presence on the surface of the erythrocyte membrane entails protection against free radicals.

  19. Synthetic aperture microwave imaging with active probing for fusion plasma diagnostics

    Shevchenko, Vladimir F.; Freethy, Simon J.; Huang, Billy K.; Vann, Roddy G. L.

    2014-08-01

    A Synthetic Aperture Microwave Imaging (SAMI) system has been designed and built to obtain 2-D images at several frequencies from fusion plasmas. SAMI uses a phased array of linearly polarised antennas. The array configuration has been optimised to achieve maximum synthetic aperture beam efficiency. The signals received by antennas are down-converted to the intermediate frequency range and then recorded in a full vector form. Full vector signals allow beam focusing and image reconstruction in both real time and a post-processing mode. SAMI can scan over 16 pre-programmed frequencies in the range of 10-35GHz with a switching time of 300ns. The system operates in 2 different modes simultaneously: both a 'passive' imaging of plasma emission and also an 'active' imaging of the back-scattered signal of the radiation launched by one of the antennas from the same array. This second mode is similar to so-called Doppler backscattering (DBS) reflectometry with 2-D resolution of the propagation velocity of turbulent structures. Both modes of operation show good performance in fusion plasma experiments on Mega Amp Spherical Tokamak (MAST). We have obtained the first ever 2-D images of BXO mode conversion windows. With active probing, first ever turbulence velocity maps have been obtained. We present an overview of the diagnostic and discuss recent results. In contrast to quasi-optical microwave imaging systems SAMI requires neither big aperture viewing ports nor large 2-D detector arrays to achieve the desired imaging resolution. The number of effective 'pixels' of the synthesized image is proportional to the number of receiving antennas squared. Thus only a small number of optimised antennas is sufficient for the majority of applications. Possible implementation of SAMI on ITERand DEMO is discussed.

  20. Low Activation Vanadium Alloys for Fusion Power Reactors - the RF Results

    Full text: The Results of development and researches of functional properties of low activation vanadium alloys (V-Ti-Cr and V-Cr-W-Zr-C systems) being developed for the cores of nuclear fusion and fission (Gen-IV, space) power reactors are presented. Scientific and technological problems of the investigations are related with enhancement of functional properties based on: 1. Special optimized thermal (TT), thermomechanical (TMT) and thermochemical (TCT) treatments of V-4Ti-4Cr alloys. 2. Development of new (V-Cr-W-Zr-C system) vanadium alloys. The TMT and TCT regimes ensuring the capability of significant (up to 2 times) enhancement of yield strength in the temperature range up to 800°C keeping relatively high plasticity reserve have been found for alloys. The results of the theoretical, modeling and simulating studies of characteristics of self-point defects and dislocations, their interactions and mobility are presented. Nuclear physics characteristics (primary radiation damage, activation, transmutation, postreactor cooling) of alloys irradiated for a long time in neutron spectra of the fusion reactor DEMO-RF (15.3 dpa/year) and fast power reactor BN-600 (80 dpa/year) are calculated. The interaction characteristics of V-4Ti-4Cr alloy with hydrogen and the influence of hydrogen on mechanical properties of the alloy (impact toughness, internal friction) have been studied. Obtained results allows one to recommend the vanadium alloys for applications in nuclear reactors at operating temperature window 300 - 800(850)°C. The planes of high-dose and high- temperature reactor tests of vanadium alloys are scheduled at material science assemblies of reactor BN-600 (2013 - 2015, doses 50 - 200 dpa, irradiation temperatures 400 - 800°C). (author)

  1. Synthetic aperture microwave imaging with active probing for fusion plasma diagnostics

    A Synthetic Aperture Microwave Imaging (SAMI) system has been designed and built to obtain 2-D images at several frequencies from fusion plasmas. SAMI uses a phased array of linearly polarised antennas. The array configuration has been optimised to achieve maximum synthetic aperture beam efficiency. The signals received by antennas are down-converted to the intermediate frequency range and then recorded in a full vector form. Full vector signals allow beam focusing and image reconstruction in both real time and a post-processing mode. SAMI can scan over 16 pre-programmed frequencies in the range of 10-35GHz with a switching time of 300ns. The system operates in 2 different modes simultaneously: both a 'passive' imaging of plasma emission and also an 'active' imaging of the back-scattered signal of the radiation launched by one of the antennas from the same array. This second mode is similar to so-called Doppler backscattering (DBS) reflectometry with 2-D resolution of the propagation velocity of turbulent structures. Both modes of operation show good performance in fusion plasma experiments on Mega Amp Spherical Tokamak (MAST). We have obtained the first ever 2-D images of BXO mode conversion windows. With active probing, first ever turbulence velocity maps have been obtained. We present an overview of the diagnostic and discuss recent results. In contrast to quasi-optical microwave imaging systems SAMI requires neither big aperture viewing ports nor large 2-D detector arrays to achieve the desired imaging resolution. The number of effective 'pixels' of the synthesized image is proportional to the number of receiving antennas squared. Thus only a small number of optimised antennas is sufficient for the majority of applications. Possible implementation of SAMI on ITERand DEMO is discussed

  2. Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger.

    Markosyan, Ruben M; Miao, Chunhui; Zheng, Yi-Min; Melikyan, Gregory B; Liu, Shan-Lu; Cohen, Fredric S

    2016-01-01

    Ebola virus (EBOV) is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein, GP, assayed by the transfer of both cytoplasmic and membrane dyes. A small molecule fusion inhibitor, a neutralizing antibody, as well as mutations in EBOV GP known to reduce viral infection, all greatly reduce fusion. By monitoring redistribution of small aqueous dyes between cells and by electrical capacitance measurements, we discovered that EBOV GP-mediated fusion pores do not readily enlarge-a marked difference from the behavior of other viral fusion proteins. EBOV GP must be cleaved by late endosome-resident cathepsins B or L in order to become fusion-competent. Cleavage of cell surface-expressed GP appears to occur in endosomes, as evidenced by the fusion block imposed by cathepsin inhibitors, agents that raise endosomal pH, or an inhibitor of anterograde trafficking. Treating effector cells with a recombinant soluble cathepsin B or thermolysin, which cleaves GP into an active form, increases the extent of fusion, suggesting that a fraction of surface-expressed GP is not cleaved. Whereas the rate of fusion is increased by a brief exposure to acidic pH, fusion does occur at neutral pH. Importantly, the extent of fusion is independent of external pH in experiments in which cathepsin activity is blocked and EBOV GP is cleaved by thermolysin. These results imply that low pH promotes fusion through the well-known pH-dependent activity of cathepsins; fusion induced by cleaved EBOV GP is a process that is fundamentally independent of pH. The cell-cell fusion system has revealed some previously unappreciated features of EBOV entry, which could not be readily elucidated in the context of endosomal entry. PMID:26730950

  3. Radiation modulation of the activity of some enzymatic systems of isolated plasma membranes in early ontogenesis

    Studied is the activity of adenilate cyclase (ADC-ase), phosphodiesterase tsAMF(FDEh) and 5'-nucleotidase of plasma membranes of liver extracted from rat embryo of the 20th day of development in norm and after the action of ionizing radiation. It is shown that γ-radiation of plasma membranes in doses from 0.1 to 100 kR decreases the ADCase activity with the more pronounced effect in the case of stimulation of isoproterenole under the conditions of large doses. The 5' nucleotidase and FDEh activity has not been changed up to 100 kR

  4. Sphingomyelinase D activity in model membranes: structural effects of in situ generation of ceramide-1-phosphate.

    Roberto P Stock

    Full Text Available The toxicity of Loxosceles spider venom has been attributed to a rare enzyme, sphingomyelinase D, which transforms sphingomyelin to ceramide-1-phosphate. The bases of its inflammatory and dermonecrotic activity, however, remain unclear. In this work the effects of ceramide-1-phosphate on model membranes were studied both by in situ generation of this lipid using a recombinant sphingomyelinase D from the spider Loxosceles laeta and by pre-mixing it with sphingomyelin and cholesterol. The systems of choice were large unilamellar vesicles for bulk studies (enzyme kinetics, fluorescence spectroscopy and dynamic light scattering and giant unilamellar vesicles for fluorescence microscopy examination using a variety of fluorescent probes. The influence of membrane lateral structure on the kinetics of enzyme activity and the consequences of enzyme activity on the structure of target membranes containing sphingomyelin were examined. The findings indicate that: 1 ceramide-1-phosphate (particularly lauroyl ceramide-1-phosphate can be incorporated into sphingomyelin bilayers in a concentration-dependent manner and generates coexistence of liquid disordered/solid ordered domains, 2 the activity of sphingomyelinase D is clearly influenced by the supramolecular organization of its substrate in membranes and, 3 in situ ceramide-1-phosphate generation by enzymatic activity profoundly alters the lateral structure and morphology of the target membranes.

  5. Multiple sources of carbonic anhydrase activity in pea thylakoids: soluble and membrane-bound forms.

    Rudenko, Natalia N; Ignatova, Lyudmila K; Ivanov, Boris N

    2007-01-01

    Carbonic anhydrase (CA) activity of pea thylakoids, thylakoid membranes enriched with photosystem I (PSI-membranes), or photosystem II (PSII-membranes) as well as both supernatant and pellet after precipitation of thylakoids treated with detergent Triton X-100 were studied. CA activity of thylakoids in the presence of varying concentrations of Triton X-100 had two maxima, at Triton/chlorophyll (triton/Chl) ratios of 0.3 and 1.0. CA activities of PSI-membranes and PSII-membranes had only one maximum each, at Triton/Chl ratio 0.3 or 1.0, respectively. Two CAs with characteristics of the membrane-bound proteins and one CA with characteristics of the soluble proteins were found in the medium after thylakoids were incubated with Triton. One of the first two CAs had mobility in PAAG after native electrophoresis the same as that of CA residing in PSI-membranes, and the other CA had mobility the same as the mobility of CA residing in PSII-membranes, but the latter was different from CA situated in PSII core-complex (Ignatova et al. 2006 Biochemistry (Moscow) 71:525-532). The properties of the "soluble" CA removed from thylakoids were different from the properties of the known soluble CAs of plant cell: apparent molecular mass was about 262 kD and it was three orders more sensitive to the specific CA inhibitor, ethoxyzolamide, than soluble stromal CA. The data are discussed as indicating the presence of, at least, four CAs in pea thylakoids. PMID:17347907

  6. [Effect of powdered activated carbon on the sludge mixed liquor characteristics and membrane fouling of MBR].

    Li, Shao-Feng; Gao, Yuan

    2011-02-01

    Effect of dosing powder activated carbon (PAC) on the characteristics of the sludge mixed liquor in membrane bioreactor (MBR) was investigated by parallel tests. And the reason that PAC mitigated membrane fouling was also explored. The results showed that PAC could decrease mixture viscosity and increase sludge particle size, which led to less trans-membrane pressure developing. Extracellular polymer substances (EPS) content, sludge specific resistance and cake layer resistance (R(c)) had a good correlation. Adding PAC could decrease EPS concentration, sludge specific resistance and then slow down the increase of R(c), which mitigated membrane fouling. Membrane pore blocking resistance (R(p)) increased exponentially with increasing of the soluble microbial products (SMP) concentration in the supernatant. Dosing PAC reduced the SMP concentration and slowed down the growth rate of R(p), which was helpful to mitigating membrane fouling. R(c) and R(p) increased along with the operation of MBRs and R(c)/R(f) (26.32% -63.16%) was always greater than R(p)/R(f) (7.89% -35.32%) which suggested the R(c) was the main factor in membrane fouling. Moreover, it was also found that controlling of dosing PAC on R(c) was better than it on R(p). PMID:21528575

  7. Comparing graphene, carbon nanotubes, and superfine powdered activated carbon as adsorptive coating materials for microfiltration membranes.

    Ellerie, Jaclyn R; Apul, Onur G; Karanfil, Tanju; Ladner, David A

    2013-10-15

    Multi-walled carbon nanotubes (MWCNTs), nano-graphene platelets (NGPs), and superfine powdered activated carbon (S-PAC) were comparatively evaluated for their applicability as adsorptive coatings on microfiltration membranes. The objective was to determine which materials were capable of contaminant removal while causing minimal flux reduction. Methylene blue and atrazine were the model contaminants. When applied as membrane coatings, MWCNTs had minimal retention capabilities for the model contaminants, and S-PAC had the fastest removal. The membrane coating approach was also compared with a stirred vessel configuration, in which the adsorbent was added to a stirred flask preceding the membrane cell. Direct application of the adsorbent to the membrane constituted a greater initial reduction in permeate concentrations of the model contaminants than with the stirred flask setup. All adsorbents except S-PAC showed flux reductions less than 5% after application as thin-layer membrane coatings, and flux recovery after membrane backwashing was greater than 90% for all materials and masses tested. PMID:23911830

  8. Avian sarcoma and leukosis virus-receptor interactions: From classical genetics to novel insights into virus-cell membrane fusion

    For over 40 years, avian sarcoma and leukosis virus (ASLV)-receptor interactions have been employed as a useful model system to study the mechanism of retroviral entry into cells. Pioneering studies on this system focused upon the genetic basis of the differential susceptibilities of different lines of chickens to infection by distinct subgroups of ASLV. These studies led to the definition of three distinct autosomal recessive genes that were predicted to encode cellular receptors for different viral subgroups. They also led to the concept of viral interference, i.e. the mechanism by which infection by one virus can render cells resistant to reinfection by other viruses that use the same cellular receptor. Here, we review the contributions that analyses of the ASLV-receptor system have made in unraveling the mechanisms of retroviral entry into cells and focus on key findings such as identification and characterization of the ASLV receptor genes and the subsequent elucidation of an unprecedented mechanism of virus-cell fusion. Since many of the initial findings on this system were published in the early volumes of Virology, this subject is especially well suited to this special anniversary issue of the journal

  9. Line tension at lipid phase boundaries as driving force for HIV fusion peptide-mediated fusion

    Yang, Sung-Tae; Kiessling, Volker; Tamm, Lukas K.

    2016-01-01

    Lipids and proteins are organized in cellular membranes in clusters, often called ‘lipid rafts'. Although raft-constituent ordered lipid domains are thought to be energetically unfavourable for membrane fusion, rafts have long been implicated in many biological fusion processes. For the case of HIV gp41-mediated membrane fusion, this apparent contradiction can be resolved by recognizing that the interfaces between ordered and disordered lipid domains are the predominant sites of fusion. Here ...

  10. Effect of various concentration of sulfuric acid for Nafion membrane activation on the performance of fuel cell

    Pujiastuti, Sri; Onggo, Holia

    2016-02-01

    This work proposes an activation treatment to Nafion 117 membrane with sulfuric acid in various concentrations. The main goal of this study is to increase the Nafion 117 membrane performance, which is determined by proton number in the membrane and membrane performance in Polymer Electrolyte Membrane Fuel Cell (PEMFC). This work was developed using sulfuric acids in four different concentrations: 1, 2, 3, and 4 M. The surface morphology and functional groups of activated membranes were studied using Scanning Electron Microscope and Fourier Transform Infrared, respectively. The proton number absorbed in membranes was observed by gravimetric measurements. The performances of activated membranes in PEMFC were studied by single cell measurements with H2/O2 operation. The experimental results showed that activation of Nafion membrane did not change its surface morphology and functional groups. The proton number increased when the concentration of sulfuric acid is increased from 1 to 3 M and from 1 to 4 M. On the other hand, there is no significant increase when the concentration of sulfuric acid was increased from 1 to 2 M. Similar trends were observed when testing activated membrane performance in PEMFC, especially for current density at 0.6 V and maximum power. It is assumed that there is a correlation between the increase of sulfuric acid concentration in activation process with the increase of proton number in the membrane that are available for facilitating of transfer protons from the anode to the cathode.

  11. Status of Safety and Environmental Activities in the US Fusion Program

    Petti, D A; Reyes, S; Cadwallader, L C; Latkowski, J F

    2004-09-02

    This paper presents an overview of recent safety efforts in both magnetic and inertial fusion energy. Safety has been a part of fusion design and operations since the inception of fusion research. Safety research and safety design support have been provided for a variety of experiments in both the magnetic and inertial fusion programs. The main safety issues are reviewed, some recent safety highlights are discussed and the programmatic impacts that safety research has had are presented. Future directions in the safety and environmental area are proposed.

  12. Status of Safety and Environmental Activities in the US Fusion Program

    David A. Petti; Susana Reyes; Lee C. Cadwallader; Jeffery F. Latkowski

    2004-09-01

    This paper presents an overview of recent safety efforts in both magnetic and inertial fusion energy. Safety has been a part of fusion design and operations since the inception of fusion research. Safety research and safety design support have been provided for a variety of experiments in both the magnetic and inertial fusion programs. The main safety issues are reviewed, some recent safety highlights are discussed and the programmatic impacts that safety research has had are presented. Future directions in the safety and environmental area are proposed.

  13. 2001 activity report of the development and research line in controlled thermonuclear fusion of the Plasma Associated Laboratory

    The year 2001 activities of the controlled thermonuclear fusion research line of the Plasma Associated Laboratory at the National Institute for Space Research - Brazil are reported. The report approaches the staff, participation in congresses, goals for the year 2002 and papers on Tokamak plasmas, plasma diagnostic, bootstraps, plasma equilibrium and diagnostic

  14. 2003 activity report of the development and research line in controlled thermonuclear fusion of the Plasma Associated Laboratory

    This document represents the 2003 activity report of the development and research line in controlled thermonuclear fusion of the Plasma Associated Laboratory - Brazil, approaching the areas of toroidal systems for magnetic confinement, plasma heating, current generation and high temperature plasma diagnostic

  15. Constitutively active IRF7/IRF3 fusion protein completely protects swine against Foot-and-Mouth Disease

    Foot-and-mouth disease (FMD) remains one of the most devastating livestock diseases around the world. Several serotype specific vaccine formulations exist but require about 5-7 days to induce protective immunity. Our previous studies have shown that a constitutively active fusion protein of porcine ...

  16. Protein kinase and phosphatase activities of thylakoid membranes

    Dephosphorylation of the 25 and 27 kDa light-harvesting Chl a/b proteins (LHCII) of the thylakoid membranes is catalyzed by a phosphatase which differs from previously reported thylakoid-bound phosphatases in having an alkaline pH optimum (9.0) and a requirement for Mg2+ ions. Dephosphorylation of the 8.3 kDa psb H gene product requires a Mg2+ ion concentration more than 200 fold higher than that for dephosphorylation of LHC II. The 8.3 kDa and 27 kDa proteins appear to be phosphorylated by two distinct kinases, which differ in substrate specificity and sensitivity to inhibitors. The plastoquinone antagonist 2,5-dibromo-3-methyl-6-isopropyl-benzoquinone (DBMIB) inhibits phosphorylation of the 27 kDa LHC II much more readily than phosphorylation of the 8.3 kDa protein. A similar pattern of inhibition is seen for two synthetic oligopeptides (MRKSATTKKAVC and ATQTLESSSRC) which are analogs of the phosphorylation sites of the two proteins. Possible modes of action of DBMIB are discussed. 45 refs., 7 figs., 3 tabs

  17. Membranolytic Activity of Bile Salts: Influence of Biological Membrane Properties and Composition

    Alfred Blume

    2007-10-01

    Full Text Available The two main steps of the membranolytic activity of detergents: 1 the partitioning of detergent molecules in the membrane and 2 the solubilisation of the membrane are systematically investigated. The interactions of two bile salt molecules, sodium cholate (NaC and sodium deoxycholate (NaDC with biological phospholipid model membranes are considered. The membranolytic activity is analysed as a function of the hydrophobicity of the bile salt, ionic strength, temperature, membrane phase properties, membrane surface charge and composition of the acyl chains of the lipids. The results are derived from calorimetric measurements (ITC, isothermal titration calorimetry. A thermodynamic model is described, taking into consideration electrostatic interactions, which is used for the calculation of the partition coefficient as well as to derive the complete thermodynamic parameters describing the interaction of detergents with biological membranes (change in enthalpy, change in free energy, change in entropy etc. The solubilisation properties are described in a so-called vesicle-to-micelle phase transition diagram. The obtained results are supplemented and confirmed by data obtained from other biophysical techniques (DSC differential scanning calorimetry, DLS dynamic light scattering, SANS small angle neutron scattering.

  18. Guanidination of notexin alters its membrane-damaging activity in response to sphingomyelin and cholesterol

    Pei-Hsiu Kao; Yi-Ling Chiou; Shinne-Ren Lin; Long-Sen Chang

    2010-12-01

    To elucidate the contribution of phospholipase A2 (PLA2) activity of notexin to its ability to perturb membranes, comparative studies on the interaction of notexin and guanidinated notexin (Gu-notexin) with egg yolk phosphatidylcholine (EYPC), EYPC/egg yolk sphingomyelin (EYSM) and EYPC/EYSM/cholesterol vesicles were conducted. EYSM notably reduced the membrane-damaging activity of notexin against EYPC vesicles, but had an insignificant influence on that of Gu-notexin. Unlike the effects noted with notexin, inactivation of PLA2 activity by EDTA led to a reduction in the ability of Gu-notexin to induce EYPC/EYSM vesicle leakage and to increase Gu-notexin-induced membrane permeability of EYPC/EYSM/cholesterol vesicles. The geometrical arrangement of notexin and Gu-notexin in contact with either EYPC/EYSM vesicles or EYPC/EYSM/cholesterol vesicles differed. Moreover, global conformation of notexin and Gu-notexin differed in either Ca2+-bound or metal-free states. These results indicate that notexin and Gu-notexin could induce membrane permeability without the involvement of PLA2 activity, and suggest that guanidination alters the membrane-bound mode of notexin on damaging phospholipid vesicles containing sphingomyelin and cholesterol.

  19. High speed laser activated membrane introduction mass spectrometric evaluation of bulk methylcyclohexane dehydrogenation catalysts

    Laser activated membrane introduction mass spectrometry (LAMIMS) is a modification of membrane introduction mass spectrometry where a silicone membrane serves as a separator between an analyte stream and the vacuum of a quadrupole mass spectrometer. Carbon paper, commonly used as a fuel cell gas diffusion layer, has been overlaid upon the silicone membrane to serve as a support for catalyst array libraries that are heated by a CO2 laser during evaluation. The LAMIMS reactor is a closed environment permitting steady state exposure of the catalyst library to the reactant stream. In this work, Pt/Al2O3 catalysts for the dehydrogenation of methylcyclohexane (MCH) to toluene have been ranked by LAMIMS. Spot-to-spot evaluation times (after preconditioning) are 1 min in this study. The data suggests that by optimization of signal-to-noise and spot-to-spot dwell times, ranking can be conducted at substantially under a minute per array spot candidate

  20. Heteronanostructure of Ag particle on titanate nanowire membrane with enhanced photocatalytic properties and bactericidal activities

    A novel seed induced method has been developed for syntheses of Ag particles on titanate nanowires, and then the heteronanostructured Ag/titanate nanowires were assembled into porous, flexible membranes. These titanate nanowires were about several hundreds micrometers in length and about 80 nm in diameter. The size of the Ag particle can be tuned within 300-700 nm. The pore size and thickness of the heteronanostructured membrane were easily controlled. An Ag/titanate nanowire membrane reactor has been developed to study the photocatalytic degradation of methamidophos in aqueous solution, and 87.0% of the methamidophos can be degraded in a concurrent filtration and photocatalytic oxidation process. The antibacterial activity was also investigated on the heteronanostructured membrane with UVA light (365 nm) irradiation, and a 99.99% satisfactory antibacterial effect on Escherichia coli was achieved.

  1. Receptor kinase-mediated control of primary active proton pumping at the plasma membrane

    Fuglsang, AT; Kristensen, A; Cuin, TA;

    2014-01-01

    and in planta with PSY1R, a receptor kinase of the plasma membrane that serves as a receptor for the peptide growth hormone PSY1. The intracellular protein kinase domain of PSY1R phosphorylates AHA2/AHA1 at Thr-881, situated in the autoinhibitory Region I of the C-terminal domain. When expressed in a yeast......Acidification of the cell wall space outside the plasma membrane is required for plant growth and is the result of proton extrusion by the plasma membrane-localized H+ -ATPases. Here we show that the major plasma membrane proton pumps in Arabidopsis, AHA1 and AHA2, interact directly in vitro...... heterologous expression system, the introduction of a negative charge at this position caused pump activation. Application of PSY1 to plant seedlings induced rapid in planta phosphorylation at Thr-881, concomitant with an instantaneous increase in proton efflux from roots. The direct interaction between AHA2...

  2. Importance of Membrane Structural Integrity for RPE65 Retinoid Isomerization Activity

    Golczak, Marcin; Kiser, Philip D.; Lodowski, David T.; Maeda, Akiko; Palczewski, Krzysztof (Case Western)

    2010-04-05

    Regeneration of visual chromophore in the vertebrate visual cycle involves the retinal pigment epithelium-specific protein RPE65, the key enzyme catalyzing the cleavage and isomerization of all-trans-retinyl fatty acid esters to 11-cis-retinol. Although RPE65 has no predicted membrane spanning domains, this protein predominantly associates with microsomal fractions isolated from bovine retinal pigment epithelium (RPE). We have re-examined the nature of RPE65 interactions with native microsomal membranes by using extraction and phase separation experiments. We observe that hydrophobic interactions are the dominant forces that promote RPE65 association with these membranes. These results are consistent with the crystallographic model of RPE65, which features a large lipophilic surface that surrounds the entrance to the catalytic site of this enzyme and likely interacts with the hydrophobic core of the endoplasmic reticulum membrane. Moreover, we report a critical role for phospholipid membranes in preserving the retinoid isomerization activity and physical properties of RPE65. Isomerase activity measured in bovine RPE was highly sensitive to phospholipase A{sup 2} treatment, but the observed decline in 11-cis-retinol production did not directly reflect inhibition by products of lipid hydrolysis. Instead, a direct correlation between the kinetics of phospholipid hydrolysis and retinoid isomerization suggests that the lipid membrane structure is critical for RPE65 enzymatic activity. We also provide evidence that RPE65 operates in a multiprotein complex with retinol dehydrogenase 5 and retinal G protein-coupled receptor in RPE microsomes. Modifications in the phospholipid environment affecting interactions with these protein components may be responsible for the alterations in retinoid metabolism observed in phospholipid-depleted RPE microsomes. Thus, our results indicate that the enzymatic activity of native RPE65 strongly depends on its membrane binding and

  3. Activation calculation and waste management for a fusion experimental breeder, FEB-E

    The engineering outline design of the fusion experimental breeder, FEB-E is reported briefly. Using an activation calculation code FDKR and its associated data library AF-DCDLIB to calculate the radioactivity, decay heat, waste disposal rating and biological hazard potential from activation products, actinides and fission products in the FEB-E. The codes and libraries used in calculation are introduced briefly, and calculation results and decay curves of related hazards after the shutdown of one-year operation of the FEB-E are given. The activation features of five candidate structural materials were also evaluated for FEB-E design. Detailed calculation and analyses of waste disposal rating and remote maintenance rating for all long-lived radionuclides were performed to identify the safety, environmental and radioactive waste characteristics of the FEB-E design. Results obtained show that the total radioactivity inventory, decay heat and BHP at shutdown are 5.74x1013MBq, 8.34 MW and 4.08x108km3 of air for 316SS structure material, respectively. The inventory of actinides important for the fuel reprocessing, such as 232U and 237Np was also investigated. It was shown that their low concentrations in FEB-E appear to be manageable

  4. Thinking in Terms of Structure-Activity-Relationships (T-SAR): A Tool to Better Understand Nanofiltration Membranes

    Stefan Stolte; Jorg Thöming; Fernández, José F.; Bernd Jastorff; Reinhold Störmann

    2011-01-01

    A frontier to be conquered in the field of membrane technology is related to the very limited scientific base for the rational and task-specific design of membranes. This is especially true for nanofiltration membranes with properties that are based on several solute-membrane interaction mechanisms. “Thinking in terms of Structure-Activity-Relationships” (T-SAR) is a methodology which applies a systematic analysis of a chemical entity based on its structural formula. However, the analysis bec...

  5. Performance of Submerged Membrane Bioreactor Combined with Powdered Activated Carbon Addition for the Treatment of an Industrial Wastewater

    Tri Widjaja; Ali Altway; Soeprijanto Soeprijanto

    2010-01-01

    Membrane technology is one of the alternative solutions to overcome industrial wastewater treatment developed nowadays. The addition of PAC (Powdered Activated Carbon) in the activated sludge using Submerged Membrane Adsorption Hybrid Bioreactor (SMAHBR) is expected to increase the organic material removal. The purpose of this study was to determine the performance of submerged membrane bioreactor and activated carbon adsorption capacity of organic materials in wastewater. This study used SIE...

  6. Energy consumption of aeration systems in membrane activation systems; Energiebedarf von Belueftungseinrichtungen in Membranbelebungsanlagen

    Krause, S.; Cornel, P. [Inst. WAR, TU Darmstadt, Darmstadt (Germany)

    2003-07-01

    Owing to the necessity of aeration aggregates, membrane activation systems have higher energy consumption than conventional activation systems. The contribution investigates the energy demand of aeration systems for oxygen supply to the biomass and for control of precipitates on submerged membranes. (orig.) [German] Membranbelebungsanlagen (MBR) zeichnen sich durch einen feststofffreien Klaeranlagenablauf aus. Zudem koennen hoehere Feststoffgehalte im Belebungsbecken erzielt werden. Jedoch ist aufgrund der zusaetzlichen Belueftungsaggregate zur Deckschichtminimierung an den Membranen und reduzierter {alpha}-Werte der Energiebedarf hoeher als in konventionellen Belebungsanlagen. Nachstehend wird der Energiebedarf der Belueftungseinrichtungen zur Sauerstoffversorgung der Biomasse und zur Deckschichtkontrolle getauchter Membranen untersucht. (orig.)

  7. Membrane attachment is key to protecting transducin GTPase activating complex from intracellular proteolysis in photoreceptors

    Gospe, Sidney M.; Sheila A Baker; Kessler, Christopher; Brucato, Martha F.; Winter, Joan R.; Burns, Marie E; Arshavsky, Vadim Y.

    2011-01-01

    The members of the R7 RGS protein sub-family are versatile regulators of G protein signaling throughout the nervous system. Recent studies indicate that they are often found in complexes with membrane anchor proteins which serve as versatile modulators of their activity, intracellular targeting and stability. One striking example is the interplay between the membrane anchor R9AP and the RGS9-1·Gβ5 GTPase activating complex responsible for the rapid inactivation of the G protein transducin in ...

  8. The multidrug transporter, P-glycoprotein, actively mediates cholesterol redistribution in the cell membrane

    Garrigues, Alexia; Escargueil, Alexandre E.; Orlowski, Stéphane

    2002-01-01

    P-glycoprotein (P-gp) is a plasma membrane ATP-binding cassette transporter, responsible for multidrug resistance in tumor cells. P-gp catalyzes the ATP hydrolysis-dependent efflux of numerous amphiphilic compounds of unrelated chemical structures. In the absence of any identified substrate, P-gp exhibits an apparently futile, basal ATPase activity. By using native membrane vesicles containing high amounts of P-gp, we show here that (i) this basal ATPase activity is tightly dependent on the p...

  9. Extracellular ATP-dependent activation of plasma membrane Ca2+ pump in HEK-293 cells

    Qi, Z.; Murase, K.; Obata, S.; Sokabe, M

    2000-01-01

    It is well known that extracellular ATP (ATPo) elevates the intracellular Ca2+ concentration ([Ca2+]i) by inducing Ca2+ influx or mobilizing Ca2+ from internal stores via activation of purinoceptors in the plasma membrane. This study shows that ATPo also activates the plasma membrane Ca2+ pumps (PMCPs) to bring the elevated [Ca2+]i back to the resting level in human embryonic kidney-293 (HEK-293) cells.The duration of ATPo-induced intracellular Ca2+ transients was significantly increased by P...

  10. Salt stress in a membrane bioreactor: Dynamics of sludge properties, membrane fouling and remediation through powdered activated carbon dosing

    Temmerman, De L.; Maere, T.; Temmink, H.; Zwijnenburg, A.; Nopens, I.

    2014-01-01

    Membrane bioreactors are a well-established technology for wastewater treatment. However, their efficiency is adversely impacted by membrane fouling, primarily inciting very conservative operations of installations that makes them less appealing from an economic perspective. This fouling propensity

  11. Fusion of protegrin-1 and plectasin to MAP30 shows significant inhibition activity against dengue virus replication.

    Hussin A Rothan

    Full Text Available Dengue virus (DENV broadly disseminates in tropical and sub-tropical countries and there are no vaccine or anti-dengue drugs available. DENV outbreaks cause serious economic burden due to infection complications that requires special medical care and hospitalization. This study presents a new strategy for inexpensive production of anti-DENV peptide-fusion protein to prevent and/or treat DENV infection. Antiviral cationic peptides protegrin-1 (PG1 and plectasin (PLSN were fused with MAP30 protein to produce recombinant antiviral peptide-fusion protein (PG1-MAP30-PLSN as inclusion bodies in E. coli. High yield production of PG1-MAP30-PLSN protein was achieved by solubilization of inclusion bodies in alkaline buffer followed by the application of appropriate refolding techniques. Antiviral PG1-MAP30-PLSN protein considerably inhibited DENV protease (NS2B-NS3pro with half-maximal inhibitory concentration (IC50 0.5±0.1 μM. The real-time proliferation assay (RTCA and the end-point proliferation assay (MTT assay showed that the maximal-nontoxic dose of the peptide-fusion protein against Vero cells is approximately 0.67±0.2 μM. The cell-based assays showed considerable inhibition of the peptide-fusion protein against binding and proliferating stages of DENV2 into the target cells. The peptide-fusion protein protected DENV2-challeged mice with 100% of survival at the dose of 50 mg/kg. In conclusion, producing recombinant antiviral peptide-fusion protein by combining short antiviral peptide with a central protein owning similar activity could be useful to minimize the overall cost of short peptide production and take advantage of its synergistic antiviral activities.

  12. Atomic Force Microscope Spectroscopy Reveals a Hemifusion Intermediate during Soluble N-Ethylmaleimide-Sensitive Factor-Attachment Protein Receptors-Mediated Membrane Fusion

    Abdulreda, Midhat H.; Bhalla, Akhil; Chapman, Edwin R.; Moy, Vincent T.

    2007-01-01

    This study investigated the effect of soluble N-ethylmaleimide-sensitive factor-attachment protein (SNAP) receptors (SNAREs) on the fusion of egg L-α-phosphatidylcholine bilayers using atomic force microscope (AFM) spectroscopy. AFM measurements of the fusion force under compression were acquired to reveal the energy landscape of the fusion process. A single main energy barrier governing the fusion process was identified in the absence and presence of SNAREs in the bilayers. Under compression...

  13. Activation and routing of membrane-tethered prohormone convertases 1 and 2.

    Bruzzaniti, A; Marx, R; Mains, R E

    1999-08-27

    Many peptide hormones and neuropeptides are processed by members of the subtilisin-like family of prohormone convertases (PCs), which are either soluble or integral membrane proteins. PC1 and PC2 are soluble PCs that are primarily localized to large dense core vesicles in neurons and endocrine cells. We examined whether PC1 and PC2 were active when expressed as membrane-tethered proteins, and how tethering to membranes alters the biosynthesis, enzymatic activity, and intracellular routing of these PCs. PC1 and PC2 chimeras were constructed using the transmembrane domain and cytoplasmic domain of the amidating enzyme, peptidylglycine alpha-amidating monooxygenase (PAM). The membrane-tethered PCs were rerouted from large dense core vesicles to the Golgi region. In addition, the chimeras were transiently expressed at the cell surface and rapidly internalized to the Golgi region in a fashion similar to PAM. Membrane-tethered PC1 and PC2 exhibited changes in pro-domain maturation rates, N-glycosylation, and in the pH and calcium optima required for maximal enzymatic activity against a fluorogenic substrate. In addition, the PC chimeras efficiently cleaved endogenous pro-opiomelanocortin to the correct bioactive peptides. The PAM transmembrane domain/cytoplasmic domain also prevented stimulated secretion of pro-opiomelanocortin products in AtT-20 cells. PMID:10455138

  14. Membrane-Active Macromolecules Resensitize NDM-1 Gram-Negative Clinical Isolates to Tetracycline Antibiotics

    Uppu, Divakara S. S. M.; Manjunath, Goutham B.; Yarlagadda, Venkateswarlu; Kaviyil, Jyothi E.; Ravikumar, Raju; Paramanandham, Krishnamoorthy; Shome, Bibek R.; Haldar, Jayanta

    2015-01-01

    Gram-negative ‘superbugs’ such as New Delhi metallo-beta-lactamase-1 (blaNDM-1) producing pathogens have become world’s major public health threats. Development of molecular strategies that can rehabilitate the ‘old antibiotics’ and halt the antibiotic resistance is a promising approach to target them. We report membrane-active macromolecules (MAMs) that restore the antibacterial efficacy (enhancement by >80-1250 fold) of tetracycline antibiotics towards blaNDM-1 Klebsiella pneumonia and blaNDM-1 Escherichia coli clinical isolates. Organismic studies showed that bacteria had an increased and faster uptake of tetracycline in the presence of MAMs which is attributed to the mechanism of re-sensitization. Moreover, bacteria did not develop resistance to MAMs and MAMs stalled the development of bacterial resistance to tetracycline. MAMs displayed membrane-active properties such as dissipation of membrane potential and membrane-permeabilization that enabled higher uptake of tetracycline in bacteria. In-vivo toxicity studies displayed good safety profiles and preliminary in-vivo antibacterial efficacy studies showed that mice treated with MAMs in combination with antibiotics had significantly decreased bacterial burden compared to the untreated mice. This report of re-instating the efficacy of the antibiotics towards blaNDM-1 pathogens using membrane-active molecules advocates their potential for synergistic co-delivery of antibiotics to combat Gram-negative superbugs. PMID:25789871

  15. E. coli a-hemolysin: a membrane-active protein toxin

    Goñi F.M.

    1998-01-01

    Full Text Available Alpha-Hemolysin is synthesized as a 1024-amino acid polypeptide, then intracellularly activated by specific fatty acylation. A second activation step takes place in the extracellular medium through binding of Ca2+ ions. Even in the absence of fatty acids and Ca2+ HlyA is an amphipathic protein, with a tendency to self-aggregation. However, Ca2+-binding appears to expose hydrophobic patches on the protein surface, facilitating both self-aggregation and irreversible insertion into membranes. The protein may somehow bind membranes in the absence of divalent cations, but only when Ca2+ (or Sr2+, or Ba2+ is bound to the toxin in aqueous suspensions, i.e., prior to its interaction with bilayers, can a-hemolysin bind irreversibly model or cell membranes in such a way that the integrity of the membrane barrier is lost, and cell or vesicle leakage ensues. Leakage is not due to the formation of proteinaceous pores, but rather to the transient disruption of the bilayer, due to the protein insertion into the outer membrane monolayer, and subsequent perturbations in the bilayer lateral tension. Protein or glycoprotein receptors for a-hemolysin may exist on the cell surface, but the toxin is also active on pure lipid bilayers.

  16. Identification and Characterization of a Proteolytically Primed Form of the Murine Coronavirus Spike Proteins after Fusion with the Target Cell

    Wicht, Oliver; Burkard, Christine; Cornelis A M de Haan; van Kuppeveld, Frank J. M.; Rottier, Peter J. M.; Bosch, Berend Jan

    2014-01-01

    Enveloped viruses carry highly specialized glycoproteins that catalyze membrane fusion under strict spatial and temporal control. To prevent premature activation after biosynthesis, viral class I fusion proteins adopt a locked conformation and require proteolytic cleavage to render them fusion-ready. This priming step may occur during virus exit from the infected cell, in the extracellular milieu or during entry at or in the next target cell. Proteolytic processing of coronavirus spike (S) fu...

  17. Activation of lysosomal function in the course of autophagy via mTORC1 suppression and autophagosome-lysosome fusion

    Jing Zhou; Shi-Hao Tan; Valérie Nicolas; Chantal Bauvy; Nai-Di Yang; Jianbin Zhang; Yuan Xue

    2013-01-01

    Lysosome is a key subcellular organelle in the execution of the autophagic process and at present little is known whether lysosomal function is controlled in the process of autophagy.In this study,we first found that suppression of mammalian target of rapamycin (mTOR) activity by starvation or two mTOR catalytic inhibitors (PP242 and Torinl),but not by an allosteric inhibitor (rapamycin),leads to activation of lysosomal function.Second,we provided evidence that activation of lysosomal function is associated with the suppression of mTOR complex 1 (mTORC1),but not mTORC2,and the mTORC1 localization to lysosomes is not directly correlated to its regulatory role in lysosomal function.Third,we examined the involvement of transcription factor EB (TFEB) and demonstrated that TFEB activation following mTORC1 suppression is necessary but not sufficient for lysosomal activation.Finally,Atg5 or Atg7deletion or blockage of the autophagosome-lysosome fusion process effectively diminished lysosomal activation,suggesting that lysosomal activation occurring in the course of autophagy is dependent on antophagosome-lysosome fusion.Taken together,this study demonstrates that in the course of autophagy,lysosomal function is upregulated via a dual mechanism involving mTORC1 suppression and autophagosome-lysosome fusion.

  18. Electrolyte Composition of Mink (Mustela vison Erythrocytes and Active Cation Transporters of the Cell Membrane

    Clausen TN

    2001-06-01

    Full Text Available Red blood cells from mink (Mustela vison were characterized with respect to their electrolyte content and their cell membranes with respect to enzymatic activity for cation transport. The intra- and extracellular concentrations of Na+, K+, Cl-, Ca2+ and Mg2+ were determined in erythrocytes and plasma, respectively. Plasma and red cell water content was determined, and molal electrolyte concentrations were calculated. Red cells from male adult mink appeared to be of the low-K+, high-Na+ type as seen in other carnivorous species. The intracellular K+ concentration is slightly higher than the extracellular one and the plasma-to-cell chemical gradient for Na+ is weak, though even the molal concentrations may differ significantly. Consistent with the high intracellular Na+ and low K+ concentrations, a very low or no ouabain-sensitive Na+,K+-ATPase activity and no K+-activated pNPPase activity were found in the plasma membrane fraction from red cells. The Cl- and Mg2+ concentrations expressed per liter cell water were significantly higher in red cells than in plasma whereas the opposite was the case with Ca2+. The distribution of Cl- thus does not seem compatible with an inside-negative membrane potential in mink erythrocytes. In spite of a steep calcium gradient across the red cell membrane, neither a calmodulin-activated Ca2+-ATPase activity nor an ATP-activated Ca2+-pNPPase activity were detectable in the plasma membrane fraction. The origin of a supposed primary Ca2+ gradient for sustaining of osmotic balance thus seems uncertain.

  19. Outgassing characteristics of F82H ferritic steel as a low activation material for fusion reactor

    Odaka, Kenji; Satou, Osamu [Hitachi Ltd., Tsuchiura, Ibaraki (Japan). Mechanical Engineering Research Lab.; Ootsuka, Michio; Abe, Tetsuya; Hara, Shigemitsu; Takatsu, Hideyuki; Enoeda, Mikio

    1997-09-01

    Outgassing characteristics of F82H ferritic steel as a low activation material for the blanket of fusion device were investigated. A test chamber was constructed by welding F82H ferritic steel plates. The inner surface of the chamber was buffed and electropolished. The test chamber was degassed by the prebaking at temperature of 350degC for 20 h in vacuum. Then outgassing rates of the test chamber were measured by the throughput method as a function of pumping time for the cases that the test chamber was baked and not baked. The typical outgassing rate after baking at 250degC for 24 h was 3 x 10{sup -9} Pa{center_dot}ms{sup -1} and it seems that this value is sufficiently small to produce pressures at least as low as 10{sup -9} Pa in the vacuum chamber made of F82H ferritic steel. In the pump-down of the test chamber without baking after exposure to air, the outgassing rate decreases with pumping time and reached 1 x 10{sup -7} Pa{center_dot}ms{sup -1} at t = 10{sup 5} s. The activation energy of hydrogen in bulk diffusion in the F82H ferritic steel was measured and found to be 7 kcal/mol. (author)

  20. Contribution of activation products to fusion accident risk: Part II. Effects of alternative materials and designs

    Comparison of accident-hazard potentials associated with neutron-activation products in fusion reactors of various designs and structural materials suffers from a number of shortcomings in the readily available hazard-index data. Neither inventories of curies nor biological hazard potentials (BHPs) are satisfactory indices of hazard even if consistently computed, and between-study inconsistencies in neutronics packages and BHP calculations further obscure the meaning of comparisons based on these measures. The authors present here the results of internally consistent calculations of radioactive inventories, BHPs, and off-site dose potentials associated with the first walls of nine reactor-design/first-wall-material combinations. A recent mirror-reactor design reduces off-site dose potentials by a factor of 2 compared to a muchstudied early tokamak, for a given first-wall material. Holding design fixed, HT-9 ferritic steel offers a factor of 2 reduction in dose potential compared to Type 316 stainless steel. By the dose-potential measure, molybdenum is the worst of the materials investigated and silicon carbide is by far the best. Hazards in realizable accidents depend not only on the hypothetical dose potentials, as calculated here, but also on the actual release fractions of first-wall (or other activated) material. Review of the theoretical and experimental evidence bearing on release fractions suggests that, for most candidate materials, high release fractions from designs containing liquid lithium cannot yet be convincingly ruled out

  1. Contribution of activation products to fusion accident risk: part II. Effects of alternative materials and designs

    Comparison of accident-hazard potentials associated with neutron-activation products in fusion reactors of various designs and structural materials suffers from a number of shortcomings in the readily available hazard-index data. Neither inventories of curies nor biological hazard potentials (BHPs) are satisfactory indices of hazard even if consistently computed, and between-study inconsistencies in neutronics packages and BHP calculations further obscure the meaning of comparisons based on these measures. We present here the results of internally consistent calculations of radioactive inventories, BHPs, and off-site dose potentials associated with the first walls of nine reactor-design/first-wall-material combinations. A recent mirror-reactor design reduces off-site dose potentials by a factor of 2 compared to a muchstudied early tokamak, for a given first-wall material. Holding design fixed, HT-9 ferritic steel offers a factor of 2 reduction in dose potential compared to Type 316 stainless steel. By the dose-potential measure, molybdenum is the worst of the materials investigated and silicon carbide is by far the best. Hazards in realizable accidents depend not only on the hypothetical dose potentials, as calculated here, but also on the actual release fractions of first-wall (or other activated) material. Review of the theoretical and experimental evidence bearing on release fractions suggests that, for most candidate materials, high release fractions from designs containing liquid lithium cannot yet be convincingly ruled out

  2. Natamycin Inhibits Vacuole Fusion at the Priming Phase via a Specific Interaction with Ergosterol▿

    te Welscher, Yvonne Maria; Jones, Lynden; van Leeuwen, Martin Richard; Dijksterhuis, Jan; de Kruijff, Ben; Eitzen, Gary; Breukink, Eefjan

    2010-01-01

    The antifungal antibiotic natamycin belongs to the family of polyene antibiotics. Its antifungal activity arises via a specific interaction with ergosterol in the plasma membrane (te Welscher et al., J. Biol. Chem. 283:6393-6401, 2008). However, this activity does not involve disruption of the membrane barrier function, a well-known property of other members of the polyene antibiotic family, such as filipin and nystatin. Here we tested the effect of natamycin on vacuole membrane fusion, which is known to be ergosterol dependent. Natamycin blocked the fusion of isolated vacuoles without compromising the barrier function of the vacuolar membrane. Sublethal doses of natamycin perturbed the cellular vacuole morphology, causing the formation of many more small vacuolar structures in yeast cells. Using vacuoles isolated from yeast strains deficient in the ergosterol biosynthesis pathway, we showed that the inhibitory activity of natamycin was dependent on the presence of specific chemical features in the structure of ergosterol that allow the binding of natamycin. We found that natamycin inhibited the priming stage of vacuole fusion. Similar results were obtained with nystatin. These results suggest a novel mode of action of natamycin and perhaps all polyene antibiotics, which involves the impairment of membrane fusion via perturbation of ergosterol-dependent priming reactions that precede membrane fusion, and they may point to an effect of natamycin on ergosterol-dependent protein function in general. PMID:20385867

  3. Natamycin inhibits vacuole fusion at the priming phase via a specific interaction with ergosterol.

    te Welscher, Yvonne Maria; Jones, Lynden; van Leeuwen, Martin Richard; Dijksterhuis, Jan; de Kruijff, Ben; Eitzen, Gary; Breukink, Eefjan

    2010-06-01

    The antifungal antibiotic natamycin belongs to the family of polyene antibiotics. Its antifungal activity arises via a specific interaction with ergosterol in the plasma membrane (te Welscher et al., J. Biol. Chem. 283:6393-6401, 2008). However, this activity does not involve disruption of the membrane barrier function, a well-known property of other members of the polyene antibiotic family, such as filipin and nystatin. Here we tested the effect of natamycin on vacuole membrane fusion, which is known to be ergosterol dependent. Natamycin blocked the fusion of isolated vacuoles without compromising the barrier function of the vacuolar membrane. Sublethal doses of natamycin perturbed the cellular vacuole morphology, causing the formation of many more small vacuolar structures in yeast cells. Using vacuoles isolated from yeast strains deficient in the ergosterol biosynthesis pathway, we showed that the inhibitory activity of natamycin was dependent on the presence of specific chemical features in the structure of ergosterol that allow the binding of natamycin. We found that natamycin inhibited the priming stage of vacuole fusion. Similar results were obtained with nystatin. These results suggest a novel mode of action of natamycin and perhaps all polyene antibiotics, which involves the impairment of membrane fusion via perturbation of ergosterol-dependent priming reactions that precede membrane fusion, and they may point to an effect of natamycin on ergosterol-dependent protein function in general. PMID:20385867

  4. Membrane Permeable Esterase–Activated Fluorescent Imaging Probe

    Kim, Youngmi; Choi, Yongdoo; Weissleder, Ralph; Tung, Ching-Hsuan

    2007-01-01

    An esterase-triggered probe 2 derived from a cyanine-based pH sensitive dye was developed for cell labeling. Permeation of probe 2 into cells and subsequent hydrolytic activation by cellular esterases result in a bright fluorescent intracellular signal.

  5. Ionophore-Based Voltammetric Ion Activity Sensing with Thin Layer Membranes.

    Cuartero, Maria; Crespo, Gaston A; Bakker, Eric

    2016-02-01

    As shown in recent work, thin layer ion-selective multi-ionophore membranes can be interrogated by cyclic voltammetry to detect the ion activity of multiple species simultaneously and selectively. Additional fundamental evidence is put forward on ion discrimination with thin multi-ionophore-based membranes with thicknesses of 200 ± 25 nm and backside contacted with poly-3-octylthiophene (POT). An anodic potential scan partially oxidizes the POT film (to POT(+)), thereby initiating the release of hydrophilic cations from the membrane phase to the sample solution at a characteristic potential. Varying concentration of added cation-exchanger demonstrates that it limits the ion transfer charge and not the deposited POT film. Voltammograms with multiple peaks are observed with each associated with the transfer of one type of ion (lithium, potassium, and sodium). Experimental conditions (thickness and composition of the membrane and concentration of the sample) are chosen that allow one to describe the system by a thermodynamic rather than kinetic model. As a consequence, apparent stability constants for sodium, potassium, and lithium (assuming 1:1 stoichiometry) with their respective ionophores are calculated and agree well with the values obtained by the potentiometric sandwich membrane technique. As an analytical application, a membrane containing three ionophores was used to determine lithium, sodium, and potassium in artificial samples at the same location and within a single voltammetric scan. Lithium and potassium were also determined in undiluted human plasma in the therapeutic concentration range. PMID:26712342

  6. Use of liposomes to evaluate the role of membrane interactions on antioxidant activity

    In this study the possibility of using liposomes as membrane mimetic systems was evaluated to estimate the antioxidant properties of oxicams and establish a relationship between the interactions of the drugs with the membrane and their consequent antioxidant activity. Different experiments were performed covering the study of the protective effect of oxicams in lipid peroxidation induced by the peroxyl radical (ROO·) derived from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) and using two fluorescence probes with distinct lipophilic properties. Lipid peroxidation using the hydrophilic probe fluorescein was evaluated in lipid and aqueous media. Lipid systems labelled with the fluorescent probe diphenylhexatriene propionic acid (DPH-PA) were used to assess the effects of the drugs on membrane peroxidation simultaneously by fluorescence intensity decay and changes in membrane fluidity by steady-state anisotropy measurements. The use of different probes and liposomes as membrane mimetic systems allowed to conclude that membrane lipoperoxidation is related not only to the scavenging characteristics of the antioxidants but also to their ability to interact with the lipid bilayers

  7. Pervaporation Separation and Catalysis Activity of Novel Zirconium Silicalite-1 Zeolite Membrane

    CHEN Pei; CHEN Xinbing; CHEN Xiangshu; AN Zhongwei; KITA Hidetoshi

    2009-01-01

    Novel zirconium silicalite-1 zeolite membrane was hydrothermally prepared on the mullite porous support at 150-185 ℃ for 40-72 h by an "in situ" method using tetraethyl orthosilicate(TEOS),zirconium butoxide (ZBOT)and tetrapropylammonium hydroxide(TPAOH)as silica source,zirconium source and organic structure directing agent,respectively.X-ray diffraction(XRD)patterns,fourier transformed infrared(FT-IR)spectra,and inductively coupled plasma-atomic emission spectrometry(ICP)of the accompanying zeolite powder confirmed that the zirconium was isomorphously incorporated into the zeolite framework.The surface chemical compositions of the obtained membrane were measured with an energy-dispersive X-ray spectral analyzer(EDS),and the membrane morphologies were observed by a scanning electron microscope(SEM).The results showed that the zeolite crystals growing on the support were zirconium silicalite-1 zeolites,and the dense membrane layer was composed of the well inter-growing zeolite crystals.The zirconium silicalite-1 zeolite membrane,which was derived from the synthesis solution having a molar ratio of 1.00SiO2:0.01ZrO2:0.17TPAOH:12OH2O,showed high ethanol w/w)system under a pervaporation condition at 60℃.Moreover,this membrane displayed pervaporation-aided catalysis activity for iso-propanol oxidation with hydrogen peroxide as oxidant,and the corresponding iso-propanol conversion was 35%.

  8. Sphingomyelinase D activity in model membranes: structural effects of in situ generation of ceramide-1-phosphate

    Stock, Roberto; Brewer, Jonathan R.; Wagner, Kerstin;

    2012-01-01

    The toxicity of Loxosceles spider venom has been attributed to a rare enzyme, sphingomyelinase D, which transforms sphingomyelin to ceramide-1-phosphate. The bases of its inflammatory and dermonecrotic activity, however, remain unclear. In this work the effects of ceramide-1-phosphate on model...... membranes were studied both by in situ generation of this lipid using a recombinant sphingomyelinase D from the spider Loxosceles laeta and by pre-mixing it with sphingomyelin and cholesterol. The systems of choice were large unilamellar vesicles for bulk studies (enzyme kinetics, fluorescence spectroscopy...... of sphingomyelinase D is clearly influenced by the supramolecular organization of its substrate in membranes and, 3) in situ ceramide-1-phosphate generation by enzymatic activity profoundly alters the lateral structure and morphology of the target membranes....

  9. Enzymatic activity of soluble and membrane tethered peptide pro-hormone convertase 1.

    Bruzzaniti, Angela; Mains, Richard E

    2002-05-01

    Pro-hormone convertases PC1 and PC2 perform endoproteolytic cleavages of precursors in peptide-containing secretory granules. PC1 and PC2 are soluble, secreted with bioactive peptides. Evolutionarily related PCs have membrane tethers, not secreted. We tethered PC1 to the transmembrane-cytoplasmic domains (CD) of a granule enzyme (peptidylglycine-alpha-amidating monooxygenase; PAM) and Golgi-localized PC8. The tethered PC1 is far more stable to elevated temperature and denaturants than soluble PC1, and more active. Both tethers allow PC1 to visit the cell surface transiently, cleaving soluble molecules outside the cell. Both membrane-bound PC1 chimeras cleave membrane PAM into soluble active fragments when PAM is expressed on adjacent cells. PMID:12084516

  10. Trans-activity of Plasma Membrane-associated Ganglioside Sialyltransferase in Mammalian Cells*

    Vilcaes, Aldo A.; Demichelis, Vanina Torres; Daniotti, Jose L.

    2011-01-01

    Gangliosides are acidic glycosphingolipids that contain sialic acid residues and are expressed in nearly all vertebrate cells. They are synthesized at the Golgi complex by a combination of glycosyltransferase activities followed by vesicular delivery to the plasma membrane, where they participate in a variety of physiological as well as pathological processes. Recently, a number of enzymes of ganglioside anabolism and catabolism have been shown to be associated with the plasma membrane. In particular, it was observed that CMP-NeuAc:GM3 sialyltransferase (Sial-T2) is able to sialylate GM3 at the plasma membrane (cis-catalytic activity). In this work, we demonstrated that plasma membrane-integrated ecto-Sial-T2 also displays a trans-catalytic activity at the cell surface of epithelial and melanoma cells. By using a highly sensitive enzyme-linked immunosorbent assay combined with confocal fluorescence microscopy, we observed that ecto-Sial-T2 was able to sialylate hydrophobically or covalently immobilized GM3 onto a solid surface. More interestingly, we observed that ecto-Sial-T2 was able to sialylate GM3 exposed on the membrane of neighboring cells by using both the exogenous and endogenous donor substrate (CMP-N-acetylneuraminic acid) available at the extracellular milieu. In addition, the trans-activity of ecto-Sial-T2 was considerably reduced when the expression of the acceptor substrate was inhibited by using a specific inhibitor of biosynthesis of glycolipids, indicating the lipidic nature of the acceptor. Our findings provide the first direct evidence that an ecto-sialyltransferase is able to trans-sialylate substrates exposed in the plasma membrane from mammalian cells, which represents a novel insight into the molecular events that regulate the local glycosphingolipid composition. PMID:21768099

  11. Trans-activity of plasma membrane-associated ganglioside sialyltransferase in mammalian cells.

    Vilcaes, Aldo A; Demichelis, Vanina Torres; Daniotti, Jose L

    2011-09-01

    Gangliosides are acidic glycosphingolipids that contain sialic acid residues and are expressed in nearly all vertebrate cells. They are synthesized at the Golgi complex by a combination of glycosyltransferase activities followed by vesicular delivery to the plasma membrane, where they participate in a variety of physiological as well as pathological processes. Recently, a number of enzymes of ganglioside anabolism and catabolism have been shown to be associated with the plasma membrane. In particular, it was observed that CMP-NeuAc:GM3 sialyltransferase (Sial-T2) is able to sialylate GM3 at the plasma membrane (cis-catalytic activity). In this work, we demonstrated that plasma membrane-integrated ecto-Sial-T2 also displays a trans-catalytic activity at the cell surface of epithelial and melanoma cells. By using a highly sensitive enzyme-linked immunosorbent assay combined with confocal fluorescence microscopy, we observed that ecto-Sial-T2 was able to sialylate hydrophobically or covalently immobilized GM3 onto a solid surface. More interestingly, we observed that ecto-Sial-T2 was able to sialylate GM3 exposed on the membrane of neighboring cells by using both the exogenous and endogenous donor substrate (CMP-N-acetylneuraminic acid) available at the extracellular milieu. In addition, the trans-activity of ecto-Sial-T2 was considerably reduced when the expression of the acceptor substrate was inhibited by using a specific inhibitor of biosynthesis of glycolipids, indicating the lipidic nature of the acceptor. Our findings provide the first direct evidence that an ecto-sialyltransferase is able to trans-sialylate substrates exposed in the plasma membrane from mammalian cells, which represents a novel insight into the molecular events that regulate the local glycosphingolipid composition. PMID:21768099

  12. Antibacterial activity on electrospun poly(lactide-co-glycolide) based membranes via Magainin II grafting

    Yüksel, Emre; Karakeçili, Ayşe, E-mail: akarakecili@eng.ankara.edu.tr

    2014-12-01

    An antimicrobial peptide (AMP), Magainin II (Mag II) was covalently immobilized on poly(lactide-co-glycolide) (PLGA) and PLGA/gelatin electrospun fibrous membranes. The surface immobilization was characterized by X-ray Photoelectron Spectroscopy (XPS). Scanning Electron Microscopy (SEM) and Atomic Force Microscopy studies showed that the surface morphology of the fibers at micron scale was not affected by the immobilization process. The antibacterial activity of the bound Mag II was tested against Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. Bacterial adhesion tests, SEM and confocal analyses revealed that the attachment and survival of bacteria were inhibited on Mag II functionalized membranes. AMP immobilization strategy was introduced as a new perspective for the modulation of antibacterial properties on PLGA based materials prepared by electrospinning. - Highlights: • PLGA and PLGA/gelatin fibrous membranes were prepared by electrospinning. • Antimicrobial peptide Mag II was successfully immobilized on PLGA based membranes. • The antibacterial activity was tested against E. coli and S. aureus. • Bacterial adhesion was inhibited on Mag II functionalized membranes.

  13. Antibacterial activity on electrospun poly(lactide-co-glycolide) based membranes via Magainin II grafting

    An antimicrobial peptide (AMP), Magainin II (Mag II) was covalently immobilized on poly(lactide-co-glycolide) (PLGA) and PLGA/gelatin electrospun fibrous membranes. The surface immobilization was characterized by X-ray Photoelectron Spectroscopy (XPS). Scanning Electron Microscopy (SEM) and Atomic Force Microscopy studies showed that the surface morphology of the fibers at micron scale was not affected by the immobilization process. The antibacterial activity of the bound Mag II was tested against Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. Bacterial adhesion tests, SEM and confocal analyses revealed that the attachment and survival of bacteria were inhibited on Mag II functionalized membranes. AMP immobilization strategy was introduced as a new perspective for the modulation of antibacterial properties on PLGA based materials prepared by electrospinning. - Highlights: • PLGA and PLGA/gelatin fibrous membranes were prepared by electrospinning. • Antimicrobial peptide Mag II was successfully immobilized on PLGA based membranes. • The antibacterial activity was tested against E. coli and S. aureus. • Bacterial adhesion was inhibited on Mag II functionalized membranes

  14. Bactericidal activity of curcumin I is associated with damaging of bacterial membrane.

    Poonam Tyagi

    Full Text Available Curcumin, an important constituent of turmeric, is known for various biological activities, primarily due to its antioxidant mechanism. The present study focused on the antibacterial activity of curcumin I, a significant component of commercial curcumin, against four genera of bacteria, including those that are Gram-positive (Staphylococcus aureus and Enterococcus faecalis and Gram-negative (Escherichia coli and Pseudomonas aeruginosa. These represent prominent human pathogens, particularly in hospital settings. Our study shows the strong antibacterial potential of curcumin I against all the tested bacteria from Gram-positive as well as Gram-negative groups. The integrity of the bacterial membrane was checked using two differential permeabilization indicating fluorescent probes, namely, propidium iodide and calcein. Both the membrane permeabilization assays confirmed membrane leakage in Gram-negative and Gram-positive bacteria on exposure to curcumin I. In addition, scanning electron microscopy and fluorescence microscopy were employed to confirm the membrane damages in bacterial cells on exposure to curcumin I. The present study confirms the broad-spectrum antibacterial nature of curcumin I, and its membrane damaging property. Findings from this study could provide impetus for further research on curcumin I regarding its antibiotic potential against rapidly emerging bacterial pathogens.

  15. A hybrid anaerobic membrane bioreactor coupled with online ultrasonic equipment for digestion of waste activated sludge.

    Xu, Meilan; Wen, Xianghua; Yu, Zhiyong; Li, Yushan; Huang, Xia

    2011-05-01

    Anaerobic membrane bioreactor and online ultrasonic equipment used to enhance membrane filtration were coupled to form a hybrid system (US-AnMBR) designed for long-term digestion of waste activated sludge. The US-AnMBR was operated under volatile solids loading rates of 1.1-3.7 gVS/L·d. After comprehensive studies on digestion performance and membrane fouling control in the US-AnMBR, the final loading rate was determined to be 2.7 gVS/L·d with 51.3% volatile solids destruction. In the US-AnMBR, the improved digestion was due to enhanced sludge disintegration, as indicated by soluble matter comparison in the supernatant and particle size distribution in the digested sludge. Maximum specific methanogenic activity revealed that ultrasound application had no negative effect on anaerobic microorganisms. Furthermore, implementing ultrasound effectively controlled membrane fouling and successfully facilitated membrane bioreactor operation. This lab-scale study demonstrates the potential feasibility and effectiveness of setting up a US-AnMBR system for sludge digestion. PMID:21421308

  16. A Novel Murine Anti-Lactoferrin Monoclonal Antibody Activates Human Polymorphonuclear Leukocytes through Membrane-Bound Lactoferrin and TLR4

    Xiao-Min Hu

    2015-01-01

    Full Text Available Soluble lactoferrin (LTF is a versatile molecule that not only regulates the iron homeostasis, but also harbors direct microbicidal and immunomodulating abilities in mammalian body fluids. In contrast, little is known about the function of membrane-bound LTF (mbLTF, although its expression on human polymorphonuclear leukocytes (huPMNs has been reported for decades. Given that LTF/anti-LTF antibodies represent a potential diagnostic/prognostic biomarker and a therapeutic target in patients with immune disorders, we wished, in the present study, to generate a novel human LTF- (huLTF- specific mAb suitable for detailed analyses on the expression and function of mbLTF as well as for deciphering the underlying mechanisms. By using the traditional hybridoma cell fusion technology, we obtained a murine IgG1 (kappa mAb, M-860, against huLTF. M-860 recognizes a conformational epitope of huLTF as it binds to natural, but not denatured, huLTF in ELISA. Moreover, M-860 detects mbLTF by FACS and captures endogenous huLTF in total cell lysates of huPMNs. Functionally, M-860 induces the activation of huPMNs partially through TLR4 but independently of phagocytosis. M-860 is thus a powerful tool to analyze the expression and function of human mbLTF, which will further our understanding of the roles of LTF in health and disease.

  17. Active Curved Polymers Form Vortex Patterns on Membranes

    Denk, Jonas; Huber, Lorenz; Reithmann, Emanuel; Frey, Erwin

    2016-04-01

    Recent in vitro experiments with FtsZ polymers show self-organization into different dynamic patterns, including structures reminiscent of the bacterial Z ring. We model FtsZ polymers as active particles moving along chiral, circular paths by Brownian dynamics simulations and a Boltzmann approach. Our two conceptually different methods point to a generic phase behavior. At intermediate particle densities, we find self-organization into vortex structures including closed rings. Moreover, we show that the dynamics at the onset of pattern formation is described by a generalized complex Ginzburg-Landau equation.

  18. Results from the CDE phase activity on neutron dosimetry for the international fusion materials irradiation facility test cell

    Esposito, B; Maruccia, G; Petrizzi, L; Bignon, G; Blandin, C; Chauffriat, S; Lebrun, A; Recroix, H; Trapp, J P; Kaschuck, Y

    2000-01-01

    The international fusion materials irradiation facility (IFMIF) project deals with the study of an accelerator-based, deuterium-lithium source, producing high energy neutrons at sufficient intensity and irradiation volume to test samples of candidate materials for fusion energy reactors. IFMIF would also provide calibration and validation of data from fission reactor and other accelerator based irradiation tests. This paper describes the activity on neutron/gamma dosimetry (necessary for the characterization of the specimens' irradiation) performed in the frame of the IFMIF conceptual design evaluation (CDE) neutronics tasks. During the previous phase (conceptual design activity (CDA)) the multifoil activation method was proposed for the measurement of the neutron fluence and spectrum and a set of suitable foils was defined. The cross section variances and covariances of this set of foils have now been used for tests on the sensitivity of the IFMIF neutron spectrum determination to cross section uncertainties...

  19. Effects of Aluminum on ATPase Activity and Lipid Composition of Plasma Membranes from Wheat Roots

    HE Long-fei; LIU You-liang; SHEN Zhen-guo; WANG Ai-qin

    2002-01-01

    The effects of aluminum on ATPase activity and lipid composition of the plasma membranes isolated from root tips of Al-tolerant (Altas 66) or Al-sensitive (Scout 66) cultivar of Triticum aestivum L.was assayed. The results showed that both cultivars had similar changes in H+ -ATPase and Ca2+ -ATPase activities after aluminum treatment. Exposure of both cultivars to 20 and 100 (mol/L aluminum for 5 d significantly decreased the activities of Ca2+ -ATPase of plasma membranes. The activities of H+-ATPasc in plasma membrane increased under 20 μmol/L aluminum and decreased at 100 μmol/L aluminum. With aluminum treatment, the PL content of plasma membrane decreased, but GL content increased. The ratio of PL to GL decreased more distinctly in Scout 66 than that in Altas 66. Treated with 20 and 100 μmol/L aluminum, linolenic acid content and the index of unsaturated fatty acids decreaced greatly in Scout 66, but the index of unsaturated fatty acids in Altas 66 increased slightly.

  20. Critical findings on the activation cascade of yeast plasma membrane H+-ATPase

    Kotyk, Arnošt; Lapathitis, Georgios; Horák, Jaroslav

    2003-01-01

    Roč. 226, č. 1 (2003), s. 175-180. ISSN 0378-1097 R&D Projects: GA ČR GA204/02/1240 Institutional research plan: CEZ:AV0Z5045916; CEZ:AV0Z5011922 Keywords : H+-ATPase * yeast plasma membrane * activation Subject RIV: CE - Biochemistry Impact factor: 1.932, year: 2003