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Sample records for surface binding demonstrated

  1. Distribution of Glycan Motifs at the Surface of Midgut Cells in the Cotton Leafworm (Spodoptera littoralis Demonstrated by Lectin Binding

    Directory of Open Access Journals (Sweden)

    Tomasz Walski

    2017-12-01

    Full Text Available Glycans are involved in many biological phenomena, including signal transduction, cell adhesion, immune response or differentiation. Although a few papers have reported on the role of glycans in the development and proper functioning of the insect midgut, no data are available regarding the localization of the glycan structures on the surface of the cells in the gut of insects. In this paper, we analyzed the spatial distribution of glycans present on the surface of the midgut cells in larvae of the cotton leafworm Spodoptera littoralis, an important agricultural pest insect worldwide. For this purpose, we established primary midgut cell cultures, probed these individual cells that are freely suspended in liquid medium with a selection of seven fluorescently labeled lectins covering a range of different carbohydrate binding specificities [mannose oligomers (GNA and HHA, GalNAc/Gal (RSA and SSA, GlcNAc (WGA and Nictaba and Neu5Ac(α-2,6Gal/GalNAc (SNA-I], and visualized the interaction of these lectins with the different zones of the midgut cells using confocal microscopy. Our analysis focused on the typical differentiated columnar cells with a microvillar brush border at their apical side, which are dominantly present in the Lepidopteran midgut and function in food digestion and absorption, and as well as on the undifferentiated stem cells that are important for midgut development and repair. Confocal microscopy analyses showed that the GalNAc/Gal-binding lectins SSA and RSA and the terminal GlcNAc-recognizing WGA bound preferentially to the apical microvillar zone of the differentiated columnar cells as compared to the basolateral pole. The reverse result was observed for the mannose-binding lectins GNA and HHA, as well as Nictaba that binds preferentially to GlcNAc oligomers. Furthermore, differences in lectin binding to the basal and lateral zones of the cell membranes of the columnar cells were apparent. In the midgut stem cells, GNA and

  2. Binding Preferences, Surface Attachment, Diffusivity, and Orientation of a Family 1 Carbohydrate-Binding Module on Cellulose

    Energy Technology Data Exchange (ETDEWEB)

    Nimlos, M. R.; Beckham, G. T.; Matthews, J. F.; Bu, L.; Himmel, M. E.; Crowley, M. F.

    2012-06-08

    Cellulase enzymes often contain carbohydrate-binding modules (CBMs) for binding to cellulose. The mechanisms by which CBMs recognize specific surfaces of cellulose and aid in deconstruction are essential to understand cellulase action. The Family 1 CBM from the Trichoderma reesei Family 7 cellobiohydrolase, Cel7A, is known to selectively bind to hydrophobic surfaces of native cellulose. It is most commonly suggested that three aromatic residues identify the planar binding face of this CBM, but several recent studies have challenged this hypothesis. Here, we use molecular simulation to study the CBM binding orientation and affinity on hydrophilic and hydrophobic cellulose surfaces. Roughly 43 {mu}s of molecular dynamics simulations were conducted, which enables statistically significant observations. We quantify the fractions of the CBMs that detach from crystal surfaces or diffuse to other surfaces, the diffusivity along the hydrophobic surface, and the overall orientation of the CBM on both hydrophobic and hydrophilic faces. The simulations demonstrate that there is a thermodynamic driving force for the Cel7A CBM to bind preferentially to the hydrophobic surface of cellulose relative to hydrophilic surfaces. In addition, the simulations demonstrate that the CBM can diffuse from hydrophilic surfaces to the hydrophobic surface, whereas the reverse transition is not observed. Lastly, our simulations suggest that the flat faces of Family 1 CBMs are the preferred binding surfaces. These results enhance our understanding of how Family 1 CBMs interact with and recognize specific cellulose surfaces and provide insights into the initial events of cellulase adsorption and diffusion on cellulose.

  3. Specific insulin binding in bovine chromaffin cells; demonstration of preferential binding to adrenalin-storing cells

    International Nuclear Information System (INIS)

    Serck-Hanssen, G.; Soevik, O.

    1987-01-01

    Insulin binding was studied in subpopulations of bovine chromaffin cells enriched in adrenalin-producing cells (A-cells) or noradrenalin-producing cells (NA-cells). Binding of 125 I-insulin was carried out at 15 0 C for 3 hrs in the absence or presence of excess unlabeled hormone. Four fractions of cells were obtained by centrifugation on a stepwise bovine serum albumin gradient. The four fractions were all shown to bind insulin in a specific manner and the highest binding was measured in the cell layers of higher densities, containing mainly A-cells. The difference in binding of insulin to the four subpopulations of chromaffin cells seemed to be related to differences in numbers of receptors as opposed to receptor affinities. The authors conclude that bovine chromaffin cells possess high affinity binding sites for insulin and that these binding sites are mainly confined to A-cells. 24 references, 2 figures, 1 table

  4. Using supramolecular binding motifs to provide precise control over the ratio and distribution of species in multiple component films grafted on surfaces: demonstration using electrochemical assembly from aryl diazonium salts.

    Science.gov (United States)

    Gui, Alicia L; Yau, Hon Man; Thomas, Donald S; Chockalingam, Muthukumar; Harper, Jason B; Gooding, J Justin

    2013-04-16

    Supramolecular interactions between two surface modification species are explored to control the ratio and distribution of these species on the resultant surface. A binary mixture of aryl diazonium salts bearing oppositely charged para-substituents (either -SO3(-) or -N(+)(Me)3), which also reduce at different potentials, has been examined on glassy carbon surfaces using cyclic voltammetry and X-ray photoelectron spectroscopy (XPS). Striking features were observed: (1) the two aryl diazonium salts in the mixed solution undergo reductive adsorption at the same potential which is distinctively less negative than the potential required for the reduction of either of the two aryl diazonium salts alone; (2) the surface ratio of the two phenyl derivatives is consistently 1:1 regardless of the ratio of the two aryl diazonium salts in the modification solutions. Homogeneous distribution of the two oppositely charged phenyl species on the modified surface has also been suggested by XPS survey spectra. Diffusion coefficient measurements by DOSY NMR and DFT based computation have indicated the association of the two aryl diazonium species in the solution, which has led to changes in the molecular orbital energies of the two species. This study highlights the potential of using intermolecular interactions to control the assembly of multicomponent thin layers.

  5. Demonstration of specific binding sites for 3H-RRR-alpha-tocopherol on human erythrocytes

    International Nuclear Information System (INIS)

    Kitabchi, A.E.; Wimalasena, J.

    1982-01-01

    Previous work from our laboratory demonstrated specific binding sites for 3 H-RRR-alpha-tocopherol ( 3 H-d alpha T) in membranes of rat adrenal cells. As tocopherol deficiency is associated with increased susceptibility of red blood cells to hemolysis, we investigated tocopherol binding sites in human RBCs. Erythrocytes were found to have specific binding sites for 3 H-d alpha T that exhibited saturability and time and cell-concentration dependence as well as reversibility of binding. Kinetic studies of binding demonstrated two binding sites--one with high affinity (Ka of 2.6 x 10(7) M-1), low capacity (7,600 sites per cell) and the other with low affinity (1.2 x 10(6) M-1), high capacity (150,000 sites per cell). In order to localize the binding sites further, RBCs were fractionated and greater than 90% of the tocopherol binding was located in the membranes. Similar to the findings in intact RBCs, the membranes exhibited two binding sites with a respective Ka of 3.3 x 10(7) M-1 and 1.5 x 10(6) M-1. Specificity data for binding demonstrated 10% binding for RRR-gamma-tocopherol, but not other tocopherol analog exhibited competition for 3 H-d alpha T binding sites. Instability data suggested a protein nature for these binding sites. Preliminary studies on Triton X-100 solubilized fractions resolved the binding sites to a major component with an Mr of 65,000 and a minor component with an Mr of 125,000. We conclude that human erythrocyte membranes contain specific binding sites for RRR-alpha-tocopherol. These sites may be of physiologic significance in the function of tocopherol on the red blood cell membrane

  6. Surface tension in soap films: revisiting a classic demonstration

    International Nuclear Information System (INIS)

    Behroozi, F

    2010-01-01

    We revisit a classic demonstration for surface tension in soap films and introduce a more striking variation of it. The demonstration shows how the film, pulling uniformly and normally on a loose string, transforms it into a circular arc under tension. The relationship between the surface tension and the string tension is analysed and presented in a useful graphical form. (letters and comments)

  7. Surface tension in soap films: revisiting a classic demonstration

    Energy Technology Data Exchange (ETDEWEB)

    Behroozi, F [Department of Physics, University of Northern Iowa, Cedar Falls, IA 50614 (United States)], E-mail: behroozi@uni.edu

    2010-01-15

    We revisit a classic demonstration for surface tension in soap films and introduce a more striking variation of it. The demonstration shows how the film, pulling uniformly and normally on a loose string, transforms it into a circular arc under tension. The relationship between the surface tension and the string tension is analysed and presented in a useful graphical form. (letters and comments)

  8. Universal binding energy relation for cleaved and structurally relaxed surfaces

    International Nuclear Information System (INIS)

    Srirangarajan, Aarti; Datta, Aditi; Gandi, Appala Naidu; Ramamurty, U; Waghmare, U V

    2014-01-01

    The universal binding energy relation (UBER), derived earlier to describe the cohesion between two rigid atomic planes, does not accurately capture the cohesive properties when the cleaved surfaces are allowed to relax. We suggest a modified functional form of UBER that is analytical and at the same time accurately models the properties of surfaces relaxed during cleavage. We demonstrate the generality as well as the validity of this modified UBER through first-principles density functional theory calculations of cleavage in a number of crystal systems. Our results show that the total energies of all the relaxed surfaces lie on a single (universal) energy surface, that is given by the proposed functional form which contains an additional length-scale associated with structural relaxation. This functional form could be used in modelling the cohesive zones in crack growth simulation studies. We find that the cohesive law (stress–displacement relation) differs significantly in the case where cracked surfaces are allowed to relax, with lower peak stresses occurring at higher displacements. (paper)

  9. Universal binding energy relation for cleaved and structurally relaxed surfaces.

    Science.gov (United States)

    Srirangarajan, Aarti; Datta, Aditi; Gandi, Appala Naidu; Ramamurty, U; Waghmare, U V

    2014-02-05

    The universal binding energy relation (UBER), derived earlier to describe the cohesion between two rigid atomic planes, does not accurately capture the cohesive properties when the cleaved surfaces are allowed to relax. We suggest a modified functional form of UBER that is analytical and at the same time accurately models the properties of surfaces relaxed during cleavage. We demonstrate the generality as well as the validity of this modified UBER through first-principles density functional theory calculations of cleavage in a number of crystal systems. Our results show that the total energies of all the relaxed surfaces lie on a single (universal) energy surface, that is given by the proposed functional form which contains an additional length-scale associated with structural relaxation. This functional form could be used in modelling the cohesive zones in crack growth simulation studies. We find that the cohesive law (stress-displacement relation) differs significantly in the case where cracked surfaces are allowed to relax, with lower peak stresses occurring at higher displacements.

  10. Surface Passivation in Empirical Tight Binding

    OpenAIRE

    He, Yu; Tan, Yaohua; Jiang, Zhengping; Povolotskyi, Michael; Klimeck, Gerhard; Kubis, Tillmann

    2015-01-01

    Empirical Tight Binding (TB) methods are widely used in atomistic device simulations. Existing TB methods to passivate dangling bonds fall into two categories: 1) Method that explicitly includes passivation atoms is limited to passivation with atoms and small molecules only. 2) Method that implicitly incorporates passivation does not distinguish passivation atom types. This work introduces an implicit passivation method that is applicable to any passivation scenario with appropriate parameter...

  11. Surface Passivation in Empirical Tight Binding

    Science.gov (United States)

    He, Yu; Tan, Yaohua; Jiang, Zhengping; Povolotskyi, Michael; Klimeck, Gerhard; Kubis, Tillmann

    2016-03-01

    Empirical Tight Binding (TB) methods are widely used in atomistic device simulations. Existing TB methods to passivate dangling bonds fall into two categories: 1) Method that explicitly includes passivation atoms is limited to passivation with atoms and small molecules only. 2) Method that implicitly incorporates passivation does not distinguish passivation atom types. This work introduces an implicit passivation method that is applicable to any passivation scenario with appropriate parameters. This method is applied to a Si quantum well and a Si ultra-thin body transistor oxidized with SiO2 in several oxidation configurations. Comparison with ab-initio results and experiments verifies the presented method. Oxidation configurations that severely hamper the transistor performance are identified. It is also shown that the commonly used implicit H atom passivation overestimates the transistor performance.

  12. Gonadal cell surface receptor for plasma retinol-binding protein

    International Nuclear Information System (INIS)

    Krishna Bhat, M.; Cama, H.R.

    1979-01-01

    A specific membrane receptor for plasma retinol-binding protein has been demonstrated in testicular cells. Prealbumin-2 did not show any specific binding to the membrane. The affinity of retinol-binding protein for receptor drastically decreases upon delivery of retinol and the retinol-binding protein does not enter the cell. The mechanism of delivery of retinol to the target cell by plasma retinol-binding protein has been investigated. The process involves two steps; direct binding of retinol-binding protein to the receptor and uptake of retinol by the target cell with a concomitant drastic reduction in the affinity of the retinol-binding protein to the receptor. Probably the second step of the process needs a cytosolic factor, possibly the cellular retinol-binding protein or an enzyme. The binding of retinol-binding protein to the receptor is saturable and reversible. The interaction shows a Ksub(d) value of 2.1x10 -10 . The specific binding of a retinol-binding protein with great affinity has been employed in the development of a method for radioassay of the receptor. The receptor level of the gonadal cell has been found to vary with the stage of differentiation. The receptor concentrations in 11-week-old birds and adult birds are comparable. Testosterone treatment of 11-week-old birds produced a substantial increase in the receptor concentration over control, while the protein content increased marginally, indicating that, probably, synthesis of the receptor is specifcally induced by testosterone during spermatogenesis, and the concentration of receptor is relatively higher before the formation of the acrosome. (Auth.)

  13. Binding of hydrophobic antigens to surfaces

    DEFF Research Database (Denmark)

    2017-01-01

    A first aspect of the present invention is a method of detecting antibodies comprising the steps of: i) providing a first group of beads comprising a surface modified with C1-C10 alkyl groups comprising amine, ammonium, ether and/or hydroxyl groups, ii) contacting said first group of beads......-antigen-antibody conjugates, and v) detecting said bead-antigen-antibody conjugates. Further aspects include an antibody detection kit, a bead-antigen conjugate and a composition comprising at least two different groups of bead-antigen-conjugates....

  14. Crystal structure of the botulinum neurotoxin type G binding domain: insight into cell surface binding.

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    Stenmark, Pål; Dong, Min; Dupuy, Jérôme; Chapman, Edwin R; Stevens, Raymond C

    2010-04-16

    Botulinum neurotoxins (BoNTs) typically bind the neuronal cell surface via dual interactions with both protein receptors and gangliosides. We present here the 1.9-A X-ray structure of the BoNT serotype G (BoNT/G) receptor binding domain (residues 868-1297) and a detailed view of protein receptor and ganglioside binding regions. The ganglioside binding motif (SxWY) has a conserved structure compared to the corresponding regions in BoNT serotype A and BoNT serotype B (BoNT/B), but several features of interactions with the hydrophilic face of the ganglioside are absent at the opposite side of the motif in the BoNT/G ganglioside binding cleft. This may significantly reduce the affinity between BoNT/G and gangliosides. BoNT/G and BoNT/B share the protein receptor synaptotagmin (Syt) I/II. The Syt binding site has a conserved hydrophobic plateau located centrally in the proposed protein receptor binding interface (Tyr1189, Phe1202, Ala1204, Pro1205, and Phe1212). Interestingly, only 5 of 14 residues that are important for binding between Syt-II and BoNT/B are conserved in BoNT/G, suggesting that the means by which BoNT/G and BoNT/B bind Syt diverges more than previously appreciated. Indeed, substitution of Syt-II Phe47 and Phe55 with alanine residues had little effect on the binding of BoNT/G, but strongly reduced the binding of BoNT/B. Furthermore, an extended solvent-exposed hydrophobic loop, located between the Syt binding site and the ganglioside binding cleft, may serve as a third membrane association and binding element to contribute to high-affinity binding to the neuronal membrane. While BoNT/G and BoNT/B are homologous to each other and both utilize Syt-I/Syt-II as their protein receptor, the precise means by which these two toxin serotypes bind to Syt appears surprisingly divergent. Copyright (c) 2010. Published by Elsevier Ltd.

  15. Reversible and Irreversible Binding of Nanoparticles to Polymeric Surfaces

    Directory of Open Access Journals (Sweden)

    Wolfgang H. Binder

    2009-01-01

    Full Text Available Reversible and irreversible binding of CdSe-nanoparticles and nanorods to polymeric surfaces via a strong, multiple hydrogen bond (= Hamilton-receptor/barbituric acid is described. Based on ROMP-copolymers, the supramolecular interaction on a thin polymer film is controlled by living polymerization methods, attaching the Hamilton-receptor in various architectures, and concentrations. Strong binding is observed with CdSe-nanoparticles and CdSe-nanorods, whose surfaces are equipped with matching barbituric acid-moieties. Addition of polar solvents, able to break the hydrogen bonds leads to the detachment of the nanoparticles from the polymeric film. Irreversible binding is observed if an azide/alkine-“click”-reaction is conducted after supramolecular recognition of the nanoparticles on the polymeric surface. Thus reversible or irreversible attachment of the nanosized objects can be achieved.

  16. Binding and orientation of fibronectin on polystyrene surfaces using immobilized bacterial adhesin-related peptides.

    Science.gov (United States)

    Klueh, U; Bryers, J D; Kreutzer, D L

    2003-10-01

    Fibronectin (FN) is known to bind to bacteria via high affinity receptors on bacterial surfaces known as adhesins. The binding of bacteria to FN is thought to have a key role in foreign device associated infections. For example, previous studies have indicated that Staphylococcus aureus adhesins bind to the 29 kDa NH(3) terminus end of FN, and thereby promote bacteria adherence to surfaces. Recently, the peptide sequences within the S. aureus adhesin molecule that are responsible for FN binding have been identified. Based on these observations, we hypothesize that functional FN can be bound and specifically oriented on polystyrene surfaces using bacterial adhesin-related (BRP-A) peptide. We further hypothesize that monoclonal antibodies that react with specific epitopes on the FN can be used to quantify both FN binding and orientation on these surfaces. Based on this hypothesis, we initiated a systematic investigation of the binding and orientation of FN on polystyrene surfaces using BRP-A peptide. To test this hypothesis, the binding and orientation of the FN to immobilized BRP-A was quantified using (125)I-FN, and monoclonal antibodies. (125)I-FN was used to quantitate FN binding to peptide-coated polystyrene surfaces. The orientation of bound FN was demonstrated by the use of monoclonal antibodies, which are reactive with the amine (N) or carboxyl (C) termini of the FN. The results of our studies demonstrated that when the BRP-A peptide was used to bind FN to surfaces that: 1. functional FN was bound to the peptide; 2. anti-C terminus antibodies bound to the peptide FN; and 3. only limited binding of anti-N terminus antibodies to peptide-bound FN occurred. We believe that the data that indicate an enhanced binding of anti-C antibodies reactive to anti-N antibodies are a result of the FN binding in an oriented manner with the N termini of FN bound tightly to the BRP-A on the polystyrene surface. Copyright 2003 Wiley Periodicals, Inc. J Biomed Mater Res 67A: 36

  17. Functional Elements on SIRPα IgV domain Mediate Cell Surface Binding to CD47

    OpenAIRE

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J.; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J.; Zen, Ke

    2006-01-01

    SIRPα and SIRPβ1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPα with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPβ1 ...

  18. Demonstration of 99mTc-labelled plasmin on the surface of ex vivo thrombi

    International Nuclear Information System (INIS)

    Tengborn, L.; Hedner, U.; Rohlin, M.; Stahlberg, F.

    1982-01-01

    In an vitro system using the Chandler model for the preparation of in vitro thrombi trace amounts of porcine or human 99mTc-labelled plasmin was found to adsorb to the surface of a preformed thrombus. A radioactive lining of the thrombus could be demonstrated using autoradiography after addition of 99mTc-labelled plasmin in concentrations of 0.04 - 0.07 or 0.4 - 0.7 CTA u/thrombus made from 2 ml whole blood (0.035 - 0.35 microM). The same pattern was found for porcine as for human plasmin. The presence of tranexamic acid in concentrations of 3 to 12 mM did not affect the binding of plasmin indicating that the plasmin binding to fibrin was independent of the lysine binding sites. Furthermore alpha 2-antiplasmin was demonstrated on/in the thrombus also when no plasmin was present indicating a binding of alpha 2-antiplasmin to the thrombus. The plasmin bound to the thrombus was proteolytically inactive. In order to obtain thrombolysis most of the alpha 2-antiplasmin in the surrounding medium had to be neutralized

  19. Equilibrium binding studies of mono, di and triisocyanide ligands on Au powder surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Ontko, Alyn [Iowa State Univ., Ames, IA (United States)

    1997-10-08

    The author`s group has previously shown that isocyanides are readily adsorbed from solutions to Au powder and bind to the Au surface in an end-on fashion through the terminal carbon. Later work demonstrated that the equilibrium constants for the reversible adsorption of electronically inequivalent isocyanides could be obtained using the Langmuir isotherm technique. This dissertation describes two projects completed which complement the initial findings of this group. Initially, several alkylisocyanides were synthesized to examine the effect of tail length on Au powder adsorption. It was observed that the length of the alkyl chain affected not only the Au surface binding affinity, but also the rate of surface saturation and saturation coverage values. Direct competition studies were also studied using a 13C-labeled isocyanide. These studies demonstrated the stabilization afforded by substrate-substrate packing forces in SAM`s formed by the longer chain isocyanides. In a second study, di and triisocyanides were synthesized to determine the effect that the length of the connecting link and the number of isocyanide groups (as points of attachment) have on Au adsorption stability. The work in this area describes the binding modes, relative binding affinities and surface coverage values for a series of flexible alkyl and xylyldiisocyanides on Au powder surfaces. This report contains only the introductory material, and general summary. Two chapters have been processed separately. 56 refs.

  20. Functional Elements on SIRPα IgV domain Mediate Cell Surface Binding to CD47

    Science.gov (United States)

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J.; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J.; Zen, Ke

    2007-01-01

    Summary SIRPα and SIRPβ1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPα with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPβ1 shares highly homologous extracellular IgV structure with SIRPα, it does not bind to CD47. In this study, we defined key amino acid residues exclusively expressing in the IgV domain of SIRPα, but not SIRPβ1, which determine the extracellular binding interaction of SIRPα to CD47. These key residues include Gln67, a small hydrophobic amino acid (Ala or Val) at the 57th position and Met102. We found that Gln67 and Ala/Val57 are critical. Mutation of either of these residues abates SIRPα directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRPα extracellular binding mediated cell interactions and cell migration. Another SIRPα-specific residue, Met102, appears to assist SIRPα IgV binding through Gln67 and Ala/Val57. An essential role of these amino acids in SIRPα binding to CD47 was further confirmed by introducing these residues into the SIRPβ1 IgV domain, which dramatically converts SIRPβ1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRPα selectively binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses. PMID:17070842

  1. Functional elements on SIRPalpha IgV domain mediate cell surface binding to CD47.

    Science.gov (United States)

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J; Yang, Yang; Zen, Ke

    2007-01-19

    SIRPalpha and SIRPbeta1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPalpha with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPbeta1 shares highly homologous extracellular IgV structure with SIRPalpha, it does not bind to CD47. Here, we defined key amino acid residues exclusively expressing in the IgV domain of SIRPalpha, but not SIRPbeta1, which determine the extracellular binding interaction of SIRPalpha to CD47. These key residues include Gln67, a small hydrophobic amino acid (Ala or Val) at the 57th position and Met102. We found that Gln67 and Ala/Val57 are critical. Mutation of either of these residues abates SIRPalpha directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRPalpha extracellular binding mediated cell interactions and cell migration. Another SIRPalpha-specific residue, Met102, appears to assist SIRPalpha IgV binding through Gln67 and Ala/Val57. An essential role of these amino acid residues in SIRPalpha binding to CD47 was further confirmed by introducing these residues into the SIRPbeta1 IgV domain, which dramatically converts SIRPbeta1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRPalpha binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses.

  2. Flow cytometric analysis of lectin binding to in vitro-cultured Perkinsus marinus surface carbohydrates

    Science.gov (United States)

    Gauthier, J.D.; Jenkins, J.A.; La Peyre, Jerome F.

    2004-01-01

    Parasite surface glycoconjugates are frequently involved in cellular recognition and colonization of the host. This study reports on the identification of Perkinsus marinus surface carbohydrates by flow cytometric analyses of fluorescein isothiocyanate-conjugated lectin binding. Lectin-binding specificity was confirmed by sugar inhibition and Kolmogorov-Smirnov statistics. Clear, measurable fluorescence peaks were discriminated, and no parasite autofluorescence was observed. Parasites (GTLA-5 and Perkinsus-1 strains) harvested during log and stationary phases of growth in a protein-free medium reacted strongly with concanavalin A and wheat germ agglutinin, which bind to glucose-mannose and N-acetyl-D-glucosamine (GlcNAc) moieties, respectively. Both P. marinus strains bound with lower intensity to Maclura pomifera agglutinin, Bauhinia purpurea agglutinin, soybean agglutinin (N-acetyl-D-galactosamine-specific lectins), peanut agglutinin (PNA) (terminal galactose specific), and Griffonia simplicifolia II (GlcNAc specific). Only background fluorescence levels were detected with Ulex europaeus agglutinin I (L-fucose specific) and Limulus polyphemus agglutinin (sialic acid specific). The lectin-binding profiles were similar for the 2 strains except for a greater relative binding intensity of PNA for Perkinsus-1 and an overall greater lectin-binding capacity of Perkinsus-1 compared with GTLA-5. Growth stage comparisons revealed increased lectin-binding intensities during stationary phase compared with log phase of growth. This is the first report of the identification of surface glycoconjugates on a Perkinsus spp. by flow cytometry and the first to demonstrate that differential surface sugar expression is growth phase and strain dependent. ?? American Society of Parasitologists 2004.

  3. Quantitative monitoring of two simultaneously binding species using Label-Enhanced surface plasmon resonance.

    Science.gov (United States)

    Eng, Lars; Garcia, Brandon L; Geisbrecht, Brian V; Hanning, Anders

    2018-02-26

    Surface plasmon resonance (SPR) is a well-established method for biomolecular interaction studies. SPR monitors the binding of molecules to a solid surface, embodied as refractive index changes close to the surface. One limitation of conventional SPR is the universal nature of the detection that results in an inability to qualitatively discriminate between different binding species. Furthermore, it is impossible to directly discriminate two species simultaneously binding to different sites on a protein, which limits the utility of SPR, for example, in the study of allosteric binders or bi-specific molecules. It is also impossible in principle to discriminate protein conformation changes from actual binding events. Here we demonstrate how Label-Enhanced SPR can be utilized to discriminate and quantitatively monitor the simultaneous binding of two different species - one dye-labeled and one unlabeled - on a standard, single-wavelength SPR instrument. This new technique increases the versatility of SPR technology by opening up application areas where the usefulness of the approach has previously been limited. Copyright © 2018 Elsevier Inc. All rights reserved.

  4. Imparting albumin-binding affinity to a human protein by mimicking the contact surface of a bacterial binding protein.

    Science.gov (United States)

    Oshiro, Satoshi; Honda, Shinya

    2014-04-18

    Attachment of a bacterial albumin-binding protein module is an attractive strategy for extending the plasma residence time of protein therapeutics. However, a protein fused with such a bacterial module could induce unfavorable immune reactions. To address this, we designed an alternative binding protein by imparting albumin-binding affinity to a human protein using molecular surface grafting. The result was a series of human-derived 6 helix-bundle proteins, one of which specifically binds to human serum albumin (HSA) with adequate affinity (KD = 100 nM). The proteins were designed by transferring key binding residues of a bacterial albumin-binding module, Finegoldia magna protein G-related albumin-binding domain (GA) module, onto the human protein scaffold. Despite 13-15 mutations, the designed proteins maintain the original secondary structure by virtue of careful grafting based on structural informatics. Competitive binding assays and thermodynamic analyses of the best binders show that the binding mode resembles that of the GA module, suggesting that the contacting surface of the GA module is mimicked well on the designed protein. These results indicate that the designed protein may act as an alternative low-risk binding module to HSA. Furthermore, molecular surface grafting in combination with structural informatics is an effective approach for avoiding deleterious mutations on a target protein and for imparting the binding function of one protein onto another.

  5. Fission Surface Power Technology Demonstration Unit Test Results

    Science.gov (United States)

    Briggs, Maxwell H.; Gibson, Marc A.; Geng, Steven M.; Sanzi, James L.

    2016-01-01

    The Fission Surface Power (FSP) Technology Demonstration Unit (TDU) is a system-level demonstration of fission power technology intended for use on manned missions to Mars. The Baseline FSP systems consists of a 190 kWt UO2 fast-spectrum reactor cooled by a primary pumped liquid metal loop. This liquid metal loop transfers heat to two intermediate liquid metal loops designed to isolate fission products in the primary loop from the balance of plant. The intermediate liquid metal loops transfer heat to four Stirling Power Conversion Units (PCU), each of which produce 12 kWe (48 kW total) and reject waste heat to two pumped water loops, which transfer the waste heat to titanium-water heat pipe radiators. The FSP TDU simulates a single leg of the baseline FSP system using an electrically heater core simulator, a single liquid metal loop, a single PCU, and a pumped water loop which rejects the waste heat to a Facility Cooling System (FCS). When operated at the nominal operating conditions (modified for low liquid metal flow) during TDU testing the PCU produced 8.9 kW of power at an efficiency of 21.7 percent resulting in a net system power of 8.1 kW and a system level efficiency of 17.2 percent. The reduction in PCU power from levels seen during electrically heated testing is the result of insufficient heat transfer from the NaK heater head to the Stirling acceptor, which could not be tested at Sunpower prior to delivery to the NASA Glenn Research Center (GRC). The maximum PCU power of 10.4 kW was achieved at the maximum liquid metal temperature of 875 K, minimum water temperature of 350 K, 1.1 kg/s liquid metal flow, 0.39 kg/s water flow, and 15.0 mm amplitude at an efficiency of 23.3 percent. This resulted in a system net power of 9.7 kW and a system efficiency of 18.7 percent.

  6. Isothermal titration calorimetry and surface plasmon resonance allow quantifying substrate binding to different binding sites of Bacillus subtilis xylanase

    DEFF Research Database (Denmark)

    Cuyvers, Sven; Dornez, Emmie; Abou Hachem, Maher

    2012-01-01

    Isothermal titration calorimetry and surface plasmon resonance were tested for their ability to study substrate binding to the active site (AS) and to the secondary binding site (SBS) of Bacillus subtilis xylanase A separately. To this end, three enzyme variants were compared. The first...

  7. Low-cost rural surface alternatives : demonstration project.

    Science.gov (United States)

    2015-06-01

    The goals of this project were to implement several stabilization methods for preventing or mitigating freeze-thaw damage to : granular surfaced roads and identify the most effective and economical methods for the soil and climate conditions of Iowa....

  8. Detection of vitamin D binding protein on the surface of cytotrophoblasts isolated from human placentae

    International Nuclear Information System (INIS)

    Nestler, J.E.; McLeod, J.F.; Kowalski, M.A.; Strauss, J.F. III; Haddad, J.G. Jr.

    1987-01-01

    Vitamin D binding protein (DBP), a Mr 56,000-58,000 alpha 2-glycoprotein, is the major serum protein involved in the transport of vitamin D sterols. Recently it has been suggested that DBP may also be involved in immunoglobulin G binding to cells. Because the trophoblast is involved in the transport of molecules such as vitamin D and immunoglobulin G to the fetus, we asked whether DBP could be detected on the surface of human placental trophoblast cells. Cytotrophoblasts purified from human term placentae were fixed and made permeant with Triton X-100 and examined by indirect immunofluorescence after incubation with a monoclonal antibody to DBP. Greater than 90% of these cells stained positively, whereas no staining was observed with nonimmune antiserum. The presence of DBP on/in the surface of cytotrophoblasts could also be demonstrated by fluorescent cytometry. When cell surface-associated proteins of cytotrophoblasts were radioiodinated, a Mr 57,000 radiolabeled protein could be immunoisolated from the cell lysate with a purified monospecific polyclonal antibody to DBP. Immunoisolation of this radiolabeled protein was prevented by the addition of excess unlabeled human DBP to the cell lysate before incubation with antibody. This Mr 57,000 radiolabeled protein could also be isolated by affinity chromatography selecting for proteins that bind to globular actin. When cytotrophoblasts were incubated with [ 35 S]methionine for 3 or 18 h, active synthesis of DBP could not be demonstrated by immunoisolation techniques. These studies demonstrate the presence of DBP on the surface of well washed, human cytotrophoblasts. This DBP may be maternally derived, since active synthesis of DBP could not be demonstrated

  9. Nucleolin: acharan sulfate–binding protein on the surface of cancer cells

    Science.gov (United States)

    Joo, Eun Ji; ten Dam, Gerdy B.; van Kuppevelt, Toin H.; Toida, Toshihiko; Linhardt, Robert J.; Kim, Yeong Shik

    2005-01-01

    Glycosaminoglycans (GAGs) are complex polysaccharides that participate in the regulation of physiological processes through the interactions with a wide variety of proteins. Acharan sulfate (AS), isolated from the giant African snail Achatina fulica, primarily consists of the repeating disaccharide structure α-D-N-acetylglucosaminyl (1→4) 2-sulfoiduronic acid. Exogenous AS was injected subcutaneously near the tumor tissue in C57BL/6 mice that had been implanted with Lewis lung carcinoma cells (LLCs). The location of AS in the tumor was assessed by staining of sectioned tissues with alcian blue and periodic acid–Schiff (PAS) reagent. In vitro assays indicated binding of cells to 50 μg/ml AS (or heparin) after a 5-h incubation. Immunofluorescence assays, using anti-AS antibody, detected AS at the cell surface. The outer-surface of LLCs were next biotinylated to identify the AS-binding proteins. Biotinylated cells were lysed, and the lysates were fractionated on the AS affinity column using a stepwise salt gradient (0, 0.1, 0.3, 0.5, 0.7, 1.0, and 2.0 M). The fractions were analyzed by SDS–PAGE with silver staining and western blotting. We focused on the proteins with high affinity for AS (eluting at 1 M NaCl) and detected only two bands by western blotting. ESI Q-TOF MS analysis of one of these bands, molecular weight ~110 kDa, showed it to be nucleolin. A phosphorylated form of nucleolin on the surface of cells acts as a cell surface receptor for a variety of ligands, including growth factors (i.e., basic fibroblast growth factor) and chemokines (i.e., midkine). These results show that nucleolin is one of several AS-binding proteins and suggest that AS might demonstrate its tumor growth inhibitory activity by binding the nucleolin receptor protein on the surface of cancer cells. PMID:15329357

  10. The Binding of Biotin to Sepharose-Avidin Column: Demonstration of the Affinity Chromatography Technique

    Science.gov (United States)

    Landman, A. D.; Landman, N. N.

    1976-01-01

    Describes a biochemistry experiment that illustrates the methodology of affinity chromatography by attaching avidin, a glycoprotein in egg white, to a Sepharose matrix in order to bind biotin-containing proteins. (MLH)

  11. The Src SH2 domain interacts dynamically with the focal adhesion kinase binding site as demonstrated by paramagnetic NMR spectroscopy.

    Science.gov (United States)

    Lindfors, Hanna E; Drijfhout, Jan Wouter; Ubbink, Marcellus

    2012-06-01

    The interaction between the tyrosine kinases Src and focal adhesion kinase (FAK) is a key step in signaling processes from focal adhesions. The phosphorylated tyrosine residue 397 in FAK is able to bind the Src SH2 domain. To establish the extent of the FAK binding motif, the binding affinity of the SH2 domain for phosphorylated and unphosphorylated FAK-derived peptides of increasing length was determined and compared with that of the internal Src SH2 binding site. It is shown that the FAK peptides have higher affinity than the internal binding site and that seven negative residues adjacent to the core SH2 binding motif increase the binding constant 30-fold. A rigid spin-label incorporated in the FAK peptides was used to establish on the basis of paramagnetic relaxation enhancement whether the peptide-protein complex is well defined. A large spread of the paramagnetic effects on the surface of the SH2 domain suggests that the peptide-protein complex exhibits dynamics, despite the high affinity of the peptide. The strong electrostatic interaction between the positive side of the SH2 domain and the negative peptide results in a high affinity but may also favor a dynamic interaction. Copyright © 2012 Wiley Periodicals, Inc.

  12. Autoradiographic demonstration of oxytocin-binding sites in the macula densa

    International Nuclear Information System (INIS)

    Stoeckel, M.E.; Freund-Mercier, M.J.

    1989-01-01

    Specific oxytocin (OT)-binding sites were localized in the rat kidney with use of a selective 125 I-labeled OT antagonist ( 125 I-OTA). High concentrations of OT binding sites were detected on the juxtaglomerular apparatus with use of the conventional film autoradiographic technique. No labeling occurred on other renal structures. The cellular localization of the OT binding sites within the juxtaglomerular apparatus was studied in light microscope autoradiography, on semithin sections from paraformaldehyde-fixed kidney slices incubated in the presence of 125 I-OTA. These preparations revealed selective labeling of the macula densa, mainly concentrated at the basal pole of the cells. Control experiments showed first that 125 I-OTA binding characteristics were not noticeably altered by prior paraformaldehyde fixation of the kidneys and second that autoradiographic detection of the binding sites was not impaired by histological treatments following binding procedures. In view of the role of the macula densa in the tubuloglomerular feedback, the putative OT receptors of this structure might mediate the stimulatory effect of OT on glomerular filtration

  13. Autoradiographic demonstration of oxytocin-binding sites in the macula densa

    Energy Technology Data Exchange (ETDEWEB)

    Stoeckel, M.E.; Freund-Mercier, M.J. (Centre National de la Recherche Scientifique, Strasbourg (France))

    1989-08-01

    Specific oxytocin (OT)-binding sites were localized in the rat kidney with use of a selective {sup 125}I-labeled OT antagonist ({sup 125}I-OTA). High concentrations of OT binding sites were detected on the juxtaglomerular apparatus with use of the conventional film autoradiographic technique. No labeling occurred on other renal structures. The cellular localization of the OT binding sites within the juxtaglomerular apparatus was studied in light microscope autoradiography, on semithin sections from paraformaldehyde-fixed kidney slices incubated in the presence of {sup 125}I-OTA. These preparations revealed selective labeling of the macula densa, mainly concentrated at the basal pole of the cells. Control experiments showed first that {sup 125}I-OTA binding characteristics were not noticeably altered by prior paraformaldehyde fixation of the kidneys and second that autoradiographic detection of the binding sites was not impaired by histological treatments following binding procedures. In view of the role of the macula densa in the tubuloglomerular feedback, the putative OT receptors of this structure might mediate the stimulatory effect of OT on glomerular filtration.

  14. Structural motif screening reveals a novel, conserved carbohydrate-binding surface in the pathogenesis-related protein PR-5d

    Directory of Open Access Journals (Sweden)

    Moffatt Barbara A

    2010-08-01

    Full Text Available Abstract Background Aromatic amino acids play a critical role in protein-glycan interactions. Clusters of surface aromatic residues and their features may therefore be useful in distinguishing glycan-binding sites as well as predicting novel glycan-binding proteins. In this work, a structural bioinformatics approach was used to screen the Protein Data Bank (PDB for coplanar aromatic motifs similar to those found in known glycan-binding proteins. Results The proteins identified in the screen were significantly associated with carbohydrate-related functions according to gene ontology (GO enrichment analysis, and predicted motifs were found frequently within novel folds and glycan-binding sites not included in the training set. In addition to numerous binding sites predicted in structural genomics proteins of unknown function, one novel prediction was a surface motif (W34/W36/W192 in the tobacco pathogenesis-related protein, PR-5d. Phylogenetic analysis revealed that the surface motif is exclusive to a subfamily of PR-5 proteins from the Solanaceae family of plants, and is absent completely in more distant homologs. To confirm PR-5d's insoluble-polysaccharide binding activity, a cellulose-pulldown assay of tobacco proteins was performed and PR-5d was identified in the cellulose-binding fraction by mass spectrometry. Conclusions Based on the combined results, we propose that the putative binding site in PR-5d may be an evolutionary adaptation of Solanaceae plants including potato, tomato, and tobacco, towards defense against cellulose-containing pathogens such as species of the deadly oomycete genus, Phytophthora. More generally, the results demonstrate that coplanar aromatic clusters on protein surfaces are a structural signature of glycan-binding proteins, and can be used to computationally predict novel glycan-binding proteins from 3 D structure.

  15. Structural motif screening reveals a novel, conserved carbohydrate-binding surface in the pathogenesis-related protein PR-5d.

    Science.gov (United States)

    Doxey, Andrew C; Cheng, Zhenyu; Moffatt, Barbara A; McConkey, Brendan J

    2010-08-03

    Aromatic amino acids play a critical role in protein-glycan interactions. Clusters of surface aromatic residues and their features may therefore be useful in distinguishing glycan-binding sites as well as predicting novel glycan-binding proteins. In this work, a structural bioinformatics approach was used to screen the Protein Data Bank (PDB) for coplanar aromatic motifs similar to those found in known glycan-binding proteins. The proteins identified in the screen were significantly associated with carbohydrate-related functions according to gene ontology (GO) enrichment analysis, and predicted motifs were found frequently within novel folds and glycan-binding sites not included in the training set. In addition to numerous binding sites predicted in structural genomics proteins of unknown function, one novel prediction was a surface motif (W34/W36/W192) in the tobacco pathogenesis-related protein, PR-5d. Phylogenetic analysis revealed that the surface motif is exclusive to a subfamily of PR-5 proteins from the Solanaceae family of plants, and is absent completely in more distant homologs. To confirm PR-5d's insoluble-polysaccharide binding activity, a cellulose-pulldown assay of tobacco proteins was performed and PR-5d was identified in the cellulose-binding fraction by mass spectrometry. Based on the combined results, we propose that the putative binding site in PR-5d may be an evolutionary adaptation of Solanaceae plants including potato, tomato, and tobacco, towards defense against cellulose-containing pathogens such as species of the deadly oomycete genus, Phytophthora. More generally, the results demonstrate that coplanar aromatic clusters on protein surfaces are a structural signature of glycan-binding proteins, and can be used to computationally predict novel glycan-binding proteins from 3 D structure.

  16. Sugar-binding sites on the surface of the carbohydrate-binding module of CBH I from Trichoderma reesei.

    Science.gov (United States)

    Tavagnacco, Letizia; Mason, Philip E; Schnupf, Udo; Pitici, Felicia; Zhong, Linghao; Himmel, Michael E; Crowley, Michael; Cesàro, Attilio; Brady, John W

    2011-05-01

    Molecular dynamics simulations were carried out for a system consisting of the carbohydrate-binding module (CBM) of the cellulase CBH I from Trichoderma reesei (Hypocrea jecorina) in a concentrated solution of β-D-glucopyranose, to determine whether there is any tendency for the sugar molecules to bind to the CBM. In spite of the general tendency of glucose to behave as an osmolyte, a marked tendency for the sugar molecules to bind to the protein was observed. However, the glucose molecules tended to bind only to specific sites on the protein. As expected, the hydrophobic face of the sugar molecules, comprising the axial H1, H3, and H5 aliphatic protons, tended to adhere to the flat faces of the three tyrosine side chains on the planar binding surface of the CBM. However, a significant tendency to bind to a groove-like feature on the upper surface of the CBM was also observed. These results would not be inconsistent with a model of the mechanism for this globular domain in which the cellodextrin chain being removed from the surface of crystalline cellulose passes over the upper surface of the CBM, presumably then available for hydrolysis in the active site tunnel of this processive cellulase. Copyright © 2011 Elsevier Ltd. All rights reserved.

  17. TAF(II)170 interacts with the concave surface of TATA-binding protein to inhibit its DNA binding activity.

    Science.gov (United States)

    Pereira, L A; van der Knaap, J A; van den Boom, V; van den Heuvel, F A; Timmers, H T

    2001-11-01

    The human RNA polymerase II transcription factor B-TFIID consists of TATA-binding protein (TBP) and the TBP-associated factor (TAF) TAF(II)170 and can rapidly redistribute over promoter DNA. Here we report the identification of human TBP-binding regions in human TAF(II)170. We have defined the TBP interaction domain of TAF(II)170 within three amino-terminal regions: residues 2 to 137, 290 to 381, and 380 to 460. Each region contains a pair of Huntington-elongation-A subunit-Tor repeats and exhibits species-specific interactions with TBP family members. Remarkably, the altered-specificity TBP mutant (TBP(AS)) containing a triple mutation in the concave surface is defective for binding the TAF(II)170 amino-terminal region of residues 1 to 504. Furthermore, within this region the TAF(II)170 residues 290 to 381 can inhibit the interaction between Drosophila TAF(II)230 (residues 2 to 81) and TBP through competition for the concave surface of TBP. Biochemical analyses of TBP binding to the TATA box indicated that TAF(II)170 region 290-381 inhibits TBP-DNA complex formation. Importantly, the TBP(AS) mutant is less sensitive to TAF(II)170 inhibition. Collectively, our results support a mechanism in which TAF(II)170 induces high-mobility DNA binding by TBP through reversible interactions with its concave DNA binding surface.

  18. Many-body dispersion effects in the binding of adsorbates on metal surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Maurer, Reinhard J. [Department of Chemistry, Yale University, New Haven, Connecticut 06520 (United States); Ruiz, Victor G.; Tkatchenko, Alexandre [Fritz-Haber-Institut der Max-Planck-Gesellschaft, Faradayweg 4-6, D-14195 Berlin (Germany)

    2015-09-14

    A correct description of electronic exchange and correlation effects for molecules in contact with extended (metal) surfaces is a challenging task for first-principles modeling. In this work, we demonstrate the importance of collective van der Waals dispersion effects beyond the pairwise approximation for organic–inorganic systems on the example of atoms, molecules, and nanostructures adsorbed on metals. We use the recently developed many-body dispersion (MBD) approach in the context of density-functional theory [Tkatchenko et al., Phys. Rev. Lett. 108, 236402 (2012) and Ambrosetti et al., J. Chem. Phys. 140, 18A508 (2014)] and assess its ability to correctly describe the binding of adsorbates on metal surfaces. We briefly review the MBD method and highlight its similarities to quantum-chemical approaches to electron correlation in a quasiparticle picture. In particular, we study the binding properties of xenon, 3,4,9,10-perylene-tetracarboxylic acid, and a graphene sheet adsorbed on the Ag(111) surface. Accounting for MBD effects, we are able to describe changes in the anisotropic polarizability tensor, improve the description of adsorbate vibrations, and correctly capture the adsorbate–surface interaction screening. Comparison to other methods and experiment reveals that inclusion of MBD effects improves adsorption energies and geometries, by reducing the overbinding typically found in pairwise additive dispersion-correction approaches.

  19. Surface passivation for tight-binding calculations of covalent solids

    International Nuclear Information System (INIS)

    Bernstein, N

    2007-01-01

    Simulation of a cluster representing a finite portion of a larger covalently bonded system requires the passivation of the cluster surface. We compute the effects of an explicit hybrid orbital passivation (EHOP) on the atomic structure in a model bulk, three-dimensional, narrow gap semiconductor, which is very different from the wide gap, quasi-one-dimensional organic molecules where most passivation schemes have been studied in detail. The EHOP approach is directly applicable to minimal atomic orbital basis methods such as tight-binding. Each broken bond is passivated by a hybrid created from an explicitly expressed linear combination of basis orbitals, chosen to represent the contribution of the missing neighbour, e.g. a sp 3 hybrid for a single bond. The method is tested by computing the forces on atoms near a point defect as a function of cluster geometry. We show that, compared to alternatives such as pseudo-hydrogen passivation, the force on an atom converges to the correct bulk limit more quickly as a function of cluster radius, and that the force is more stable with respect to perturbations in the position of the cluster centre. The EHOP method also obviates the need for parameterizing the interactions between the system atoms and the passivating atoms. The method is useful for cluster calculations of non-periodic defects in large systems and for hybrid schemes that simulate large systems by treating finite regions with a quantum-mechanical model, coupled to an interatomic potential description of the rest of the system

  20. Surface displaced alfa-enolase of Lactobacillus plantarum is a fibronectin binding protein

    Directory of Open Access Journals (Sweden)

    Muscariello Lidia

    2009-02-01

    Full Text Available Abstract Background Lactic acid bacteria of the genus Lactobacillus and Bifidobacterium are one of the most important health promoting groups of the human intestinal microbiota. Their protective role within the gut consists in out competing invading pathogens for ecological niches and metabolic substrates. Among the features necessary to provide health benefits, commensal microorganisms must have the ability to adhere to human intestinal cells and consequently to colonize the gut. Studies on mechanisms mediating adhesion of lactobacilli to human intestinal cells showed that factors involved in the interaction vary mostly among different species and strains, mainly regarding interaction between bacterial adhesins and extracellular matrix or mucus proteins. We have investigated the adhesive properties of Lactobacillus plantarum, a member of the human microbiota of healthy individuals. Results We show the identification of a Lactobacillus plantarum LM3 cell surface protein (48 kDa, which specifically binds to human fibronectin (Fn, an extracellular matrix protein. By means of mass spectrometric analysis this protein was identified as the product of the L. plantarum enoA1 gene, coding the EnoA1 alfa-enolase. Surface localization of EnoA1 was proved by immune electron microscopy. In the mutant strain LM3-CC1, carrying the enoA1 null mutation, the 48 kDa adhesin was not anymore detectable neither by anti-enolase Western blot nor by Fn-overlay immunoblotting assay. Moreover, by an adhesion assay we show that LM3-CC1 cells bind to fibronectin-coated surfaces less efficiently than wild type cells, thus demonstrating the significance of the surface displaced EnoA1 protein for the L. plantarum LM3 adhesion to fibronectin. Conclusion Adhesion to host tissues represents a crucial early step in the colonization process of either pathogens or commensal bacteria. We demonstrated the involvement of the L. plantarum Eno A1 alfa-enolase in Fn-binding, by studying

  1. Aflatoxin Toxicity Reduction in Feed by Enhanced Binding to Surface-Modified Clay Additives

    Science.gov (United States)

    Jaynes, William F.; Zartman, Richard E.

    2011-01-01

    Animal feeding studies have demonstrated that clay additives, such as bentonites, can bind aflatoxins in ingested feed and reduce or eliminate the toxicity. Bentonite deposits are found throughout the world and mostly consist of expandable smectite minerals, such as montmorillonite. The surfaces of smectite minerals can be treated with organic compounds to create surface-modified clays that more readily bind some contaminants than the untreated clay. Montmorillonites treated with organic cations, such as hexadecyltrimethylammonium (HDTMA) and phenyltrimethylammonium (PTMA), more effectively remove organic contaminants, such as benzene and toluene, from water than untreated clay. Similarly, montmorillonite treated with PTMA (Kd = 24,100) retained more aflatoxin B1 (AfB1) from aqueous corn flour than untreated montmorillonite (Kd = 944). Feed additives that reduced aflatoxin toxicity in animal feeding studies adsorbed more AfB1 from aqueous corn flour than feed additives that were less effective. The organic cations HDTMA and PTMA are considered toxic and would not be suitable for clay additives used in feed or food, but other non-toxic or nutrient compounds can be used to prepare surface-modified clays. Montmorillonite (SWy) treated with choline (Kd = 13,800) and carnitine (Kd = 3960) adsorbed much more AfB1 from aqueous corn flour than the untreated clay (Kd = 944). A choline-treated clay prepared from a reduced-charge, high-charge montmorillonite (Kd = 20,100) adsorbed more AfB1 than the choline-treated high-charge montmorillonite (Kd = 1340) or the untreated montmorillonite (Kd = 293). Surface-modified clay additives prepared using low-charge smectites and nutrient or non-toxic organic compounds might be used to more effectively bind aflatoxins in contaminated feed or food and prevent toxicity. PMID:22069725

  2. Binding modes and functional surface of anti-mammalian scorpion α-toxins to sodium channels.

    Science.gov (United States)

    Chen, Rong; Chung, Shin-Ho

    2012-10-02

    Scorpion α-toxins bind to the voltage-sensing domains of voltage-gated sodium (Na(V)) channels and interfere with the inactivation mechanisms. The functional surface of α-toxins has been shown to contain an NC-domain consisting of the five-residue turn (positions 8-12) and the C-terminus (positions 56-64) and a core-domain centered on the residue 18. The NC- and core-domains are interconnected by the linker-domain (positions 8-18). Here with atomistic molecular dynamics simulations, we examine the binding modes between two α-toxins, the anti-mammalian AahII and the anti-insect LqhαIT, and the voltage-sensing domain of rat Na(V)1.2, a subtype of Na(V) channels expressed in nerve cells. Both toxins are docked to the extracellular side of the voltage-sensing domain of Na(V)1.2 using molecular dynamics simulations, with the linker-domain assumed to wedge into the binding pocket. Several salt bridges and hydrophobic clusters are observed to form between the NC- and core-domains of the toxins and Na(V)1.2 and stabilize the toxin-channel complexes. The binding modes predicted are consistent with available mutagenesis data and can readily explain the relative affinities of AahII and LqhαIT for Na(V)1.2. The dissociation constants for the two toxin-channel complexes are derived, which compare favorably with experiment. Our models demonstrate that the functional surface of anti-mammalian scorpion α-toxins is centered on the linker-domain, similar to that of β-toxins.

  3. Detection of Biomolecular Binding Through Enhancement of Localized Surface Plasmon Resonance (LSPR by Gold Nanoparticles

    Directory of Open Access Journals (Sweden)

    Min-Gon Kim

    2009-03-01

    Full Text Available To amplify the difference in localized surface plasmon resonance (LSPR spectra of gold nano-islands due to intermolecular binding events, gold nanoparticles were used. LSPR-based optical biosensors consisting of gold nano-islands were readily made on glass substrates using evaporation and heat treatment. Streptavidin (STA and biotinylated bovine serum albumin (Bio-BSA were chosen as the model receptor and the model analyte, respectively, to demonstrate the effectiveness of this detection method. Using this model system, we were able to enhance the sensitivity in monitoring the binding of Bio-BSA to gold nano-island surfaces functionalized with STA through the addition of gold nanoparticle-STA conjugates. In addition, SU-8 well chips with gold nano-island surfaces were fabricated through a conventional UV patterning method and were then utilized for image detection using the attenuated total reflection mode. These results suggest that the gold nano-island well chip may have the potential to be used for multiple and simultaneous detection of various bio-substances.

  4. Selective Binding, Self-Assembly and Nanopatterning of the Creutz-Taube Ion on Surfaces

    Directory of Open Access Journals (Sweden)

    Qingling Hang

    2009-02-01

    Full Text Available The surface attachment properties of the Creutz-Taube ion, i.e., [(NH35Ru(pyrazineRu(NH35]5+, on both hydrophilic and hydrophobic types of surfaces were investigated using X-ray photoelectron spectroscopy (XPS. The results indicated that the Creutz-Taube ions only bound to hydrophilic surfaces, such as SiO2 and –OH terminated organic SAMs on gold substrates. No attachment of the ions on hydrophobic surfaces such as –CH3 terminated organic SAMs and poly(methylmethacrylate (PMMA thin films covered gold or SiO2 substrates was observed. Further ellipsometric, atomic force microscopy (AFM and time-dependent XPS studies suggested that the attached cations could form an inorganic analog of the self-assembled monolayer on SiO2 substrate with a “lying-down” orientation. The strong electrostatic interaction between the highly charged cations and the anionic SiO2 surface was believed to account for these observations. Based on its selective binding property, patterning of wide (~200 nm and narrow (~35 nm lines of the Creutz-Taube ions on SiO2 surface were demonstrated through PMMA electron resist masks written by electron beam lithography (EBL.

  5. In vivo autoradiographic demonstration of β-adrenergic binding sites in adult rat type II alveolar epithelial cells

    International Nuclear Information System (INIS)

    Smith, D.M.; Sidhu, M.K.

    1984-01-01

    Adult male rats were injected intravenously with the muscarinic binding probe 3 H-Quinuclidinyl benzilate (QNB) or the β-adrenergic probe 3 H-dihydroalprenolol (DHA). Other rats were pre-treated with an intraperitoneal injection of a 500-fold excess of L-isoproterenol prior to the DHA. Light microscopic autoradiography of 0.5 μm sections of lung from the QNB group demonstrated very little labelling even after 6 months of exposure. In constrast, trachealis smooth muscle from these animals contained substantial labelling. Autoradiographs of lung from rats injected with DHA demonstrated labelling which was well localized over alveolar septa and concentrated over the cytoplasm of type II cells. Quantitative analysis of labelling in the DHA groups indicated a significant reduction of labelling in animals treated with L-isoproterenol prior to DHA, in both the alveolar parenchyma in general and over type II cells. The results of this study provide morphologic evidence for the uptake and specific binding of β-adrenergic antagonists by the adult lung in vivo, while failing to demonstrate similar binding of a muscarinic probe. In addition, the results demonstrate specific β-adrenergic receptors on type II cells in vivo and substantiate the view of a direct effect of β-adrenergic agonists on alveolar type II cells

  6. Demonstration of epidermal growth factor binding sites in the adult rat pancreas by light microscopic autoradiography

    International Nuclear Information System (INIS)

    Chabot, J.G.; Walker, P.; Pelletier, G.

    1987-01-01

    The distribution of epidermal growth factor (EGF) receptors was studied in the pancreas using light microscopic autoradiography, which was performed at different time intervals (2-60 min) after injecting 125 I-labeled EGF intravenously into the adult rat. In the exocrine pancreas, a labeling was found to occur over the pyramidal cells of the acini and cells lining the intercalated ducts. Moreover, substantial binding of EGF to cells of the islets of Langerhans was also revealed. At the 2-min time interval, most silver grains were found at the periphery of the target cells. The localization, as well as the diminution of silver grains over the cytoplasm of these cells, between 7 and 60 min, suggested the internalization and degradation of 125 I-labeled EGF. Control experiments indicated that the autoradiography reaction was due to specific interaction of 125 I-labeled EGF with its receptor. These results clearly indicate that EGF receptors are present in the acinar cells and the cells of intercalated ducts of the exocrine pancreas, as well as the cells of the endocrine pancreas. Finding that there are EGF binding sites in pancreatic acinar cells supports the physiological role of EGF in the regulation of pancreatic exocrine function. The presence of EGF receptors in cells of the islets of Langerhans suggests that EGF may play a role in the regulation of the endocrine pancreas

  7. Tailoring odorant-binding protein coatings characteristics for surface acoustic wave biosensor development

    Energy Technology Data Exchange (ETDEWEB)

    Di Pietrantonio, F., E-mail: fabio.dp@idasc.cnr.it [Institute of Acoustics and Sensors “O. M. Corbino”, National Research Council of Italy, Via del Fosso del Cavaliere 100, 00133 Rome (Italy); Benetti, M. [Institute of Acoustics and Sensors “O. M. Corbino”, National Research Council of Italy, Via del Fosso del Cavaliere 100, 00133 Rome (Italy); Dinca, V. [National Institute for Lasers, Plasma and Radiation Physics, 409 Atomistilor Street, PO Box MG-16, 077125 Magurele (Romania); Cannatà, D. [Institute of Acoustics and Sensors “O. M. Corbino”, National Research Council of Italy, Via del Fosso del Cavaliere 100, 00133 Rome (Italy); Verona, E. [Institute for Photonics and Nanotechnologies, National Research Council of Italy, Via del Cineto Romano 42, 00156 Rome (Italy); D’Auria, S. [Institute of Protein Biochemistry, National Research Council of Italy, Via Pietro Castellino 111, 80131 Naples (Italy); Dinescu, M. [National Institute for Lasers, Plasma and Radiation Physics, 409 Atomistilor Street, PO Box MG-16, 077125 Magurele (Romania)

    2014-05-01

    In this study, wild type bovine odorant-binding proteins (wtbOBPs) were deposited by matrix-assisted pulsed laser evaporation (MAPLE) and utilized as active material on surface acoustic wave (SAW) biosensors. Fourier transform infrared spectroscopy (FTIR), and atomic force microscopy (AFM) were used to determine the chemical, morphological characteristics of the protein thin films. The FTIR data demonstrates that the functional groups of wtbOBPs do not suffer significant changes in the MAPLE-deposited films when compared to the reference one. The topographical studies show that the homogeneity, density and the roughness of the coatings are related mainly to the laser parameters (fluence and number of pulses). SAW biosensor responses to different concentrations of R-(–)-1-octen-3-ol (octenol) and R-(–)-carvone (carvone) were evaluated. The obtained sensitivities, achieved through the optimization of deposition parameters, demonstrated that MAPLE is a promising deposition technique for SAW biosensor implementation.

  8. Adherence of platelets to in situ albumin-binding surfaces under flow conditions: role of surface-adsorbed albumin

    International Nuclear Information System (INIS)

    Guha Thakurta, Sanjukta; Miller, Robert; Subramanian, Anuradha

    2012-01-01

    Surfaces that preferentially bind human serum albumin (HSA) were generated by grafting albumin-binding linear peptide (LP1) onto silicon surfaces. The research aim was to evaluate the adsorption pattern of proteins and the adhesion of platelets from platelet-poor plasma and platelet-rich plasma, respectively, by albumin-binding surfaces under physiological shear rate (96 and 319 s −1 ) conditions. Bound proteins were quantified using enzyme-linked immunosorbent assays (ELISAs) and two-dimensional gel electrophoresis. A ratio of ∼1000:100:1 of adsorbed HSA, human immunoglobulin (HIgG) and human fibrinogen (HFib) was noted, respectively, on LP1-functionalized surfaces, and a ratio of ∼5:2:1 of the same was noted on control surfaces, as confirmed by ELISAs. The surface-adsorbed von Willebrand factor was undetectable by sensitive ELISAs. The amount of adhered platelets correlated with the ratio of adsorbed HSA/HFib. Platelet morphology was more rounded on LP1-functionalized surfaces when compared to control surfaces. The platelet adhesion response on albumin-binding surfaces can be explained by the reduction in the co-adsorption of other plasma proteins in a surface environment where there is an excess of albumin molecules, coupled with restrictions in the conformational transitions of other surface-adsorbed proteins into hemostatically active forms. (paper)

  9. Binding Ligand Prediction for Proteins Using Partial Matching of Local Surface Patches

    Directory of Open Access Journals (Sweden)

    Lee Sael

    2010-12-01

    Full Text Available Functional elucidation of uncharacterized protein structures is an important task in bioinformatics. We report our new approach for structure-based function prediction which captures local surface features of ligand binding pockets. Function of proteins, specifically, binding ligands of proteins, can be predicted by finding similar local surface regions of known proteins. To enable partial comparison of binding sites in proteins, a weighted bipartite matching algorithm is used to match pairs of surface patches. The surface patches are encoded with the 3D Zernike descriptors. Unlike the existing methods which compare global characteristics of the protein fold or the global pocket shape, the local surface patch method can find functional similarity between non-homologous proteins and binding pockets for flexible ligand molecules. The proposed method improves prediction results over global pocket shape-based method which was previously developed by our group.

  10. Binding ligand prediction for proteins using partial matching of local surface patches.

    Science.gov (United States)

    Sael, Lee; Kihara, Daisuke

    2010-01-01

    Functional elucidation of uncharacterized protein structures is an important task in bioinformatics. We report our new approach for structure-based function prediction which captures local surface features of ligand binding pockets. Function of proteins, specifically, binding ligands of proteins, can be predicted by finding similar local surface regions of known proteins. To enable partial comparison of binding sites in proteins, a weighted bipartite matching algorithm is used to match pairs of surface patches. The surface patches are encoded with the 3D Zernike descriptors. Unlike the existing methods which compare global characteristics of the protein fold or the global pocket shape, the local surface patch method can find functional similarity between non-homologous proteins and binding pockets for flexible ligand molecules. The proposed method improves prediction results over global pocket shape-based method which was previously developed by our group.

  11. Distinctive receptor binding properties of the surface glycoprotein of a natural Feline Leukemia Virus isolate with unusual disease spectrum

    Directory of Open Access Journals (Sweden)

    Albritton Lorraine M

    2011-05-01

    Full Text Available Abstract Background Feline leukemia virus (FeLV-945, a member of the FeLV-A subgroup, was previously isolated from a cohort of naturally infected cats. An unusual multicentric lymphoma of non-T-cell origin was observed in natural and experimental infection with FeLV-945. Previous studies implicated the FeLV-945 surface glycoprotein (SU as a determinant of disease outcome by an as yet unknown mechanism. The present studies demonstrate that FeLV-945 SU confers distinctive properties of binding to the cell surface receptor. Results Virions bearing the FeLV-945 Env protein were observed to bind the cell surface receptor with significantly increased efficiency, as was soluble FeLV-945 SU protein, as compared to the corresponding virions or soluble protein from a prototype FeLV-A isolate. SU proteins cloned from other cohort isolates exhibited increased binding efficiency comparable to or greater than FeLV-945 SU. Mutational analysis implicated a domain containing variable region B (VRB to be the major determinant of increased receptor binding, and identified a single residue, valine 186, to be responsible for the effect. Conclusions The FeLV-945 SU protein binds its cell surface receptor, feTHTR1, with significantly greater efficiency than does that of prototype FeLV-A (FeLV-A/61E when present on the surface of virus particles or in soluble form, demonstrating a 2-fold difference in the relative dissociation constant. The results implicate a single residue, valine 186, as the major determinant of increased binding affinity. Computational modeling suggests a molecular mechanism by which residue 186 interacts with the receptor-binding domain through residue glutamine 110 to effect increased binding affinity. Through its increased receptor binding affinity, FeLV-945 SU might function in pathogenesis by increasing the rate of virus entry and spread in vivo, or by facilitating entry into a novel target cell with a low receptor density.

  12. Surface binding sites in carbohydrate active enzymes: An emerging picture of structural and functional diversity

    DEFF Research Database (Denmark)

    Svensson, Birte; Cockburn, Darrell

    2013-01-01

    is not universal and is in fact rare among some families of enzymes. In some cases an alternative to possessing a CBM is for the enzyme to bind to the substrate at a site on the catalytic domain, but away from the active site. Such a site is termed a surface (or secondary) binding site (SBS). SBSs have been...

  13. Human IgG lacking effector functions demonstrate lower FcRn-binding and reduced transplacental transport.

    Science.gov (United States)

    Stapleton, Nigel M; Armstrong-Fisher, Sylvia S; Andersen, Jan Terje; van der Schoot, C Ellen; Porter, Charlene; Page, Kenneth R; Falconer, Donald; de Haas, Masja; Williamson, Lorna M; Clark, Michael R; Vidarsson, Gestur; Armour, Kathryn L

    2018-03-01

    We have previously generated human IgG1 antibodies that were engineered for reduced binding to the classical Fcγ receptors (FcγRI-III) and C1q, thereby eliminating their destructive effector functions (constant region G1Δnab). In their potential use as blocking agents, favorable binding to the neonatal Fc receptor (FcRn) is important to preserve the long half-life typical of IgG. An ability to cross the placenta, which is also mediated, at least in part, by FcRn is desirable in some indications, such as feto-maternal alloimmune disorders. Here, we show that G1Δnab mutants retain pH-dependent binding to human FcRn but that the amino acid alterations reduce the affinity of the IgG1:FcRn interaction by 2.0-fold and 1.6-fold for the two antibodies investigated. The transport of the modified G1Δnab mutants across monolayers of human cell lines expressing FcRn was approximately 75% of the wild-type, except that no difference was observed with human umbilical vein endothelial cells. G1Δnab mutation also reduced transport in an ex vivo placenta model. In conclusion, we demonstrate that, although the G1Δnab mutations are away from the FcRn-binding site, they have long-distance effects, modulating FcRn binding and transcellular transport. Our findings have implications for the design of therapeutic human IgG with tailored effector functions. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. An Allergen Portrait Gallery: Representative Structures and an Overview of IgE Binding Surfaces

    Directory of Open Access Journals (Sweden)

    Ovidiu Ivanciuc

    2010-10-01

    Full Text Available Recent progress in the biochemical classification and structural determination of allergens and allergen–antibody complexes has enhanced our understanding of the molecular determinants of allergenicity. Databases of allergens and their epitopes have facilitated the clustering of allergens according to their sequences and, more recently, their structures. Groups of similar sequences are identified for allergenic proteins from diverse sources, and all allergens are classified into a limited number of protein structural families. A gallery of experimental structures selected from the protein classes with the largest number of allergens demonstrate the structural diversity of the allergen universe. Further comparison of these structures and identification of areas that are different from innocuous proteins within the same protein family can be used to identify features specific to known allergens. Experimental and computational results related to the determination of IgE binding surfaces and methods to define allergen-specific motifs are highlighted.

  15. Co-binding of pharmaceutical compounds at mineral surfaces: Molecular investigations of dimer formation at goethite/water interfaces

    OpenAIRE

    Xu , Jing; Marsac , Rémi; Costa , Dominique; Cheng , Wei; Wu , Feng; Boily , Jean-François; Hanna , Khalil

    2017-01-01

    International audience; The emergence of antibiotic and anti-inflammatory agents in aquatic and terrestrial systems is becoming a serious threat to human and animal health worldwide. Because pharmaceutical compounds rarely exist individually in nature, interactions between various compounds can have unforeseen effects on their binding to mineral surfaces. This work demonstrates this important possibility for the case of two typical antibiotic and anti-inflammatory agents (nalidixic acid (NA) ...

  16. Affinity purification of human granulocyte macrophage colony-stimulating factor receptor alpha-chain. Demonstration of binding by photoaffinity labeling

    International Nuclear Information System (INIS)

    Chiba, S.; Shibuya, K.; Miyazono, K.; Tojo, A.; Oka, Y.; Miyagawa, K.; Takaku, F.

    1990-01-01

    The human granulocyte macrophage colony-stimulating factor (GM-CSF) receptor alpha-chain, a low affinity component of the receptor, was solubilized and affinity-purified from human placenta using biotinylated GM-CSF. Scatchard analysis of 125 I-GM-CSF binding to the placental membrane extract disclosed that the GM-CSF receptor had a dissociation constant (Kd) of 0.5-0.8 nM, corresponding to the Kd value of the GM-CSF receptor alpha-chain on the intact placental membrane. Affinity labeling of the solubilized protein using a photoreactive cross-linking agent, N-hydroxysuccinimidyl-4-azidobenzoate (HSAB), demonstrated a single specific band of 70-95 kDa representing a ligand-receptor complex. Approximately 2 g of the placental membrane extract was subjected to a biotinylated GM-CSF-fixed streptavidin-agarose column, resulting in a single major band at 70 kDa on a silver-stained sodium dodecyl sulfate gel. The radioiodination for the purified material disclosed that the purified protein had an approximate molecular mass of 70 kDa and a pI of 6.6. Binding activity of the purified material was demonstrated by photoaffinity labeling using HSAB- 125 I-GM-CSF, producing a similar specific band at 70-95 kDa as was demonstrated for the crude protein

  17. Visualisation of variable binding pockets on protein surfaces by probabilistic analysis of related structure sets

    Directory of Open Access Journals (Sweden)

    Ashford Paul

    2012-03-01

    Full Text Available Abstract Background Protein structures provide a valuable resource for rational drug design. For a protein with no known ligand, computational tools can predict surface pockets that are of suitable size and shape to accommodate a complementary small-molecule drug. However, pocket prediction against single static structures may miss features of pockets that arise from proteins' dynamic behaviour. In particular, ligand-binding conformations can be observed as transiently populated states of the apo protein, so it is possible to gain insight into ligand-bound forms by considering conformational variation in apo proteins. This variation can be explored by considering sets of related structures: computationally generated conformers, solution NMR ensembles, multiple crystal structures, homologues or homology models. It is non-trivial to compare pockets, either from different programs or across sets of structures. For a single structure, difficulties arise in defining particular pocket's boundaries. For a set of conformationally distinct structures the challenge is how to make reasonable comparisons between them given that a perfect structural alignment is not possible. Results We have developed a computational method, Provar, that provides a consistent representation of predicted binding pockets across sets of related protein structures. The outputs are probabilities that each atom or residue of the protein borders a predicted pocket. These probabilities can be readily visualised on a protein using existing molecular graphics software. We show how Provar simplifies comparison of the outputs of different pocket prediction algorithms, of pockets across multiple simulated conformations and between homologous structures. We demonstrate the benefits of use of multiple structures for protein-ligand and protein-protein interface analysis on a set of complexes and consider three case studies in detail: i analysis of a kinase superfamily highlights the

  18. Visualisation of variable binding pockets on protein surfaces by probabilistic analysis of related structure sets.

    Science.gov (United States)

    Ashford, Paul; Moss, David S; Alex, Alexander; Yeap, Siew K; Povia, Alice; Nobeli, Irene; Williams, Mark A

    2012-03-14

    Protein structures provide a valuable resource for rational drug design. For a protein with no known ligand, computational tools can predict surface pockets that are of suitable size and shape to accommodate a complementary small-molecule drug. However, pocket prediction against single static structures may miss features of pockets that arise from proteins' dynamic behaviour. In particular, ligand-binding conformations can be observed as transiently populated states of the apo protein, so it is possible to gain insight into ligand-bound forms by considering conformational variation in apo proteins. This variation can be explored by considering sets of related structures: computationally generated conformers, solution NMR ensembles, multiple crystal structures, homologues or homology models. It is non-trivial to compare pockets, either from different programs or across sets of structures. For a single structure, difficulties arise in defining particular pocket's boundaries. For a set of conformationally distinct structures the challenge is how to make reasonable comparisons between them given that a perfect structural alignment is not possible. We have developed a computational method, Provar, that provides a consistent representation of predicted binding pockets across sets of related protein structures. The outputs are probabilities that each atom or residue of the protein borders a predicted pocket. These probabilities can be readily visualised on a protein using existing molecular graphics software. We show how Provar simplifies comparison of the outputs of different pocket prediction algorithms, of pockets across multiple simulated conformations and between homologous structures. We demonstrate the benefits of use of multiple structures for protein-ligand and protein-protein interface analysis on a set of complexes and consider three case studies in detail: i) analysis of a kinase superfamily highlights the conserved occurrence of surface pockets at the active

  19. Glycosaminoglycans mediate retention of the poxvirus type I interferon binding protein at the cell surface to locally block interferon antiviral responses

    Science.gov (United States)

    Montanuy, Imma; Alejo, Ali; Alcami, Antonio

    2011-01-01

    Eradication of smallpox was accomplished 30 yr ago, but poxviral infections still represent a public health concern due to the potential release of variola virus or the emergence of zoonotic poxviruses, such as monkeypox virus. A critical determinant of poxvirus virulence is the inhibition of interferons (IFNs) by the virus-encoded type I IFN-binding protein (IFNα/βBP). This immunomodulatory protein is secreted and has the unique property of interacting with the cell surface in order to prevent IFN-mediated antiviral responses. However, the mechanism of its attachment to the cell surface remains unknown. Using surface plasmon resonance and cell-binding assays, we report that the IFNα/βBP from vaccinia virus, the smallpox vaccine, interacts with cell surface glycosaminoglycans (GAGs). Analysis of the contribution of different regions of the protein to cell surface binding demonstrated that clusters of basic residues in the first immunoglobulin domain mediate GAG interactions. Furthermore, mutation of the GAG-interaction motifs does not affect its IFN-binding and -blocking capacity. Functional conservation of GAG-binding sites is demonstrated for the IFNα/βBP from variola and monkeypox viruses, extending our understanding of immune modulation by the most virulent human poxviruses. These results are relevant for the design of improved vaccines and intervention strategies.—Montanuy, I., Alejo, A., Alcami, A. Glycosaminoglycans mediate retention of the poxvirus type I interferon binding protein at the cell surface to locally block interferon antiviral responses. PMID:21372110

  20. The influence of surface integrin binding patterns on specific biomaterial-cell interactions

    Science.gov (United States)

    Beranek, Maggi Marie

    As the future of biomaterials progresses toward bioactivity, the biomaterial surface must control non-specific protein adsorption and encourage selective protein and cell adsorption. Integrins alphavbeta3, alpha 1beta1, alpha5beta1 and alpha Mbeta2 are expressed on cells involved in endothelialization, inflammation, and intimal hyperplasia. These cellular events play a vital role in biomaterial biocompatibility, especially in the vascular environment. The overall hypothesis of these studies is that biomaterial surfaces exhibit selective integrin binding, which then specifies differential cell binding. To test this hypothesis, four specific aims were developed. The first aim was designed to determine whether metal and polymeric biomaterials exhibit selective integrin binding. The tested materials included 316L stainless steel, nitinol, gold, Elgiloy RTM, poly(D, L-lactide-co-glycolide), polycarbonate urethane and expanded polytetrafluoroethylene. Discrete integrin binding patterns were detected microscopically using integrin specific fluorescent antibodies. Stainless steel exhibited high level integrin alpha1beta 1 and low level integrin alphaMbeta2 binding pattern. This suggests that this metal surface should selectively encourage endothelial cell to inflammatory cell binding. In contrast, gold bound ten times the amount of integrin alphaMbeta2 compared to integrin alpha1beta1, which should encourage inflammatory cell adhesion. The 65/35 poly(D, L-lactide-co-glycolide) was the only polymeric biomaterial tested that had integrin binding levels comparable to metal biomaterials. Based on these observations, a combinational biomaterial with a surface pattern of 65/35 poly(D, L-lactide-co-glycolide) dots on a 316L stainless steel background was created. A pattern of high level integrin alpha1beta1 binding and low level integrin alpha Mbeta2 binding on this combinational surface indicates that this surface should selectively favor endothelial cell binding. In the second

  1. Reverse Non-Equilibrium Molecular Dynamics Demonstrate That Surface Passivation Controls Thermal Transport at Semiconductor-Solvent Interfaces.

    Science.gov (United States)

    Hannah, Daniel C; Gezelter, J Daniel; Schaller, Richard D; Schatz, George C

    2015-06-23

    We examine the role played by surface structure and passivation in thermal transport at semiconductor/organic interfaces. Such interfaces dominate thermal transport in semiconductor nanomaterials owing to material dimensions much smaller than the bulk phonon mean free path. Utilizing reverse nonequilibrium molecular dynamics simulations, we calculate the interfacial thermal conductance (G) between a hexane solvent and chemically passivated wurtzite CdSe surfaces. In particular, we examine the dependence of G on the CdSe slab thickness, the particular exposed crystal facet, and the extent of surface passivation. Our results indicate a nonmonotonic dependence of G on ligand-grafting density, with interfaces generally exhibiting higher thermal conductance for increasing surface coverage up to ∼0.08 ligands/Å(2) (75-100% of a monolayer, depending on the particular exposed facet) and decreasing for still higher coverages. By analyzing orientational ordering and solvent penetration into the ligand layer, we show that a balance of competing effects is responsible for this nonmonotonic dependence. Although the various unpassivated CdSe surfaces exhibit similar G values, the crystal structure of an exposed facet nevertheless plays an important role in determining the interfacial thermal conductance of passivated surfaces, as the density of binding sites on a surface determines the ligand-grafting densities that may ultimately be achieved. We demonstrate that surface passivation can increase G relative to a bare surface by roughly 1 order of magnitude and that, for a given extent of passivation, thermal conductance can vary by up to a factor of ∼2 between different surfaces, suggesting that appropriately tailored nanostructures may direct heat flow in an anisotropic fashion for interface-limited thermal transport.

  2. Low-cost rural surface alternatives : demonstration project : [tech transfer summary].

    Science.gov (United States)

    2015-06-01

    Identify the most effective and economical methods for preventing or : mitigating freeze-thaw damage to granular surfaced roads in seasonally : cold regions : Construct demonstration test sections using several stabilization : methods recomme...

  3. Nanopatterned submicron pores as a shield for nonspecific binding in surface plasmon resonance-based sensing

    NARCIS (Netherlands)

    Raz, Sabina Rebe; Marchesini, Gerardo R.; Bremer, Maria G. E. G.; Colpo, Pascal; Garcia, Cesar Pascual; Guidetti, Guido; Norde, Willem; Rossi, Francois

    2012-01-01

    We present a novel approach to tackle the most common drawback of using surface plasmon resonance for analyte screening in complex biological matrices - the nonspecific binding to the sensor chip surface. By using a perforated membrane supported by a polymeric gel structure at the evanescent wave

  4. Nanopatterned submicron pores as a shield for nonspecific binding in surface plasmon resonance-based sensing

    NARCIS (Netherlands)

    Rebe-Raz, S.; Marchesini, G.R.; Bremer, M.G.E.G.; Colpo, P.; Garcia, C.P.; Guidetti, G.; Norde, W.; Rossi, F.

    2012-01-01

    We present a novel approach to tackle the most common drawback of using surface plasmon resonance for analyte screening in complex biological matrices – the nonspecific binding to the sensor chip surface. By using a perforated membrane supported by a polymeric gel structure at the evanescent wave

  5. Detecting Local Ligand-Binding Site Similarity in Non-Homologous Proteins by Surface Patch Comparison

    Science.gov (United States)

    Sael, Lee; Kihara, Daisuke

    2012-01-01

    Functional elucidation of proteins is one of the essential tasks in biology. Function of a protein, specifically, small ligand molecules that bind to a protein, can be predicted by finding similar local surface regions in binding sites of known proteins. Here, we developed an alignment free local surface comparison method for predicting a ligand molecule which binds to a query protein. The algorithm, named Patch-Surfer, represents a binding pocket as a combination of segmented surface patches, each of which is characterized by its geometrical shape, the electrostatic potential, the hydrophobicity, and the concaveness. Representing a pocket by a set of patches is effective to absorb difference of global pocket shape while capturing local similarity of pockets. The shape and the physicochemical properties of surface patches are represented using the 3D Zernike descriptor, which is a series expansion of mathematical 3D function. Two pockets are compared using a modified weighted bipartite matching algorithm, which matches similar patches from the two pockets. Patch-Surfer was benchmarked on three datasets, which consist in total of 390 proteins that bind to one of 21 ligands. Patch-Surfer showed superior performance to existing methods including a global pocket comparison method, Pocket-Surfer, which we have previously introduced. Particularly, as intended, the accuracy showed large improvement for flexible ligand molecules, which bind to pockets in different conformations. PMID:22275074

  6. Detecting local ligand-binding site similarity in nonhomologous proteins by surface patch comparison.

    Science.gov (United States)

    Sael, Lee; Kihara, Daisuke

    2012-04-01

    Functional elucidation of proteins is one of the essential tasks in biology. Function of a protein, specifically, small ligand molecules that bind to a protein, can be predicted by finding similar local surface regions in binding sites of known proteins. Here, we developed an alignment free local surface comparison method for predicting a ligand molecule which binds to a query protein. The algorithm, named Patch-Surfer, represents a binding pocket as a combination of segmented surface patches, each of which is characterized by its geometrical shape, the electrostatic potential, the hydrophobicity, and the concaveness. Representing a pocket by a set of patches is effective to absorb difference of global pocket shape while capturing local similarity of pockets. The shape and the physicochemical properties of surface patches are represented using the 3D Zernike descriptor, which is a series expansion of mathematical 3D function. Two pockets are compared using a modified weighted bipartite matching algorithm, which matches similar patches from the two pockets. Patch-Surfer was benchmarked on three datasets, which consist in total of 390 proteins that bind to one of 21 ligands. Patch-Surfer showed superior performance to existing methods including a global pocket comparison method, Pocket-Surfer, which we have previously introduced. Particularly, as intended, the accuracy showed large improvement for flexible ligand molecules, which bind to pockets in different conformations. Copyright © 2011 Wiley Periodicals, Inc.

  7. The "Chocolate Experiment"--A Demonstration of Radiation Absorption by Different Colored Surfaces

    Science.gov (United States)

    Fung, Dennis

    2015-01-01

    In the typical "cookbook" experiment comparing the radiation absorption rates of different colored surfaces, students' hands are commonly used as a measurement instrument to demonstrate that dull black and silvery surfaces are good and poor absorbers of radiation, respectively. However, college students are often skeptical about using…

  8. Road surface erosion on the Jackson Demonstration State Forest: results of a pilot study

    Science.gov (United States)

    Brian Barrett; Rosemary Kosaka; David. Tomberlin

    2012-01-01

    This paper presents results of a 3 year pilot study of surface erosion on forest roads in the Jackson Demonstration State Forest in California’s coastal redwood region. Ten road segments representing a range of surface, grade, and ditch conditions were selected for the study. At each segment, settling basins with tipping buckets were installed to measure...

  9. Development of a Surface Plasmon Resonance Assay for the Characterization of Small-Molecule Binding Kinetics and Mechanism of Binding to Kynurenine 3-Monooxygenase.

    Science.gov (United States)

    Poda, Suresh B; Kobayashi, Masakazu; Nachane, Ruta; Menon, Veena; Gandhi, Adarsh S; Budac, David P; Li, Guiying; Campbell, Brian M; Tagmose, Lena

    2015-10-01

    Kynurenine 3-monooxygenase (KMO), a pivotal enzyme in the kynurenine pathway, was identified as a potential therapeutic target for treating neurodegenerative and psychiatric disorders. In this article, we describe a surface plasmon resonance (SPR) assay that delivers both kinetics and the mechanism of binding (MoB) data, enabling a detailed characterization of KMO inhibitors for the enzyme in real time. SPR assay development included optimization of the protein construct and the buffer conditions. The stability and inhibitor binding activity of the immobilized KMO were significantly improved when the experiments were performed at 10°C using a buffer containing 0.05% n-dodecyl-β-d-maltoside (DDM) as the detergent. The KD values of the known KMO inhibitors (UPF648 and RO61-8048) from the SPR assay were in good accordance with the biochemical LC/MS/MS assay. Also, the SPR assay was able to differentiate the binding kinetics (k(a) and k(d)) of the selected unknown KMO inhibitors. For example, the inhibitors that showed comparable IC50 values in the LC/MS/MS assay displayed differences in their residence time (τ = 1/k(d)) in the SPR assay. To better define the MoB of the inhibitors to KMO, an SPR-based competition assay was developed, which demonstrated that both UPF648 and RO61-8048 bound to the substrate-binding site. These results demonstrate the potential of the SPR assay for characterizing the affinity, the kinetics, and the MoB profiles of the KMO inhibitors.

  10. PsB multiprotein complex of Dictyostelium discoideum. Demonstration of cellulose binding activity and order of protein subunit assembly.

    Science.gov (United States)

    McGuire, V; Alexander, S

    1996-06-14

    The differentiated spores of Dictyostelium are surrounded by an extracellular matrix, the spore coat, which protects them from environmental factors allowing them to remain viable for extended periods of time. This presumably is a major evolutionary advantage. This unique extracellular matrix is composed of cellulose and glycoproteins. Previous work has shown that some of these spore coat glycoproteins exist as a preassembled multiprotein complex (the PsB multiprotein complex) which is stored in the prespore vesicles (Watson, N., McGuire, V., and Alexander, S (1994) J. Cell Sci. 107, 2567-2579). Later in development, the complex is synchronously secreted from the prespore vesicles and incorporated into the spore coat. We now have shown that the PsB complex has a specific in vitro cellulose binding activity. The analysis of mutants lacking individual subunits of the PsB complex revealed the relative order of assembly of the subunit proteins and demonstrated that the protein subunits must be assembled for cellulose binding activity. These results provide a biochemical explanation for the localization of this multiprotein complex in the spore coat.

  11. Effects of Multiwalled Carbon Nanotube Surface Modification and Purification on Bovine Serum Albumin Binding and Biological Responses

    Directory of Open Access Journals (Sweden)

    Wei Bai

    2016-01-01

    Full Text Available Carboxylation of multiwalled carbon nanotubes (MWCNTs has been used to improve solubility in aqueous systems and for further functionalization with biologically active moieties for biomedical uses. An important consideration is that oxidation debris is generated during the process of carboxylation, which can be removed by base washing. We hypothesized that surface modification as well as purification by debris removal may alter physicochemical properties of MWCNTs and their ability to bind proteins. We utilized pristine MWCNT, carboxylated MWCNTs (F-MWCNTs, and base-washed carboxylated MWCNTs (BW-F-MWCNTs to examine formation of a bovine serum albumin (BSA protein corona and impact on biological responses. We found that carboxylation increased the capability of F-MWCNTs to bind BSA, and base washing further increased this binding. Functionalization increased cellular uptake by rat aortic endothelial cells (RAEC and mouse macrophages (RAW264.7, while base washing showed results similar to the functionalized analog. Interestingly, BSA binding downregulated mRNA levels of interleukin-6 (IL-6 and heme oxygenase 1 (Hmox1 in RAEC cells but upregulated the expression of IL-6 and Hmox1 in RAW264.7 cells. Overall, our study demonstrated that surface modification as well as further purification impacted the interaction of MWCNTs with proteins and subsequent cellular responses.

  12. Lectin binding profiles of SSEA-4 enriched, pluripotent human embryonic stem cell surfaces

    Science.gov (United States)

    Venable, Alison; Mitalipova, Maisam; Lyons, Ian; Jones, Karen; Shin, Soojung; Pierce, Michael; Stice, Steven

    2005-01-01

    Background Pluripotent human embryonic stem cells (hESCs) have the potential to form every cell type in the body. These cells must be appropriately characterized prior to differentiation studies or when defining characteristics of the pluripotent state. Some developmentally regulated cell surface antigens identified by monoclonal antibodies in a variety of species and stem cell types have proven to be side chains of membrane glycolipids and glycoproteins. Therefore, to examine hESC surfaces for other potential pluripotent markers, we used a panel of 14 lectins, which were chosen based on their specificity for a variety of carbohydrates and carbohydrate linkages, along with stage specific embryonic antigen-4 (SSEA-4), to determine binding quantitation by flow cytometry and binding localization in adherent colonies by immunocytochemistry. Results Enriching cells for SSEA-4 expression increased the percentage of SSEA-4 positive cells to 98–99%. Using enriched high SSEA-4-expressing hESCs, we then analyzed the binding percentages of selected lectins and found a large variation in binding percentages ranging from 4% to 99% binding. Lycopersicon (tomato)esculetum lectin (TL), Ricinus communis agglutinin (RCA), and Concanavalin A (Con A) bound to SSEA-4 positive regions of hESCs and with similar binding percentages as SSEA-4. In contrast, we found Dolichos biflorus agglutinin (DBA) and Lotus tetragonolobus lectin (LTL) did not bind to hESCs while Phaseolus vulgaris leuco-agglutinin (PHA-L), Vicia villosa agglutinin (VVA), Ulex europaeus agglutinin (UEA), Phaseolus vulgaris erythro-agglutinin (PHA-E), and Maackia amurensis agglutinin (MAA) bound partially to hESCs. These binding percentages correlated well with immunocytochemistry results. Conclusion Our results provide information about types of carbohydrates and carbohydrate linkages found on pluripotent hESC surfaces. We propose that TL, RCA and Con A may be used as markers that are associated with the pluripotent

  13. Lectin binding profiles of SSEA-4 enriched, pluripotent human embryonic stem cell surfaces

    Directory of Open Access Journals (Sweden)

    Shin Soojung

    2005-07-01

    Full Text Available Abstract Background Pluripotent human embryonic stem cells (hESCs have the potential to form every cell type in the body. These cells must be appropriately characterized prior to differentiation studies or when defining characteristics of the pluripotent state. Some developmentally regulated cell surface antigens identified by monoclonal antibodies in a variety of species and stem cell types have proven to be side chains of membrane glycolipids and glycoproteins. Therefore, to examine hESC surfaces for other potential pluripotent markers, we used a panel of 14 lectins, which were chosen based on their specificity for a variety of carbohydrates and carbohydrate linkages, along with stage specific embryonic antigen-4 (SSEA-4, to determine binding quantitation by flow cytometry and binding localization in adherent colonies by immunocytochemistry. Results Enriching cells for SSEA-4 expression increased the percentage of SSEA-4 positive cells to 98–99%. Using enriched high SSEA-4-expressing hESCs, we then analyzed the binding percentages of selected lectins and found a large variation in binding percentages ranging from 4% to 99% binding. Lycopersicon (tomatoesculetum lectin (TL, Ricinus communis agglutinin (RCA, and Concanavalin A (Con A bound to SSEA-4 positive regions of hESCs and with similar binding percentages as SSEA-4. In contrast, we found Dolichos biflorus agglutinin (DBA and Lotus tetragonolobus lectin (LTL did not bind to hESCs while Phaseolus vulgaris leuco-agglutinin (PHA-L, Vicia villosa agglutinin (VVA, Ulex europaeus agglutinin (UEA, Phaseolus vulgaris erythro-agglutinin (PHA-E, and Maackia amurensis agglutinin (MAA bound partially to hESCs. These binding percentages correlated well with immunocytochemistry results. Conclusion Our results provide information about types of carbohydrates and carbohydrate linkages found on pluripotent hESC surfaces. We propose that TL, RCA and Con A may be used as markers that are associated with the

  14. Real-Time Ligand Binding Pocket Database Search Using Local Surface Descriptors

    Science.gov (United States)

    Chikhi, Rayan; Sael, Lee; Kihara, Daisuke

    2010-01-01

    Due to the increasing number of structures of unknown function accumulated by ongoing structural genomics projects, there is an urgent need for computational methods for characterizing protein tertiary structures. As functions of many of these proteins are not easily predicted by conventional sequence database searches, a legitimate strategy is to utilize structure information in function characterization. Of a particular interest is prediction of ligand binding to a protein, as ligand molecule recognition is a major part of molecular function of proteins. Predicting whether a ligand molecule binds a protein is a complex problem due to the physical nature of protein-ligand interactions and the flexibility of both binding sites and ligand molecules. However, geometric and physicochemical complementarity is observed between the ligand and its binding site in many cases. Therefore, ligand molecules which bind to a local surface site in a protein can be predicted by finding similar local pockets of known binding ligands in the structure database. Here, we present two representations of ligand binding pockets and utilize them for ligand binding prediction by pocket shape comparison. These representations are based on mapping of surface properties of binding pockets, which are compactly described either by the two dimensional pseudo-Zernike moments or the 3D Zernike descriptors. These compact representations allow a fast real-time pocket searching against a database. Thorough benchmark study employing two different datasets show that our representations are competitive with the other existing methods. Limitations and potentials of the shape-based methods as well as possible improvements are discussed. PMID:20455259

  15. Antibacterial surfaces by adsorptive binding of polyvinyl-sulphonate-stabilized silver nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Vasilev, Krasimir; Sah, Vasu R; Goreham, Renee V; Short, Robert D [Mawson Institute, University of South Australia, Mawson Lakes Campus, Mawson Lakes, Adelaide, SA 5095 (Australia); Ndi, Chi; Griesser, Hans J, E-mail: Krasimir.vasilev@unisa.edu.au [Ian Wark Research Institute, University of South Australia, Mawson Lakes, Adelaide, SA 5095 (Australia)

    2010-05-28

    This paper presents a novel and facile method for the generation of efficient antibacterial coatings which can be applied to practically any type of substrate. Silver nanoparticles were stabilized with an adsorbed surface layer of polyvinyl sulphonate (PVS). This steric layer provided excellent colloidal stability, preventing aggregation over periods of months. PVS-coated silver nanoparticles were bound onto amine-containing surfaces, here produced by deposition of an allylamine plasma polymer thin film onto various substrates. SEM imaging showed no aggregation upon surface binding of the nanoparticles; they were well dispersed on amine surfaces. Such nanoparticle-coated surfaces were found to be effective in preventing attachment of Staphylococcus epidermidis bacteria and also in preventing biofilm formation. Combined with the ability of plasma polymerization to apply the thin polymeric binding layer onto a wide range of materials, this method appears promising for the fabrication of a wide range of infection-resistant biomedical devices.

  16. Identification of distant drug off-targets by direct superposition of binding pocket surfaces.

    Science.gov (United States)

    Schumann, Marcel; Armen, Roger S

    2013-01-01

    Correctly predicting off-targets for a given molecular structure, which would have the ability to bind a large range of ligands, is both particularly difficult and important if they share no significant sequence or fold similarity with the respective molecular target ("distant off-targets"). A novel approach for identification of off-targets by direct superposition of protein binding pocket surfaces is presented and applied to a set of well-studied and highly relevant drug targets, including representative kinases and nuclear hormone receptors. The entire Protein Data Bank is searched for similar binding pockets and convincing distant off-target candidates were identified that share no significant sequence or fold similarity with the respective target structure. These putative target off-target pairs are further supported by the existence of compounds that bind strongly to both with high topological similarity, and in some cases, literature examples of individual compounds that bind to both. Also, our results clearly show that it is possible for binding pockets to exhibit a striking surface similarity, while the respective off-target shares neither significant sequence nor significant fold similarity with the respective molecular target ("distant off-target").

  17. An Energy Conservation Approach to Adsorbate-Induced Surface Stress and the Extraction of Binding Energy Using Nanomechanics

    Energy Technology Data Exchange (ETDEWEB)

    Pinnaduwage, Lal A [ORNL; Boiadjiev, Vassil I [ORNL; Fernando, G. W. [University of Connecticut, Storrs; Hawk, J. E. [Oak Ridge National Laboratory (ORNL); Wijewardhana, L.C. R. [University of Cincinnati; Gehl, Anthony C [ORNL

    2008-01-01

    Microcantilevers are ideally-suited for the study of surface phenomena due to their large surface-to-volume ratios, which amplify surface effects. We show that when guest molecules bind to atoms/molecules on a microcantilever surface, the released binding energy is retained in the host surface, leading to a metastable state where the excess energy on the surface is manifested as an increase in surface stress leading to the bending of the microcantilever. When the excess energy is released, the microcantilever relaxes back to the original state, and the relaxation time depends on the particular binding process involved. Such experiments were conducted for three binding processes in vapor phase experiments: physisorption, hydrogen bonding, and chemisorption. To our knowledge, such an energy conservation approach has not been taken into account in adsorbate-induced surface effect investigations. Furthermore, these experiments illustrate that detailed molecular-level information on binding energies can be extracted from this simple micromechanical sensor.

  18. Effects of sodium on cell surface and intracellular 3H-naloxone binding sites

    International Nuclear Information System (INIS)

    Pollack, A.E.; Wooten, G.F.

    1987-01-01

    The binding of the opiate antagonist 3 H-naloxone was examined in rat whole brain homogenates and in crude subcellular fractions of these homogenates (nuclear, synaptosomal, and mitochondrial fractions) using buffers that approximated intra- (low sodium concentration) and extracellular (high sodium concentration) fluids. Saturation studies showed a two-fold decrease in the dissociation constant (Kd) in all subcellular fractions examined in extracellular buffer compared to intracellular buffer. In contrast, there was no significant effect of the buffers on the Bmax. Thus, 3 H-naloxone did not distinguish between binding sites present on cell surface and intracellular tissues in these two buffers. These results show that the sodium effect of opiate antagonist binding is probably not a function of altered selection of intra- and extracellular binding sites. 17 references, 2 tables

  19. The binding of cytochrome c to neuroglobin: A docking and surface plasmon resonance study

    DEFF Research Database (Denmark)

    Bønding, Signe Helbo; Henty, K.; Dingley, A.J.

    2008-01-01

    is associated with a small unfavourable enthalpy change (1.9 kcal mol-1) and a moderately large, favourable entropy change (14.8 cal mol-1 deg-1). The sensitivity of the binding constant to the presence of salt suggests that the complex formation involves electrostatic interactions....... one major binding site for cytochrome c to neuroglobin. The results yield a plausible structure for the most likely complex structure in which the hemes of each protein are in close contact. NMR analysis identifies the formation of a weak complex in which the heme group of cytochrome c is involved....... surface plasmon resonance studies provide a value of 45 μM for the equilibrium constant for cytochrome c binding to neuroglobin, which increases significantly as the ionic strength of the solution increases. The temperature dependence of the binding constant indicates that the complex formation...

  20. Low level chemiluminescence measurement of the binding of 8-methoxypsoralen to proteins and lymphocytic surfaces

    International Nuclear Information System (INIS)

    Lange, B.

    1980-01-01

    Photochemotherapy with 8-methoxypsoralen (8-MOP) and longwave ultraviolet light is beneficial in such different disorders like psoriasis, lichen planus, and mykosis fungoides. In contrast to a widely accepted hypothesis 8-MOP does not solely bind to nucleic acid, but also to certain proteins. The mechanism of this binding as well as the precise binding area are unknown. Therefore the UV-provoked reactions of 8-MOP with a lipid mixture, a glucosaminoglycan solution, a protein solution, and lymphocyte suspensions, respectively were investigated using low level chemiluminescence (LLCL). It was found an 8-MOP concentration-dependent decrease of LLCL intensity in the lymphocyte suspensions (10 3 to 10 4 cells/μl). This effect is result of the diminution of the photoactive 8-MOP content of the solution. 8-MOP binds quickly and in the course of a free radical reaction to lymphocytic surfaces and coincidentally loses its potency to start LLCL-detectable free radical chain responses. (author)

  1. Selectivity of the surface binding site (SBS) on barley starch synthase I

    DEFF Research Database (Denmark)

    Wilkens, Casper; Cuesta-Seijo, Jose A.; Palcic, Monica

    2014-01-01

    Starch synthase I (SSI) from various sources has been shown to preferentially elongate branch chains of degree of polymerisation (DP) from 6–7 to produce chains of DP 8–12. In the recently determined crystal structure of barley starch synthase I (HvSSI) a so-called surface binding site (SBS) was ...

  2. Autodisplay of an avidin with biotin-binding activity on the surface of Escherichia coli.

    Science.gov (United States)

    Pardavé-Alejandre, H D; Alvarado-Yaah, J E; Pompa-Mera, E N; Muñoz-Medina, J E; Sárquiz-Martínez, B; Santacruz-Tinoco, C E; Manning-Cela, R G; Ortíz-Navarrete, V; López-Macías, C; González-Bonilla, C R

    2018-03-01

    To display a recombinant avidin fused to the autotransporter ShdA to bind biotinylated molecules on the surface of Escherichia coli. Two chimeric protein constructs containing avidin fused to the autotransporter ShdA were expressed on the surface of Escherichia coli DH5α. One fusion protein contained 476 amino acids of the ShdA α and β domains, whereas the second consisted of a 314 amino acid from α and truncated β domains. Protein production was verified by SDS-PAGE using an antibody to the molecular FLAG-tag. The surface display of the avidin-shdA fusion protein was confirmed by confocal microscopy and flow cytometry analysis, and the biotin-binding activity was evaluated by fluorescence microscopy and flow cytometry using biotin-4-fluorescein and biotinylated-ovalbumin (OVA). Expression of a recombinant avidin with biotin-binding activity on the surface of E. coli was achieved using the autotransporter ShdA. This system is an alternative to bind biotinylated molecules to E. coli.

  3. Analysis of myelomonocytic leukemic differentiation by a cell surface marker panel including a fucose-binding lectin from Lotus tetragonolobus.

    Science.gov (United States)

    Elias, L; Van Epps, D E

    1984-06-01

    The fucose-binding lectin from Lotus tetragonolobus ( FBL -L) has been previously shown to bind specifically to normal cells of the myeloid and monocytic lineages. The purpose of this study was to explore the utility of fluoresceinated FBL -L as a leukemia differentiation marker in conjunction with a panel of other frequently used surface markers (Fc receptor, HLA-DR, OKM1, and antimonocyte antibody). FBL -L reacted with leukemic cells in 8/9 cases of clinically recognized acute myeloid leukemia, including myeloid blast crisis of chronic granulocytic leukemia, 3/3 cases of chronic phase chronic myelogenous leukemia, and in 2/7 cases of clinically undifferentiated acute leukemia. Correlations were noted between reactivity with FBL -L, and DR and Fc receptor expression. Among continuous cell lines, FBL -L bound with high intensity to a majority of HL-60 and U937 cells. The less well differentiated myeloblast cell lines, KG-1, KG1a , and HL-60 blast II, exhibited less FBL -L binding than HL-60 and U937. A moderate proportion of K562 cells exhibited low level binding of FBL -L. Several lymphoblastic cell lines exhibited a pattern of low intensity binding that was distinguishable from the high intensity binding pattern of the myeloblastic lines. FBL -L reactivity of U937 was enhanced by induction of differentiation with leukocyte conditioned medium, but not dimethylsulfoxide. Such treatments induced contrasting patterns of change of HL-60 and U937 when labeled with OKM1, alpha-Mono, and HLA-DR. These studies demonstrate the application of FBL -L to analysis and quantitation of myelomonocytic leukemic differentiation.

  4. Binding of a cementum attachment protein to extracellular matrix components and to dental surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Pitaru, S; Hekmati, H [Department of Oral Biology, Goldschleger School of Dental Medicine, Tel Aviv University (Israel); Savion, N [Goldschleger Eye Institute, Sackler School of Medicine, Tel Aviv University (Israel); Olsen, S; Narayanan, S A [Department of Pathology, School of Medicine, University of Washington, Seattle, Washington (United States)

    1992-01-01

    Cementum proteins (CP) have been shown to mediate cell attachment. Among these, a 55 kDa protein was isolated. The purpose of the present study was to assess the capacity of CP to bind to non-demineralized and demineralized root surfaces and to support cell attachment to dentin. CP were prepared by sequential extraction of bovine cementum with 25 mM EDTA, 0.5 M acetic acid followed by 4 M guanidine HCl. The latter was subjected to ion exchange chromatography on a DEAE-3SW column and eluted stepwise with a 0-0.5 M NaCl gradient. CP were labelled with [sup 125]I and the capacity of [sup 125]I-CP to bind to mineralized and partially demineralized dentin, synthetic hydroxyapatite, collagen, fibronectin and fibrillar collagen-fibronectin cimplex was assessed. It was found that CP bind specifically to mineralized dentin and synthetic hydroxyapatite but not to demineralized dentin. The specific binding was 60% of the total binding. SDS-PAGE analysis of the proteins bound to dentin indicated that the main bound protein had a molecular weight of 55 kDa. CP exhibited high affinity for fibronectin (k[sub D] = 1.56 x 10[sup -10] M) and fibronectincollagen complex, but their binding to either molecular or fibrillar collagen was negligible. It is suggested that CP may play an important role in the attachment of cells of the periodontium to cementum extracellular matrix during homeostasis and regeneration. (au).

  5. Octasaccharide is the minimal length unit required for efficient binding of cyclophilin B to heparin and cell surface heparan sulphate.

    Science.gov (United States)

    Vanpouille, Christophe; Denys, Agnès; Carpentier, Mathieu; Pakula, Rachel; Mazurier, Joël; Allain, Fabrice

    2004-09-01

    Cyclophilin B (CyPB) is a heparin-binding protein first identified as a receptor for cyclosporin A. In previous studies, we reported that CyPB triggers chemotaxis and integrin-mediated adhesion of T-lymphocytes by way of interaction with two types of binding sites. The first site corresponds to a signalling receptor; the second site has been identified as heparan sulphate (HS) and appears crucial to induce cell adhesion. Characterization of the HS-binding unit is critical to understand the requirement of HS in pro-adhesive activity of CyPB. By using a strategy based on gel mobility shift assays with fluorophore-labelled oligosaccharides, we demonstrated that the minimal heparin unit required for efficient binding of CyPB is an octasaccharide. The mutants CyPB(KKK-) [where KKK- refers to the substitutions K3A(Lys3-->Ala)/K4A/K5A] and CyPB(DeltaYFD) (where Tyr14-Phe-Asp16 has been deleted) failed to interact with octasaccharides, confirming that the Y14FD16 and K3KK5 clusters are required for CyPB binding. Molecular modelling revealed that both clusters are spatially arranged so that they may act synergistically to form a binding site for the octasaccharide. We then demonstrated that heparin-derived octasaccharides and higher degree of polymerization oligosaccharides inhibited the interaction between CyPB and fluorophore-labelled HS chains purified from T-lymphocytes, and strongly reduced the HS-dependent pro-adhesive activity of CyPB. However, oligosaccharides or heparin were unable to restore adhesion of heparinase-treated T-lymphocytes, indicating that HS has to be present on the cell membrane to support the pro-adhesive activity of CyPB. Altogether, these results demonstrate that the octasaccharide is likely to be the minimal length unit required for efficient binding of CyPB to cell surface HS and consequent HS-dependent cell responses.

  6. Inverse gas chromatography as a method for determination of surface properties of binding materials

    Science.gov (United States)

    Yu, Jihai; Lu, Xiaolei; Yang, Chunxia; Du, Baoli; Wang, Shuxian; Ye, Zhengmao

    2017-09-01

    Inverse gas chromatography (IGC) is a promising measurement technique for investigating the surface properties of binding materials, which are the major influence element for the adsorption performance of superplasticizer. In this work, using the IGC method, blast furnace slag (BFS), sulphoaluminate cement (SAC) and portland cement (P·O) are employed to systematically evaluate the corresponding dispersive component (γsd), specific surface free energy (γsab), and acid-base properties. The obtained results show that γsd contributes to a major section of the surface free energy in the three binding materials, suggesting they are of a relatively low polarity. Compared to the two kinds of cements, the BFS possesses the highest dispersive and specific surface free energies (the values are 45.01 mJ/m2 and 11.68 mJ/m2, respectively), and also exhibits a wider distribution range of γsd, indicating their surfaces are heterogeneous. For acid-base properties, the results indicate the surfaces of three samples are basic in nature. In addition, the adsorption investigation shows that per unit surface of BFS adsorbs the most superplasticizer molecules, which indicates the higher surface free energies is beneficial to the superplasticizer adsorption.

  7. Hydration behavior at the ice-binding surface of the Tenebrio molitor antifreeze protein.

    Science.gov (United States)

    Midya, Uday Sankar; Bandyopadhyay, Sanjoy

    2014-05-08

    Molecular dynamics (MD) simulations have been carried out at two different temperatures (300 and 220 K) to study the conformational rigidity of the hyperactive Tenebrio molitor antifreeze protein (TmAFP) in aqueous medium and the structural arrangements of water molecules hydrating its surface. It is found that irrespective of the temperature the ice-binding surface (IBS) of the protein is relatively more rigid than its nonice-binding surface (NIBS). The presence of a set of regularly arranged internally bound water molecules is found to play an important role in maintaining the flat rigid nature of the IBS. Importantly, the calculations reveal that the strategically located hydroxyl oxygens of the threonine (Thr) residues in the IBS influence the arrangements of five sets of ordered waters around it on two parallel planes that closely resemble the basal plane of ice. As a result, these waters can register well with the ice basal plane, thereby allowing the IBS to preferentially bind at the ice interface and inhibit its growth. This provides a possible molecular reason behind the ice-binding activity of TmAFP at the basal plane of ice.

  8. Facile Photoimmobilization of Proteins onto Low-Binding PEG-Coated Polymer Surfaces

    DEFF Research Database (Denmark)

    Larsen, Esben Kjær Unmack; Mikkelsen, Morten Bo Lindholm; Larsen, Niels Bent

    2014-01-01

    was verified for both enzymes and antibodies, and their presence on the surface was confirmed by X-ray photoelectron spectroscopy (XPS) and confocal fluorescence microscopy. Conjugation of capture antibody onto the PEG coating was employed for a simplified ELISA protocol without the need for blocking uncoated...... surface areas, showing ng/mL sensitivity to a cytokine antigen target. Moreover, spatially patterned attachment of fluorescently labeled protein onto the low-binding PEG-coated surface was achieved with a projection lithography system that enabled the creation of micrometer-sized protein features....

  9. Human IgG lacking effector functions demonstrate lower FcRn-binding and reduced transplacental transport

    NARCIS (Netherlands)

    Stapleton, Nigel M.; Armstrong-Fisher, Sylvia S.; Andersen, Jan Terje; van der Schoot, C. Ellen; Porter, Charlene; Page, Kenneth R.; Falconer, Donald; de Haas, Masja; Williamson, Lorna M.; Clark, Michael R.; Vidarsson, Gestur; Armour, Kathryn L.

    2018-01-01

    We have previously generated human IgG1 antibodies that were engineered for reduced binding to the classical Fcγ receptors (FcγRI-III) and C1q, thereby eliminating their destructive effector functions (constant region G1Δnab). In their potential use as blocking agents, favorable binding to the

  10. Short term memory for single surface features and bindings in ageing: A replication study.

    Science.gov (United States)

    Isella, Valeria; Molteni, Federica; Mapelli, Cristina; Ferrarese, Carlo

    2015-06-01

    In the present study we replicated a previous experiment investigating visuo-spatial short term memory binding in young and older healthy individuals, in the attempt to verify the pattern of impairment that can be observed in normal elderly for short term memory for single items vs short term memory for bindings. Assessing a larger sample size (25 young and 25 older subjects), using a more appropriate measure of accuracy for a change detection task (A'), and adding the evaluation of speed of performance, we confirmed that old normals show a decline in short term memory for bindings of shape and colour that is of comparable extent, and not major, to the decline in memory for single shapes and single colours. The absence of a specific deficit of short term memory for conjunctions of surface features seems to distinguish cognitive ageing from Alzheimer's Disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Use of computational modeling in preparation and evaluation of surface imprinted xerogels for binding tetracycline

    International Nuclear Information System (INIS)

    Pace, Samantha J.; Nguyen, Eric; Baria, Maximillian P.; Mojica, Elmer-Rico E.

    2015-01-01

    Linear alkyl alkoxysilanes (methoxy and ethoxy-based) of varying length were used in preparing tetracycline surface imprinted silica xerogels by the sol–gel process. The resulting xerogels were characterized in terms of binding tetracycline (TC) by using tritium-labeled TC. Results showed preferential binding in the ethoxysilane based xerogels in comparison to methoxysilane based xerogels. A computational approach using the interaction energy (IE) between TC and each alkyl alkoxysilane was deduced as a rational way of predicting the formulation that would provide the best analytical performance for a given molecularly imprinted xerogel (MIX). Hartree-Fock calculations revealed an increase in IE as the length of the carbon chain increases until an optimum value at C6 in both alkoxysilanes. This is consistent with the experimental results wherein the C6 xerogel formulation has the highest imprinting factor. These results show the potential of using computational modeling as a rational way of preparing surface imprinted materials. (author)

  12. Gaussian-Based Smooth Dielectric Function: A Surface-Free Approach for Modeling Macromolecular Binding in Solvents

    Directory of Open Access Journals (Sweden)

    Arghya Chakravorty

    2018-03-01

    Full Text Available Conventional modeling techniques to model macromolecular solvation and its effect on binding in the framework of Poisson-Boltzmann based implicit solvent models make use of a geometrically defined surface to depict the separation of macromolecular interior (low dielectric constant from the solvent phase (high dielectric constant. Though this simplification saves time and computational resources without significantly compromising the accuracy of free energy calculations, it bypasses some of the key physio-chemical properties of the solute-solvent interface, e.g., the altered flexibility of water molecules and that of side chains at the interface, which results in dielectric properties different from both bulk water and macromolecular interior, respectively. Here we present a Gaussian-based smooth dielectric model, an inhomogeneous dielectric distribution model that mimics the effect of macromolecular flexibility and captures the altered properties of surface bound water molecules. Thus, the model delivers a smooth transition of dielectric properties from the macromolecular interior to the solvent phase, eliminating any unphysical surface separating the two phases. Using various examples of macromolecular binding, we demonstrate its utility and illustrate the comparison with the conventional 2-dielectric model. We also showcase some additional abilities of this model, viz. to account for the effect of electrolytes in the solution and to render the distribution profile of water across a lipid membrane.

  13. Microassay for measurement of binding of radiolabelled ligands to cell surface molecules

    International Nuclear Information System (INIS)

    Woof, J.M.; Burton, D.R.

    1988-01-01

    An improved technique for measuring the binding of radiolabelled ligands to cell surface molecules has been developed by modification of a procedure using centrifugation through a water-immiscible oil to separate free and cell-bound ligand. It maximises the percentage of ligand bound since cell-bound and free ligand can be separated easily and reproducibly even when very small reaction volumes are used. This permits low levels of ligand radiolabelling and relatively low numbers of cells to be used

  14. Thorium binding by biochar fibres derived from Luffa Cylindrica after controlled surface oxidation

    Science.gov (United States)

    Liatsou, Ioanna; Christodoulou, Eleni; Paschalidis, Ioannis

    2017-04-01

    Controlled surface modification of biochar fibres derived from Luffa Cylindrica sponges has been carried out by nitric acid and the degree of oxidation could be controlled by changing the acid concentration or the reaction time. The extent of surface oxidation has been quantified by acid-base titration and FTIR-spectroscopy. Furthermore, thorium binding has been studied as a function of various parameters and the experimental results show that even under strong acidic conditions the relative sorption is above 70% and the sorption capacity of the biochar fibres for Th(IV) at pH 3 is qmax= 70 gṡkg-1.

  15. Effect of fullerenol surface chemistry on nanoparticle binding-induced protein misfolding

    Science.gov (United States)

    Radic, Slaven; Nedumpully-Govindan, Praveen; Chen, Ran; Salonen, Emppu; Brown, Jared M.; Ke, Pu Chun; Ding, Feng

    2014-06-01

    Fullerene and its derivatives with different surface chemistry have great potential in biomedical applications. Accordingly, it is important to delineate the impact of these carbon-based nanoparticles on protein structure, dynamics, and subsequently function. Here, we focused on the effect of hydroxylation -- a common strategy for solubilizing and functionalizing fullerene -- on protein-nanoparticle interactions using a model protein, ubiquitin. We applied a set of complementary computational modeling methods, including docking and molecular dynamics simulations with both explicit and implicit solvent, to illustrate the impact of hydroxylated fullerenes on the structure and dynamics of ubiquitin. We found that all derivatives bound to the model protein. Specifically, the more hydrophilic nanoparticles with a higher number of hydroxyl groups bound to the surface of the protein via hydrogen bonds, which stabilized the protein without inducing large conformational changes in the protein structure. In contrast, fullerene derivatives with a smaller number of hydroxyl groups buried their hydrophobic surface inside the protein, thereby causing protein denaturation. Overall, our results revealed a distinct role of surface chemistry on nanoparticle-protein binding and binding-induced protein misfolding.Fullerene and its derivatives with different surface chemistry have great potential in biomedical applications. Accordingly, it is important to delineate the impact of these carbon-based nanoparticles on protein structure, dynamics, and subsequently function. Here, we focused on the effect of hydroxylation -- a common strategy for solubilizing and functionalizing fullerene -- on protein-nanoparticle interactions using a model protein, ubiquitin. We applied a set of complementary computational modeling methods, including docking and molecular dynamics simulations with both explicit and implicit solvent, to illustrate the impact of hydroxylated fullerenes on the structure and

  16. Fermi Surface Manipulation by External Magnetic Field Demonstrated for a Prototypical Ferromagnet

    Directory of Open Access Journals (Sweden)

    E. Młyńczak

    2016-12-01

    Full Text Available We consider the details of the near-surface electronic band structure of a prototypical ferromagnet, Fe(001. Using high-resolution angle-resolved photoemission spectroscopy, we demonstrate openings of the spin-orbit-induced electronic band gaps near the Fermi level. The band gaps, and thus the Fermi surface, can be manipulated by changing the remanent magnetization direction. The effect is of the order of ΔE=100  meV and Δk=0.1  Å^{−1}. We show that the observed dispersions are dominated by the bulk band structure. First-principles calculations and one-step photoemission calculations suggest that the effect is related to changes in the electronic ground state and not caused by the photoemission process itself. The symmetry of the effect indicates that the observed electronic bulk states are influenced by the presence of the surface, which might be understood as related to a Rashba-type effect. By pinpointing the regions in the electronic band structure where the switchable band gaps occur, we demonstrate the significance of spin-orbit interaction even for elements as light as 3d ferromagnets. These results set a new paradigm for the investigations of spin-orbit effects in the spintronic materials. The same methodology could be used in the bottom-up design of the devices based on the switching of spin-orbit gaps such as electric-field control of magnetic anisotropy or tunneling anisotropic magnetoresistance.

  17. LIGSITEcsc: predicting ligand binding sites using the Connolly surface and degree of conservation

    Directory of Open Access Journals (Sweden)

    Schroeder Michael

    2006-09-01

    Full Text Available Abstract Background Identifying pockets on protein surfaces is of great importance for many structure-based drug design applications and protein-ligand docking algorithms. Over the last ten years, many geometric methods for the prediction of ligand-binding sites have been developed. Results We present LIGSITEcsc, an extension and implementation of the LIGSITE algorithm. LIGSITEcsc is based on the notion of surface-solvent-surface events and the degree of conservation of the involved surface residues. We compare our algorithm to four other approaches, LIGSITE, CAST, PASS, and SURFNET, and evaluate all on a dataset of 48 unbound/bound structures and 210 bound-structures. LIGSITEcsc performs slightly better than the other tools and achieves a success rate of 71% and 75%, respectively. Conclusion The use of the Connolly surface leads to slight improvements, the prediction re-ranking by conservation to significant improvements of the binding site predictions. A web server for LIGSITEcsc and its source code is available at scoppi.biotec.tu-dresden.de/pocket.

  18. Second Harmonic Correlation Spectroscopy: Theory and Principles for Determining Surface Binding Kinetics.

    Science.gov (United States)

    Sly, Krystal L; Conboy, John C

    2017-06-01

    A novel application of second harmonic correlation spectroscopy (SHCS) for the direct determination of molecular adsorption and desorption kinetics to a surface is discussed in detail. The surface-specific nature of second harmonic generation (SHG) provides an efficient means to determine the kinetic rates of adsorption and desorption of molecular species to an interface without interference from bulk diffusion, which is a significant limitation of fluorescence correlation spectroscopy (FCS). The underlying principles of SHCS for the determination of surface binding kinetics are presented, including the role of optical coherence and optical heterodyne mixing. These properties of SHCS are extremely advantageous and lead to an increase in the signal-to-noise (S/N) of the correlation data, increasing the sensitivity of the technique. The influence of experimental parameters, including the uniformity of the TEM00 laser beam, the overall photon flux, and collection time are also discussed, and are shown to significantly affect the S/N of the correlation data. Second harmonic correlation spectroscopy is a powerful, surface-specific, and label-free alternative to other correlation spectroscopic methods for examining surface binding kinetics.

  19. Sensing (un)binding events via surface plasmons: effects of resonator geometry

    Science.gov (United States)

    Antosiewicz, Tomasz J.; Claudio, Virginia; Käll, Mikael

    2016-04-01

    The resonance conditions of localized surface plasmon resonances (LSPRs) can be perturbed in any number ways making plasmon nanoresonators viable tools in detection of e.g. phase changes, pH, gasses, and single molecules. Precise measurement via LSPR of molecular concentrations hinge on the ability to confidently count the number of molecules attached to a metal resonator and ideally to track binding and unbinding events in real-time. These two requirements make it necessary to rigorously quantify relations between the number of bound molecules and response of plasmonic sensors. This endeavor is hindered on the one hand by a spatially varying response of a given plasmonic nanosensor. On the other hand movement of molecules is determined by stochastic effects (Brownian motion) as well as deterministic flow, if present, in microfluidic channels. The combination of molecular dynamics and the electromagnetic response of the LSPR yield an uncertainty which is little understood and whose effect is often disregarded in quantitative sensing experiments. Using a combination of electromagnetic finite-difference time-domain (FDTD) calculations of the plasmon resonance peak shift of various metal nanosensors (disk, cone, rod, dimer) and stochastic diffusion-reaction simulations of biomolecular interactions on a sensor surface we clarify the interplay between position dependent binding probability and inhomogeneous sensitivity distribution. We show, how the statistical characteristics of the total signal upon molecular binding are determined. The proposed methodology is, in general, applicable to any sensor and any transduction mechanism, although the specifics of implementation will vary depending on circumstances. In this work we focus on elucidating how the interplay between electromagnetic and stochastic effects impacts the feasibility of employing particular shapes of plasmonic sensors for real-time monitoring of individual binding reactions or sensing low concentrations

  20. NOx Binding and Dissociation: Enhanced Ferroelectric Surface Chemistry by Catalytic Monolayers

    Science.gov (United States)

    Kakekhani, Arvin; Ismail-Beigi, Sohrab

    2013-03-01

    NOx molecules are regulated air pollutants produced during automotive combustion. As part of an effort to design viable catalysts for NOx decomposition operating at higher temperatures that would allow for improved fuel efficiency, we examine NOx chemistry on ferroelectric perovskite surfaces. Changing the direction of ferroelectric polarization can modify surface electronic properties and may lead to switchable surface chemistry. Here, we describe our recent work on potentially enhanced surface chemistry using catalytic RuO2 monolayers on perovskite ferroelectric substrates. In addition to thermodynamic stabilization of the RuO2 layer, we present results on the polarization-dependent binding of NO, O2, N2, and atomic O and N. We present results showing that one key problem with current catalysts, involving the difficulty of releasing dissociation products (especially oxygen), can be ameliorated by this method. Primary support from Toyota Motor Engineering and Manufacturing, North America, Inc.

  1. Surface-enhanced Raman spectroscopy competitive binding biosensor development utilizing surface modification of silver nanocubes and a citrulline aptamer

    Science.gov (United States)

    Walton, Brian M.; Jackson, George W.; Deutz, Nicolaas; Cote, Gerard

    2017-07-01

    A point-of-care (PoC) device with the ability to detect biomarkers at low concentrations in bodily fluids would have an enormous potential for medical diagnostics outside the central laboratory. One method to monitor analytes at low concentrations is by using surface-enhanced Raman spectroscopy (SERS). In this preliminary study toward using SERS for PoC biosensing, the surface of colloidal silver (Ag) nanocubes has been modified to test the feasibility of a competitive binding SERS assay utilizing aptamers against citrulline. Specifically, Ag nanocubes were functionalized with mercaptobenzoic acid, as well as a heterobifunctional polyethylene glycol linker that forms an amide bond with the amino acid citrulline. After the functionalization, the nanocubes were characterized by zeta-potential, transmission electron microscopy images, ultraviolet/visible spectroscopy, and by SERS. The citrulline aptamers were developed and tested using backscattering interferometry. The data show that our surface modification method does work and that the functionalized nanoparticles can be detected using SERS down to a 24.5 picomolar level. Last, we used microscale thermophoresis to show that the aptamers bind to citrulline with at least a 50 times stronger affinity than other amino acids.

  2. Demonstrating electromagnetic control of free-surface, liquid-metal flows relevant to fusion reactors

    Science.gov (United States)

    Hvasta, M. G.; Kolemen, E.; Fisher, A. E.; Ji, H.

    2018-01-01

    Plasma-facing components (PFC’s) made from solid materials may not be able to withstand the large heat and particle fluxes that will be produced within next-generation fusion reactors. To address the shortcomings of solid PFC’s, a variety of liquid-metal (LM) PFC concepts have been proposed. Many of the suggested LM-PFC designs rely on electromagnetic restraint (Lorentz force) to keep free-surface, liquid-metal flows adhered to the interior surfaces of a fusion reactor. However, there is very little, if any, experimental data demonstrating that free-surface, LM-PFC’s can actually be electromagnetically controlled. Therefore, in this study, electrical currents were injected into a free-surface liquid-metal that was flowing through a uniform magnetic field. The resultant Lorentz force generated within the liquid-metal affected the velocity and depth of the flow in a controllable manner that closely matched theoretical predictions. These results show the promise of electromagnetic control for LM-PFC’s and suggest that electromagnetic control could be further developed to adjust liquid-metal nozzle output, prevent splashing within a tokamak, and alter heat transfer properties for a wide-range of liquid-metal systems.

  3. The `Chocolate Experiment' - A Demonstration of Radiation Absorption by Different Colored Surfaces

    Science.gov (United States)

    Fung, Dennis

    2015-12-01

    In the typical "cookbook" experiment comparing the radiation absorption rates of different colored surfaces, students' hands are commonly used as a measurement instrument to demonstrate that dull black and silvery surfaces are good and poor absorbers of radiation, respectively. However, college students are often skeptical about using their bare hands in this experiment because they learned in early science lessons that skin is not a reliable detector of heat transfer. Moreover, when the experiment is conducted in a school laboratory, it is often difficult for students to perceive the slight differences in heat transfer on the dull black and silvery aluminum leaves attached to their hands. Rather than replacing students' bare hands with such sophisticated apparatus as a data logger and temperature probe, I suggest using a simple (and delicious!) low-cost instrument, i.e., chocolate, which simply melts when it receives radiation.

  4. Delay/Disruption Tolerant Networks (DTN): Testing and Demonstration for Lunar Surface Applications

    Science.gov (United States)

    2009-01-01

    This slide presentation reviews the testing of the Delay/Disruption Tolerant Network (DTN) designed for use with Lunar Surface applications. This is being done through the DTN experimental Network (DEN), that permit access and testing by other NASA centers, DTN team members and protocol developers. The objective of this work is to demonstrate DTN for high return applications in lunar scenarios, provide DEN connectivity with analogs of Constellation elements, emulators, and other resources from DTN Team Members, serve as a wireless communications staging ground for remote analog excursions and enable testing of detailed communication scenarios and evaluation of network performance. Three scenarios for DTN on the Lunar surface are reviewed: Motion imagery, Voice and sensor telemetry, and Navigation telemetry.

  5. An energy conservation approach to adsorbate-induced surface stress and the extraction of binding energy using nanomechanics

    Science.gov (United States)

    Pinnaduwage, Lal A.; Boiadjiev, Vassil I.; Hawk, John E.; Gehl, Anthony C.; Fernando, Gayanath W.; Rohana Wijewardhana, L. C.

    2008-03-01

    Surface stress induced by molecular adsorption in three different binding processes has been studied experimentally using a microcantilever sensor. A comprehensive free-energy analysis based on an energy conservation approach is proposed to explain the experimental observations. We show that when guest molecules bind to atoms/molecules on a microcantilever surface, the released binding energy is retained in the host surface, leading to a metastable state where the excess energy on the surface is manifested as an increase in surface stress leading to the bending of the microcantilever. The released binding energy appears to be almost exclusively channeled to the surface energy, and energy distribution to other channels, including heat, appears to be inactive for this micromechanical system. When this excess surface energy is released, the microcantilever relaxes back to the original state, and the relaxation time depends on the particular binding process involved. Such vapor phase experiments were conducted for three binding processes: physisorption, hydrogen bonding, and chemisorption. Binding energies for these three processes were also estimated.

  6. An energy conservation approach to adsorbate-induced surface stress and the extraction of binding energy using nanomechanics

    Energy Technology Data Exchange (ETDEWEB)

    Pinnaduwage, Lal A; Boiadjiev, Vassil I; Hawk, John E; Gehl, Anthony C [Oak Ridge National Laboratory, PO Box 2008, Oak Ridge, TN 37831-6122 (United States); Fernando, Gayanath W [Physics Department, University of Connecticut, Storrs, CT 06269 (United States); Wijewardhana, L C Rohana [Physics Department, University of Cincinnati, Cincinnati, OH 45221 (United States)

    2008-03-12

    Surface stress induced by molecular adsorption in three different binding processes has been studied experimentally using a microcantilever sensor. A comprehensive free-energy analysis based on an energy conservation approach is proposed to explain the experimental observations. We show that when guest molecules bind to atoms/molecules on a microcantilever surface, the released binding energy is retained in the host surface, leading to a metastable state where the excess energy on the surface is manifested as an increase in surface stress leading to the bending of the microcantilever. The released binding energy appears to be almost exclusively channeled to the surface energy, and energy distribution to other channels, including heat, appears to be inactive for this micromechanical system. When this excess surface energy is released, the microcantilever relaxes back to the original state, and the relaxation time depends on the particular binding process involved. Such vapor phase experiments were conducted for three binding processes: physisorption, hydrogen bonding, and chemisorption. Binding energies for these three processes were also estimated.

  7. Experimental demonstration of tunable directional excitation of surface plasmon polaritons with a subwavelength metallic double slit

    Science.gov (United States)

    Li, Xiaowei; Tan, Qiaofeng; Bai, Benfeng; Jin, Guofan

    2011-06-01

    We demonstrate experimentally the directional excitation of surface plasmon polaritons (SPPs) on a metal film by a subwavelength double slit under backside illumination, based on the interference of SPPs generated by the two slits. By varying the incident angle, the SPPs can be tunably directed into two opposite propagating directions with a predetermined splitting ratio. Under certain incident angle, unidirectional SPP excitation can be achieved. This compact directional SPP coupler is potentially useful for many on-chip applications. As an example, we show the integration of the double-slit couplers with SPP Bragg mirrors, which can effectively realize selective coupling of SPPs into different ports in an integrated plasmonic chip.

  8. Development and Demonstration of Sustainable Surface Infrastructure for Moon/Mars Exploration

    Science.gov (United States)

    Sanders, Gerald B.; Larson, William E.; Picard, Martin

    2011-01-01

    For long-term human exploration of the Moon and Mars to be practical, affordable, and sustainable, future missions must be able to identify and utilize resources at the site of exploration. The ability to characterize, extract, processes, and separate products from local material, known as In-Situ Resource Utilization (ISRU), can provide significant reductions in launch mass, logistics, and development costs while reducing risk through increased mission flexibility and protection as well as increased mission capabilities in the areas of power and transportation. Making mission critical consumables like propellants, fuel cell reagents and life support gases, as well as in-situ crew/hardware protection and energy storage capabilities can significantly enhance robotic and human science and exploration missions, however other mission systems need to be designed to interface with and utilize these in-situ developed products and services from the start or the benefits will be minimized or eliminated. This requires a level of surface and transportation system development coordination not typically utilized during early technology and system development activities. An approach being utilized by the US National Aeronautics and Space Administration and the Canadian Space Agency has been to utilize joint analogue field demonstrations to focus technology development activities to demonstrate and integrate new and potentially game changing. mission critical capabilities that would enable an affordable and sustainable surface infrastructure for lunar and Mars robotic and human exploration. Two analogue field tests performed in November 2008 and February 2010 demonstrated first generation capabilities for lunar resource prospecting, exploration site preparation, and oxygen extraction from regolith while initiating integration with mobility, science, fuel cell power, and propulsion disciplines. A third analogue field test currently planned for June 2012 will continue and expand

  9. Characterisation of peptide microarrays for studying antibody-antigen binding using surface plasmon resonance imagery.

    Directory of Open Access Journals (Sweden)

    Claude Nogues

    Full Text Available BACKGROUND: Non-specific binding to biosensor surfaces is a major obstacle to quantitative analysis of selective retention of analytes at immobilized target molecules. Although a range of chemical antifouling monolayers has been developed to address this problem, many macromolecular interactions still remain refractory to analysis due to the prevalent high degree of non-specific binding. We describe how we use the dynamic process of the formation of self assembling monolayers and optimise physical and chemical properties thus reducing considerably non-specific binding and allowing analysis of specific binding of analytes to immobilized target molecules. METHODOLOGY/PRINCIPAL FINDINGS: We illustrate this approach by the production of specific protein arrays for the analysis of interactions between the 65kDa isoform of human glutamate decarboxylase (GAD65 and a human monoclonal antibody. Our data illustrate that we have effectively eliminated non-specific interactions with the surface containing the immobilised GAD65 molecules. The findings have several implications. First, this approach obviates the dubious process of background subtraction and gives access to more accurate kinetic and equilibrium values that are no longer contaminated by multiphase non-specific binding. Second, an enhanced signal to noise ratio increases not only the sensitivity but also confidence in the use of SPR to generate kinetic constants that may then be inserted into van't Hoff type analyses to provide comparative DeltaG, DeltaS and DeltaH values, making this an efficient, rapid and competitive alternative to ITC measurements used in drug and macromolecular-interaction mechanistic studies. Third, the accuracy of the measurements allows the application of more intricate interaction models than simple Langmuir monophasic binding. CONCLUSIONS: The detection and measurement of antibody binding by the type 1 diabetes autoantigen GAD65 represents an example of an antibody

  10. Predicting Ligand Binding Sites on Protein Surfaces by 3-Dimensional Probability Density Distributions of Interacting Atoms

    Science.gov (United States)

    Jian, Jhih-Wei; Elumalai, Pavadai; Pitti, Thejkiran; Wu, Chih Yuan; Tsai, Keng-Chang; Chang, Jeng-Yih; Peng, Hung-Pin; Yang, An-Suei

    2016-01-01

    Predicting ligand binding sites (LBSs) on protein structures, which are obtained either from experimental or computational methods, is a useful first step in functional annotation or structure-based drug design for the protein structures. In this work, the structure-based machine learning algorithm ISMBLab-LIG was developed to predict LBSs on protein surfaces with input attributes derived from the three-dimensional probability density maps of interacting atoms, which were reconstructed on the query protein surfaces and were relatively insensitive to local conformational variations of the tentative ligand binding sites. The prediction accuracy of the ISMBLab-LIG predictors is comparable to that of the best LBS predictors benchmarked on several well-established testing datasets. More importantly, the ISMBLab-LIG algorithm has substantial tolerance to the prediction uncertainties of computationally derived protein structure models. As such, the method is particularly useful for predicting LBSs not only on experimental protein structures without known LBS templates in the database but also on computationally predicted model protein structures with structural uncertainties in the tentative ligand binding sites. PMID:27513851

  11. Decontamination and radioactivity measurement on building surfaces related to dismantling of Japan power demonstration reactor (JPDR)

    International Nuclear Information System (INIS)

    Hatakeyama, Mutsuo; Tachibana, Mitsuo; Yanagihara, Satoshi

    1997-12-01

    In the final stage of dismantling activities for decommissioning a nuclear power plant, building structures have to be demolished to release the site for unrestricted use. Since building structures are generally made from massive reinforced concrete materials, it is not a rational way to treat all concrete materials arising from its demolition as radioactive waste. Segregation of radioactive parts from building structures is therefore indispensable. The rational procedures were studied for demolition of building structures by treating arising waste as non-radioactive materials, based on the concept established by Nuclear Safety Commission, then these were implemented in the following way by the JPDR dismantling demonstration project. Areas of the JPDR facilities are categorized into two groups : possibly contaminated areas, and possibly non-contaminated areas, based on the document of the reactor operation. Radioactivity on the building surfaces was then measured to confirm that the qualitative categorization is reasonable. After that, building surfaces were decontaminated in such a way that the contaminated layers were removed with enough margin to separate radioactive parts from non-radioactive building structures. Thought it might be possible to demolish the building structures by treating arising waste as non-radioactive materials, confirmation survey for radioactivity was conducted to show that there is no artificial radioactive nuclides produced by operation in the facility. This report describes the procedures studied on measurement of radioactivity and decontamination, and the results of its implementation in the JPDR dismantling demonstration project. (author)

  12. Effect of structural modulation and thickness of a graphene overlayer on the binding energy of the Rashba-type surface state of Ir(111)

    International Nuclear Information System (INIS)

    Sánchez-Barriga, J; Marchenko, D; Rader, O; Varykhalov, A; Bihlmayer, G; Wortmann, D

    2013-01-01

    The Ir(111) surface is known to host a surface state with a giant spin–orbit splitting due to the Rashba effect. This surface state is stable even in air when Ir is protected with an epitaxial graphene overlayer. In the present paper, we reveal an effect allowing one to tune the binding energy of this spin-split surface state up and down and demonstrate the practical application of this effect by two different approaches. The first approach is related to a decoration of the moiré pattern of single-layer graphene on Ir(111) by self-assembled nanoclusters of different compositions. The clusters locally pin graphene to the Ir substrate and enhance the amplitude of its structural corrugation, which, in turn, leads to an increase in the surface state binding energy. The second approach is related to the synthesis of few-layer graphene on Ir(111) by segregation of carbon. Additional graphene layers induce a shift of the Ir surface state towards lower binding energies and bring it almost to the Fermi level. Based on density functional calculations performed for the graphene/Ir(111) system, we show that in both cases the effect causing the binding energy shifts is intimately related to the distance between graphene and the Ir surface, which is subject to change due to deposition of clusters or by increasing the amount of graphene overlayers. In contrast, the observed spin–orbit splitting of the Ir(111) surface state remains remarkably robust and constant in all cases. Our theoretical analysis reveals that such stability can be explained by the localization properties of the Ir surface state that is a deep surface resonance. (paper)

  13. A simplified ALARA approach to demonstration of compliance with surface contaminated object regulatory requirements

    International Nuclear Information System (INIS)

    Pope, R.B.; Shappert, L.B.; Michelhaugh, R.D.; Boyle, R.W.; Cook, J.C.

    1998-02-01

    The US Department of Transportation (DOT) and the US Nuclear Regulatory Commission (NRC) have jointly prepared a comprehensive set of draft guidance for consignors and inspectors to use when applying the newly imposed regulatory requirements for low specific activity (LSA) material and surface contaminated objects (SCOs). The guidance is being developed to facilitate compliance with the new LSA material and SCO requirements, not to impose additional requirements. These new requirements represent, in some areas, significant departures from the manner in which packaging and transportation of these materials and objects were previously controlled. On occasion, it may be appropriate to use conservative approaches to demonstrate compliance with some of the requirements, ensuring that personnel are not exposed to radiation at unnecessary levels, so that exposures are kept as low as reasonably achievable (ALARA). In the draft guidance, one such approach would assist consignors preparing a shipment of a large number of SCOs in demonstrating compliance without unnecessarily exposing personnel. In applying this approach, users need to demonstrate that four conditions are met. These four conditions are used to categorize non-activated, contaminated objects as SCO-2. It is expected that, by applying this approach, it will be possible to categorize a large number of small contaminated objects as SCO-2 without the need for detailed, quantitative measurements of fixed, accessible contamination, or of total (fixed and non-fixed) contamination on inaccessible surfaces. The method, which is based upon reasoned argument coupled with limited measurements and the application of a sum of fractions rule, is described and examples of its use are provided

  14. Woody vegetation and succession on the Fonde surface mine demonstration area, Bell County, Kentucky

    International Nuclear Information System (INIS)

    Wade, G.L.; Thompson, R.L.

    1999-01-01

    The long term impact of surface mining on vegetation and plant succession has always been of concern to environmentalists and residents of Appalachia. The Fonde Surface Mine Demonstration Area is a 7.3-ha, NE-NW-aspect contour coal mine at an elevation of 562 m. It was reclaimed in 1965 to show state-of-the-art surface mine reclamation techniques consistent with then-current law and regulations after coal mining in 1959 and 1963. The mine spoils were lightly graded to control erosion and crates a bench with water control and two sediment ponds. Soil pH ranged from 2.8 to 5.9. About 80 percent of the mine was planted with 18 tree and shrub species including plantations of mixed pine, mixed hardwoods, black locust, and shrubs for wildlife. In a complete floristic inventory conducted 25 years later, the authors found the woody flora consisted of 34 families, 53 genera, and 70 species including 7 exotics. This inventory of the Fonde mine shows that a diverse forest vegetation can be reestablished after extreme disturbances in Appalachia. Black locust, yellow poplar, and Virginia pine reproduction varied significantly among plantation types. Canopy tree species significantly affected ground layer cover, total species richness, number of tree seedling species, and total number of tree seedlings present. Mine soil type affected ground layer percent cover and total species richness. Pre-SMCRA (Surface Mining Control and Reclamation Act of 1977) reclaimed and inventoried mines can be used to evaluate biodiversity on post-SMCRA mines

  15. Woody vegetation and succession on the Fonde surface mine demonstration area, Bell County, Kentucky

    Energy Technology Data Exchange (ETDEWEB)

    Wade, G.L.; Thompson, R.L.

    1999-07-01

    The long term impact of surface mining on vegetation and plant succession has always been of concern to environmentalists and residents of Appalachia. The Fonde Surface Mine Demonstration Area is a 7.3-ha, NE-NW-aspect contour coal mine at an elevation of 562 m. It was reclaimed in 1965 to show state-of-the-art surface mine reclamation techniques consistent with then-current law and regulations after coal mining in 1959 and 1963. The mine spoils were lightly graded to control erosion and crates a bench with water control and two sediment ponds. Soil pH ranged from 2.8 to 5.9. About 80 percent of the mine was planted with 18 tree and shrub species including plantations of mixed pine, mixed hardwoods, black locust, and shrubs for wildlife. In a complete floristic inventory conducted 25 years later, the authors found the woody flora consisted of 34 families, 53 genera, and 70 species including 7 exotics. This inventory of the Fonde mine shows that a diverse forest vegetation can be reestablished after extreme disturbances in Appalachia. Black locust, yellow poplar, and Virginia pine reproduction varied significantly among plantation types. Canopy tree species significantly affected ground layer cover, total species richness, number of tree seedling species, and total number of tree seedlings present. Mine soil type affected ground layer percent cover and total species richness. Pre-SMCRA (Surface Mining Control and Reclamation Act of 1977) reclaimed and inventoried mines can be used to evaluate biodiversity on post-SMCRA mines.

  16. Analysis of surface binding sites (SBSs) in carbohydrate active enzymes with focus on glycoside hydrolase families 13 and 77

    DEFF Research Database (Denmark)

    Cockburn, Darrell; Wilkens, Casper; Ruzanski, Christian

    2014-01-01

    Surface binding sites (SBSs) interact with carbohydrates outside of the enzyme active site. They are frequently situated on catalytic domains and are distinct from carbohydrate binding modules (CBMs). SBSs are found in a variety of enzymes and often seen in crystal structures. Notably about half ...

  17. Positron emission tomography with (18F)methylspiperone demonstrates D2 dopamine receptor binding differences of clozapine and haloperidol

    International Nuclear Information System (INIS)

    Karbe, H.; Wienhard, K.; Huber, M.; Herholz, K.; Heiss, W.D.; Hamacher, K.; Coenen, H.H.; Stoecklin, G.; Loevenich, A.

    1991-01-01

    Four schizophrenic patients were investigated with dynamic positron emission tomography (PET) using ( 18 F)fluorodeoxyglucose (FDG) and ( 18 F)methylspiperone (MSP) as tracers. Two schizophrenics were on haloperidol therapy at the time of MSP PET. The other two schizophrenics were treated with clozapine, in one of them MSP PET was carried out twice with different daily doses (100 mg and 450mg respectively). Neuroleptic serum levels were measured in all patients. Results were compared with MSP PET of two drugfree male control subjects and with a previous fluoroethylspiperone (FESP) study of normals. Three hours after tracer injection specific binding of MSP was observed in the striatum in all cases. The striatum to cerebellum ratio was used to estimate the degree of neuroleptic-caused striatal D 2 dopamine receptor occupancy. In the haloperidol treated patients MSP binding was significantly decreased, whereas in the clozapine treated patients striatum to cerebellum ratio was normal. Even the increase of clozapine dose in the same patient had no influence on this ratio. Despite the smaller number of patients the study shows for the first time in humans that striatal MSP binding reflects the different D 2 dopamine receptor affinities of clozapine and haloperidol. (authors)

  18. Binding and distribution studies in the SENCAR mouse of compounds demonstrating a route-dependent tumorigenic effect

    International Nuclear Information System (INIS)

    Carlson, G.P.; Fossa, A.A.; Morse, M.A.; Weaver, P.M.

    1986-01-01

    Previous investigators have determined that benzo(a)pyrene [B(a)P] was much more effective in causing skin papillomas if applied topically than when administered orally in the initiation-promotion assay in SENCAR mouse. Conversely, urethane and acrylamide caused a higher percentage of mice to develop papillomas and induced more tumors per mouse when given orally. In an attempt to understand the reason for this discrepancy in route dependency, 3 H-benzo(a)pyrene, 14 C-acrylamide were administered as single doses orally or topically to male SENCAR mice. Distribution in skin, stomach, liver, and lung was determined for time periods up to 48 hr. The binding of these compounds to DNA, RNA, and protein in these tissues was determined 6 and 48 hr after administration. For all three compounds, high concentrations were found in the skin following topical application, but very little material reached this target organ following oral administration. In contrast, the internal organs generally contained more material after oral administration. The binding of label compounds to DNA, RNA, and protein generally reflected the distribution data, thus more compound was bound in the stomach, liver, and lung after oral administration compared to topical application, whereas the opposite was true for the skin. This finding was particularly evident for B(a)P. The results suggest that differences in distribution to the skin and binding to macromolecules following oral or topical administration cannot explain the greater tumorigenicity of urethane and acrylamide after oral administration in the SENCAR mouse

  19. Photoelectron binding energy shifts observed during oxidation of group IIA, IIIA and IVA elemental surfaces

    International Nuclear Information System (INIS)

    Heide, P.A.W. van der

    2006-01-01

    An extensive re-evaluation of XPS binding energies (BE's) and binding energy shifts (ΔBE's) from metals, oxides and the carbonates of the group II, III and IVA elements (exceptions are Be, Mg and Hf) has been carried out using a substrate specific BE referencing approach. From this, O-1s BE's are found to fall into surface oxide, bulk oxide and carbonate groupings, with bulk oxides showing the lowest BE's followed by surface oxides (+∼1.5 eV) and then carbonates (+∼3.0 eV). The O-1s BE's from the bulk oxides also appear to scale with 1/d, where d is inter-atomic distance. The same is noted in the ΔBE's observed from the metallic counterparts during oxidation of the elemental surfaces. This, and the decreasing BE exhibited by Ca, Sr and Ba on oxidation is explained within the charge potential model as resulting from competing inter- and intra-atomic effects, and is shown to be consistent with partial covalency arguments utilizing Madulung potentials. The ΔBE's also fall into groups according to the elements location in the periodic table, i.e. s, p or d block. These trends open up the possibility of approximating ΔBE's arising from initial and final state effects, and bond distances

  20. Post-Spaceflight (STS-135 Mouse Splenocytes Demonstrate Altered Activation Properties and Surface Molecule Expression.

    Directory of Open Access Journals (Sweden)

    Shen-An Hwang

    Full Text Available Alterations in immune function have been documented during or post-spaceflight and in ground based models of microgravity. Identification of immune parameters that are dysregulated during spaceflight is an important step in mitigating crew health risks during deep space missions. The in vitro analysis of leukocyte activity post-spaceflight in both human and animal species is primarily focused on lymphocytic function. This report completes a broader spectrum analysis of mouse lymphocyte and monocyte changes post 13 days orbital flight (mission STS-135. Analysis includes an examination in surface markers for cell activation, and antigen presentation and co-stimulatory molecules. Cytokine production was measured after stimulation with T-cell mitogen or TLR-2, TLR-4, or TLR-5 agonists. Splenocyte surface marker analysis immediate post-spaceflight and after in vitro culture demonstrated unique changes in phenotypic populations between the flight mice and matched treatment ground controls. Post-spaceflight splenocytes (flight splenocytes had lower expression intensity of CD4+CD25+ and CD8+CD25+ cells, lower percentage of CD11c+MHC II+ cells, and higher percentage of CD11c+MHC I+ populations compared to ground controls. The flight splenocytes demonstrated an increase in phagocytic activity. Stimulation with ConA led to decrease in CD4+ population but increased CD4+CD25+ cells compared to ground controls. Culturing with TLR agonists led to a decrease in CD11c+ population in splenocytes isolated from flight mice compared to ground controls. Consequently, flight splenocytes with or without TLR-agonist stimulation showed a decrease in CD11c+MHC I+, CD11c+MHC II+, and CD11c+CD86+ cells compared to ground controls. Production of IFN-γ was decreased and IL-2 was increased from ConA stimulated flight splenocytes. This study demonstrated that expression of surface molecules can be affected by conditions of spaceflight and impaired responsiveness persists under

  1. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

    Directory of Open Access Journals (Sweden)

    Mariya Tarazanova

    2017-09-01

    Full Text Available Surface properties of bacteria are determined by the molecular composition of the cell wall and they are important for interactions of cells with their environment. Well-known examples of bacterial interactions with surfaces are biofilm formation and the fermentation of solid materials like food and feed. Lactococcus lactis is broadly used for the fermentation of cheese and buttermilk and it is primarily isolated from either plant material or the dairy environment. In this study, we characterized surface hydrophobicity, charge, emulsification properties, and the attachment to milk proteins of 55 L. lactis strains in stationary and exponential growth phases. The attachment to milk protein was assessed through a newly developed flow cytometry-based protocol. Besides finding a high degree of biodiversity, phenotype-genotype matching allowed the identification of candidate genes involved in the modification of the cell surface. Overexpression and gene deletion analysis allowed to verify the predictions for three identified proteins that altered surface hydrophobicity and attachment of milk proteins. The data also showed that lactococci isolated from a dairy environment bind higher amounts of milk proteins when compared to plant isolates. It remains to be determined whether the alteration of surface properties also has potential to alter starter culture functionalities.

  2. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

    Science.gov (United States)

    Tarazanova, Mariya; Huppertz, Thom; Beerthuyzen, Marke; van Schalkwijk, Saskia; Janssen, Patrick; Wels, Michiel; Kok, Jan; Bachmann, Herwig

    2017-01-01

    Surface properties of bacteria are determined by the molecular composition of the cell wall and they are important for interactions of cells with their environment. Well-known examples of bacterial interactions with surfaces are biofilm formation and the fermentation of solid materials like food and feed. Lactococcus lactis is broadly used for the fermentation of cheese and buttermilk and it is primarily isolated from either plant material or the dairy environment. In this study, we characterized surface hydrophobicity, charge, emulsification properties, and the attachment to milk proteins of 55 L. lactis strains in stationary and exponential growth phases. The attachment to milk protein was assessed through a newly developed flow cytometry-based protocol. Besides finding a high degree of biodiversity, phenotype-genotype matching allowed the identification of candidate genes involved in the modification of the cell surface. Overexpression and gene deletion analysis allowed to verify the predictions for three identified proteins that altered surface hydrophobicity and attachment of milk proteins. The data also showed that lactococci isolated from a dairy environment bind higher amounts of milk proteins when compared to plant isolates. It remains to be determined whether the alteration of surface properties also has potential to alter starter culture functionalities. PMID:28936202

  3. Three-dimensional structures of the mammalian multidrug resistance P-glycoprotein demonstrate major conformational changes in the transmembrane domains upon nucleotide binding.

    Science.gov (United States)

    Rosenberg, Mark F; Kamis, Alhaji Bukar; Callaghan, Richard; Higgins, Christopher F; Ford, Robert C

    2003-03-07

    P-glycoprotein is an ATP-binding cassette transporter that is associated with multidrug resistance and the failure of chemotherapy in human patients. We have previously shown, based on two-dimensional projection maps, that P-glycoprotein undergoes conformational changes upon binding of nucleotide to the intracellular nucleotide binding domains. Here we present the three-dimensional structures of P-glycoprotein in the presence and absence of nucleotide, at a resolution limit of approximately 2 nm, determined by electron crystallography of negatively stained crystals. The data reveal a major reorganization of the transmembrane domains throughout the entire depth of the membrane upon binding of nucleotide. In the absence of nucleotide, the two transmembrane domains form a single barrel 5-6 nm in diameter and about 5 nm deep with a central pore that is open to the extracellular surface and spans much of the membrane depth. Upon binding nucleotide, the transmembrane domains reorganize into three compact domains that are each 2-3 nm in diameter and 5-6 nm deep. This reorganization opens the central pore along its length in a manner that could allow access of hydrophobic drugs (transport substrates) directly from the lipid bilayer to the central pore of the transporter.

  4. Weakly hydrated surfaces and the binding interactions of small biological solutes.

    Science.gov (United States)

    Brady, John W; Tavagnacco, Letizia; Ehrlich, Laurent; Chen, Mo; Schnupf, Udo; Himmel, Michael E; Saboungi, Marie-Louise; Cesàro, Attilio

    2012-04-01

    Extended planar hydrophobic surfaces, such as are found in the side chains of the amino acids histidine, phenylalanine, tyrosine, and tryptophan, exhibit an affinity for the weakly hydrated faces of glucopyranose. In addition, molecular species such as these, including indole, caffeine, and imidazole, exhibit a weak tendency to pair together by hydrophobic stacking in aqueous solution. These interactions can be partially understood in terms of recent models for the hydration of extended hydrophobic faces and should provide insight into the architecture of sugar-binding sites in proteins.

  5. Analysis of Surface Binding Sites (SBS) within GH62, GH13, and GH77

    DEFF Research Database (Denmark)

    Wilkens, Casper; Cockburn, Darrell; Andersen, Susan

    2015-01-01

    Certain interactions between carbohydrate active enzymes and polysaccharides involve surface binding sites (SBS) situated on catalytic domains outside of the active site. We recently undertook to develop a toolbox for SBS identification and characterization. In affinity gel electrophoresis (AGE...... of the reported SBSs. In GH13 SBSs have been seen in 17 subfamilies including SBSs with highly diverse functions in the same enzyme. Circumstantial evidence is provided for an SBS in the GH77 MalQ from Escherichia coli, the bacterial orthologue of Arabidopsis DPE2 involved in starch metabolism. Furthermore...

  6. A demonstration of adjoint methods for multi-dimensional remote sensing of the atmosphere and surface

    Science.gov (United States)

    Martin, William G. K.; Hasekamp, Otto P.

    2018-01-01

    In previous work, we derived the adjoint method as a computationally efficient path to three-dimensional (3D) retrievals of clouds and aerosols. In this paper we will demonstrate the use of adjoint methods for retrieving two-dimensional (2D) fields of cloud extinction. The demonstration uses a new 2D radiative transfer solver (FSDOM). This radiation code was augmented with adjoint methods to allow efficient derivative calculations needed to retrieve cloud and surface properties from multi-angle reflectance measurements. The code was then used in three synthetic retrieval studies. Our retrieval algorithm adjusts the cloud extinction field and surface albedo to minimize the measurement misfit function with a gradient-based, quasi-Newton approach. At each step we compute the value of the misfit function and its gradient with two calls to the solver FSDOM. First we solve the forward radiative transfer equation to compute the residual misfit with measurements, and second we solve the adjoint radiative transfer equation to compute the gradient of the misfit function with respect to all unknowns. The synthetic retrieval studies verify that adjoint methods are scalable to retrieval problems with many measurements and unknowns. We can retrieve the vertically-integrated optical depth of moderately thick clouds as a function of the horizontal coordinate. It is also possible to retrieve the vertical profile of clouds that are separated by clear regions. The vertical profile retrievals improve for smaller cloud fractions. This leads to the conclusion that cloud edges actually increase the amount of information that is available for retrieving the vertical profile of clouds. However, to exploit this information one must retrieve the horizontally heterogeneous cloud properties with a 2D (or 3D) model. This prototype shows that adjoint methods can efficiently compute the gradient of the misfit function. This work paves the way for the application of similar methods to 3D remote

  7. T Tank Farm Interim Surface Barrier Demonstration -- Vadose Zone Monitoring FY07 Report

    International Nuclear Information System (INIS)

    Zhang, Z. F.; Strickland, Christopher E.; Keller, Jason M.; Wittreich, Curtis D.; Sydnor, Harold A.

    2008-01-01

    CH2M HILL Hanford Group, Inc. is currently in the process of constructing a temporary surface barrier over a portion of the T Tank Farm as part of the T farm Interim Surface Barrier Demonstration Project. The surface barrier is designed to prevent the infiltration of precipitation into the contaminated soil zone created by the Tank T-106 leak and minimize movement of the contamination. As part of the demonstration effort, vadose zone moisture monitoring is being performed to assess the effectiveness of the barrier at reducing soil moisture. A solar-powered and remotely-controlled system was installed to continuously monitor soil water conditions in four instrument nests (i.e., A, B, C, and D) and the site meteorological condition. Each instrument nest was composed of a capacitance probe with multiple sensors, multiple heat-dissipation units, a neutron probe access tube and a datalogger. Nests A and B also contained a drain gauge each. The principle variables monitored for this purpose are soil-water content, soil-water pressure, and soil-water flux. In addition to these, soil temperature, precipitation, and air temperature are measured. Data from each of the dataloggers were transmitted remotely to the receiving computer. The neutron probe access tube was used to perform quarterly manual measurements of soil-water content using a neutron probe. This monitoring system was used to assess the soil water conditions in the soil outside and within the footprint of the surface barrier to be emplaced in the Hanford T Tank Farm. Data to date is baseline under the condition without the interim surface barrier in place. All the instruments except the two drain gauges were functional in FY07. The capacitance-probe measurements showed that the soil-moisture content at relatively shallow depths (e.g., 0.6 and 0.9 m) was increasing since October 2006 and reached the highest in early January 2007 followed by a slight decrease. Soil-moisture contents at the depths of 1.3 m and

  8. Implementation of a Surface Electromyography-Based Upper Extremity Exoskeleton Controller Using Learning from Demonstration

    Science.gov (United States)

    Arenas, Ana M.; Sun, Tingxiao

    2018-01-01

    Upper-extremity exoskeletons have demonstrated potential as augmentative, assistive, and rehabilitative devices. Typical control of upper-extremity exoskeletons have relied on switches, force/torque sensors, and surface electromyography (sEMG), but these systems are usually reactionary, and/or rely on entirely hand-tuned parameters. sEMG-based systems may be able to provide anticipatory control, since they interface directly with muscle signals, but typically require expert placement of sensors on muscle bodies. We present an implementation of an adaptive sEMG-based exoskeleton controller that learns a mapping between muscle activation and the desired system state during interaction with a user, generating a personalized sEMG feature classifier to allow for anticipatory control. This system is robust to novice placement of sEMG sensors, as well as subdermal muscle shifts. We validate this method with 18 subjects using a thumb exoskeleton to complete a book-placement task. This learning-from-demonstration system for exoskeleton control allows for very short training times, as well as the potential for improvement in intent recognition over time, and adaptation to physiological changes in the user, such as those due to fatigue. PMID:29401754

  9. Implementation of a Surface Electromyography-Based Upper Extremity Exoskeleton Controller Using Learning from Demonstration

    Directory of Open Access Journals (Sweden)

    Ho Chit Siu

    2018-02-01

    Full Text Available Upper-extremity exoskeletons have demonstrated potential as augmentative, assistive, and rehabilitative devices. Typical control of upper-extremity exoskeletons have relied on switches, force/torque sensors, and surface electromyography (sEMG, but these systems are usually reactionary, and/or rely on entirely hand-tuned parameters. sEMG-based systems may be able to provide anticipatory control, since they interface directly with muscle signals, but typically require expert placement of sensors on muscle bodies. We present an implementation of an adaptive sEMG-based exoskeleton controller that learns a mapping between muscle activation and the desired system state during interaction with a user, generating a personalized sEMG feature classifier to allow for anticipatory control. This system is robust to novice placement of sEMG sensors, as well as subdermal muscle shifts. We validate this method with 18 subjects using a thumb exoskeleton to complete a book-placement task. This learning-from-demonstration system for exoskeleton control allows for very short training times, as well as the potential for improvement in intent recognition over time, and adaptation to physiological changes in the user, such as those due to fatigue.

  10. A demonstration of adjoint methods for multi-dimensional remote sensing of the atmosphere and surface

    International Nuclear Information System (INIS)

    Martin, William G.K.; Hasekamp, Otto P.

    2018-01-01

    Highlights: • We demonstrate adjoint methods for atmospheric remote sensing in a two-dimensional setting. • Searchlight functions are used to handle the singularity of measurement response functions. • Adjoint methods require two radiative transfer calculations to evaluate the measurement misfit function and its derivatives with respect to all unknown parameters. • Synthetic retrieval studies show the scalability of adjoint methods to problems with thousands of measurements and unknown parameters. • Adjoint methods and the searchlight function technique are generalizable to 3D remote sensing. - Abstract: In previous work, we derived the adjoint method as a computationally efficient path to three-dimensional (3D) retrievals of clouds and aerosols. In this paper we will demonstrate the use of adjoint methods for retrieving two-dimensional (2D) fields of cloud extinction. The demonstration uses a new 2D radiative transfer solver (FSDOM). This radiation code was augmented with adjoint methods to allow efficient derivative calculations needed to retrieve cloud and surface properties from multi-angle reflectance measurements. The code was then used in three synthetic retrieval studies. Our retrieval algorithm adjusts the cloud extinction field and surface albedo to minimize the measurement misfit function with a gradient-based, quasi-Newton approach. At each step we compute the value of the misfit function and its gradient with two calls to the solver FSDOM. First we solve the forward radiative transfer equation to compute the residual misfit with measurements, and second we solve the adjoint radiative transfer equation to compute the gradient of the misfit function with respect to all unknowns. The synthetic retrieval studies verify that adjoint methods are scalable to retrieval problems with many measurements and unknowns. We can retrieve the vertically-integrated optical depth of moderately thick clouds as a function of the horizontal coordinate. It is also

  11. MULTIPLE IMAGING TECHNIQUES DEMONSTRATE THE MANIPULATION OF SURFACES TO REDUCE BACTERIAL CONTAMINATION

    Science.gov (United States)

    Surface imaging techniques were combined to determine appropriate manipulation of technologically important surfaces for commercial applications. Stainless steel surfaces were engineered to reduce bacterial contamination, biofilm formation, and corrosion during product processing...

  12. Decorin core protein (decoron) shape complements collagen fibril surface structure and mediates its binding.

    Science.gov (United States)

    Orgel, Joseph P R O; Eid, Aya; Antipova, Olga; Bella, Jordi; Scott, John E

    2009-09-15

    Decorin is the archetypal small leucine rich repeat proteoglycan of the vertebrate extracellular matrix (ECM). With its glycosaminoglycuronan chain, it is responsible for stabilizing inter-fibrillar organization. Type I collagen is the predominant member of the fibrillar collagen family, fulfilling both organizational and structural roles in animal ECMs. In this study, interactions between decoron (the decorin core protein) and binding sites in the d and e(1) bands of the type I collagen fibril were investigated through molecular modeling of their respective X-ray diffraction structures. Previously, it was proposed that a model-based, highly curved concave decoron interacts with a single collagen molecule, which would form extensive van der Waals contacts and give rise to strong non-specific binding. However, the large well-ordered aggregate that is the collagen fibril places significant restraints on modes of ligand binding and necessitates multi-collagen molecular contacts. We present here a relatively high-resolution model of the decoron-fibril collagen complex. We find that the respective crystal structures complement each other well, although it is the monomeric form of decoron that shows the most appropriate shape complementarity with the fibril surface and favorable calculated energies of interaction. One molecule of decoron interacts with four to six collagen molecules, and the binding specificity relies on a large number of hydrogen bonds and electrostatic interactions, primarily with the collagen motifs KXGDRGE and AKGDRGE (d and e(1) bands). This work helps us to understand collagen-decorin interactions and the molecular architecture of the fibrillar ECM in health and disease.

  13. Decorin core protein (decoron shape complements collagen fibril surface structure and mediates its binding.

    Directory of Open Access Journals (Sweden)

    Joseph P R O Orgel

    2009-09-01

    Full Text Available Decorin is the archetypal small leucine rich repeat proteoglycan of the vertebrate extracellular matrix (ECM. With its glycosaminoglycuronan chain, it is responsible for stabilizing inter-fibrillar organization. Type I collagen is the predominant member of the fibrillar collagen family, fulfilling both organizational and structural roles in animal ECMs. In this study, interactions between decoron (the decorin core protein and binding sites in the d and e(1 bands of the type I collagen fibril were investigated through molecular modeling of their respective X-ray diffraction structures. Previously, it was proposed that a model-based, highly curved concave decoron interacts with a single collagen molecule, which would form extensive van der Waals contacts and give rise to strong non-specific binding. However, the large well-ordered aggregate that is the collagen fibril places significant restraints on modes of ligand binding and necessitates multi-collagen molecular contacts. We present here a relatively high-resolution model of the decoron-fibril collagen complex. We find that the respective crystal structures complement each other well, although it is the monomeric form of decoron that shows the most appropriate shape complementarity with the fibril surface and favorable calculated energies of interaction. One molecule of decoron interacts with four to six collagen molecules, and the binding specificity relies on a large number of hydrogen bonds and electrostatic interactions, primarily with the collagen motifs KXGDRGE and AKGDRGE (d and e(1 bands. This work helps us to understand collagen-decorin interactions and the molecular architecture of the fibrillar ECM in health and disease.

  14. A trained demonstrator has a calming effect on naïve horses when crossing a novel surface

    DEFF Research Database (Denmark)

    Rørvang, Maria Vilain; Ahrendt, Line Peerstrup; Christensen, Janne Winther

    2015-01-01

    horse being led five times across a novel surface. Immediately afterwards the observer horses were given the opportunity to cross the novel surface themselves to obtain food on the other side. Controls (n = 11) were allowed to observe the demonstrator eating on the opposite side of the novel surface...

  15. Quantitative analysis of rat Ig (sub)classes binding to cell surface antigens

    International Nuclear Information System (INIS)

    Nilsson, R.; Brodin, T.; Sjoegren, H.-O.

    1982-01-01

    An indirect 125 I-labeled protein A assay for detection of cell surface-bound rat immunoglobulins is presented. The assay is quantitative and rapid and detects as little as 1 ng of cell surface-bound Ig. It discriminates between antibodies belonging to different IgG subclasses, IgM and IgA. The authors describe the production and specificity control of the reagents used and show that the test can be used for quantitative analysis. A large number of sera from untreated rats are tested to evaluate the frequency of falsely positive responses and variation due to age, sex and strain of rat. With this test it is relatively easy to quantitate the binding of classes and subclasses of rat immunoglobulins in a small volume (6 μl) of untreated serum. (Auth.)

  16. Lsa63, a newly identified surface protein of Leptospira interrogans binds laminin and collagen IV.

    Science.gov (United States)

    Vieira, Monica L; de Morais, Zenaide M; Gonçales, Amane P; Romero, Eliete C; Vasconcellos, Silvio A; Nascimento, Ana L T O

    2010-01-01

    Leptospira interrogans is the etiological agent of leptospirosis, a zoonotic disease that affects populations worldwide. We have identified in proteomic studies a protein that is encoded by the gene LIC10314 and expressed in virulent strain of L. interrogans serovar Pomona. This protein was predicted to be surface exposed by PSORT program and contains a p83/100 domain identified by BLAST analysis that is conserved in protein antigens of several strains of Borrelia and Treponema spp. The proteins containing this domain have been claimed antigen candidates for serodiagnosis of Lyme borreliosis. Thus, we have cloned the LIC10314 and expressed the protein in Escherichia coli BL21-SI strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. This protein is conserved among several species of pathogenic Leptospira and absent in the saprophytic strain L. biflexa. We confirm by liquid-phase immunofluorescence assays with living organisms that this protein is most likely a new surface leptospiral protein. The ability of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC10314, named Lsa63 (Leptospiral surface adhesin of 63kDa), binds strongly to laminin and collagen IV in a dose-dependent and saturable fashion. In addition, Lsa63 is probably expressed during infection since it was recognized by antibodies of serum samples of confirmed-leptospirosis patients in convalescent phase of the disease. Altogether, the data suggests that this novel identified surface protein may be involved in leptospiral pathogenesis. 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

  17. Roles of multiple surface sites, long substrate binding clefts, and carbohydrate binding modules in the action of amylolytic enzymes on polysaccharide substrates

    DEFF Research Database (Denmark)

    Nielsen, Morten Munch; Seo, E.S.; Dilokpimol, Adiphol

    2008-01-01

    Germinating barley seeds contain multiple forms of alpha-amylase, which are subject to both differential gene expression and differential degradation as part of the repertoire of starch-degrading enzymes. The alpha-amylases are endo-acting and possess a long substrate binding cleft with a charact......Germinating barley seeds contain multiple forms of alpha-amylase, which are subject to both differential gene expression and differential degradation as part of the repertoire of starch-degrading enzymes. The alpha-amylases are endo-acting and possess a long substrate binding cleft...... will address surface sites in both barley alpha-amylase 1 and in the related isozyme 2....

  18. The E1 beta-subunit of pyruvate dehydrogenase is surface-expressed in Lactobacillus plantarum and binds fibronectin.

    Science.gov (United States)

    Vastano, Valeria; Salzillo, Marzia; Siciliano, Rosa A; Muscariello, Lidia; Sacco, Margherita; Marasco, Rosangela

    2014-01-01

    Lactobacillus plantarum is among the species with a probiotic activity. Adhesion of probiotic bacteria to host tissues is an important principle for strain selection, because it represents a crucial step in the colonization process of either pathogens or commensals. Most bacterial adhesins are proteins, and a major target for them is fibronectin, an extracellular matrix glycoprotein. In this study we demonstrate that PDHB, a component of the pyruvate dehydrogenase complex, is a factor contributing to fibronectin-binding in L. plantarum LM3. By means of fibronectin overlay immunoblotting assay, we identified a L. plantarum LM3 surface protein with apparent molecular mass of 35 kDa. Mass spectrometric analysis shows that this protein is the pyruvate dehydrogenase E1 beta-subunit (PDHB). The corresponding pdhB gene is located in a 4-gene cluster encoding pyruvate dehydrogenase. In LM3-B1, carrying a null mutation in pdhB, the 35 kDa adhesin was not anymore detectable by immunoblotting assay. Nevertheless, the pdhB null mutation did not abolish pdhA, pdhC, and pdhD transcription in LM3-B1. By adhesion assays, we show that LM3-B1 cells bind to immobilized fibronectin less efficiently than wild type cells. Moreover, we show that pdhB expression is negatively regulated by the CcpA protein and is induced by bile. Copyright © 2013. Published by Elsevier GmbH.

  19. Endocytosis of wheat germ agglutinin binding sites from the cell surface into a tubular endosomal network.

    Science.gov (United States)

    Raub, T J; Koroly, M J; Roberts, R M

    1990-04-01

    By using fluorescence and electron microscopy, the endocytic pathway encountered by cell surface components after they had bound wheat germ agglutinin (WGA) was visualized. The majority of these components are thought to consist of sialylated glycoproteins (HMWAG) that represent a subpopulation of the total cell surface proteins but most of the externally disposed plasma membrane proteins of the cell. Examination of semi-thin sections by medium- and high-voltage electron microscopy revealed the three-dimensional organization of vesicular and tubular endosomes. Binding of either fluorescein isothiocyanate-, horseradish peroxidase-, or ferritin-conjugated WGA to cells at 4 degrees C showed that the HMWAG were distributed uniformly over the cell surface. Warming of surface-labeled cells to 37 degrees C resulted in the endocytosis of WGA into peripheral endosomes via invagination of regions of both coated and uncoated membrane. The peripheral endosome appeared as isolated complexes comprising a vesicular element (300-400 nm diam.) surrounded by and continuous with tubular cisternae (45-60 nm diam.), which did not interconnect the endosomes. After 30 min or more label also became localized in a network of anastomosing tubules (45-60 nm diam.) that were located in the centrosomal region of the cell. Endocytosed WGA-HMWAG complexes did not become associated with cisternae of the Golgi apparatus, although tubular and vesicular endosomes were noted in the vicinity of the trans-Golgi region. The accumulation of WGA-HMWAG in the endosomes within the centrosomal region was inhibited when cells were incubated at 18 degrees C. None of these compartments contained acid phosphatase activity, a result that is consistent with other data that the HMWAG do not pass through lysosomes initially. The kinetics of labeling were consistent with the interpretation that recycling of most of the WGA binding surface glycoproteins occurred rapidly from early peripheral endosomes followed by the

  20. Testing of an Annular Linear Induction Pump for the Fission Surface Power Technology Demonstration Unit

    Science.gov (United States)

    Polzin, K. A.; Pearson, J. B.; Webster, K.; Godfoy, T. J.; Bossard, J. A.

    2013-01-01

    Results of performance testing of an annular linear induction pump that has been designed for integration into a fission surface power technology demonstration unit are presented. The pump electromagnetically pushes liquid metal (NaK) through a specially-designed apparatus that permits quantification of pump performance over a range of operating conditions. Testing was conducted for frequencies of 40, 55, and 70 Hz, liquid metal temperatures of 125, 325, and 525 C, and input voltages from 30 to 120 V. Pump performance spanned a range of flow rates from roughly 0.3 to 3.1 L/s (4.8 to 49 gpm), and pressure heads of <1 to 104 kPa (<0.15 to 15 psi). The maximum efficiency measured during testing was 5.4%. At the technology demonstration unit operating temperature of 525 C the pump operated over a narrower envelope, with flow rates from 0.3 to 2.75 L/s (4.8 to 43.6 gpm), developed pressure heads from <1 to 55 kPa (<0.15 to 8 psi), and a maximum efficiency of 3.5%. The pump was supplied with three-phase power at 40 and 55 Hz using a variable-frequency motor drive, while power at 55 and 70 Hz was supplied using a variable-frequency power supply. Measured performance of the pump at 55 Hz using either supply exhibited good quantitative agreement. For a given temperature, the peak in efficiency occurred at different flow rates as the frequency was changed, but the maximum value of efficiency was relative insensitive within 0.3% over the frequency range tested, including a scan from 45 to 78 Hz. The objectives of the FSP technology project are as follows:5 • Develop FSP concepts that meet expected surface power requirements at reasonable cost with added benefits over other options. • Establish a nonnuclear hardware-based technical foundation for FSP design concepts to reduce overall development risk. • Reduce the cost uncertainties for FSP and establish greater credibility for flight system cost estimates. • Generate the key nonnuclear products to allow Agency

  1. Binding characteristics of thrombin-activatable fibrinolysis inhibitor to streptococcal surface collagen-like proteins A and B

    NARCIS (Netherlands)

    Seron, Mercedes Valls; Plug, Tom; Marquart, J. Arnoud; Marx, Pauline F.; Herwald, Heiko; de Groot, Philip G.; Meijers, Joost C. M.

    2011-01-01

    Streptococcus pyogenes is the causative agent in a wide range of diseases in humans. Thrombin-activatable fibrinolysis inhibitor (TAFI) binds to collagen-like proteins ScIA and ScIB at the surface of S. pyogenes. Activation of TAFI at this surface redirects inflammation from a transient to chronic

  2. Selecting for Fast Protein-Protein Association As Demonstrated on a Random TEM1 Yeast Library Binding BLIP.

    Science.gov (United States)

    Cohen-Khait, Ruth; Schreiber, Gideon

    2018-04-27

    Protein-protein interactions mediate the vast majority of cellular processes. Though protein interactions obey basic chemical principles also within the cell, the in vivo physiological environment may not allow for equilibrium to be reached. Thus, in vitro measured thermodynamic affinity may not provide a complete picture of protein interactions in the biological context. Binding kinetics composed of the association and dissociation rate constants are relevant and important in the cell. Therefore, changes in protein-protein interaction kinetics have a significant impact on the in vivo activity of the proteins. The common protocol for the selection of tighter binders from a mutant library selects for protein complexes with slower dissociation rate constants. Here we describe a method to specifically select for variants with faster association rate constants by using pre-equilibrium selection, starting from a large random library. Toward this end, we refine the selection conditions of a TEM1-β-lactamase library against its natural nanomolar affinity binder β-lactamase inhibitor protein (BLIP). The optimal selection conditions depend on the ligand concentration and on the incubation time. In addition, we show that a second sort of the library helps to separate signal from noise, resulting in a higher percent of faster binders in the selected library. Fast associating protein variants are of particular interest for drug development and other biotechnological applications.

  3. Surface-associated plasminogen binding of Cryptococcus neoformans promotes extracellular matrix invasion.

    Directory of Open Access Journals (Sweden)

    Jamal Stie

    2009-06-01

    Full Text Available The fungal pathogen Cryptococcus neoformans is a leading cause of illness and death in persons with predisposing factors, including: malignancies, solid organ transplants, and corticosteroid use. C. neoformans is ubiquitous in the environment and enters into the lungs via inhalation, where it can disseminate through the bloodstream and penetrate the central nervous system (CNS, resulting in a difficult to treat and often-fatal infection of the brain, called meningoencephalitis. Plasminogen is a highly abundant protein found in the plasma component of blood and is necessary for the degradation of fibrin, collagen, and other structural components of tissues. This fibrinolytic system is utilized by cancer cells during metastasis and several pathogenic species of bacteria have been found to manipulate the host plasminogen system to facilitate invasion of tissues during infection by modifying the activation of this process through the binding of plasminogen at their surface.The invasion of the brain and the central nervous system by penetration of the protective blood-brain barrier is a prerequisite to the establishment of meningoencephalitis by the opportunistic fungal pathogen C. neoformans. In this study, we examined the ability of C. neoformans to subvert the host plasminogen system to facilitate tissue barrier invasion. Through a combination of biochemical, cell biology, and proteomic approaches, we have shown that C. neoformans utilizes the host plasminogen system to cross tissue barriers, providing support for the hypothesis that plasminogen-binding may contribute to the invasion of the blood-brain barrier by penetration of the brain endothelial cells and underlying matrix. In addition, we have identified the cell wall-associated proteins that serve as plasminogen receptors and characterized both the plasminogen-binding and plasmin-activation potential for this significant human pathogen.The results of this study provide evidence for the

  4. Communication: Photoinduced carbon dioxide binding with surface-functionalized silicon quantum dots

    Science.gov (United States)

    Douglas-Gallardo, Oscar A.; Sánchez, Cristián Gabriel; Vöhringer-Martinez, Esteban

    2018-04-01

    Nowadays, the search for efficient methods able to reduce the high atmospheric carbon dioxide concentration has turned into a very dynamic research area. Several environmental problems have been closely associated with the high atmospheric level of this greenhouse gas. Here, a novel system based on the use of surface-functionalized silicon quantum dots (sf-SiQDs) is theoretically proposed as a versatile device to bind carbon dioxide. Within this approach, carbon dioxide trapping is modulated by a photoinduced charge redistribution between the capping molecule and the silicon quantum dots (SiQDs). The chemical and electronic properties of the proposed SiQDs have been studied with a Density Functional Theory and Density Functional Tight-Binding (DFTB) approach along with a time-dependent model based on the DFTB framework. To the best of our knowledge, this is the first report that proposes and explores the potential application of a versatile and friendly device based on the use of sf-SiQDs for photochemically activated carbon dioxide fixation.

  5. Cyclophilin B mediates cyclosporin A incorporation in human blood T-lymphocytes through the specific binding of complexed drug to the cell surface.

    Science.gov (United States)

    Allain, F; Denys, A; Spik, G

    1996-07-15

    Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein located within intracellular vesicles and released in biological fluids. We recently reported the specific binding of this protein to T-cell surface receptor which is internalized even in the presence of CsA. These results suggest that CyPB might target the drug to lymphocytes and consequently modify its activity. To verify this hypothesis, we have first investigated the binding capacity and internalization of the CsA-CyPB complex in human peripheral blood T-lymphocytes and secondly compared the inhibitory effect of both free and CyPB-complexed CsA on the CD3-induced activation and proliferation of T-cells. Here, we present evidence that both the CsA-CyPB complex and free CyPB bind to the T-lymphocyte surface, with similar values of Kd and number of sites. At 37 degrees C, the complex is internalized but, in contrast to the protein, the drug is accumulated within the cell. Moreover, CyPB receptors are internalized together with the ligand and rapidly recycled to the cell surface. Finally, we demonstrate that CyPB-complexed CsA remains as efficient as uncomplexed CsA and that CyPB enhances the immunosuppressive activity of the drug. Taken together, our results support the hypothesis that surface CyPB receptors may be related to the selective and variable action of CsA, through specific binding and targeting of the CyPB-CsA complex to peripheral blood T-lymphocytes.

  6. Fractal binding and dissociation kinetics of lecithin cholesterol acyl transferase (LCAT), a heart-related compound, on biosensor surfaces

    Science.gov (United States)

    Doke, Atul M.; Sadana, Ajit

    2006-05-01

    A fractal analysis is presented for the binding and dissociation of different heart-related compounds in solution to receptors immobilized on biosensor surfaces. The data analyzed include LCAT (lecithin cholesterol acyl transferase) concentrations in solution to egg-white apoA-I rHDL immobilized on a biosensor chip surface.1 Single- and dual- fractal models were employed to fit the data. Values of the binding and the dissociation rate coefficient(s), affinity values, and the fractal dimensions were obtained from the regression analysis provided by Corel Quattro Pro 8.0 (Corel Corporation Limited).2 The binding rate coefficients are quite sensitive to the degree of heterogeneity on the sensor chip surface. Predictive equations are developed for the binding rate coefficient as a function of the degree of heterogeneity present on the sensor chip surface and on the LCAT concentration in solution, and for the affinity as a function of the ratio of fractal dimensions present in the binding and the dissociation phases. The analysis presented provided physical insights into these analyte-receptor reactions occurring on different biosensor surfaces.

  7. A novel carbohydrate-binding surface layer protein from the hyperthermophilic archaeon Pyrococcus horikoshii.

    Science.gov (United States)

    Goda, Shuichiro; Koga, Tomoyuki; Yamashita, Kenichiro; Kuriura, Ryo; Ueda, Toshifumi

    2018-04-08

    In Archaea and Bacteria, surface layer (S-layer) proteins form the cell envelope and are involved in cell protection. In the present study, a putative S-layer protein was purified from the crude extract of Pyrococcus horikoshii using affinity chromatography. The S-layer gene was cloned and expressed in Escherichia coli. Isothermal titration calorimetry analyses showed that the S-layer protein bound N-acetylglucosamine and induced agglutination of the gram-positive bacterium Micrococcus lysodeikticus. The protein comprised a 21-mer structure, with a molecular mass of 1,340 kDa, as determined using small-angle X-ray scattering. This protein showed high thermal stability, with a midpoint of thermal denaturation of 79 °C in dynamic light scattering experiments. This is the first description of the carbohydrate-binding archaeal S-layer protein and its characteristics.

  8. Incorporation of the purified epstein barr virus/C3d receptor (CR2) into liposomes and demonstration of its dual ligand binding functions

    International Nuclear Information System (INIS)

    Mold, C.; Cooper, N.R.; Nemerow, G.R.

    1986-01-01

    The 145-kDA molecule that has been identified as the C3d receptor CR2 was isolated from lysates of Raji cells by affinity chromatography by using the monoclonal antibody (MoAb)HB-5. The purified protein was incorporated into 14 C-phosphatidylcholine liposomes by deoxycholate dialysis followed by flotation on discontinuous sucrose gradients. Incorporation of the receptor was verified by testing the gradient fractions for CR2 by an enzyme-linked immunosorbent assay. Liposomes were shown to be unilamellar vesicles ranging in diameter from 25 to 100 nm by electron microscopy. The external orientation of CR2 in the membranes was demonstrated by immunoelectron microscopy. The functional activities of liposomes containing CR2 and liposomes without protein were compared. CR2 liposomes bound to EC3d, but not to E, and this binding was inhibited by the anti-CR2 MoAb OKB7 and by a MoAb specific for C3d. Control liposomes failed to bind to either E or EC3D. The ability of CR2 to function as a receptor for Epstein Barr virus (EBV) was tested in two ways. First, CR2 liposomes bound to B95-8, a cell line expressing EBV membrane antigens, but not to B95-8 cells treated with the viral DNA polymerase inhibitor phosphonoformic acid. Second, liposomes containing CR2 were shown by ultracentrifugal analyses to bind directly to purified EBV, and this binding was also inhibited by OKB7. Control liposomes did not bind to B95-8 cells or to EBV. These findings show that CR2 purified from detergent extracts of Raji cells can be reconstituted into lipid membranes with maintenance of its dual functions as a receptor for C3d and EBV

  9. Incorporation of the purified epstein barr virus/C3d receptor (CR2) into liposomes and demonstration of its dual ligand binding functions

    Energy Technology Data Exchange (ETDEWEB)

    Mold, C.; Cooper, N.R.; Nemerow, G.R.

    1986-06-01

    The 145-kDA molecule that has been identified as the C3d receptor CR2 was isolated from lysates of Raji cells by affinity chromatography by using the monoclonal antibody (MoAb)HB-5. The purified protein was incorporated into /sup 14/C-phosphatidylcholine liposomes by deoxycholate dialysis followed by flotation on discontinuous sucrose gradients. Incorporation of the receptor was verified by testing the gradient fractions for CR2 by an enzyme-linked immunosorbent assay. Liposomes were shown to be unilamellar vesicles ranging in diameter from 25 to 100 nm by electron microscopy. The external orientation of CR2 in the membranes was demonstrated by immunoelectron microscopy. The functional activities of liposomes containing CR2 and liposomes without protein were compared. CR2 liposomes bound to EC3d, but not to E, and this binding was inhibited by the anti-CR2 MoAb OKB7 and by a MoAb specific for C3d. Control liposomes failed to bind to either E or EC3D. The ability of CR2 to function as a receptor for Epstein Barr virus (EBV) was tested in two ways. First, CR2 liposomes bound to B95-8, a cell line expressing EBV membrane antigens, but not to B95-8 cells treated with the viral DNA polymerase inhibitor phosphonoformic acid. Second, liposomes containing CR2 were shown by ultracentrifugal analyses to bind directly to purified EBV, and this binding was also inhibited by OKB7. Control liposomes did not bind to B95-8 cells or to EBV. These findings show that CR2 purified from detergent extracts of Raji cells can be reconstituted into lipid membranes with maintenance of its dual functions as a receptor for C3d and EBV.

  10. Effects of synthetic cohesin-containing scaffold protein architecture on binding dockerin-enzyme fusions on the surface of Lactococcus lactis

    Directory of Open Access Journals (Sweden)

    Wieczorek Andrew S

    2012-12-01

    Full Text Available Abstract Background The microbial synthesis of fuels, commodity chemicals, and bioactive compounds necessitates the assemblage of multiple enzyme activities to carry out sequential chemical reactions, often via substrate channeling by means of multi-domain or multi-enzyme complexes. Engineering the controlled incorporation of enzymes in recombinant protein complexes is therefore of interest. The cellulosome of Clostridium thermocellum is an extracellular enzyme complex that efficiently hydrolyzes crystalline cellulose. Enzymes interact with protein scaffolds via type 1 dockerin/cohesin interactions, while scaffolds in turn bind surface anchor proteins by means of type 2 dockerin/cohesin interactions, which demonstrate a different binding specificity than their type 1 counterparts. Recombinant chimeric scaffold proteins containing cohesins of different specificity allow binding of multiple enzymes to specific sites within an engineered complex. Results We report the successful display of engineered chimeric scaffold proteins containing both type 1 and type 2 cohesins on the surface of Lactococcus lactis cells. The chimeric scaffold proteins were able to form complexes with the Escherichia coli β-glucuronidase fused to either type 1 or type 2 dockerin, and differences in binding efficiencies were correlated with scaffold architecture. We used E. coli β-galactosidase, also fused to type 1 or type 2 dockerins, to demonstrate the targeted incorporation of two enzymes into the complexes. The simultaneous binding of enzyme pairs each containing a different dockerin resulted in bi-enzymatic complexes tethered to the cell surface. The sequential binding of the two enzymes yielded insights into parameters affecting assembly of the complex such as protein size and position within the scaffold. Conclusions The spatial organization of enzymes into complexes is an important strategy for increasing the efficiency of biochemical pathways. In this study

  11. Changes in cell surface structure by viral transformation studied by binding of lectins differing in sugar specificity.

    Science.gov (United States)

    Tsuda, M; Kurokawa, T; Takeuchi, M; Sugino, Y

    1975-10-01

    Changes in cell surface structure by viral transformation were studied by examining changes in the binding of various lectins differing in carbohydrate specificities. Binding of lectins was assayed directly using cells grown in coverslips. The following 125I-lectins were used: Concanavalin-A (specific for glucose and mannose), wheat germ agglutinin (specific for N-acetylglucosamine), castor bean agglutinin (specific for galactose), Wistaria floribunda agglutinin (specific for N-acetylgalactosamine), and soybean agglutinin (specific for N-acetyl-galactosamine). Cells for a clone, SS7, transformed by bovine adenovirus type-3, were found to bind 5 to 6 times more Wistaria floribunda agglutinin than the normal counterpart cells (clone C31, from C3H mouse kidney). In contrast, the binding of soybean agglutinin, which has a sugar specificity similar to Wistaria floribunda agglutinin, to normal and transformed cells was similar. The binding of wheat germ agglutinin and castor bean agglutinin, respectively, to normal and transformed cells was also similar. However, normal cells bound twice as much concanavalin-A as transformed cells. Only half as much Wistaria floribunda agglutinin was bound to transformed cells when they had been dispersed with EDTA. These changes in the number of lectin binding sites on transformation are thought to reflect alteration of the cell surface structure. The amount of lectins bound per cell decreased with increase in cell density, especially in the case of binding of Wistaria floribunda agglutinin to normal cells.

  12. 40 CFR 63.5755 - How do I demonstrate compliance with the aluminum recreational boat surface coating spray gun...

    Science.gov (United States)

    2010-07-01

    ... the aluminum recreational boat surface coating spray gun cleaning work practice standards? 63.5755... surface coating spray gun cleaning work practice standards? You must demonstrate compliance with the aluminum coating spray gun cleaning work practice standards by meeting the requirements of paragraph (a) or...

  13. Fine definition of the CXCR4-binding region on the V3 loop of feline immunodeficiency virus surface glycoprotein.

    Directory of Open Access Journals (Sweden)

    Qiong-Ying Hu

    2010-05-01

    Full Text Available The chemokine receptor CXCR4 is shared by primary and laboratory-adapted strains of feline immunodeficiency virus (FIV for viral entry. Our previous studies implicated a contiguous nine-amino-acid region of the V3 loop of the FIV envelope surface as important in CXCR4 binding and virus entry. The binding is specific for CXCR4 since it can be inhibited by AMD3100, a selective CXCR4 inhibitor. Additional site-directed mutagenesis was used to further reveal the key residues. Binding studies indicated that basic residues R395, K397, R399 as well as N398 are critical for CXCR4 binding. The effect of other amino acid residues on receptor binding depends on the type of amino acid residue substituted. The binding study results were confirmed on human CXCR4-expressing SupT1 cells and correlated with entry efficiency using a virus entry assay. Amino acid residues critical for CXCR4 are not critical for interactions with the primary binding receptor CD134, which has an equivalent role as CD4 for HIV-1 binding. The ELISA results show that W394 and W400 are crucial for the recognition by neutralizing anti-V3 antibodies. Since certain strains of HIV-1 also use CXCR4 as the entry receptor, the findings make the feline model attractive for development of broad-based entry antagonists and for study of the molecular mechanism of receptor/virus interactions.

  14. Binding of chloroquine to ionic micelles: Effect of pH and micellar surface charge

    Energy Technology Data Exchange (ETDEWEB)

    Souza Santos, Marcela de, E-mail: marcelafarmausp77@gmail.com [Departamento de Física e Química, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Avenida do Café, s/n, Ribeirão Preto, São Paulo 14040-903 (Brazil); Perpétua Freire de Morais Del Lama, Maria, E-mail: mpemdel@fcfrp.usp.br [Departamento de Física e Química, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Avenida do Café, s/n, Ribeirão Preto, São Paulo 14040-903 (Brazil); Instituto Nacional de Ciência e Tecnologia de Bioanalítica, Departamento de Química Analítica, Universidade Estadual de Campinas, Cidade Universitária Zeferino Vaz, s/n, Campinas, São Paulo 13083-970 (Brazil); Siuiti Ito, Amando, E-mail: amandosi@ffclrp.usp.br [Departamento de Física, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Avenida Bandeirantes, 3900, Ribeirão Preto, São Paulo 14040-901 (Brazil); and others

    2014-03-15

    The pharmacological action of chloroquine relies on its ability to cross biological membranes in order to accumulate inside lysosomes. The present work aimed at understanding the basis for the interaction between different chloroquine species and ionic micelles of opposite charges, the latter used as a simple membrane model. The sensitivity of absorbance and fluorescence of chloroquine to changes in its local environment was used to probe its interaction with cetyltrimethylammonium micelles presenting bromide (CTAB) and sulfate (CTAS) as counterions, in addition to dodecyl sulfate micelles bearing sodium (SDS) and tetramethylammonium (TMADS) counterions. Counterion exchange was shown to have little effect on drug–micelle interaction. Chloroquine first dissociation constant (pKa{sub 1}) shifted to opposite directions when anionic and cationic micelles were compared. Chloroquine binding constants (K{sub b}) revealed that electrostatic forces mediate charged drug–micelle association, whereas hydrophobic interactions allowed neutral chloroquine to associate with anionic and cationic micelles. Fluorescence quenching studies indicated that monoprotonated chloroquine is inserted deeper into the micelle surface of anionic micelles than its neutral form, the latter being less exposed to the aqueous phase when associated with cationic over anionic assemblies. The findings provide further evidence that chloroquine–micelle interaction is driven by a tight interplay between the drug form and the micellar surface charge, which can have a major effect on the drug biological activity. -- Highlights: • Chloroquine (CQ) pKa{sub 1} increased for SDS micelles and decreased for CTAB micelles. • CQ is solubilized to the surface of both CTAB and SDS micelles. • Monoprotonated CQ is buried deeper into SDS micelles than neutral CQ. • Neutral CQ is less exposed to aqueous phase in CTAB over SDS micelles. • Local pH and micellar surface charge mediate interaction of CQ with

  15. Bee venom phospholipase A2 as a membrane-binding vector for cell surface display or internalization of soluble proteins.

    Science.gov (United States)

    Babon, Aurélie; Wurceldorf, Thibault; Almunia, Christine; Pichard, Sylvain; Chenal, Alexandre; Buhot, Cécile; Beaumelle, Bruno; Gillet, Daniel

    2016-06-15

    We showed that bee venom phospholipase A2 can be used as a membrane-binding vector to anchor to the surface of cells a soluble protein fused to its C-terminus. ZZ, a two-domain derivative of staphylococcal protein A capable of binding constant regions of antibodies was fused to the C-terminus of the phospholipase or to a mutant devoid of enzymatic activity. The fusion proteins bound to the surface of cells and could themselves bind IgGs. Their fate depended on the cell type to which they bound. On the A431 carcinoma cell line the proteins remained exposed on the cell surface. In contrast, on human dendritic cells the proteins were internalized into early endosomes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Surface monitoring of microseismicity at the Decatur, Illinois, CO2 sequestration demonstration site

    Science.gov (United States)

    Kaven, Joern; Hickman, Stephen H.; McGarr, Arthur F.; Ellsworth, William L.

    2015-01-01

    Sequestration of CO2 into subsurface reservoirs can play an important role in limiting future emission of CO2 into the atmosphere (e.g., Benson and Cole, 2008). For geologic sequestration to become a viable option to reduce greenhouse gas emissions, large-volume injection of supercritical CO2 into deep sedimentary formations is required. These formations offer large pore volumes and good pore connectivity and are abundant (Bachu, 2003; U.S. Geological Survey Geologic Carbon Dioxide Storage Resources Assessment Team, 2013). However, hazards associated with injection of CO2 into deep formations require evaluation before widespread sequestration can be adopted safely (Zoback and Gorelick, 2012). One of these hazards is the potential to induce seismicity on pre-existing faults or fractures. If these faults or fractures are large and critically stressed, seismic events can occur with magnitudes large enough to pose a hazard to surface installations and, possibly more critical, the seal integrity of the cap rock. The Decatur, Illinois, carbon capture and storage (CCS) demonstration site is the first, and to date, only CCS project in the United States that injects a large volume of supercritical CO2 into a regionally extensive, undisturbed saline formation. The first phase of the Decatur CCS project was completed in November 2014 after injecting a million metric tons of supercritical CO2 over three years. This phase was led by the Illinois State Geological Survey (ISGS) and included seismic monitoring using deep borehole sensors, with a few sensors installed within the injection horizon. Although the deep borehole network provides a more comprehensive seismic catalog than is presented in this paper, these deep data are not publically available. We contend that for monitoring induced microseismicity as a possible seismic hazard and to elucidate the general patterns of microseismicity, the U.S. Geological Survey (USGS) surface and shallow borehole network described below

  17. Experimental demonstration of CMOS-compatible long-range dielectric-loaded surface plasmon-polariton waveguides (LR-DLSPPWs)

    DEFF Research Database (Denmark)

    Zektzer, Roy; Desiatov, Boris; Mazurski, Noa

    2015-01-01

    We demonstrate the design, fabrication and experimental characterization of long-range dielectric-loaded surface plasmon-polariton waveguides (LR-DLSPPWs) that are compatible with complementary metal-oxide semiconductor (CMOS) technology. The demonstrated waveguides feature good mode confinement...

  18. Alkaline phosphatase binds tenaciously to titanium; implications for biological surface evaluation following bone implant retrieval.

    Science.gov (United States)

    Mansell, J P; Shiel, A I; Harwood, C; Stephens, D

    2017-07-01

    Enhancing the performance and longevity of titanium (Ti) implants continues to be a significant developmental theme in contemporary biomaterials design. Our specific focus pertains to the surface functionalisation of Ti using the bioactive lipid, lysophosphatidic acid (LPA) and certain phosphatase-resistant analogues of LPA. Coating survivorship to a plethora of testing regimens is required to align with due regulatory process before novel biomaterials can enter clinical trials. One of the key acceptance criteria is coating retention to the physical stresses experienced during implantation. In assessing coating stability to insertion into porcine bone we found that a subsequent in vitro assessment to confirm coating persistence was masked by abundant alkaline phosphatase (ALP) contamination adsorbed to the metal surface. Herein we report that ALP can bind to Ti in a matter of minutes by simply immersing Ti samples in aqueous solutions of the enzyme. We strongly discourage the in vitro monitoring of osteoblast and stromal cell ALP expression when assessing bioactive coating survivorship following Ti implant retrieval form native bone tissue. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Coating magnesium hydroxide on surface of carbon microspheres and interface binding with poly (ethylene terephthalate) matrix

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Baoxia [Key Laboratory of Interface Science and Engineering in Advanced Materials, Taiyuan University of Technology, Ministry of Education, Taiyuan 030024 (China); Research Center on Advanced Materials Science and Technology, Taiyuan University of Technology, Taiyuan 030024 (China); College of Textile Engineering, Taiyuan University of Technology, Yuci 030600 (China); Niu, Mei, E-mail: niumei@tyut.edu.cn [Key Laboratory of Interface Science and Engineering in Advanced Materials, Taiyuan University of Technology, Ministry of Education, Taiyuan 030024 (China); College of Textile Engineering, Taiyuan University of Technology, Yuci 030600 (China); Yang, Yongzhen, E-mail: yyztyut@126.com [Key Laboratory of Interface Science and Engineering in Advanced Materials, Taiyuan University of Technology, Ministry of Education, Taiyuan 030024 (China); Research Center on Advanced Materials Science and Technology, Taiyuan University of Technology, Taiyuan 030024 (China); Bai, Jie; Song, Yinghao; Peng, Yun [College of Textile Engineering, Taiyuan University of Technology, Yuci 030600 (China); Liu, Xuguang [Key Laboratory of Interface Science and Engineering in Advanced Materials, Taiyuan University of Technology, Ministry of Education, Taiyuan 030024 (China); College of Chemistry and Chemical Engineering, Taiyuan University of Technology, Taiyuan 030024 (China)

    2017-08-01

    Highlights: • Magnesium hydroxide (MH) as a capsule wall was firstly coated on the surface of carbon microspheres (CMSs) to obtain MH@CMSs by liquid phase deposition method. • An organic layer of 3-Aminopropyltriethoxysilane (APTS) was then introduced on the surface of MH@CMSs. • The formed two layers provided the FMH@CMSs/PET with good mechanical and flame-retardant properties. - Abstract: In this account, magnesium hydroxide (MH) employed as a capsule wall was firstly coated on the surface of carbon microspheres (CMSs) to obtain MH@CMSs using liquid phase deposition, then was modified by 3-Aminopropyltriethoxysilane (APTS) to form FMH@CMSs. To investigate the interface binding forces, a series of PET composites was prepared by melt compounding with MH@CMSs or FMH@CMSs. Field-emission scanning electron microscopy (FESEM), Transmission electron microscopy (TEM), Fourier-transform Infrared spectrometry (FTIR) and X-ray photoelectron spectroscopy (XPS) were used to characterize the morphology, chemical structure, and effect of functionalization of CMSs. The coating degree and thermal stability were investigated by thermogravimetry analysis. The results showed that CMSs were coated by an inorganic shell layer of MH as a capsule wall. On the other hand, MH@CMSs were coated with an organic layer of APTS. When compared to MH@CMSs, the interface binding forces between FMH@CMSs and PET matrix were significantly improved, and the tensile strength of FMH@CMSs/PET was higher than that of MH@CMSs/PET. At 1 wt% mass fraction of FMH@CMSs, the limiting oxygen index (LOI) value of PET composites increased from 21% to 27.6% following a V-0 rating. The tensile strength of FMH@CMSs/PET increased by 66.2% to reach 47.20 MPa, a value nearly similar to that of PET. Overall, the formed two layers provided the FMH@CMSs/PET with good mechanical and flame-retardant properties, which would broaden their scope of application.

  20. The Non-Specific Binding of Fluorescent-Labeled MiRNAs on Cell Surface by Hydrophobic Interaction.

    Science.gov (United States)

    Lu, Ting; Lin, Zongwei; Ren, Jianwei; Yao, Peng; Wang, Xiaowei; Wang, Zhe; Zhang, Qunye

    2016-01-01

    MicroRNAs are small noncoding RNAs about 22 nt long that play key roles in almost all biological processes and diseases. The fluorescent labeling and lipofection are two common methods for changing the levels and locating the position of cellular miRNAs. Despite many studies about the mechanism of DNA/RNA lipofection, little is known about the characteristics, mechanisms and specificity of lipofection of fluorescent-labeled miRNAs. Therefore, miRNAs labeled with different fluorescent dyes were transfected into adherent and suspension cells using lipofection reagent. Then, the non-specific binding and its mechanism were investigated by flow cytometer and laser confocal microscopy. The results showed that miRNAs labeled with Cy5 (cyanine fluorescent dye) could firmly bind to the surface of adherent cells (Hela) and suspended cells (K562) even without lipofection reagent. The binding of miRNAs labeled with FAM (carboxyl fluorescein) to K562 cells was obvious, but it was not significant in Hela cells. After lipofectamine reagent was added, most of the fluorescently labeled miRNAs binding to the surface of Hela cells were transfected into intra-cell because of the high transfection efficiency, however, most of them were still binding to the surface of K562 cells. Moreover, the high-salt buffer which could destroy the electrostatic interactions did not affect the above-mentioned non-specific binding, but the organic solvent which could destroy the hydrophobic interactions eliminated it. These results implied that the fluorescent-labeled miRNAs could non-specifically bind to the cell surface by hydrophobic interaction. It would lead to significant errors in the estimation of transfection efficiency only according to the cellular fluorescence intensity. Therefore, other methods to evaluate the transfection efficiency and more appropriate fluorescent dyes should be used according to the cell types for the accuracy of results.

  1. The Non-Specific Binding of Fluorescent-Labeled MiRNAs on Cell Surface by Hydrophobic Interaction.

    Directory of Open Access Journals (Sweden)

    Ting Lu

    Full Text Available MicroRNAs are small noncoding RNAs about 22 nt long that play key roles in almost all biological processes and diseases. The fluorescent labeling and lipofection are two common methods for changing the levels and locating the position of cellular miRNAs. Despite many studies about the mechanism of DNA/RNA lipofection, little is known about the characteristics, mechanisms and specificity of lipofection of fluorescent-labeled miRNAs.Therefore, miRNAs labeled with different fluorescent dyes were transfected into adherent and suspension cells using lipofection reagent. Then, the non-specific binding and its mechanism were investigated by flow cytometer and laser confocal microscopy. The results showed that miRNAs labeled with Cy5 (cyanine fluorescent dye could firmly bind to the surface of adherent cells (Hela and suspended cells (K562 even without lipofection reagent. The binding of miRNAs labeled with FAM (carboxyl fluorescein to K562 cells was obvious, but it was not significant in Hela cells. After lipofectamine reagent was added, most of the fluorescently labeled miRNAs binding to the surface of Hela cells were transfected into intra-cell because of the high transfection efficiency, however, most of them were still binding to the surface of K562 cells. Moreover, the high-salt buffer which could destroy the electrostatic interactions did not affect the above-mentioned non-specific binding, but the organic solvent which could destroy the hydrophobic interactions eliminated it.These results implied that the fluorescent-labeled miRNAs could non-specifically bind to the cell surface by hydrophobic interaction. It would lead to significant errors in the estimation of transfection efficiency only according to the cellular fluorescence intensity. Therefore, other methods to evaluate the transfection efficiency and more appropriate fluorescent dyes should be used according to the cell types for the accuracy of results.

  2. Demonstration of pulmonary {beta}{sub 2}-adrenergic receptor binding in vivo with [{sup 18}F]fluoroethyl-fenoterol in a guinea pig model

    Energy Technology Data Exchange (ETDEWEB)

    Helisch, A.; Schirrmacher, E.; Schirrmacher, R.; Buchholz, H.G.; Bartenstein, P. [University Hospital, Department of Nuclear Medicine, Mainz (Germany); Thews, O.; Dillenburg, W.; Tillmanns, J. [University of Mainz, Institute of Physiology and Pathophysiology, Mainz (Germany); Hoehnemann, S.; Roesch, F. [University of Mainz, Institute of Nuclear Chemistry, Mainz (Germany); Wessler, I. [University of Mainz, Institute of Pharmacology, Mainz (Germany); Buhl, R. [University Hospital, Pulmonary Division, Mainz (Germany)

    2005-11-01

    The new {beta}{sub 2} radioligand (R,R)(S,S) 5-(2-(2-[4-(2-[{sup 18}F]fluoroethoxy)phenyl]-1-methylethylamino)-1-hydroxyethyl)-benzene-1,3-diol ([{sup 18}F]FE-fenoterol; [{sup 18}F]FEFE), a fluoroethylated derivative of racemic fenoterol, was evaluated in vivo and ex vivo using a guinea pig model. Dynamic PET studies over 60 min with [{sup 18}F]FEFE were performed in nine Hartley guinea pigs in which a baseline (group 1, n=3), a predose (group 2, n=3; 2 mg/kg fenoterol 5 min prior to injection of [{sup 18}F]FEFE) or a displacement study (group 3, n=3; 2 mg/kg fenoterol 5 min post injection of [{sup 18}F]FEFE) was conducted. In all animal groups, the lungs could be visualised and semi-quantified separately by calculating uptake ratios to non-specific binding in the neck area. Premedication with non-radioactive fenoterol and displacement tests showed significant reduction of lung uptake, by 94% and 76%, respectively. These data demonstrate specific binding of the new radioligand to the pulmonary {beta}{sub 2}-receptors in accordance with ex vivo measurements. Therefore, [{sup 18}F]FEFE seems to be suitable for the in vivo visualisation and quantification of the pulmonary {beta}{sub 2}-receptor binding in this animal model. (orig.)

  3. Mhp182 (P102) binds fibronectin and contributes to the recruitment of plasmin(ogen) to the Mycoplasma hyopneumoniae cell surface.

    Science.gov (United States)

    Seymour, Lisa M; Jenkins, Cheryl; Deutscher, Ania T; Raymond, Benjamin B A; Padula, Matthew P; Tacchi, Jessica L; Bogema, Daniel R; Eamens, Graeme J; Woolley, Lauren K; Dixon, Nicholas E; Walker, Mark J; Djordjevic, Steven P

    2012-01-01

    Mycoplasma hyopneumoniae is a major, economically damaging respiratory pathogen. Although M. hyopneumoniae cells bind plasminogen, the identification of plasminogen-binding surface proteins and the biological ramifications of acquiring plasminogen requires further investigation. mhp182 encodes a highly expressed 102 kDa protein (P102) that undergoes proteolytic processing to generate surface-located N-terminal 60 kDa (P60) and C-terminal 42 kDa (P42) proteins of unknown function. We show that recombinant P102 (rP102) binds plasminogen at physiologically relevant concentrations (K(D) ~ 76 nM) increasing the susceptibility of plasmin(ogen) to activation by tissue-specific plasminogen activator (tPA). Recombinant proteins constructed to mimic P60 (rP60) and P42 (rP42) also bound plasminogen at physiologically significant levels. M. hyopneumoniae surface-bound plasminogen was activated by tPA and is able to degrade fibrinogen, demonstrating the biological functionality of M. hyopneumoniae-bound plasmin(ogen) upon activation. Plasmin(ogen) was readily detected in porcine ciliated airways and plasmin levels were consistently higher in bronchoalveolar lavage fluid from M. hyopneumoniae-infected animals. Additionally, rP102 and rP42 bind fibronectin with K(D) s of 26 and 33 nM respectively and recombinant P102 proteins promote adherence to porcine kidney epithelial-like cells. The multifunctional binding ability of P102 and activation of M. hyopneumoniae-sequestered plasmin(ogen) by an exogenous activator suggests P102 plays an important role in virulence. © 2011 Blackwell Publishing Ltd.

  4. Experimental demonstration of Martian soil simulant removal from a surface using a pulsed plasma jet

    Science.gov (United States)

    Ticoş, C. M.; Scurtu, A.; Toader, D.; Banu, N.

    2015-03-01

    A plasma jet produced in a small coaxial plasma gun operated at voltages up to 2 kV and working in pure carbon dioxide (CO2) at a few Torr is used to remove Martian soil simulant from a surface. A capacitor with 0.5 mF is charged up from a high voltage source and supplies the power to the coaxial electrodes. The muzzle of the coaxial plasma gun is placed at a few millimeters near the dusty surface and the jet is fired parallel with the surface. Removal of dust is imaged in real time with a high speed camera. Mars regolith simulant JSC-Mars-1A with particle sizes up to 5 mm is used on different types of surfaces made of aluminium, cotton fabric, polyethylene, cardboard, and phenolic.

  5. Experimental demonstration of Martian soil simulant removal from a surface using a pulsed plasma jet.

    Science.gov (United States)

    Ticoş, C M; Scurtu, A; Toader, D; Banu, N

    2015-03-01

    A plasma jet produced in a small coaxial plasma gun operated at voltages up to 2 kV and working in pure carbon dioxide (CO2) at a few Torr is used to remove Martian soil simulant from a surface. A capacitor with 0.5 mF is charged up from a high voltage source and supplies the power to the coaxial electrodes. The muzzle of the coaxial plasma gun is placed at a few millimeters near the dusty surface and the jet is fired parallel with the surface. Removal of dust is imaged in real time with a high speed camera. Mars regolith simulant JSC-Mars-1A with particle sizes up to 5 mm is used on different types of surfaces made of aluminium, cotton fabric, polyethylene, cardboard, and phenolic.

  6. Reversible surface binding of cadmium and lead by lactic acid and bifidobacteria.

    Science.gov (United States)

    Teemu, Halttunen; Seppo, Salminen; Jussi, Meriluoto; Raija, Tahvonen; Kalle, Lertola

    2008-07-15

    Extensive cadmium and lead contamination of water has been reported to occur locally as a result of human activities. Lactic acid bacteria have been reported to remove cadmium and lead from water. The aim of this work was to clarify the mechanisms of cadmium and lead removal from water. In addition, the effect of other metals, reversibility of binding and recyclability of the biomass was assessed. Based on our earlier data, the two most promising lactic acid bacteria, Lactobacillus fermentum ME3 and Bifidobacterium longum 46, were selected for these experiments. The results showed that the presence of other cationic metals and blocking of carboxyl and phosphoryl groups reduced cadmium and lead removal. These results suggest involvement of electrostatic interactions in cadmium and lead removal, and support our earlier findings. Transmission electron micrographs showed large deposits of lead on the bacterial surface suggesting formation of metallic lead precipitates. Both cadmium and lead removal were reversible processes established by full recovery of removed metal after desorption with dilute solutions of EDTA and HNO(3). Resorption capacity of both biomasses tested was reduced after regeneration with 10 mM EDTA and 15 mM HNO(3). Taken together, the results suggest involvement of several reversible mechanisms such as ion exchange and precipitation in cadmium and lead binding by lactic acid bacteria. The results show that specific lactic acid bacteria have the potential for removal of cadmium and lead from water although reduction in resorption capacity after regeneration of the biomass may form a problem. Since the studies so far have mainly focused on removal of single metals from pure water, metal removal in conditions of natural waters should be assessed in further experiments.

  7. Results from the search-lidar demonstrator project for detection of small Sea-Surface targets

    NARCIS (Netherlands)

    Heuvel, J.C. van den; Putten, F.J.M. van; Cohen, L.H.; Kemp, R.A.W.; Franssen, G.C.

    2009-01-01

    Coastal surveillance and naval operations in the littoral both have to deal with the threat of small sea-surface targets. These targets have a low radar cross-section and a low velocity that makes them hard to detect by radar. Typical threats include jet skis, FIAC's, and speedboats. Previous lidar

  8. Search-Lidar Demonstrator for Detection of Small Sea-Surface Targets

    NARCIS (Netherlands)

    Heuvel, J.C. van den; Bekman, H.H.P.T.; Putten, F.J.M. van; Cohen, L.H.; Schleijpen, H.M.A.

    2008-01-01

    Coastal surveillance and naval operations in the littoral both have to deal with the threat of small sea-surface targets. These targets have a low radar cross-section and a low velocity that makes them hard to detect by radar. Typical threats include jet skis, FIAC’s, and speedboats. Lidar

  9. Characterization of sperm surface protein patterns of ejaculated and capacitated boar sperm, with the detection of ZP binding candidates

    Czech Academy of Sciences Publication Activity Database

    Zigo, Michal; Jonáková, Věra; Šulc, Miroslav; Maňásková-Postlerová, Pavla

    2013-01-01

    Roč. 61, oct (2013), s. 322-328 ISSN 0141-8130 R&D Projects: GA ČR GAP503/12/1834 Institutional research plan: CEZ:AV0Z50520701 Institutional support: RVO:61388971 Keywords : Sperm surface protein * Zona pellucida-binding receptors * PKDREJ protein Subject RIV: CE - Biochemistry Impact factor: 3.096, year: 2013

  10. Dispersion relation and self-collimation frequency of spoof surface plasmon using tight binding model

    International Nuclear Information System (INIS)

    Bhattacharya, Sayak; Shah, Kushal

    2015-01-01

    The analytical dispersion relation of spoof surface plasmon (SSP) is known only in the low-frequency limit and thus cannot be used to describe various practically important characteristics of SSP in the high-frequency limit (such as multimodal nature, anisotropic propagation, self-collimation). In this article, we consider a square lattice of holes made on a perfect electric conductor and derive a closed form expression of the SSP dispersion relation in the high-frequency limit using a tight binding model. Instead of using prior knowledge of the band diagram along the entire first Brillouin zone (BZ) edge, we analytically determine the hopping parameters by using the eigenfrequencies only at the three high-symmetry points of the square lattice. Using this dispersion relation, we derive an expression for the self-collimation frequency of SSP. We show that this analytical formulation is also applicable to dielectric photonic crystals and can be used to predict the frequencies corresponding to centimetre-scale supercollimation and second band self-collimation in these structures. Finally, we show that our analytical results are in agreement with the simulation results for both SSP and photonic crystals. (paper)

  11. BB0347, from the lyme disease spirochete Borrelia burgdorferi, is surface exposed and interacts with the CS1 heparin-binding domain of human fibronectin.

    Directory of Open Access Journals (Sweden)

    Robert A Gaultney

    Full Text Available The causative agent of Lyme disease, Borrelia burgdorferi, codes for several known fibronectin-binding proteins. Fibronectin a common the target of diverse bacterial pathogens, and has been shown to be essential in allowing for the development of certain disease states. Another borrelial protein, BB0347, has sequence similarity with these other known fibronectin-binding proteins, and may be important in Lyme disease pathogenesis. Herein, we perform an initial characterization of BB0347 via the use of molecular and biochemical techniques. We found that BB0347 is expressed, produced, and presented on the outer surface of intact B. burgdorferi. We also demonstrate that BB0347 has the potential to be important in Lyme disease progression, and have begun to characterize the nature of the interaction between human fibronectin and this bacterial protein. Further work is needed to define the role of this protein in the borrelial infection process.

  12. Curcumin stably interacts with DNA hairpin through minor groove binding and demonstrates enhanced cytotoxicity in combination with FdU nucleotides.

    Science.gov (United States)

    Ghosh, Supratim; Mallick, Sumana; Das, Upasana; Verma, Ajay; Pal, Uttam; Chatterjee, Sabyasachi; Nandy, Abhishek; Saha, Krishna D; Maiti, Nakul Chandra; Baishya, Bikash; Suresh Kumar, G; Gmeiner, William H

    2018-03-01

    We report, based on biophysical studies and molecular mechanical calculations that curcumin binds DNA hairpin in the minor groove adjacent to the loop region forming a stable complex. UV-Vis and fluorescence spectroscopy indicated interaction of curcumin with DNA hairpin. In this novel binding motif, two ɣ H of curcumin heptadiene chain are closely positioned to the A 16 -H8 and A 17 -H8, while G 12 -H8 is located in the close proximity of curcumin α H. Molecular dynamics (MD) simulations suggest, the complex is stabilized by noncovalent forces including; π-π stacking, H-bonding and hydrophobic interactions. Nuclear magnetic resonance (NMR) spectroscopy in combination with molecular dynamics simulations indicated curcumin is bound in the minor groove, while circular dichroism (CD) spectra suggested minute enhancement in base stacking and a little change in DNA helicity, without significant conformational change of DNA hairpin structure. The DNA:curcumin complex formed with FdU nucleotides rather than Thymidine, demonstrated enhanced cytotoxicity towards oral cancer cells relative to the only FdU substituted hairpin. Fluorescence co-localization demonstrated stability of the complex in biologically relevant conditions, including its cellular uptake. Acridine orange/EtBr staining further confirmed the enhanced cytotoxic effects of the complex, suggesting apoptosis as mode of cell death. Thus, curcumin can be noncovalently complexed to small DNA hairpin for cellular delivery and the complex showed increased cytotoxicity in combination with FdU nucleotides, demonstrating its potential for advanced cancer therapy. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Equilibrium and kinetics of Sin Nombre hantavirus binding at DAF/CD55 functionalized bead surfaces.

    Science.gov (United States)

    Buranda, Tione; Swanson, Scarlett; Bondu, Virginie; Schaefer, Leah; Maclean, James; Mo, Zhenzhen; Wycoff, Keith; Belle, Archana; Hjelle, Brian

    2014-03-10

    Decay accelerating factor (DAF/CD55) is targeted by many pathogens for cell entry. It has been implicated as a co-receptor for hantaviruses. To examine the binding of hantaviruses to DAF, we describe the use of Protein G beads for binding human IgG Fc domain-functionalized DAF ((DAF)₂-Fc). When mixed with Protein G beads the resulting DAF beads can be used as a generalizable platform for measuring kinetic and equilibrium binding constants of DAF binding targets. The hantavirus interaction has high affinity (24-30 nM; k(on) ~ 10⁵ M⁻¹ s⁻¹, k(off) ~ 0.0045 s⁻¹). The bivalent (DAF)₂-Fc/SNV data agree with hantavirus binding to DAF expressed on Tanoue B cells (K(d) = 14.0 nM). Monovalent affinity interaction between SNV and recombinant DAF of 58.0 nM is determined from competition binding. This study serves a dual purpose of presenting a convenient and quantitative approach of measuring binding affinities between DAF and the many cognate viral and bacterial ligands and providing new data on the binding constant of DAF and Sin Nombre hantavirus. Knowledge of the equilibrium binding constant allows for the determination of the relative fractions of bound and free virus particles in cell entry assays. This is important for drug discovery assays for cell entry inhibitors.

  14. Modified cytokeratins expressed on the surface of carcinoma cells undergo endocytosis upon binding of human monoclonal antibody and its recombinant Fab fragment

    DEFF Research Database (Denmark)

    Ditzel, H J; Garrigues, U; Andersen, C B

    1997-01-01

    display selection and the human Fab fragment was expressed in bacteria. Analysis by confocal laser scanning microscopy demonstrated that COU-1 bound in a uniform punctate pattern to the surface of viable carcinoma cells stained at 4 degrees C, and binding increased significantly when cells were cultured...... was significantly reduced. Similar results were obtained using intact IgM COU-1 and the recombinant Fab fragment. Immunohistological studies indicated that COU-1, in contrast to murine monoclonal antibodies against normal cytokeratin 8 and 18, could differentiate between malignant and normal colon epithelia...

  15. Nucleolin inhibits Fas ligand binding and suppresses Fas-mediated apoptosis in vivo via a surface nucleolin-Fas complex.

    Science.gov (United States)

    Wise, Jillian F; Berkova, Zuzana; Mathur, Rohit; Zhu, Haifeng; Braun, Frank K; Tao, Rong-Hua; Sabichi, Anita L; Ao, Xue; Maeng, Hoyoung; Samaniego, Felipe

    2013-06-06

    Resistance to Fas-mediated apoptosis is associated with poor cancer outcomes and chemoresistance. To elucidate potential mechanisms of defective Fas signaling, we screened primary lymphoma cell extracts for Fas-associated proteins that would have the potential to regulate Fas signaling. An activation-resistant Fas complex selectively included nucleolin. We confirmed the presence of nucleolin-Fas complexes in B-cell lymphoma cells and primary tissues, and the absence of such complexes in B-lymphocytes from healthy donors. RNA-binding domain 4 and the glycine/arginine-rich domain of nucleolin were essential for its association with Fas. Nucleolin colocalized with Fas on the surface of B-cell lymphoma cells. Nucleolin knockdown sensitized BJAB cells to Fas ligand (FasL)-induced and Fas agonistic antibody-induced apoptosis through enhanced binding, suggesting that nucleolin blocks the FasL-Fas interaction. Mice transfected with nucleolin were protected from the lethal effects of agonistic anti-mouse Fas antibody (Jo2) and had lower rates of hepatocyte apoptosis, compared with vector and a non-Fas-binding mutant of nucleolin. Our results show that cell surface nucleolin binds Fas, inhibits ligand binding, and thus prevents induction of Fas-mediated apoptosis in B-cell lymphomas and may serve as a new therapeutic target.

  16. Surface plasmon resonance imaging reveals multiple binding modes of Agrobacterium transformation mediator VirE2 to ssDNA.

    Science.gov (United States)

    Kim, Sanghyun; Zbaida, David; Elbaum, Michael; Leh, Hervé; Nogues, Claude; Buckle, Malcolm

    2015-07-27

    VirE2 is the major secreted protein of Agrobacterium tumefaciens in its genetic transformation of plant hosts. It is co-expressed with a small acidic chaperone VirE1, which prevents VirE2 oligomerization. After secretion into the host cell, VirE2 serves functions similar to a viral capsid in protecting the single-stranded transferred DNA en route to the nucleus. Binding of VirE2 to ssDNA is strongly cooperative and depends moreover on protein-protein interactions. In order to isolate the protein-DNA interactions, imaging surface plasmon resonance (SPRi) studies were conducted using surface-immobilized DNA substrates of length comparable to the protein-binding footprint. Binding curves revealed an important influence of substrate rigidity with a notable preference for poly-T sequences and absence of binding to both poly-A and double-stranded DNA fragments. Dissociation at high salt concentration confirmed the electrostatic nature of the interaction. VirE1-VirE2 heterodimers also bound to ssDNA, though by a different mechanism that was insensitive to high salt. Neither VirE2 nor VirE1-VirE2 followed the Langmuir isotherm expected for reversible monomeric binding. The differences reflect the cooperative self-interactions of VirE2 that are suppressed by VirE1. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  17. Simulation analysis of the cellulase Cel7A carbohydrate binding module on the surface of the cellulose Iβ

    Energy Technology Data Exchange (ETDEWEB)

    Alekozai, Emal M. [Univ. of Heidelberg (Germany); Univ. of Tennessee, Knoxville, TN (United States); GhattyVenkataKrishna, Pavan K. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Uberbacher, Edward C. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Crowley, Michael F. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); National Renewable Energy Lab. (NREL), Golden, CO (United States); Smith, Jeremy C. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Tennessee, Knoxville, TN (United States); Cheng, Xiaolin [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Tennessee, Knoxville, TN (United States)

    2013-08-22

    The Family 7 cellobiohydrolase (Cel7A) from Trichoderma reesei consists of a carbohydrate-binding module (CBM) joined by a linker to a catalytic domain. Cellulose hydrolysis is limited by the accessibility of Cel7A to crystalline substrates, which is perceived to be primarily mediated by the CBM. The binding of CBM to the cellulose I fiber is characterized by combined Brownian dynamics (BD) and molecular dynamics (MD) simulations. Our results confirm that CBM prefers to dock to the hydrophobic than to the hydrophilic fiber faces. Both electrostatic (ES) and van der Waals (VDW) interactions are required for achieving the observed binding preference. The VDW interactions play a more important role in stabilizing the CBM-fiber binding, whereas the ES interactions contribute through the formation of a number of hydrogen bonds between the CBM and the fiber. At long distances, an ES steering effect is also observed that tends to align the CBM in an antiparallel manner relative to the fiber axis. Moreover, the MD results reveal hindered diffusion of the CBM on all fiber surfaces. The binding of the CBM to the hydrophobic surfaces is found to involve partial dewetting at the CBM-fiber interface coupled with local structural arrangements of the protein. The present simulation results complement and rationalize a large body of previous work and provide detailed insights into the mechanism of the CBM-cellulose fiber interactions.

  18. Demonstration of Cold 40Ca+ Ions Confined in a Microscopic Surface-Electrode Ion Trap

    International Nuclear Information System (INIS)

    Chen Liang; Wan Wei; Xie Yi; Wu Hao-Yu; Zhou Fei; Feng Mang

    2013-01-01

    40 Ca + ions are successfully confined, under the cooling of a red-detuned laser, in a home-built microscopic surface-electrode (MSE) trap. With all electrodes deposited on a low-rf-loss substrate, our 500-μm-scale MSE trap is designed involving three potential wells and manufactured by the standard technique of the printed circuit board. Both linear and two-dimensional crystals of 40 Ca + are observed in the trap after preliminary micromotion compensation is carried out. The development of the MSE trap aims at large-scale trapped-ion quantum information processing

  19. A semi-grand canonical Monte Carlo simulation model for ion binding to ionizable surfaces: proton binding of carboxylated latex particles as a case study.

    Science.gov (United States)

    Madurga, Sergio; Rey-Castro, Carlos; Pastor, Isabel; Vilaseca, Eudald; David, Calin; Garcés, Josep Lluís; Puy, Jaume; Mas, Francesc

    2011-11-14

    In this paper, we present a computer simulation study of the ion binding process at an ionizable surface using a semi-grand canonical Monte Carlo method that models the surface as a discrete distribution of charged and neutral functional groups in equilibrium with explicit ions modelled in the context of the primitive model. The parameters of the simulation model were tuned and checked by comparison with experimental titrations of carboxylated latex particles in the presence of different ionic strengths of monovalent ions. The titration of these particles was analysed by calculating the degree of dissociation of the latex functional groups vs. pH curves at different background salt concentrations. As the charge of the titrated surface changes during the simulation, a procedure to keep the electroneutrality of the system is required. Here, two approaches are used with the choice depending on the ion selected to maintain electroneutrality: counterion or coion procedures. We compare and discuss the difference between the procedures. The simulations also provided a microscopic description of the electrostatic double layer (EDL) structure as a function of pH and ionic strength. The results allow us to quantify the effect of the size of the background salt ions and of the surface functional groups on the degree of dissociation. The non-homogeneous structure of the EDL was revealed by plotting the counterion density profiles around charged and neutral surface functional groups. © 2011 American Institute of Physics

  20. T Tank Farm Interim Surface Barrier Demonstration--Vadose Zone Monitoring Plan

    International Nuclear Information System (INIS)

    Zhang, Z. F.; Keller, Jason M.; Strickland, Christopher E.

    2007-01-01

    The Hanford Site has 149 underground single-shell tanks that store hazardous radioactive waste. Many of these tanks and their associated infrastructure (e.g., pipelines, diversion boxes) have leaked. Some of the leaked waste has entered the groundwater. The largest known leak occurred from the T-106 Tank in 1973. Many of the contaminants from that leak still reside within the vadose zone beneath the T Tank Farm. CH2M Hill Hanford Group, Inc. seeks to minimize movement of this residual contaminant plume by placing an interim barrier on the surface. Such a barrier is expected to prevent infiltrating water from reaching the plume and moving it further. A plan has been prepared to monitor and determine the effectiveness of the interim surface barrier. Soil water content and water pressure will be monitored using off-the-shelf equipment that can be installed by the hydraulic hammer technique. In fiscal year 2006, two instrument nests were installed. Each instrument nest contains a neutron probe access tube, a capacitance probe, four heat-dissipation units, and a drain gauge to measure soil water flux. A meteorological station has been installed outside of the fence. In fiscal year 2007, two additional instrument nests are planned to be installed beneath the proposed barrier.

  1. The salivary scavenger and agglutinin (SALSA binds MBL and regulates the lectin pathway of complement in solution and on surfaces

    Directory of Open Access Journals (Sweden)

    Martin eParnov Reichhardt

    2012-07-01

    Full Text Available The scavenger receptor cysteine-rich (SRCR protein SALSA, also known as gp340, salivary agglutinin (SAG and deleted in malignant brain tumor 1 (DMBT1, is a 340 kDa glycoprotein expressed on mucosal surfaces and secreted into several body fluids. SALSA binds to a broad variety of microbes and endogenous ligands, such as complement factor C1q, surfactant proteins D and A (SP-D and SP-A and IgA. Our search for novel ligands of SALSA by direct protein-interaction studies led to the identification of mannan binding lectin (MBL as a new binding partner. We observed that surface-associated SALSA activates complement via binding of MBL. On the other hand, soluble SALSA was found to inhibit C. albicans-induced complement activation. Thus, SALSA has a dual complement regulatory function. It activates the lectin pathway when bound to a surface and inhibits it when free in the fluid-phase. These activities are mediated via a direct interaction with MBL.

  2. Surface modification by cold-plasma technique for dental implants—Bio-functionalization with binding pharmaceuticals

    Directory of Open Access Journals (Sweden)

    Masao Yoshinari

    2011-08-01

    At the bone tissue/implant interface, a thin calcium phosphate coating and rapid heating with infrared radiation were effective in controlling the dissolution without cracking the coating. These thin calcium phosphate coatings may directly promote osteogenisis, but also enable immobilization and subsequent drug delivery system (DDS of bisphosphonates. Simvastatin is also an effective candidate that is reported to increase the expression of BMP-2. The thin-film of hexamethyldisiloxane (HMDSO was plasma-polymerized onto titanium, and then HMDSO surface was activated by O2-plasma treatment. A quartz crystal microbalance (QCM-D technique demonstrated that simvastatin was immobilized on the plasma-treated surfaces due to introduction of O2-functional groups. At the soft tissue/implant interface, multi-grooved surface topographies and utilizing the adhesive proteins such as fibronectin or laminin-5 may help in providing a biological seal around the implant. At the oral fluid/implant interface, an alumina coating, F+-implantation and immobilization of anti-microbial peptides were responsible for inhibiting the biofilm accumulation.

  3. Photo-induced electrochemical anodization of p-type silicon: achievement and demonstration of long term surface stability

    International Nuclear Information System (INIS)

    Dhanekar, Saakshi; Islam, S S; Harsh

    2012-01-01

    Surface stability is achieved and demonstrated by porous silicon (PS) fabricated using a wavelength-dependent photo-electrochemical (PEC) anodization technique. During anodization, the photon flux for all wavelengths was kept constant while only the effect of light wavelength on the surface morphology of PS was investigated. PS optical sensors were realized, characterized and tested using a photoluminescence (PL) quenching technique. An aliphatic chain of alcohols (methanol to n-octanol) was detected in the range of 10–200 ppm. Long term surface stability was observed from samples prepared under red (750–620 nm) and green illumination (570–495 nm), where the PL quenching cycles evoke the possibility of using PS for stable sensor device applications. This study provides a route for preparing highly sensitive organic vapour sensors with a precise selection of the fabrication parameters and demonstrating their prolonged performance. (paper)

  4. T-TY Tank Farm Interim Surface Barrier Demonstration - Vadose Zone Monitoring Plan

    International Nuclear Information System (INIS)

    Zhang, Z.F.; Strickland, Christopher E.; Field, Jim G.; Parker, Danny L.

    2010-01-01

    The Hanford Site has 149 underground single-shell tanks that store hazardous radioactive waste. Many of these tanks and their associated infrastructure (e.g., pipelines, diversion boxes) have leaked. Some of the leaked waste has entered the groundwater. The largest known leak occurred from the T-106 Tank of the 241-T Tank Farm in 1973. Five tanks are assumed to have leaked in the TY Farm. Many of the contaminants from those leaks still reside within the vadose zone within the T and TY Tank Farms. The Department of Energy's Office of River Protection seeks to minimize the movement of these contaminant plumes by placing interim barriers on the ground surface. Such barriers are expected to prevent infiltrating water from reaching the plumes and moving them further. The soil water regime is monitored to determine the effectiveness of the interim surface barriers. Soil-water content and water pressure are monitored using off-the-shelf equipment that can be installed by the hydraulic hammer technique. Four instrument nests were installed in the T Farm in fiscal year (FY) 2006 and FY2007; two nests were installed in the TY Farm in FY2010. Each instrument nest contains a neutron probe access tube, a capacitance probe, and four heat-dissipation units. A meteorological station has been installed at the north side of the fence of the T Farm. This document summarizes the monitoring methods, the instrument calibration and installation, and the vadose zone monitoring plan for interim barriers in T farm and TY Farm.

  5. Binding of p-mercaptobenzoic acid and adenine to gold-coated electroless etched silicon nanowires studied by surface-enhanced Raman scattering.

    Science.gov (United States)

    Mohaček-Grošev, Vlasta; Gebavi, Hrvoje; Bonifacio, Alois; Sergo, Valter; Daković, Marko; Bajuk-Bogdanović, Danica

    2018-04-10

    Modern diagnostic tools ever aim to reduce the amount of analyte and the time needed for obtaining the result. Surface-enhanced Raman spectroscopy is a method that could satisfy both of these requirements, provided that for each analyte an adequate substrate is found. Here we demonstrate the ability of gold-sputtered silicon nanowires (SiNW) to bind p-mercaptobenzoic acid in 10 -3 , 10 -4 and 10 -5 M and adenine in 30 and 100μM concentrations. Based on the normal mode analysis, presented here for the first time, the binding of p-mercaptobenzoic acid is deduced. The intensity enhancement of the 1106cm -1 band is explained by involvement of the CS stretching deformation, and the appearance of the broad 300cm -1 band attributed to SAu stretching mode. Adenine SERS spectra demonstrate the existence of the 7H tautomer since the strongest band observed is at 736cm -1 . The adenine binding is likely to occur in several ways, because the number of observed bands in the 1200-1600cm -1 interval exceeds the number of observed bands in the normal Raman spectrum of the free molecule. Copyright © 2018 Elsevier B.V. All rights reserved.

  6. The OmpL37 surface-exposed protein is expressed by pathogenic Leptospira during infection and binds skin and vascular elastin.

    Science.gov (United States)

    Pinne, Marija; Choy, Henry A; Haake, David A

    2010-09-07

    Pathogenic Leptospira spp. shed in the urine of reservoir hosts into freshwater can be transmitted to a susceptible host through skin abrasions or mucous membranes causing leptospirosis. The infection process involves the ability of leptospires to adhere to cell surface and extracellular matrix components, a crucial step for dissemination and colonization of host tissues. Therefore, the elucidation of novel mediators of host-pathogen interaction is important in the discovery of virulence factors involved in the pathogenesis of leptospirosis. In this study, we assess the functional roles of transmembrane outer membrane proteins OmpL36 (LIC13166), OmpL37 (LIC12263), and OmpL47 (LIC13050), which we recently identified on the leptospiral surface. We determine the capacity of these proteins to bind to host tissue components by enzyme-linked immunosorbent assay. OmpL37 binds elastin preferentially, exhibiting dose-dependent, saturating binding to human skin (K(d), 104±19 nM) and aortic elastin (K(d), 152±27 nM). It also binds fibrinogen (K(d), 244±15 nM), fibrinogen fragment D (K(d), 132±30 nM), plasma fibronectin (K(d), 359±68 nM), and murine laminin (K(d), 410±81 nM). The binding to human skin elastin by both recombinant OmpL37 and live Leptospira interrogans is specifically enhanced by rabbit antiserum for OmpL37, suggesting the involvement of OmpL37 in leptospiral binding to elastin and also the possibility that host-generated antibodies may promote rather than inhibit the adherence of leptospires to elastin-rich tissues. Further, we demonstrate that OmpL37 is recognized by acute and convalescent leptospirosis patient sera and also by Leptospira-infected hamster sera. Finally, OmpL37 protein is detected in pathogenic Leptospira serovars and not in saprophytic Leptospira. Thus, OmpL37 is a novel elastin-binding protein of pathogenic Leptospira that may be promoting attachment of Leptospira to host tissues.

  7. Rotavirus NSP4 is secreted from infected cells as an oligomeric lipoprotein and binds to glycosaminoglycans on the surface of non-infected cells

    Directory of Open Access Journals (Sweden)

    Didsbury Alicia

    2011-12-01

    Full Text Available Abstract Background Nonstructural glycoprotein 4 (NSP4 encoded by rotavirus is the only viral protein currently believed to function as an enterotoxin. NSP4 is synthesized as an intracellular transmembrane glycoprotein and as such is essential for virus assembly. Infection of polarized Caco-2 cells with rotavirus also results in the secretion of glycosylated NSP4 apparently in a soluble form despite retention of its transmembrane domain. We have examined the structure, solubility and cell-binding properties of this secreted form of NSP4 to further understand the biochemical basis for its enterotoxic function. We show here that NSP4 is secreted as discrete detergent-sensitive oligomers in a complex with phospholipids and demonstrate that this secreted form of NSP4 can bind to glycosaminoglycans present on the surface of a range of different cell types. Methods NSP4 was purified from the medium of infected cells after ultracentrifugation and ultrafiltration by successive lectin-affinity and ion exchange chromatography. Oligomerisation of NSP4 was examined by density gradient centrifugation and chemical crosslinking and the lipid content was assessed by analytical thin layer chromatography and flame ionization detection. Binding of NSP4 to various cell lines was measured using a flow cytometric-based assay. Results Secreted NSP4 formed oligomers that contained phospholipid but dissociated to a dimeric species in the presence of non-ionic detergent. The purified glycoprotein binds to the surface of various non-infected cells of distinct lineage. Binding of NSP4 to HT-29, a cell line of intestinal origin, is saturable and independent of divalent cations. Complementary biochemical approaches reveal that NSP4 binds to sulfated glycosaminoglycans on the plasma membrane. Conclusion Our study is the first to analyze an authentic (i.e. non-recombinant form of NSP4 that is secreted from virus-infected cells. Despite retention of the transmembrane domain

  8. Molecular binding mechanisms of aqueous cadmium and lead to siderophores, bacteria and mineral surfaces

    Science.gov (United States)

    Mishra, Bhoopesh

    Recent studies have shown that diverse groups of bacteria adsorb metals to similar extents and uptake can be modeled using a universal adsorption model. In this study, XAFS has been used to resolve whether binding sites determined for single species systems are responsible for adsorption in more complex natural bacterial assemblages. Results obtained from a series of XAFS experiments on pure Gram positive and Gram negative bacterial strains and consortia of bacteria as a function of pH and Cd loading suggests that every bacterial strain has a complex physiology and they are all slightly different from each other. Nevertheless from the metal adsorption chemistry point of view, the main difference between them lies in the site ratio of three fundamental sites only - carboxyl, phosphoryl and sulfide. Two completely different consortia of bacteria (obtained from natural river water, and soil system with severe organic contamination) were successfully modeled in the pH range 3.4--7.8 using the EXAFS models developed for single species systems. Results thus obtained can potentially have very high impact on the modeling of the complex bacterial systems in realistic geological settings, leading to further refinement and development of robust remediation strategies for metal contamination at macroscopic level. In another study, solution speciation of Pb and Cd with DFO-B has been examined using a combination of techniques (ICP, TOC, thermodynamic modeling and XAFS). Results indicate that Pb does not complex with DFO-B at all until about pH 3.5, but forms a totally caged structure at pH 7.5. At intermediate pH conditions, mixture of species (one and two hydroxamate groups complexed) is formed. Cd on the other hand, does not complex until pH 5, forms intermediate complexes at pH 8 and is totally chelated at pH 9. Further studies were conducted for Pb sorption to mineral surface kaolinite with and without DFO-B. In the absence of DFO-B, results suggest outer sphere and inner

  9. Cell surface binding and uptake of arginine- and lysine-rich penetratin peptides in absence and presence of proteoglycans

    KAUST Repository

    Åmand, Helene L.

    2012-11-01

    Cell surface proteoglycans (PGs) appear to promote uptake of arginine-rich cell-penetrating peptides (CPPs), but their exact functions are unclear. To address if there is specificity in the interactions of arginines and PGs leading to improved internalization, we used flow cytometry to examine uptake in relation to cell surface binding for penetratin and two arginine/lysine substituted variants (PenArg and PenLys) in wildtype CHO-K1 and PG-deficient A745 cells. All peptides were more efficiently internalized into CHO-K1 than into A745, but their cell surface binding was independent of cell type. Thus, PGs promote internalization of cationic peptides, irrespective of the chemical nature of their positive charges. Uptake of each peptide was linearly dependent on its cell surface binding, and affinity is thus important for efficiency. However, the gradients of these linear dependencies varied significantly. Thus each peptide\\'s ability to stimulate uptake once bound to the cell surface is reliant on formation of specific uptake-promoting interactions. Heparin affinity chromatography and clustering experiments showed that penetratin and PenArg binding to sulfated sugars is stabilized by hydrophobic interactions and result in clustering, whereas PenLys only interacts through electrostatic attraction. This may have implications for the molecular mechanisms behind arginine-specific uptake stimulation as penetratin and PenArg are more efficiently internalized than PenLys upon interaction with PGs. However, PenArg is also least affected by removal of PGs. This indicates that an increased arginine content not only improve PG-dependent uptake but also that PenArg is more adaptable as it can use several portals of entry into the cell. © 2012 Elsevier B.V.

  10. Screening Mixtures of Small Molecules for Binding to Multiple Sites on the Surface Tetanus Toxin C Fragment by Bioaffinity NMR

    International Nuclear Information System (INIS)

    Cosman, M; Zeller, L; Lightstone, F C; Krishnan, V V; Balhorn, R

    2002-01-01

    The clostridial neurotoxins include the closely related tetanus (TeNT) and botulinum (BoNT) toxins. Botulinum toxin is used to treat severe muscle disorders and as a cosmetic wrinkle reducer. Large quantities of botulinum toxin have also been produced by terrorists for use as a biological weapon. Because there are no known antidotes for these toxins, they thus pose a potential threat to human health whether by an accidental overdose or by a hostile deployment. Thus, the discovery of high specificity and affinity compounds that can inhibit their binding to neural cells can be used as antidotes or in the design of chemical detectors. Using the crystal structure of the C fragment of the tetanus toxin (TetC), which is the cell recognition and cell surface binding domain, and the computational program DOCK, sets of small molecules have been predicted to bind to two different sites located on the surface of this protein. While Site-1 is common to the TeNT and BoNTs, Site-2 is unique to TeNT. Pairs of these molecules from each site can then be linked together synthetically to thereby increase the specificity and affinity for this toxin. Electrospray ionization mass spectroscopy was used to experimentally screen each compound for binding. Mixtures containing binders were further screened for activity under biologically relevant conditions using nuclear magnetic resonance (NMR) methods. The screening of mixtures of compounds offers increased efficiency and throughput as compared to testing single compounds and can also evaluate how possible structural changes induced by the binding of one ligand can influence the binding of the second ligand. In addition, competitive binding experiments with mixtures containing ligands predicted to bind the same site could identify the best binder for that site. NMR transfer nuclear Overhauser effect (trNOE) confirm that TetC binds doxorubicin but that this molecule is displaced by N-acetylneuraminic acid (sialic acid) in a mixture that

  11. Characterization, cell-surface expression and ligand-binding properties of different truncated N-terminal extracellular domains of the ionotropic glutamate receptor subunit GluR1.

    Science.gov (United States)

    McIlhinney, R A; Molnár, E

    1996-04-01

    To identify the location of the first transmembrane segment of the GluR1 glutamate receptor subunit artificial stop codons have been introduced into the N-terminal domain at amino acid positions 442, 510, and 563, namely just before and spanning the proposed first two transmembrane regions. The resultant truncated N-terminal fragments of GluR1, termed NT1, NT2, and NT3 respectively were expressed in Cos-7 cells and their cellular distribution and cell-surface expression analysed using an N-terminal antibody to GluR1. All of the fragments were fully glycosylated and were found to be associated with cell membranes but none was secreted. Differential extraction of the cell membranes indicated that both NT1 and NT2 behave as peripheral membrane proteins. In contrast NT3, like the full subunit, has integral membrane protein properties. Furthermore only NT3 is expressed at the cell surface as determined by immunofluorescence and cell-surface biotinylation. Protease protection assays indicated that only NT3 had a cytoplasmic tail. Binding studies using the selective ligand [(3)H]alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate ([(3)H]AMPA) demonstrated that NT3 does not bind ligand. Together these results indicate that the first transmembrane domain of the GluR1 subunit lies between residues 509 and 562, that the N-terminal domain alone cannot form a functional ligand-binding site and that this domain can be targeted to the cell surface provided that it has a transmembrane-spanning region.

  12. Atomistic modeling determination of placeholder binding energy of Ti, C, and N atoms on a-Fe (100) surfaces

    International Nuclear Information System (INIS)

    Wei, X J; Liu, Y P; Han, S P

    2015-01-01

    A Fe(100) surface containing Ti, C, and N was constructed and optimized to study the placeholder binding energy of the Ti, C, and N surface atoms; this was achieved by searching the transition state with the LST (linear synchronous transit) method of the CASTEP (Cambridge Serial Total Energy Package) module. Also, the authors analyzed electron structures to determine how Ti, C, and N atoms strengthen the Fe(100) surface. The results show that when Ti, C, or N atoms take placeholder alone, or simultaneously at the Fe(100) surface, the structure stability is at its best. When including Ti, C, and N as solid solutions on the Fe(100) surface, orbital electrons of Fe3d, Ti3d, C2p, and N2p hybridize near the Fermi level; the number of electronic bonding peaks increase and bonding capacity enhances. Also, a large amount of covalent bonds formed. Covalent bonds and metallic bond coexisted. (paper)

  13. Crystal structure of the gamma-2 herpesvirus LANA DNA binding domain identifies charged surface residues which impact viral latency.

    Directory of Open Access Journals (Sweden)

    Bruno Correia

    Full Text Available Latency-associated nuclear antigen (LANA mediates γ2-herpesvirus genome persistence and regulates transcription. We describe the crystal structure of the murine gammaherpesvirus-68 LANA C-terminal domain at 2.2 Å resolution. The structure reveals an alpha-beta fold that assembles as a dimer, reminiscent of Epstein-Barr virus EBNA1. A predicted DNA binding surface is present and opposite this interface is a positive electrostatic patch. Targeted DNA recognition substitutions eliminated DNA binding, while certain charged patch mutations reduced bromodomain protein, BRD4, binding. Virus containing LANA abolished for DNA binding was incapable of viable latent infection in mice. Virus with mutations at the charged patch periphery exhibited substantial deficiency in expansion of latent infection, while central region substitutions had little effect. This deficiency was independent of BRD4. These results elucidate the LANA DNA binding domain structure and reveal a unique charged region that exerts a critical role in viral latent infection, likely acting through a host cell protein(s.

  14. Mutations on the DNA binding surface of TBP discriminate between yeast TATA and TATA-less gene transcription.

    Science.gov (United States)

    Kamenova, Ivanka; Warfield, Linda; Hahn, Steven

    2014-08-01

    Most RNA polymerase (Pol) II promoters lack a TATA element, yet nearly all Pol II transcription requires TATA binding protein (TBP). While the TBP-TATA interaction is critical for transcription at TATA-containing promoters, it has been unclear whether TBP sequence-specific DNA contacts are required for transcription at TATA-less genes. Transcription factor IID (TFIID), the TBP-containing coactivator that functions at most TATA-less genes, recognizes short sequence-specific promoter elements in metazoans, but analogous promoter elements have not been identified in Saccharomyces cerevisiae. We generated a set of mutations in the yeast TBP DNA binding surface and found that most support growth of yeast. Both in vivo and in vitro, many of these mutations are specifically defective for transcription of two TATA-containing genes with only minor defects in transcription of two TATA-less, TFIID-dependent genes. TBP binds several TATA-less promoters with apparent high affinity, but our results suggest that this binding is not important for transcription activity. Our results are consistent with the model that sequence-specific TBP-DNA contacts are not important at yeast TATA-less genes and suggest that other general transcription factors or coactivator subunits are responsible for recognition of TATA-less promoters. Our results also explain why yeast TBP derivatives defective for TATA binding appear defective in activated transcription. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  15. Safety standards for near surface disposal and the safety case and supporting safety assessment for demonstrating compliance with the standards

    International Nuclear Information System (INIS)

    Metcalf, P.

    2003-01-01

    The report presents the safety standards for near surface disposal (ICRP guidance and IAEA standards) and the safety case and supporting safety assessment for demonstrating compliance with the standards. Special attention is paid to the recommendations for disposal of long-lived solid radioactive waste. The requirements are based on the principle for the same level of protection of future individuals as for the current generation. Two types of exposure are considered: human intrusion and natural processes and protection measures are discussed. Safety requirements for near surface disposal are discussed including requirements for protection of human health and environment, requirements or safety assessments, waste acceptance and requirements etc

  16. Radiochemical and immunohistochemical detection of low density lipoprotein surface binding by lymphocytes

    International Nuclear Information System (INIS)

    Melzner, I.; Hambitzer, R.; Haferkamp, O.

    1983-01-01

    Human peripheral blood lymphocytes bind and take up low density lipoprotein (LDL) by receptor-mediated endocytosis. The binding of LDL was determiend by incubation with 125 I-LDL and an immunohistochemical assay. By both techniques a diminished rate of binding was found when cells were freshly isolated from the blood, but increased 5 to 10 fold when lymphocytes were incubated in lipoprotein-deficient medium for 72 hours. In addition, it was shown immunohistochemically that only few ceels showed an LDL-dependent fluorescent labelling: approximately 5 to 10 % of the freshly isolated lymphocytes and 40 to 50 % of the cells incubated for 72 hours under lipoprotein-free conditions. The present data indicate that not only the high affinity LDL receptor described by Goldstein and Braun may be involved in the uptake of cholesterol by lymphocytes, but also other binding sites, which may have immunological function in some lymphocyte subpopulations. (author)

  17. Deciphering the Arginine-binding preferences at the substrate-binding groove of Ser/Thr kinases by computational surface mapping.

    Directory of Open Access Journals (Sweden)

    Avraham Ben-Shimon

    2011-11-01

    Full Text Available Protein kinases are key signaling enzymes that catalyze the transfer of γ-phosphate from an ATP molecule to a phospho-accepting residue in the substrate. Unraveling the molecular features that govern the preference of kinases for particular residues flanking the phosphoacceptor is important for understanding kinase specificities toward their substrates and for designing substrate-like peptidic inhibitors. We applied ANCHORSmap, a new fragment-based computational approach for mapping amino acid side chains on protein surfaces, to predict and characterize the preference of kinases toward Arginine binding. We focus on positions P-2 and P-5, commonly occupied by Arginine (Arg in substrates of basophilic Ser/Thr kinases. The method accurately identified all the P-2/P-5 Arg binding sites previously determined by X-ray crystallography and produced Arg preferences that corresponded to those experimentally found by peptide arrays. The predicted Arg-binding positions and their associated pockets were analyzed in terms of shape, physicochemical properties, amino acid composition, and in-silico mutagenesis, providing structural rationalization for previously unexplained trends in kinase preferences toward Arg moieties. This methodology sheds light on several kinases that were described in the literature as having non-trivial preferences for Arg, and provides some surprising departures from the prevailing views regarding residues that determine kinase specificity toward Arg. In particular, we found that the preference for a P-5 Arg is not necessarily governed by the 170/230 acidic pair, as was previously assumed, but by several different pairs of acidic residues, selected from positions 133, 169, and 230 (PKA numbering. The acidic residue at position 230 serves as a pivotal element in recognizing Arg from both the P-2 and P-5 positions.

  18. BL-7010 demonstrates specific binding to gliadin and reduces gluten-associated pathology in a chronic mouse model of gliadin sensitivity.

    Directory of Open Access Journals (Sweden)

    Justin L McCarville

    Full Text Available Celiac disease (CD is an autoimmune disorder in individuals that carry DQ2 or DQ8 MHC class II haplotypes, triggered by the ingestion of gluten. There is no current treatment other than a gluten-free diet (GFD. We have previously shown that the BL-7010 copolymer poly(hydroxyethyl methacrylate-co-styrene sulfonate (P(HEMA-co-SS binds with higher efficiency to gliadin than to other proteins present in the small intestine, ameliorating gliadin-induced pathology in the HLA-HCD4/DQ8 model of gluten sensitivity. The aim of this study was to investigate the efficiency of two batches of BL-7010 to interact with gliadin, essential vitamins and digestive enzymes not previously tested, and to assess the ability of the copolymer to reduce gluten-associated pathology using the NOD-DQ8 mouse model, which exhibits more significant small intestinal damage when challenged with gluten than HCD4/DQ8 mice. In addition, the safety and systemic exposure of BL-7010 was evaluated in vivo (in rats and in vitro (genetic toxicity studies. In vitro binding data showed that BL-7010 interacted with high affinity with gliadin and that BL-7010 had no interaction with the tested vitamins and digestive enzymes. BL-7010 was effective at preventing gluten-induced decreases in villus-to-crypt ratios, intraepithelial lymphocytosis and alterations in paracellular permeability and putative anion transporter-1 mRNA expression in the small intestine. In rats, BL-7010 was well-tolerated and safe following 14 days of daily repeated administration of 3000 mg/kg. BL-7010 did not exhibit any mutagenic effect in the genetic toxicity studies. Using complementary animal models and chronic gluten exposure the results demonstrate that administration of BL-7010 is effective and safe and that it is able to decrease pathology associated with gliadin sensitization warranting the progression to Phase I trials in humans.

  19. Use of surface plasmon resonance (SPR) to study the dissociation and polysaccharide binding of casein micelles and caseins.

    Science.gov (United States)

    Thompson, Abby K; Singh, Harjinder; Dalgleish, Douglas G

    2010-11-24

    Tests were made to determine whether surface plasmon resonance (SPR) could be used as a technique to study the dissociation properties of bovine casein micelles or of sodium caseinate and the interactions between these protein particles and different polysaccharides. Surfaces of bound micelles or caseinate were made, and the changes in refractive index of these layers were used to define changes in the structures of the chemisorbed material. The technique appears to have some potential for studying details of the dissociation of casein micelles and of the binding of different polysaccharides to caseins.

  20. Insights into cellulase-lignin non-specific binding revealed by computational redesign of the surface of green fluorescent protein.

    Science.gov (United States)

    Haarmeyer, Carolyn N; Smith, Matthew D; Chundawat, Shishir P S; Sammond, Deanne; Whitehead, Timothy A

    2017-04-01

    Biological-mediated conversion of pretreated lignocellulosic biomass to biofuels and biochemicals is a promising avenue toward energy sustainability. However, a critical impediment to the commercialization of cellulosic biofuel production is the high cost of cellulase enzymes needed to deconstruct biomass into fermentable sugars. One major factor driving cost is cellulase adsorption and inactivation in the presence of lignin, yet we currently have a poor understanding of the protein structure-function relationships driving this adsorption. In this work, we have systematically investigated the role of protein surface potential on lignin adsorption using a model monomeric fluorescent protein. We have designed and experimentally characterized 16 model protein variants spanning the physiological range of net charge (-24 to +16 total charges) and total charge density (0.28-0.40 charges per sequence length) typical for natural proteins. Protein designs were expressed, purified, and subjected to in silico and in vitro biophysical measurements to evaluate the relationship between protein surface potential and lignin adsorption properties. The designs were comparable to model fluorescent protein in terms of thermostability and heterologous expression yield, although the majority of the designs unexpectedly formed homodimers. Protein adsorption to lignin was studied at two different temperatures using Quartz Crystal Microbalance with Dissipation Monitoring and a subtractive mass balance assay. We found a weak correlation between protein net charge and protein-binding capacity to lignin. No other single characteristic, including apparent melting temperature and 2nd virial coefficient, showed correlation with lignin binding. Analysis of an unrelated cellulase dataset with mutations localized to a family I carbohydrate-binding module showed a similar correlation between net charge and lignin binding capacity. Overall, our study provides strategies to identify highly active, low

  1. Well-oriented ZZ–PS-tag with high Fc-binding onto polystyrene surface for controlled immobilization of capture antibodies

    International Nuclear Information System (INIS)

    Tang, Jin-Bao; Sun, Xi-Feng; Yang, Hong-Ming; Zhang, Bao-Gang; Li, Zhi-Jian; Lin, Zhi-Juan; Gao, Zhi-Qin

    2013-01-01

    Graphical abstract: -- Highlights: •A versatile platform for immobilizing functionally intact IgG is proposed. •The mechanism relies on properly oriented ZZ–PS-tag onto a hydrophilic PS surface. •The oriented ZZ–PS-tag presents ∼fivefold higher IgG-binding activity. •The platform shows tenfold higher sensitivity and a wider linear range in ELISA. -- Abstract: The site specificity and bioactivity retention of antibodies immobilized on a solid substrate are crucial requirements for solid phase immunoassays. A fusion protein between an immunoglobulin G (IgG)-binding protein (ZZ protein) and a polystyrene-binding peptide (PS-tag) was constructed, and then used to develop a simple method for the oriented immobilization of the ZZ protein onto a PS support by the specific attachment of the PS-tag onto a hydrophilic PS. The orientation of intact IgG was achieved via the interaction of the ZZ protein and the constant fragment (Fc), thereby displayed the Fab fragment for binding antigen. The interaction between rabbit IgG anti-horseradish peroxidase (anti-HRP) and its binding partner HRP was analyzed. Results showed that the oriented ZZ–PS-tag yielded an IgG-binding activity that is fivefold higher than that produced by the passive immobilization of the ZZ protein. The advantage of the proposed immunoassay strategy was demonstrated through an enzyme-linked immunosorbent assay, in which monoclonal mouse anti-goat IgG and HRP-conjugated rabbit F(ab′) 2 anti-goat IgG were used to detect goat IgG. The ZZ–PS-tag presented a tenfold higher sensitivity and a wider linear range than did the passively immobilized ZZ protein. The proposed approach may be an attractive strategy for a broad range of applications involving the oriented immobilization of intact IgGs onto PS supports, in which only one type of phi-PS (ZZ–PS-tag) surface is used

  2. Well-oriented ZZ–PS-tag with high Fc-binding onto polystyrene surface for controlled immobilization of capture antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Jin-Bao, E-mail: tangjinbao@yahoo.com.cn [School of Pharmacy and Biology, Weifang Medical University, Weifang 261053 (China); Sun, Xi-Feng [Clinical Laboratory, Weifang People' s Hospital, Weifang 261041 (China); Yang, Hong-Ming [School of Pharmacy and Biology, Weifang Medical University, Weifang 261053 (China); Zhang, Bao-Gang [School of Basic Medicine, Weifang Medical University, Weifang 261053 (China); Li, Zhi-Jian [School of Pharmacy and Biology, Weifang Medical University, Weifang 261053 (China); Lin, Zhi-Juan [School of Basic Medicine, Weifang Medical University, Weifang 261053 (China); Gao, Zhi-Qin, E-mail: zhiqingao@yahoo.cn [School of Pharmacy and Biology, Weifang Medical University, Weifang 261053 (China)

    2013-05-07

    Graphical abstract: -- Highlights: •A versatile platform for immobilizing functionally intact IgG is proposed. •The mechanism relies on properly oriented ZZ–PS-tag onto a hydrophilic PS surface. •The oriented ZZ–PS-tag presents ∼fivefold higher IgG-binding activity. •The platform shows tenfold higher sensitivity and a wider linear range in ELISA. -- Abstract: The site specificity and bioactivity retention of antibodies immobilized on a solid substrate are crucial requirements for solid phase immunoassays. A fusion protein between an immunoglobulin G (IgG)-binding protein (ZZ protein) and a polystyrene-binding peptide (PS-tag) was constructed, and then used to develop a simple method for the oriented immobilization of the ZZ protein onto a PS support by the specific attachment of the PS-tag onto a hydrophilic PS. The orientation of intact IgG was achieved via the interaction of the ZZ protein and the constant fragment (Fc), thereby displayed the Fab fragment for binding antigen. The interaction between rabbit IgG anti-horseradish peroxidase (anti-HRP) and its binding partner HRP was analyzed. Results showed that the oriented ZZ–PS-tag yielded an IgG-binding activity that is fivefold higher than that produced by the passive immobilization of the ZZ protein. The advantage of the proposed immunoassay strategy was demonstrated through an enzyme-linked immunosorbent assay, in which monoclonal mouse anti-goat IgG and HRP-conjugated rabbit F(ab′){sub 2} anti-goat IgG were used to detect goat IgG. The ZZ–PS-tag presented a tenfold higher sensitivity and a wider linear range than did the passively immobilized ZZ protein. The proposed approach may be an attractive strategy for a broad range of applications involving the oriented immobilization of intact IgGs onto PS supports, in which only one type of phi-PS (ZZ–PS-tag) surface is used.

  3. Selective binding of oligonucleotide on TiO2 surfaces modified by swift heavy ion beam lithography

    International Nuclear Information System (INIS)

    Vicente Pérez-Girón, J.; Hirtz, M.; McAtamney, C.; Bell, A.P.; Antonio Mas, J.; Jaafar, M.; Luis, O. de; Fuchs, H.

    2014-01-01

    We have used swift heavy-ion beam based lithography to create patterned bio-functional surfaces on rutile TiO 2 single crystals. The applied lithography method generates a permanent and well defined periodic structure of micrometre sized square holes having nanostructured TiO 2 surfaces, presenting different physical and chemical properties compared to the surrounding rutile single crystal surface. On the patterned substrates selective binding of oligonucleotides molecules is possible at the surfaces of the holes. This immobilisation process is only being controlled by UV light exposure. The patterned transparent substrates are compatible with fluorescence detection techniques, are mechanically robust, have a high tolerance to extreme chemical and temperature environments, and apparently do not degrade after ten cycles of use. These qualities make the patterned TiO 2 substrates useful for potential biosensor applications

  4. Selective binding of oligonucleotide on TiO{sub 2} surfaces modified by swift heavy ion beam lithography

    Energy Technology Data Exchange (ETDEWEB)

    Vicente Pérez-Girón, J. [Nanoate, S.L. C/Poeta Rafael Morales 2, San Sebastian de los Reyes, 28702 Madrid (Spain); Emerging Viruses Department Heinrich Pette Institute, Hamburg 20251 (Germany); Hirtz, M. [Institute of Nanotechnology (INT) and Karlsruhe Nano Micro Facility (KNMF), Karlsruhe Institute of Technology - KIT, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany); McAtamney, C.; Bell, A.P. [Advanced Microscopy Laboratory, CRANN, Trinity College Dublin, Dublin 2 (Ireland); Antonio Mas, J. [Laboratorio de Genómica del Centro de Apoyo Tecnológico, Universidad Rey Juan Carlos, Campus de Alcorcón 28922, Madrid (Spain); Jaafar, M. [Nanoate, S.L. C/Poeta Rafael Morales 2, San Sebastian de los Reyes, 28702 Madrid (Spain); Departamento de Física de la Materia Condensada, Facultad de Ciencias, Universidad Autónoma de Madrid, Campus de Cantoblanco, 28049 Madrid (Spain); Luis, O. de [Nanoate, S.L. C/Poeta Rafael Morales 2, San Sebastian de los Reyes, 28702 Madrid (Spain); Departamento de Bioquímica, Fisiología y Genética Molecular, Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos, Campus de Alcorcón, 28922 Madrid (Spain); Fuchs, H. [Institute of Nanotechnology (INT) and Karlsruhe Nano Micro Facility (KNMF), Karlsruhe Institute of Technology - KIT, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany); Physical Institute and Center for Nanotechnology (CeNTech), Wilhelm-Klemm-Straße 10, University of Münster (Germany); and others

    2014-11-15

    We have used swift heavy-ion beam based lithography to create patterned bio-functional surfaces on rutile TiO{sub 2} single crystals. The applied lithography method generates a permanent and well defined periodic structure of micrometre sized square holes having nanostructured TiO{sub 2} surfaces, presenting different physical and chemical properties compared to the surrounding rutile single crystal surface. On the patterned substrates selective binding of oligonucleotides molecules is possible at the surfaces of the holes. This immobilisation process is only being controlled by UV light exposure. The patterned transparent substrates are compatible with fluorescence detection techniques, are mechanically robust, have a high tolerance to extreme chemical and temperature environments, and apparently do not degrade after ten cycles of use. These qualities make the patterned TiO{sub 2} substrates useful for potential biosensor applications.

  5. Ribosomal protein L5 has a highly twisted concave surface and flexible arms responsible for rRNA binding.

    Science.gov (United States)

    Nakashima, T; Yao, M; Kawamura, S; Iwasaki, K; Kimura, M; Tanaka, I

    2001-05-01

    Ribosomal protein L5 is a 5S rRNA binding protein in the large subunit and plays an essential role in the promotion of a particular conformation of 5S rRNA. The crystal structure of the ribosomal protein L5 from Bacillus stearothermophilus has been determined at 1.8 A resolution. The molecule consists of a five-stranded antiparallel beta-sheet and four alpha-helices, which fold in a way that is topologically similar to the ribonucleoprotein (RNP) domain. The molecular shape and electrostatic representation suggest that the concave surface and loop regions are involved in 5S rRNA binding. To identify amino acid residues responsible for 5S rRNA binding, we made use of Ala-scanning mutagenesis of evolutionarily conserved amino acids occurring in the beta-strands and loop regions. The mutations of Asn37 at the beta1-strand and Gln63 at the loop between helix 2 and beta3-strand as well as that of Phe77 at the tip of the loop structure between the beta2- and beta3-strands caused a significant reduction in 5S rRNA binding. In addition, the mutations of Thr90 on the beta3-strand and Ile141 and Asp144 at the loop between beta4- and beta5-strands moderately reduced the 5S rRNA-binding affinity. Comparison of these results with the more recently analyzed structure of the 50S subunit from Haloarcula marismortui suggests that there are significant differences in the structure at N- and C-terminal regions and probably in the 5S rRNA binding.

  6. Binding of Thrombin-Activated Platelets to a Fibrin Scaffold through αIIbβ3 Evokes Phosphatidylserine Exposure on Their Cell Surface

    Science.gov (United States)

    Brzoska, Tomasz; Suzuki, Yuko; Mogami, Hideo; Sano, Hideto; Urano, Tetsumei

    2013-01-01

    Recently, by employing intra-vital confocal microscopy, we demonstrated that platelets expose phosphatidylserine (PS) and fibrin accumulate only in the center of the thrombus but not in its periphery. To address the question how exposure of platelet anionic phospholipids is regulated within the thrombus, an in-vitro experiment using diluted platelet-rich plasma was employed, in which the fibrin network was formed in the presence of platelets, and PS exposure on the platelet surface was analyzed using Confocal Laser Scanning Microscopy. Almost all platelets exposed PS after treatment with tissue factor, thrombin or ionomycin. Argatroban abrogated fibrin network formation in all samples, however, platelet PS exposure was inhibited only in tissue factor- and thrombin-treated samples but not in ionomycin-treated samples. FK633, an αIIbβ3 antagonist, and cytochalasin B impaired platelet binding to the fibrin scaffold and significantly reduced PS exposure evoked by thrombin. Gly-Pro-Arg-Pro amide abrogated not only fibrin network formation, but also PS exposure on platelets without suppressing platelet binding to fibrin/fibrinogen. These results suggest that outside-in signals in platelets generated by their binding to the rigid fibrin network are essential for PS exposure after thrombin treatment. PMID:23383331

  7. Binding of thrombin-activated platelets to a fibrin scaffold through α(IIb)β₃ evokes phosphatidylserine exposure on their cell surface.

    Science.gov (United States)

    Brzoska, Tomasz; Suzuki, Yuko; Mogami, Hideo; Sano, Hideto; Urano, Tetsumei

    2013-01-01

    Recently, by employing intra-vital confocal microscopy, we demonstrated that platelets expose phosphatidylserine (PS) and fibrin accumulate only in the center of the thrombus but not in its periphery. To address the question how exposure of platelet anionic phospholipids is regulated within the thrombus, an in-vitro experiment using diluted platelet-rich plasma was employed, in which the fibrin network was formed in the presence of platelets, and PS exposure on the platelet surface was analyzed using Confocal Laser Scanning Microscopy. Almost all platelets exposed PS after treatment with tissue factor, thrombin or ionomycin. Argatroban abrogated fibrin network formation in all samples, however, platelet PS exposure was inhibited only in tissue factor- and thrombin-treated samples but not in ionomycin-treated samples. FK633, an α(IIb)β₃ antagonist, and cytochalasin B impaired platelet binding to the fibrin scaffold and significantly reduced PS exposure evoked by thrombin. Gly-Pro-Arg-Pro amide abrogated not only fibrin network formation, but also PS exposure on platelets without suppressing platelet binding to fibrin/fibrinogen. These results suggest that outside-in signals in platelets generated by their binding to the rigid fibrin network are essential for PS exposure after thrombin treatment.

  8. Binding of thrombin-activated platelets to a fibrin scaffold through α(IIbβ₃ evokes phosphatidylserine exposure on their cell surface.

    Directory of Open Access Journals (Sweden)

    Tomasz Brzoska

    Full Text Available Recently, by employing intra-vital confocal microscopy, we demonstrated that platelets expose phosphatidylserine (PS and fibrin accumulate only in the center of the thrombus but not in its periphery. To address the question how exposure of platelet anionic phospholipids is regulated within the thrombus, an in-vitro experiment using diluted platelet-rich plasma was employed, in which the fibrin network was formed in the presence of platelets, and PS exposure on the platelet surface was analyzed using Confocal Laser Scanning Microscopy. Almost all platelets exposed PS after treatment with tissue factor, thrombin or ionomycin. Argatroban abrogated fibrin network formation in all samples, however, platelet PS exposure was inhibited only in tissue factor- and thrombin-treated samples but not in ionomycin-treated samples. FK633, an α(IIbβ₃ antagonist, and cytochalasin B impaired platelet binding to the fibrin scaffold and significantly reduced PS exposure evoked by thrombin. Gly-Pro-Arg-Pro amide abrogated not only fibrin network formation, but also PS exposure on platelets without suppressing platelet binding to fibrin/fibrinogen. These results suggest that outside-in signals in platelets generated by their binding to the rigid fibrin network are essential for PS exposure after thrombin treatment.

  9. Surface complement C3 fragments and cellular binding of microparticles in patients with SLE

    DEFF Research Database (Denmark)

    Winberg, Line Kjær; Nielsen, Claus Henrik; Jacobsen, Søren

    2017-01-01

    Objectives: To examine microparticles (MPs) from patients with SLE and healthy controls (HCs) by determining the cellular origin of the MPs, quantifying attached fragments of complement component 3 (C3) and assessing the ability of MPs to bind to circulating phagocytes and erythrocytes. These fea......Objectives: To examine microparticles (MPs) from patients with SLE and healthy controls (HCs) by determining the cellular origin of the MPs, quantifying attached fragments of complement component 3 (C3) and assessing the ability of MPs to bind to circulating phagocytes and erythrocytes...

  10. Copper tolerance in Frankia sp. strain EuI1c involves surface binding and copper transport.

    Science.gov (United States)

    Rehan, Medhat; Furnholm, Teal; Finethy, Ryan H; Chu, Feixia; El-Fadly, Gomaah; Tisa, Louis S

    2014-09-01

    Several Frankia strains have been shown to be copper-tolerant. The mechanism of their copper tolerance was investigated for Frankia sp. strain EuI1c. Copper binding was shown by binding studies. Unusual globular structures were observed on the surface of the bacterium. These globular structures were composed of aggregates containing many relatively smaller "leaf-like" structures. Scanning electron microscopy with energy-dispersive X-ray (SEM-EDAX) analysis of these structures indicated elevated copper and phosphate levels compared to the control cells. Fourier transform infrared spectroscopy (FTIR) analysis indicated an increase in extracellular phosphate on the cell surface of copper-stressed cells. Bioinformatics' analysis of the Frankia sp. strain EuI1c genome revealed five potential cop genes: copA, copZ, copC, copCD, and copD. Experiments with Frankia sp. strain EuI1c using qRT-PCR indicated an increase in messenger RNA (mRNA) levels of the five cop genes upon Cu(2+) stress. After 5 days of Cu(2+) stress, the copA, copZ, copC, copCD, and copD mRNA levels increased 25-, 8-, 18-, 18-, and 25-fold, respectively. The protein profile of Cu(2+)-stressed Frankia sp. strain EuI1c cells revealed the upregulation of a 36.7 kDa protein that was identified as FraEuI1c_1092 (sulfate-binding periplasmic transport protein). Homologues of this gene were only present in the genomes of the Cu(2+)-resistant Frankia strains (EuI1c, DC12, and CN3). These data indicate that copper tolerance by Frankia sp. strain EuI1c involved the binding of copper to the cell surface and transport proteins.

  11. Binding of the urokinase-type plasminogen activator to its cell surface receptor is inhibited by low doses of suramin

    DEFF Research Database (Denmark)

    Behrendt, N; Rønne, E; Danø, K

    1993-01-01

    micrograms/ml when using U937 cells and a ligand concentration of 0.3 nM. This concentration of the drug is well below the serum levels found in suramin-treated patients. Inhibition of binding was also demonstrated at the molecular level, using chemical cross-linking or an enzyme-linked immunosorbent assay...... to the anti-invasive properties of suramin by destroying the cellular potential for localized plasminogen activation and proteolytic matrix degradation....

  12. Degradation of the starch components amylopectin and amylose by barley α-amylase 1: Role of surface binding site 2

    DEFF Research Database (Denmark)

    Nielsen, Jonas Willum; Kramhøft, Birte; Bozonnet, Sophie

    2012-01-01

    Barley α-amylase isozyme 1 (AMY1, EC 3.2.1.1) contains two surface binding sites, SBS1 and SBS2, involved in the degradation of starch granules. The distinct role of SBS1 and SBS2 remains to be fully understood. Mutational analysis of Tyr-380 situated at SBS2 previously revealed that Tyr-380...... is required for binding of the amylose helix mimic, β-cyclodextrin. Also, mutant enzymes altered at position 380 displayed reduced binding to starch granules. Similarly, binding of wild type AMY1 to starch granules was suppressed in the presence of β-cyclodextrin. We investigated the role of SBS2 by comparing...... kinetic properties of the wild type AMY1 and the Y380A mutant enzyme in hydrolysis of amylopectin, amylose and β-limit dextrin, and the inhibition by β-cyclodextrin. Progress curves of the release of reducing ends revealed a bi-exponential hydrolysis of amylopectin and β-limit dextrin, whereas hydrolysis...

  13. Surface binding sites in amylase have distinct roles in recognition of starch structure motifs and degradation

    DEFF Research Database (Denmark)

    Cockburn, Darrell; Nielsen, Morten M.; Christiansen, Camilla

    2015-01-01

    degrading enzymes and critically important for their function. The affinity towards a variety of starch granules as well as soluble poly- and oligosaccharides of barley alpha-amylase 1 (AMY1) wild-type and mutants of two SBSs (SBS1 and SBS2) was investigated using Langmuir binding analysis, confocal laser...

  14. The ligand-binding domain of the cell surface receptor for urokinase-type plasminogen activator

    DEFF Research Database (Denmark)

    Behrendt, N; Ploug, M; Patthy, L

    1991-01-01

    with the internal repeats of u-PAR constitute the extracellular part of Ly-6 antigens and of the squid glycoprotein Sgp-2. Like u-PAR, these proteins are attached to the membrane by a glycosyl-phosphatidylinositol anchor. The hydrophilic, ligand-binding u-PAR domain identified in the present study has potential...

  15. Human milk containing specific secretory IgA inhibits binding of Giardia lamblia to nylon and glass surfaces.

    Science.gov (United States)

    Samra, H K; Ganguly, N K; Mahajan, R C

    1991-06-01

    The effects of human milk, containing specific secretory IgA, on the adherence of Giardia lamblia trophozoites in the presence and in the absence of intestinal mucus in vitro were studied. It was found that the trophozoites treated with breast milk, containing specific secretory IgA to G. lamblia, showed a significant decrease (p less than 0.01) in adherence to nylon fibre columns and glass surfaces than did trophozoites treated with milk containing no SIgA antibodies. The adherence to glass surfaces was significantly more (p less than 0.01) in the presence of intestinal mucus than when the mucus was absent. Milk that did not contain specific secretory SIgA to G. lamblia did not decrease the adherence to glass surfaces either in the presence or in the absence of mucus. The fluorescence study revealed the binding of specific secretory IgA on the trophozoite surface. The results suggest that binding of SIgA antibodies in milk to G. lamblia trophozoites inhibits parasite adherence, thus protecting against this infection in breast-fed babies.

  16. Physical adsorption vs. chemical binding of undecylenic acid on porous silicon surface: a comparative study of differently functionalized materials

    Energy Technology Data Exchange (ETDEWEB)

    Salonen, J.; Lehto, V.P. [University of Turku (Finland). Department of Physics; Chirvony, V.; Matveeva, E. [Nanophotonics Technology Center, Technical University of Valencia (Spain); Pastor, E.

    2009-07-15

    To imply miscibility to porous silicon (PSi) used for biomedical purposes a number of functionalization methods are employed. In order to distinguish between a non-specific surfactant-like interaction (physical sorption) and chemical binding of unsaturated chemicals (undecylenic acid, UD) to H-terminated PSi surface we studied the two differently treated materials. Differential scanning calorimetry (DSC) and thermogravimetry (TGA), BET and FTIR measurements were performed with the PSi powder samples (n+ doped). Changes in surface area, weight loss, calorific effect and chemical composition that accompanied the thermal treatment have shown that the physisorbed UD molecules undergo a chemical process (binding) with the Si-H{sub x} surface groups at about 150 C in both, N{sub 2} inert atmosphere and in a synthetic air, oxidative atmosphere. Controlled conversion of physically sorbed molecules to the chemically attached ones is discussed with respect to methods of surface modification of PSi materials for increasing their biocompatibility. (copyright 2009 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  17. Plasminogen Binding Proteins and Plasmin Generation on the Surface of Leptospira spp.: The Contribution to the Bacteria-Host Interactions

    Directory of Open Access Journals (Sweden)

    Monica L. Vieira

    2012-01-01

    Full Text Available Leptospirosis is considered a neglected infectious disease of human and veterinary concern. Although extensive investigations on host-pathogen interactions have been pursued by several research groups, mechanisms of infection, invasion and persistence of pathogenic Leptospira spp. remain to be elucidated. We have reported the ability of leptospires to bind human plasminogen (PLG and to generate enzimatically active plasmin (PLA on the bacteria surface. PLA-coated Leptospira can degrade immobilized ECM molecules, an activity with implications in host tissue penetration. Moreover, we have identified and characterized several proteins that may act as PLG-binding receptors, each of them competent to generate active plasmin. The PLA activity associated to the outer surface of Leptospira could hamper the host immune attack by conferring the bacteria some benefit during infection. The PLA-coated leptospires obstruct complement C3b and IgG depositions on the bacterial surface, most probably through degradation. The decrease of leptospiral opsonization might be an important aspect of the immune evasion strategy. We believe that the presence of PLA on the leptospiral surface may (i facilitate host tissue penetration, (ii help the bacteria to evade the immune system and, as a consequence, (iii permit Leptospira to reach secondary sites of infection.

  18. Modifications of nano-titania surface for in vitro evaluations of hemolysis, cytotoxicity, and nonspecific protein binding

    Energy Technology Data Exchange (ETDEWEB)

    Datta, Aparna, E-mail: adatta.research@gmail.com [Jadavpur University, School of Materials Science and Nanotechnology (India); Dasgupta, Sayantan [NRS Medical College and Hospital, Department of Biochemistry (India); Mukherjee, Siddhartha [Jadavpur University, Department of Metallurgical and Material Engineering (India)

    2017-04-15

    In the past decade, a variety of drug carriers based on mesoporous silica nanoparticles has been extensively reported. However, their biocompatibility still remains debatable, which motivated us to explore the porous nanostructures of other metal oxides, for example titanium dioxide (TiO{sub 2}), as potential drug delivery vehicles. Herein, we report the in vitro hemolysis, cytotoxicity, and protein binding of TiO{sub 2} nanoparticles, synthesized by a sol–gel method. The surface of the TiO{sub 2} nanoparticles was modified with hydroxyl, amine, or thiol containing moieties to examine the influence of surface functional groups on the toxicity and protein binding aspects of the nanoparticles. Our study revealed the superior hemocompatibility of pristine, as well as functionalized TiO{sub 2} nanoparticles, compared to that of mesoporous silica, the present gold standard. Among the functional groups studied, aminosilane moieties on the TiO{sub 2} surface substantially reduced the degree of hemolysis (down to 5%). Further, cytotoxicity studies by MTT assay suggested that surface functional moieties play a crucial role in determining the biocompatibility of the nanoparticles. The presence of NH{sub 2}– functional groups on the TiO{sub 2} nanoparticle surface enhanced the cell viability by almost 28% as compared to its native counterpart (at 100 μg/ml), which was in agreement with the hemolysis assay. Finally, nonspecific protein adsorption on functionalized TiO{sub 2} surfaces was examined using human serum albumin and it was found that negatively charged surface moieties, like –OH and –SH, could mitigate protein adsorption to a significant extent.

  19. Modifications of nano-titania surface for in vitro evaluations of hemolysis, cytotoxicity, and nonspecific protein binding

    Science.gov (United States)

    Datta, Aparna; Dasgupta, Sayantan; Mukherjee, Siddhartha

    2017-04-01

    In the past decade, a variety of drug carriers based on mesoporous silica nanoparticles has been extensively reported. However, their biocompatibility still remains debatable, which motivated us to explore the porous nanostructures of other metal oxides, for example titanium dioxide (TiO2), as potential drug delivery vehicles. Herein, we report the in vitro hemolysis, cytotoxicity, and protein binding of TiO2 nanoparticles, synthesized by a sol-gel method. The surface of the TiO2 nanoparticles was modified with hydroxyl, amine, or thiol containing moieties to examine the influence of surface functional groups on the toxicity and protein binding aspects of the nanoparticles. Our study revealed the superior hemocompatibility of pristine, as well as functionalized TiO2 nanoparticles, compared to that of mesoporous silica, the present gold standard. Among the functional groups studied, aminosilane moieties on the TiO2 surface substantially reduced the degree of hemolysis (down to 5%). Further, cytotoxicity studies by MTT assay suggested that surface functional moieties play a crucial role in determining the biocompatibility of the nanoparticles. The presence of NH2- functional groups on the TiO2 nanoparticle surface enhanced the cell viability by almost 28% as compared to its native counterpart (at 100 μg/ml), which was in agreement with the hemolysis assay. Finally, nonspecific protein adsorption on functionalized TiO2 surfaces was examined using human serum albumin and it was found that negatively charged surface moieties, like -OH and -SH, could mitigate protein adsorption to a significant extent.

  20. Constructing polyatomic potential energy surfaces by interpolating diabatic Hamiltonian matrices with demonstration on green fluorescent protein chromophore

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jae Woo; Rhee, Young Min, E-mail: ymrhee@postech.ac.kr [Center for Self-assembly and Complexity, Institute for Basic Science (IBS), Pohang 790-784 (Korea, Republic of); Department of Chemistry, Pohang University of Science and Technology (POSTECH), Pohang 790-784 (Korea, Republic of)

    2014-04-28

    Simulating molecular dynamics directly on quantum chemically obtained potential energy surfaces is generally time consuming. The cost becomes overwhelming especially when excited state dynamics is aimed with multiple electronic states. The interpolated potential has been suggested as a remedy for the cost issue in various simulation settings ranging from fast gas phase reactions of small molecules to relatively slow condensed phase dynamics with complex surrounding. Here, we present a scheme for interpolating multiple electronic surfaces of a relatively large molecule, with an intention of applying it to studying nonadiabatic behaviors. The scheme starts with adiabatic potential information and its diabatic transformation, both of which can be readily obtained, in principle, with quantum chemical calculations. The adiabatic energies and their derivatives on each interpolation center are combined with the derivative coupling vectors to generate the corresponding diabatic Hamiltonian and its derivatives, and they are subsequently adopted in producing a globally defined diabatic Hamiltonian function. As a demonstration, we employ the scheme to build an interpolated Hamiltonian of a relatively large chromophore, para-hydroxybenzylidene imidazolinone, in reference to its all-atom analytical surface model. We show that the interpolation is indeed reliable enough to reproduce important features of the reference surface model, such as its adiabatic energies and derivative couplings. In addition, nonadiabatic surface hopping simulations with interpolation yield population transfer dynamics that is well in accord with the result generated with the reference analytic surface. With these, we conclude by suggesting that the interpolation of diabatic Hamiltonians will be applicable for studying nonadiabatic behaviors of sizeable molecules.

  1. Demonstration of surface plasmons in metal island films and the effect of the surrounding medium--An undergraduate experiment

    Energy Technology Data Exchange (ETDEWEB)

    Orfanides, P. [Department of Physics, The University of Memphis, Memphis, Tennessee 38152 (United States); Buckner, T. F. [Department of Physics, The University of Memphis, Memphis, Tennessee 38152 (United States); Buncick, M. C. [Department of Physics, The University of Memphis, Memphis, Tennessee 38152 (United States); Meriaudeau, F. [LE21, 12 rue de la fonderie, 71200 Le Creusot, (France); Ferrell, T. L. [Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831 (United States)

    2000-10-01

    We present a demonstration of the surface plasmon phenomenon as it occurs in thin metal island films. The metal films are deposited on glass microscope slides. The effect of the surface plasmon resonance may be observed visually on the slide without further apparatus. Heating the film changes the shape of the islands and therefore the resonant frequency of the surface plasmon and changes the color of the film. Placing the film in a dielectric medium changes the resonance condition for the surface plasmon again and changes the color again. We show this by coating the slides with commercially available liquids with different indices of refraction. We present a theoretical model that assumes the islands are oblate spheroids. There are enough details given so that the equations can be programed and the theoretical optical absorbance can be reproduced. We also present a modification to the theory so that the shift in resonant frequency can be calculated when the spheroids are immersed in the index fluids. We describe our apparatus for making thin films and our optical spectrometer system. We then present optical absorbance measurements of thin films of both Ag and Au in air and in two liquids with different indices of refraction. (c) 2000 American Association of Physics Teachers.

  2. A Density Functional Tight Binding Study of Acetic Acid Adsorption on Crystalline and Amorphous Surfaces of Titania

    Directory of Open Access Journals (Sweden)

    Sergei Manzhos

    2015-02-01

    Full Text Available We present a comparative density functional tight binding study of an organic molecule attachment to TiO2 via a carboxylic group, with the example of acetic acid. For the first time, binding to low-energy surfaces of crystalline anatase (101, rutile (110 and (B-TiO2 (001, as well as to the surface of amorphous (a- TiO2 is compared with the same computational setup. On all surfaces, bidentate configurations are identified as providing the strongest adsorption energy, Eads = −1.93, −2.49 and −1.09 eV for anatase, rutile and (B-TiO2, respectively. For monodentate configurations, the strongest Eads = −1.06, −1.11 and −0.86 eV for anatase, rutile and (B-TiO2, respectively. Multiple monodentate and bidentate configurations are identified on a-TiO2 with a distribution of adsorption energies and with the lowest energy configuration having stronger bonding than that of the crystalline counterparts, with Eads up to −4.92 eV for bidentate and −1.83 eV for monodentate adsorption. Amorphous TiO2 can therefore be used to achieve strong anchoring of organic molecules, such as dyes, that bind via a -COOH group. While the presence of the surface leads to a contraction of the band gap vs. the bulk, molecular adsorption caused no appreciable effect on the band structure around the gap in any of the systems.

  3. Specific binding of a naturally occurring amyloidogenic fragment of Streptococcus mutans adhesin P1 to intact P1 on the cell surface characterized by solid state NMR spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Wenxing; Bhatt, Avni [University of Florida, Department of Biochemistry and Molecular Biology, College of Medicine (United States); Smith, Adam N. [University of Florida, Department of Chemistry, College of Liberal Arts and Sciences (United States); Crowley, Paula J.; Brady, L. Jeannine, E-mail: jbrady@dental.ufl.edu [University of Florida, Department of Oral Biology, College of Dentistry (United States); Long, Joanna R., E-mail: jrlong@ufl.edu [University of Florida, Department of Biochemistry and Molecular Biology, College of Medicine (United States)

    2016-02-15

    The P1 adhesin (aka Antigen I/II or PAc) of the cariogenic bacterium Streptococcus mutans is a cell surface-localized protein involved in sucrose-independent adhesion and colonization of the tooth surface. The immunoreactive and adhesive properties of S. mutans suggest an unusual functional quaternary ultrastructure comprised of intact P1 covalently attached to the cell wall and interacting with non-covalently associated proteolytic fragments thereof, particularly the ∼57-kDa C-terminal fragment C123 previously identified as Antigen II. S. mutans is capable of amyloid formation when grown in a biofilm and P1 is among its amyloidogenic proteins. The C123 fragment of P1 readily forms amyloid fibers in vitro suggesting it may play a role in the formation of functional amyloid during biofilm development. Using wild-type and P1-deficient strains of S. mutans, we demonstrate that solid state NMR (ssNMR) spectroscopy can be used to (1) globally characterize cell walls isolated from a Gram-positive bacterium and (2) characterize the specific binding of heterologously expressed, isotopically-enriched C123 to cell wall-anchored P1. Our results lay the groundwork for future high-resolution characterization of the C123/P1 ultrastructure and subsequent steps in biofilm formation via ssNMR spectroscopy, and they support an emerging model of S. mutans colonization whereby quaternary P1-C123 interactions confer adhesive properties important to binding to immobilized human salivary agglutinin.

  4. Specific binding of a naturally occurring amyloidogenic fragment of Streptococcus mutans adhesin P1 to intact P1 on the cell surface characterized by solid state NMR spectroscopy

    International Nuclear Information System (INIS)

    Tang, Wenxing; Bhatt, Avni; Smith, Adam N.; Crowley, Paula J.; Brady, L. Jeannine; Long, Joanna R.

    2016-01-01

    The P1 adhesin (aka Antigen I/II or PAc) of the cariogenic bacterium Streptococcus mutans is a cell surface-localized protein involved in sucrose-independent adhesion and colonization of the tooth surface. The immunoreactive and adhesive properties of S. mutans suggest an unusual functional quaternary ultrastructure comprised of intact P1 covalently attached to the cell wall and interacting with non-covalently associated proteolytic fragments thereof, particularly the ∼57-kDa C-terminal fragment C123 previously identified as Antigen II. S. mutans is capable of amyloid formation when grown in a biofilm and P1 is among its amyloidogenic proteins. The C123 fragment of P1 readily forms amyloid fibers in vitro suggesting it may play a role in the formation of functional amyloid during biofilm development. Using wild-type and P1-deficient strains of S. mutans, we demonstrate that solid state NMR (ssNMR) spectroscopy can be used to (1) globally characterize cell walls isolated from a Gram-positive bacterium and (2) characterize the specific binding of heterologously expressed, isotopically-enriched C123 to cell wall-anchored P1. Our results lay the groundwork for future high-resolution characterization of the C123/P1 ultrastructure and subsequent steps in biofilm formation via ssNMR spectroscopy, and they support an emerging model of S. mutans colonization whereby quaternary P1-C123 interactions confer adhesive properties important to binding to immobilized human salivary agglutinin

  5. Tamm-Horsfall Glycoprotein Enhances PMN Phagocytosis by Binding to Cell Surface-Expressed Lactoferrin and Cathepsin G That Activates MAP Kinase Pathway

    Directory of Open Access Journals (Sweden)

    Chia-Li Yu

    2011-03-01

    Full Text Available The molecular basis of polymorphonuclear neutrophil (PMN phagocytosis-enhancing activity (PEA by human purified urinary Tamm-Horsfall glyco- protein (THP has not been elucidated. In this study, we found human THP bound to lactoferrin (LF and cathepsin G (CG expressed on the surface of PMN, identified by a proteomic study with MALDI-TOF- LC/LC/mass spectrometric analysis. Pre-incubation of 10% SDS-PAGE electrophoresed PMN lysates with monoclonal anti-LF or anti-CG antibody reduced the binding with THP. To elucidate the signaling pathway of THP on PMN activation, we found THP enhanced ERK1/2 phosphorylation, reduced p38 MAP kinase phosphorylation, but had no effect on DNA binding of the five NF-kB family members in PMN. To further clarify whether the carbohydrate-side chains or protein-core structure in THP molecule is responsible for THP-PEA, THP was cleaved by different degrading enzymes with carbohydrate specificity (neuraminidase and β-galactosidase, protein specificity (V8 protease and proteinase K or glycoconjugate specificity (carboxylpeptidase Y and O-sialoglycoprotein endopeptidase. We clearly demonstrated that the intact protein-core structure in THP molecule was more important for THP-PEA than carbohydrate-side chains. Putting these results together, we conclude that THP adheres to surface-expressed LF and CG on PMN and transduces signaling via the MAP kinase pathway to enhance PMN phagocytosis.

  6. Specific binding of a naturally occurring amyloidogenic fragment of Streptococcus mutans adhesin P1 to intact P1 on the cell surface characterized by solid state NMR spectroscopy.

    Science.gov (United States)

    Tang, Wenxing; Bhatt, Avni; Smith, Adam N; Crowley, Paula J; Brady, L Jeannine; Long, Joanna R

    2016-02-01

    The P1 adhesin (aka Antigen I/II or PAc) of the cariogenic bacterium Streptococcus mutans is a cell surface-localized protein involved in sucrose-independent adhesion and colonization of the tooth surface. The immunoreactive and adhesive properties of S. mutans suggest an unusual functional quaternary ultrastructure comprised of intact P1 covalently attached to the cell wall and interacting with non-covalently associated proteolytic fragments thereof, particularly the ~57-kDa C-terminal fragment C123 previously identified as Antigen II. S. mutans is capable of amyloid formation when grown in a biofilm and P1 is among its amyloidogenic proteins. The C123 fragment of P1 readily forms amyloid fibers in vitro suggesting it may play a role in the formation of functional amyloid during biofilm development. Using wild-type and P1-deficient strains of S. mutans, we demonstrate that solid state NMR (ssNMR) spectroscopy can be used to (1) globally characterize cell walls isolated from a Gram-positive bacterium and (2) characterize the specific binding of heterologously expressed, isotopically-enriched C123 to cell wall-anchored P1. Our results lay the groundwork for future high-resolution characterization of the C123/P1 ultrastructure and subsequent steps in biofilm formation via ssNMR spectroscopy, and they support an emerging model of S. mutans colonization whereby quaternary P1-C123 interactions confer adhesive properties important to binding to immobilized human salivary agglutinin.

  7. Exposed and hidden lectin-binding epitopes at the surface of Borrelia burgdorferi

    Czech Academy of Sciences Publication Activity Database

    Stoitsova, S. R.; Grubhoffer, Libor; Nebesářová, Jana

    2003-01-01

    Roč. 48, č. 5 (2003), s. 654-658 ISSN 0015-5632 R&D Projects: GA AV ČR IAA6022001 Grant - others:National Research Council at the Ministry of Education and Science(BG) K-709/97 Institutional research plan: CEZ:AV0Z6022909 Keywords : Borrelia burgdorferi * lectin-binding epitopes Subject RIV: EE - Microbiology, Virology Impact factor: 0.857, year: 2003

  8. Characterization of a viral phosphoprotein binding site on the surface of the respiratory syncytial nucleoprotein.

    Science.gov (United States)

    Galloux, Marie; Tarus, Bogdan; Blazevic, Ilfad; Fix, Jenna; Duquerroy, Stéphane; Eléouët, Jean-François

    2012-08-01

    The human respiratory syncytial virus (HRSV) genome is composed of a negative-sense single-stranded RNA that is tightly associated with the nucleoprotein (N). This ribonucleoprotein (RNP) complex is the template for replication and transcription by the viral RNA-dependent RNA polymerase. RNP recognition by the viral polymerase involves a specific interaction between the C-terminal domain of the phosphoprotein (P) (P(CTD)) and N. However, the P binding region on N remains to be identified. In this study, glutathione S-transferase (GST) pulldown assays were used to identify the N-terminal core domain of HRSV N (N(NTD)) as a P binding domain. A biochemical characterization of the P(CTD) and molecular modeling of the N(NTD) allowed us to define four potential candidate pockets on N (pocket I [PI] to PIV) as hydrophobic sites surrounded by positively charged regions, which could constitute sites complementary to the P(CTD) interaction domain. The role of selected amino acids in the recognition of the N-RNA complex by P was first screened for by site-directed mutagenesis using a polymerase activity assay, based on an HRSV minigenome containing a luciferase reporter gene. When changed to Ala, most of the residues of PI were found to be critical for viral RNA synthesis, with the R132A mutant having the strongest effect. These mutations also reduced or abolished in vitro and in vivo P-N interactions, as determined by GST pulldown and immunoprecipitation experiments. The pocket formed by these residues is critical for P binding to the N-RNA complex, is specific for pneumovirus N proteins, and is clearly distinct from the P binding sites identified so far for other nonsegmented negative-strand viruses.

  9. The Glycolytic Enzyme Triosephosphate Isomerase of Trichomonas vaginalis Is a Surface-Associated Protein Induced by Glucose That Functions as a Laminin- and Fibronectin-Binding Protein.

    Science.gov (United States)

    Miranda-Ozuna, Jesús F T; Hernández-García, Mar S; Brieba, Luis G; Benítez-Cardoza, Claudia G; Ortega-López, Jaime; González-Robles, Arturo; Arroyo, Rossana

    2016-10-01

    Triosephosphate isomerase of Trichomonas vaginalis (TvTIM) is a 27-kDa cytoplasmic protein encoded by two genes, tvtim1 and tvtim2, that participates in glucose metabolism. TvTIM is also localized to the parasite surface. Thus, the goal of this study was to identify the novel functions of the surface-associated TvTIM in T. vaginalis and to assess the effect of glucose as an environmental factor that regulates its expression and localization. Reverse transcription-PCR (RT-PCR) showed that the tvtim genes were differentially expressed in response to glucose concentration. tvtim1 was overexpressed under glucose-restricted (GR) conditions, whereas tvtim2 was overexpressed under glucose-rich, or high-glucose (HG), conditions. Western blot and indirect immunofluorescence assays also showed that glucose positively affected the amount and surface localization of TvTIM in T. vaginalis Affinity ligand assays demonstrated that the recombinant TvTIM1 and TvTIM2 proteins bound to laminin (Lm) and fibronectin (Fn) but not to plasminogen. Moreover, higher levels of adherence to Lm and Fn were detected in parasites grown under HG conditions than in those grown under GR conditions. Furthermore, pretreatment of trichomonads with an anti-TvTIMr polyclonal antibody or pretreatment of Lm- or Fn-coated wells with both recombinant proteins (TvTIM1r and TvTIM2r) specifically reduced the binding of live parasites to Lm and Fn in a concentration-dependent manner. Moreover, T. vaginalis was exposed to different glucose concentrations during vaginal infection of women with trichomoniasis. Our data indicate that TvTIM is a surface-associated protein under HG conditions that mediates specific binding to Lm and Fn as a novel virulence factor of T. vaginalis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  10. The 1.7 Å X-ray crystal structure of the porcine factor VIII C2 domain and binding analysis to anti-human C2 domain antibodies and phospholipid surfaces.

    Directory of Open Access Journals (Sweden)

    Caileen M Brison

    Full Text Available The factor VIII C2 domain is essential for binding to activated platelet surfaces as well as the cofactor activity of factor VIII in blood coagulation. Inhibitory antibodies against the C2 domain commonly develop following factor VIII replacement therapy for hemophilia A patients, or they may spontaneously arise in cases of acquired hemophilia. Porcine factor VIII is an effective therapeutic for hemophilia patients with inhibitor due to its low cross-reactivity; however, the molecular basis for this behavior is poorly understood. In this study, the X-ray crystal structure of the porcine factor VIII C2 domain was determined, and superposition of the human and porcine C2 domains demonstrates that most surface-exposed differences cluster on the face harboring the "non-classical" antibody epitopes. Furthermore, antibody-binding results illustrate that the "classical" 3E6 antibody can bind both the human and porcine C2 domains, although the inhibitory titer to human factor VIII is 41 Bethesda Units (BU/mg IgG versus 0.8 BU/mg IgG to porcine factor VIII, while the non-classical G99 antibody does not bind to the porcine C2 domain nor inhibit porcine factor VIII activity. Further structural analysis of differences between the electrostatic surface potentials suggest that the C2 domain binds to the negatively charged phospholipid surfaces of activated platelets primarily through the 3E6 epitope region. In contrast, the G99 face, which contains residue 2227, should be distal to the membrane surface. Phospholipid binding assays indicate that both porcine and human factor VIII C2 domains bind with comparable affinities, and the human K2227A and K2227E mutants bind to phospholipid surfaces with similar affinities as well. Lastly, the G99 IgG bound to PS-immobilized factor VIII C2 domain with an apparent dissociation constant of 15.5 nM, whereas 3E6 antibody binding to PS-bound C2 domain was not observed.

  11. A surface plasmon resonance immunosensor for detecting a dioxin precursor using a gold binding polypeptide

    DEFF Research Database (Denmark)

    Soh, N; Tokuda, T.; Watanabe, T.

    2003-01-01

    A surface plasmon resonance (SPR) based biosensor was developed for monitoring 2,4-dichlorophenol, a known dioxin precursor, using an indirect competitive immunoassay. The SPR sensor was fabricated by immobilizing a gold-thin layer on the surface of an SPR sensor chip with an anti-(2,4-dichloroph......A surface plasmon resonance (SPR) based biosensor was developed for monitoring 2,4-dichlorophenol, a known dioxin precursor, using an indirect competitive immunoassay. The SPR sensor was fabricated by immobilizing a gold-thin layer on the surface of an SPR sensor chip with an anti-(2...

  12. Polyethylene imine/graphene oxide layer-by-layer surface functionalization for significantly improved limit of detection and binding kinetics of immunoassays on acrylate surfaces.

    Science.gov (United States)

    Miyazaki, Celina M; Mishra, Rohit; Kinahan, David J; Ferreira, Marystela; Ducrée, Jens

    2017-10-01

    Antibody immobilization on polymeric substrates is a key manufacturing step for microfluidic devices that implement sample-to-answer automation of immunoassays. In this work, a simple and versatile method to bio-functionalize poly(methylmethacrylate) (PMMA), a common material of such "Lab-on-a-Chip" systems, is proposed; using the Layer-by-Layer (LbL) technique, we assemble nanostructured thin films of poly(ethylene imine) (PEI) and graphene oxide (GO). The wettability of PMMA surfaces was significantly augmented by the surface treatment with (PEI/GO) 5 film, with an 81% reduction of the contact angle, while the surface roughness increased by 600%, thus clearly enhancing wettability and antibody binding capacity. When applied to enzyme-linked immunosorbent assays (ELISAs), the limit of detection of PMMA surface was notably improved from 340pgmL -1 on commercial grade polystyrene (PS) and 230pgmL -1 on plain PMMA surfaces to 130pgmL -1 on (PEI/GO) 5 treated PMMA. Furthermore, the accelerated antibody adsorption kinetics on the LbL films of GO allowed to substantially shorten incubation times, e.g. for anti-rat IgG adsorption from 2h down to 15min on conventional and treated surfaces, respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Direct demonstration of ATP-dependent release of SecA from a translocating preprotein by surface plasmon resonance

    NARCIS (Netherlands)

    de Keyzer, J; van der Does, C; Kloosterman, TG; Driessen, AJM

    2003-01-01

    Translocase mediates the transport of preproteins across the inner membrane of Escherichia coli. SecA binds with high affinity to the membrane-embedded protein-conducting SecYEG complex and serves as both a receptor for secretory proteins and as an ATP-driven molecular motor. Cycles of ATP binding

  14. Demonstration of a reduction in muscarinic receptor binding in early Alzheimer's disease using iodine-123 dexetimide single-photon emission tomography

    International Nuclear Information System (INIS)

    Claus, J.J.; Dubois, E.A.; Booij, J.; Habraken, J.; Munck, J.C. van; Herk, M. van; Verbeeten, B. Jr.; Royen, E.A. van

    1997-01-01

    Decreased muscarinic receptor binding has been suggested in single-photon emission tomography (SPET) studies of Alzheimer's disease. However, it remains unclear whether these changes are present in mildly demented patients, and the role of cortical atrophy in receptor binding assessment has not been investigated. We studied muscarinic receptor binding normalized to neostriatum with SPET using [ 123 I[4-iododexetimide in five mildly affected patients with probable Alzheimer's disease and in five age-matched control subjects. Region of interest (ROI) analysis was performed in a consensus procedure blind to clinical diagnosis using matched magnetic resonance (MRI) images. Cortical atrophy was assessed by calculating percentages of cerebrospinal fluid in each ROI. An observer study with three observers was conducted to validate this method. Alzheimer patients showed statistically significantly less [ 123 I[4-iododexetimide binding in left temporal and right temporo-parietal cortex compared with controls, independent of age, sex and cortical atrophy. Mean intra-observer variability was 3.6% and inter-observer results showed consistent differences in [ 123 I[4-iododexetimide binding between observers. However, differences between patients and controls were comparable among observers and statistically significant in the same regions as in the consensus procedure. Using an MRI-SPET matching technique, we conclude that [ 123 I[4-iododexetimide binding is reduced in patients with mild probable Alzheimer's disease in areas of temporal and temporo-parietal cortex. (orig.). With 1 fig., 4 tabs

  15. Cell Surface Binding and Internalization of Aβ Modulated by Degree of Aggregation

    Directory of Open Access Journals (Sweden)

    David A. Bateman

    2011-01-01

    Full Text Available The amyloid peptides, Aβ40 and Aβ42, are generated through endoproteolytic cleavage of the amyloid precursor protein. Here we have developed a model to investigate the interaction of living cells with various forms of aggregated Aβ40/42. After incubation at endosomal pH 6, we observed a variety of Aβ conformations after 3 (Aβ3, 24 (Aβ24, and 90 hours (Aβ90. Both Aβ4224 and Aβ4024 were observed to rapidly bind and internalize into differentiated PC12 cells, leading to accumulation in the lysosome. In contrast, Aβ40/4290 were both found to only weakly associate with cells, but were observed as the most aggregated using dynamic light scattering and thioflavin-T. Internalization of Aβ40/4224 was inhibited with treatment of monodansylcadaverine, an endocytosis inhibitor. These studies indicate that the ability of Aβ40/42 to bind and internalize into living cells increases with degree of aggregation until it reaches a maximum beyond which its ability to interact with cells diminishes drastically.

  16. Screening Effect of PEG on Avidin Binding to Liposome Surface Receptors

    DEFF Research Database (Denmark)

    Kaasgaard, Thomas; Mouritsen, Ole G.; Jørgensen, Kent

    2000-01-01

    This study investigates the screening effect of poly(ethylene glycol)-phospholipids (PE-PEG) on the interaction of avidin with PEGylated liposomes containing surface-bound biotin ligands. The influence of grafting density and lipopolymer chain length is examined. A simple fluorescence assay....... Furthermore. it is found that none of the lipopolymers completely prevents avidin from reaching the surface-bound biotin ligands....

  17. Thermodynamic analysis of water molecules at the surface of proteins and applications to binding site prediction and characterization.

    Science.gov (United States)

    Beuming, Thijs; Che, Ye; Abel, Robert; Kim, Byungchan; Shanmugasundaram, Veerabahu; Sherman, Woody

    2012-03-01

    Water plays an essential role in determining the structure and function of all biological systems. Recent methodological advances allow for an accurate and efficient estimation of the thermodynamic properties of water molecules at the surface of proteins. In this work, we characterize these thermodynamic properties and relate them to various structural and functional characteristics of the protein. We find that high-energy hydration sites often exist near protein motifs typically characterized as hydrophilic, such as backbone amide groups. We also find that waters around alpha helices and beta sheets tend to be less stable than waters around loops. Furthermore, we find no significant correlation between the hydration site-free energy and the solvent accessible surface area of the site. In addition, we find that the distribution of high-energy hydration sites on the protein surface can be used to identify the location of binding sites and that binding sites of druggable targets tend to have a greater density of thermodynamically unstable hydration sites. Using this information, we characterize the FKBP12 protein and show good agreement between fragment screening hit rates from NMR spectroscopy and hydration site energetics. Finally, we show that water molecules observed in crystal structures are less stable on average than bulk water as a consequence of the high degree of spatial localization, thereby resulting in a significant loss in entropy. These findings should help to better understand the characteristics of waters at the surface of proteins and are expected to lead to insights that can guide structure-based drug design efforts. Copyright © 2011 Wiley Periodicals, Inc.

  18. Improving density functional tight binding predictions of free energy surfaces for peptide condensation reactions in solution

    Science.gov (United States)

    Kroonblawd, Matthew; Goldman, Nir

    First principles molecular dynamics using highly accurate density functional theory (DFT) is a common tool for predicting chemistry, but the accessible time and space scales are often orders of magnitude beyond the resolution of experiments. Semi-empirical methods such as density functional tight binding (DFTB) offer up to a thousand-fold reduction in required CPU hours and can approach experimental scales. However, standard DFTB parameter sets lack good transferability and calibration for a particular system is usually necessary. Force matching the pairwise repulsive energy term in DFTB to short DFT trajectories can improve the former's accuracy for chemistry that is fast relative to DFT simulation times (Contract DE-AC52-07NA27344.

  19. Improving Density Functional Tight Binding Predictions of Free Energy Surfaces for Slow Chemical Reactions in Solution

    Science.gov (United States)

    Kroonblawd, Matthew; Goldman, Nir

    2017-06-01

    First principles molecular dynamics using highly accurate density functional theory (DFT) is a common tool for predicting chemistry, but the accessible time and space scales are often orders of magnitude beyond the resolution of experiments. Semi-empirical methods such as density functional tight binding (DFTB) offer up to a thousand-fold reduction in required CPU hours and can approach experimental scales. However, standard DFTB parameter sets lack good transferability and calibration for a particular system is usually necessary. Force matching the pairwise repulsive energy term in DFTB to short DFT trajectories can improve the former's accuracy for reactions that are fast relative to DFT simulation times (Contract DE-AC52-07NA27344.

  20. Co-expression of TIMP-1 and its cell surface binding partner CD63 in glioblastomas

    DEFF Research Database (Denmark)

    Aaberg-Jessen, Charlotte; Sørensen, Mia D.; Matos, Ana L.S.A.

    2018-01-01

    scoring. CD63 expression in tumor-associated microglia/macrophages was examined by double-immunofluorescence with ionized calcium-binding adapter molecule 1 (Iba1). The association between CD63 and TIMP-1 was investigated using previously obtained TIMP-1 data from our astrocytoma cohort. Cellular co-expression...... of CD63 was widely distributed in astrocytomas with a significantly increased level in glioblastomas. CD63 levels did not significantly correlate with patient survival at a protein level, and CD63 did not augment the prognostic significance of TIMP-1. Up to 38% of the CD63+ cells expressed Iba1; however......, Iba1 did not appear to impact the prognostic value of CD63. A significant correlation was found between TIMP-1 and CD63, and the TIMP-1 and CD63 proteins were co-expressed at the cellular level and located in close molecular proximity, suggesting that TIMP-1 and CD63 could be co...

  1. Surface binding sites (SBSs), mechanism and regulation of enzymes degrading amylopectin and α-limit dextrins

    DEFF Research Database (Denmark)

    Møller, Marie Sofie; Cockburn, Darrell; Nielsen, Jonas W.

    2013-01-01

    into barley seed α-amylase 1 (AMY1) and limit dextrinase (LD) includes i. kinetics of bi-exponential amylopectin hydrolysis by AMY1, one reaction having low Km (8 μg/mL) and high kcat (57 s-1) and the other high Km (97 μg/mL) and low kcat (23 s-1). β-Cyclodextrin (β-CD) inhibits the first reaction by binding...... to an SBS (SBS2) on domain C with Kd = 70 μM, which for the SBS2 Y380A mutant increases to 1.4 mM. SBS2 thus has a role in the fast, high-affinity component of amylopectin degradation. ii. The N-terminal domain of LD, the debranching enzyme in germinating seeds, shows distant structural similarity...

  2. Developing a Novel Hydrogen Sponge with Ideal Binding Energy and High Surface Area for Practical Hydrogen Storage

    Energy Technology Data Exchange (ETDEWEB)

    Chung, T. C. Mike

    2018-04-19

    This Phase I (5 quarters) research project was to examine the validity of a new class of boron-containing polymer (B-polymer) frameworks, serving as the adsorbents for the practical onboard H2 storage applications. Three B-polymer frameworks were synthesized and investigated, which include B-poly(butyenylstyrene) (B-PBS) framework (A), B-poly(phenyldiacetyene) (B-PPDA) framework (B), and B-poly(phenyltriacetylene) (B-PPTA) framework (C). They are 2-D polymer structures with the repeating cyclic units that spontaneously form open morphology and the B-doped (p-type) π-electrons delocalized surfaces. The ideal B-polymer framework shall exhibit open micropores (pore size in the range of 1-1.5nm) with high surface area (>3000 m2/g), and the B-dopants in the conjugated framework shall provide high surface energy for interacting with H2 molecules (an ideal H2 binding energy in the range of 15-25 kJ/mol). The pore size distribution and H2 binding energy were investigated at both Penn State and NREL laboratories. So far, the experimental results show the successful synthesis of B-polymer frameworks with the relatively well-defined planar (2-D) structures. The intrinsically formed porous morphology exhibits a broad pore size distribution (in the range of 0.5-10 nm) with specific surface area (~1000 m2/g). The miss-alignment between 2-D layers may block some micropore channels and limit gas diffusion throughout the entire matrix. In addition, the 2-D planar conjugated structure may also allow free π-electrons delocalization throughout the framework, which significantly reduces the acidity of B-moieties (electron-deficiency).The resulting 2-D B-polymer frameworks only exhibit a small increase of H2 binding energy in the range of 8-9 KJ/mole (quite constant over the whole sorption range).

  3. The Covalent Binding of Photosensitive Dyes to Monocrystalline Silicon Surface and Their Spectral Response

    Institute of Scientific and Technical Information of China (English)

    郭志新; 郝纪祥; 张祖训; 曹子祥

    1993-01-01

    A chemical method is proposed to bond photo-sensitive dyes directly to the surface of polished monocrystalline silicon. A methincyanine dye and a trimethincyanine dye have been bonded covalently onto silicon surface through Si—N bond, which are characterized by XPS technique and laser Raman spectra. Photovoltaic effect has been observed with the In/dye/n-Si sandwich devices composed of the dye-bonded n-Si wafers. Significant spectral response shows the characteristic absorptance maxima of the bonded dyes.

  4. An Extended Surface Loop on Toxoplasma gondii Apical Membrane Antigen 1 (AMA1 Governs Ligand Binding Selectivity.

    Directory of Open Access Journals (Sweden)

    Michelle L Parker

    Full Text Available Apicomplexan parasites are the causative agents of globally prevalent diseases including malaria and toxoplasmosis. These obligate intracellular pathogens have evolved a sophisticated host cell invasion strategy that relies on a parasite-host cell junction anchored by interactions between apical membrane antigens (AMAs on the parasite surface and rhoptry neck 2 (RON2 proteins discharged from the parasite and embedded in the host cell membrane. Key to formation of the AMA1-RON2 complex is displacement of an extended surface loop on AMA1 called the DII loop. While conformational flexibility of the DII loop is required to expose the mature RON2 binding groove, a definitive role of this substructure has not been elucidated. To establish a role of the DII loop in Toxoplasma gondii AMA1, we engineered a form of the protein where the mobile portion of the loop was replaced with a short Gly-Ser linker (TgAMA1ΔDIIloop. Isothermal titration calorimetry measurements with a panel of RON2 peptides revealed an influential role for the DII loop in governing selectivity. Most notably, an Eimeria tenella RON2 (EtRON2 peptide that showed only weak binding to TgAMA1 bound with high affinity to TgAMA1ΔDIIloop. To define the molecular basis for the differential binding, we determined the crystal structure of TgAMA1ΔDIIloop in complex with the EtRON2 peptide. When analyzed in the context of existing AMA1-RON2 structures, spatially distinct anchor points in the AMA1 groove were identified that, when engaged, appear to provide the necessary traction to outcompete the DII loop. Collectively, these data support a model where the AMA1 DII loop serves as a structural gatekeeper to selectively filter out ligands otherwise capable of binding with high affinity in the AMA1 apical groove. These data also highlight the importance of considering the functional implications of the DII loop in the ongoing development of therapeutic intervention strategies targeting the AMA1-RON

  5. Domain interplay in the urokinase receptor. Requirement for the third domain in high affinity ligand binding and demonstration of ligand contact sites in distinct receptor domains

    DEFF Research Database (Denmark)

    Behrendt, N; Ronne, E; Dano, K

    1996-01-01

    by chemical cross-linking, but quantitative binding/competition studies showed that the apparent ligand affinity was 100- to 1000-fold lower than that of the intact suPAR. This loss of affinity was comparable with the loss found after cleavage between the first domain (D1) and D(2 + 3), using chymotrypsin...

  6. Selective albumin-binding surfaces modified with a thrombin-inhibiting peptide.

    Science.gov (United States)

    Freitas, Sidónio C; Maia, Sílvia; Figueiredo, Ana C; Gomes, Paula; Pereira, Pedro J B; Barbosa, Mário A; Martins, M Cristina L

    2014-03-01

    Blood-contacting medical devices have been associated with severe clinical complications, such as thrombus formation, triggered by the activation of the coagulation cascade due to the adsorption of certain plasma proteins on the surface of biomaterials. Hence, the coating of such surfaces with antithrombotic agents has been used to increase biomaterial haemocompatibility. Biomaterial-induced clotting may also be decreased by albumin adsorption from blood plasma in a selective and reversible way, since this protein is not involved in the coagulation cascade. In this context, this paper reports that the immobilization of the thrombin inhibitor D-Phe-Pro-D-Arg-D-Thr-CONH2 (fPrt) onto nanostructured surfaces induces selective and reversible adsorption of albumin, delaying the clotting time when compared to peptide-free surfaces. fPrt, synthesized with two glycine residues attached to the N-terminus (GGfPrt), was covalently immobilized onto self-assembled monolayers (SAMs) having different ratios of carboxylate-hexa(ethylene glycol)- and tri(ethylene glycol)-terminated thiols (EG6-COOH/EG3) that were specifically designed to control GGfPrt orientation, exposure and density at the molecular level. In solution, GGfPrt was able to inactivate the enzymatic activity of thrombin and to delay plasma clotting time in a concentration-dependent way. After surface immobilization, and independently of its concentration, GGfPrt lost its selectivity to thrombin and its capacity to inhibit thrombin enzymatic activity against the chromogenic substrate n-p-tosyl-Gly-Pro-Arg-p-nitroanilide. Nevertheless, surfaces with low concentrations of GGfPrt could delay the capacity of adsorbed thrombin to cleave fibrinogen. In contrast, GGfPrt immobilized in high concentrations was found to induce the procoagulant activity of the adsorbed thrombin. However, all surfaces containing GGfPrt have a plasma clotting time similar to the negative control (empty polystyrene wells), showing resistance to

  7. Studies on the pathogenesis of Aleutian disease of mink. X. demonstration of immune complexes by the /sup 125/I-C 1 q binding test after experimental infection

    Energy Technology Data Exchange (ETDEWEB)

    Mueller-Peddinghaus, R [Kali-Chemie Pharma G.m.b.H., Hannover (Germany, F.R.). Abt. fuer Experimentelle Pathologie; Meyer zu Schwabedissen, H [Medizinische Hochschule Hannover (Germany, F.R.). Abt. fuer Klinische Immunologie und Bluttransfusionswesen; Kalden, J R [Erlangen-Nuernberg Univ., Erlangen (Germany, F.R.). Inst. und Poliklinik fuer Klinische Immunologie; Trautwein, G; Ueberschaer, S [Tieraerztliche Hochschule Hannover (Germany, F.R.). Inst. fuer Pathologie

    1980-01-01

    Aleutian disease (AD) of mink most closely resembles systemic lupus erythematosus (SLE) in man; both are immune complex disease. In experimental AD serum immune complexes are determined by the /sup 125/J-C 1 q-binding test using human C 1 q. Mink (n = 12) infected intraperitoneally with Aleutian disease virus (ADV), grown in fetal mink kidney cells, developed during the course of infection a mean of /sup 125/I-C 1 q serum binding equivalent to 3.62 +- 1.68 mg./ml. aggr. HGG. (aggregated human immunoglobulin). Sera of mink (n = 8) which were infected with ADV grown in L-cells showed a less marked /sup 125/I-C 1 q binding with a mean equivalent to 2.52 +- 1.43 mg./ml. aggr. HGG. In contrast control animals (n = 8) treated with non-ADV-infected mink epidermal fibroblasts or Eagle's minimal essential medium substituted with fetal calf serum only bound /sup 125/I-C 1 q equivalent to 1.02 +- 0.99 mg./ml. aggr. HGG. In mink infected with ADV propagated in fetal mink kidney cells a constant increase in the /sup 125/I-C 1 q serum binding occurred from the 4th to the 7th and 13th week after ADV infection. Mink which were infected with ADV propagated in mouse L-cells exhibited a different pattern of the /sup 125/I-C 1 q serum binding capacity with a sharp increase from the 4th to the 7th week, followed by a decline towards the 13th week post infection. The serum /sup 125/I-C 1 q binding capacity of all experimental animal groups exhibited at different times of the experiment a significant correlation with the presence of hypergammaglobulinaemia and raised ADV-antibody titers. From the data obtained it appears that the /sup 125/I-C 1 q binding test, utilizing human C 1 q, is a suitable method for the detection of circulating serum immune complexes in mink during the course of ADV-infection.

  8. Effects of DDT and Triclosan on Tumor-cell Binding Capacity and Cell-Surface Protein Expression of Human Natural Killer Cells

    Science.gov (United States)

    Hurd-Brown, Tasia; Udoji, Felicia; Martin, Tamara; Whalen, Margaret M.

    2012-01-01

    1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT) and triclosan (TCS) are organochlorine (OC) compounds that contaminate the environment, are found in human blood, and have been shown to decrease the tumor-cell killing (lytic) function of human natural killer (NK) cells. NK cells defend against tumor cells and virally infected cells. They bind to these targets, utilizing a variety of cell surface proteins. This study examined concentrations of DDT and TCS that decrease lytic function for alteration of NK binding to tumor targets. Levels of either compound that caused loss of binding function were then examined for effects on expression of cell-surface proteins needed for binding. NK cells exposed to 2.5 μM DDT for 24 h (which caused a greater than 55% loss of lytic function) showed a decrease in NK binding function of about 22%, and a decrease in CD16 cell-surface protein of 20%. NK cells exposed to 5 μM TCS for 24 h showed a decrease in ability to bind tumor cells of 37% and a decrease in expression of CD56 of about 34%. This same treatment caused a decrease in lytic function of greater than 87%. These results indicated that only a portion of the loss of NK lytic function seen with exposures to these compounds could be accounted for by loss of binding function. They also showed that loss of binding function is accompanied by a loss cell-surface proteins important in binding function. PMID:22729613

  9. Modelling of microbial polyhydroxyalkanoate surface binding protein PhaP for rational mutagenesis.

    Science.gov (United States)

    Zhao, Hongyu; Yao, Zhenyu; Chen, Xiangbin; Wang, Xinquan; Chen, Guo-Qiang

    2017-11-01

    Phasins are unusual amphiphilic proteins that bind to microbial polyhydroxyalkanoate (PHA) granules in nature and show great potential for various applications in biotechnology and medicine. Despite their remarkable diversity, only the crystal structure of PhaP A h from Aeromonas hydrophila has been solved to date. Based on the structure of PhaP A h , homology models of PhaP A z from Azotobacter sp. FA-8 and PhaP TD from Halomonas bluephagenesis TD were successfully established, allowing rational mutagenesis to be conducted to enhance the stability and surfactant properties of these proteins. PhaP A z mutants, including PhaP A z Q38L and PhaP A z Q78L, as well as PhaP TD mutants, including PhaP TD Q38M and PhaP TD Q72M, showed better emulsification properties and improved thermostability (6-10°C higher melting temperatures) compared with their wild-type homologues under the same conditions. Importantly, the established PhaP homology-modelling approach, based on the high-resolution structure of PhaP A h , can be generalized to facilitate the study of other PhaP members. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  10. Fragment library screening identifies hits that bind to the non-catalytic surface of Pseudomonas aeruginosa DsbA1

    Science.gov (United States)

    Headey, Stephen J.; Vazirani, Mansha; Shouldice, Stephen R.; Coinçon, Mathieu; Tay, Stephanie; Morton, Craig J.; Simpson, Jamie S.; Martin, Jennifer L.

    2017-01-01

    At a time when the antibiotic drug discovery pipeline has stalled, antibiotic resistance is accelerating with catastrophic implications for our ability to treat bacterial infections. Globally we face the prospect of a future when common infections can once again kill. Anti-virulence approaches that target the capacity of the bacterium to cause disease rather than the growth or survival of the bacterium itself offer a tantalizing prospect of novel antimicrobials. They may also reduce the propensity to induce resistance by removing the strong selection pressure imparted by bactericidal or bacteriostatic agents. In the human pathogen Pseudomonas aeruginosa, disulfide bond protein A (PaDsbA1) plays a central role in the oxidative folding of virulence factors and is therefore an attractive target for the development of new anti-virulence antimicrobials. Using a fragment-based approach we have identified small molecules that bind to PaDsbA1. The fragment hits show selective binding to PaDsbA1 over the DsbA protein from Escherichia coli, suggesting that developing species-specific narrow-spectrum inhibitors of DsbA enzymes may be feasible. Structures of a co-complex of PaDsbA1 with the highest affinity fragment identified in the screen reveal that the fragment binds on the non-catalytic surface of the protein at a domain interface. This biophysical and structural data represent a starting point in the development of higher affinity compounds, which will be assessed for their potential as selective PaDsbA1 inhibitors. PMID:28346540

  11. New strategy for enhancement of microbial viability in simulated gastric conditions based on display of starch-binding domain on cell surface.

    Science.gov (United States)

    Tarahomjoo, Shirin; Katakura, Yoshio; Shioya, Suteaki

    2008-05-01

    The C-terminal region of the peptidoglycan hydrolase (CPH) of Lactococcus lactis IL1403 fused to the linker region and the starch-binding domain (SBD) of the *-amylase of Streptococcus bovis 148 was produced intracellularly in Escherichia coli. The fusion protein (CPH-SBD) was able to bind to the cell surface of Lactobacillus casei NRRL B-441 and to corn starch. Therefore, adhesion of cells to corn starch was mediated by the fusion protein. At a cell density of 10(9) cfu/ml and a starch concentration of 5 mg/ml, CPH-SBD-displaying L. casei cells aggregated with corn starch, whereas the free cells of L. casei did not form any aggregates with corn starch. After incubation in simulated gastric juice (pH 3.0, 1 h), the survival percentages of free cells, amylose-coated free cells, and free cells mixed with corn starch were 0.074%, 7.2%, and 3.1% respectively. When CPH-SBD-displaying bacteria aggregated with corn starch, their survival percentage was 8% higher than that of free cells mixed with corn starch. The survival of the amylose-coated CPH-SBD-displaying L. casei cells was comparable to that of amylose-coated free cells, whereas the survival percentage of amylose-coated aggregates of CPH-SBD-displaying bacteria with corn starch was 28% higher than that of amylose-coated mixture of free cells with corn starch. These results demonstrate the potential usefulness of the cell-surface display technique for enhancement of the delivery of viable microorganisms to the intestinal tract.

  12. Layer-by-Layer Heparinization of the Cell Surface by Using Heparin-Binding Peptide Functionalized Human Serum Albumin.

    Science.gov (United States)

    Song, Guowei; Hu, Yaning; Liu, Yusheng; Jiang, Rui

    2018-05-20

    Layer-by-layer heparinization of therapeutic cells prior to transplantation is an effective way to inhibit the instant blood-mediated inflammatory reactions (IBMIRs), which are the major cause of early cell graft loss during post-transplantation. Here, a conjugate of heparin-binding peptide (HBP) and human serum albumin (HSA), HBP-HSA, was synthesized by using heterobifunctional crosslinker. After the first heparin layer was coated on human umbilical vein endothelial cells (HUVECs) by means of the HBP-polyethylene glycol-phospholipid conjugate, HBP-HSA and heparin were then applied to the cell surface sequentially to form multiple layers. The immobilization and retention of heparin were analyzed by confocal microscopy and flow cytometry, respectively, and the cytotoxity of HBP-HSA was further evaluated by cell viability assay. Results indicated that heparin was successfully introduced to the cell surface in a layer-by-layer way and retained for at least 24 h, while the cytotoxity of HBP-HSA was negligible at the working concentration. Accordingly, this conjugate provides a promising method for co-immobilization of heparin and HSA to the cell surface under physiological conditions with improved biocompatibility.

  13. Observation of core-level binding energy shifts between (100) surface and bulk atoms of epitaxial CuInSe{sub 2}

    Energy Technology Data Exchange (ETDEWEB)

    Nelson, A.J. [Colorado School of Mines, Golden, CO (United States); Berry, G.; Rockett, A. [Univ. of Illinois, Urbana-Champaign, IL (United States)] [and others

    1997-04-01

    Core-level and valence band photoemission from semiconductors has been shown to exhibit binding energy differences between surface atoms and bulk atoms, thus allowing one to unambiguously distinguish between the two atomic positions. Quite clearly, surface atoms experience a potential different from the bulk due to the lower coordination number - a characteristic feature of any surface is the incomplete atomic coordination. Theoretical accounts of this phenomena are well documented in the literature for III-V and II-VI semiconductors. However, surface state energies corresponding to the equilibrium geometry of (100) and (111) surfaces of Cu-based ternary chalcopyrite semiconductors have not been calculated or experimental determined. These compounds are generating great interest for optoelectronic and photovoltaic applications, and are an isoelectronic analog of the II-VI binary compound semiconductors. Surface core-level binding energy shifts depend on the surface cohesive energies, and surface cohesive energies are related to surface structure. For ternary compound semiconductor surfaces, such as CuInSe{sub 2}, one has the possibility of variations in surface stoichiometry. Applying standard thermodynamical calculations which consider the number of individual surface atoms and their respective chemical potentials should allow one to qualitatively determine the magnitude of surface core-level shifts and, consequently, surface state energies.

  14. Observation of core-level binding energy shifts between (100) surface and bulk atoms of epitaxial CuInSe2

    International Nuclear Information System (INIS)

    Nelson, A.J.; Berry, G.; Rockett, A.

    1997-01-01

    Core-level and valence band photoemission from semiconductors has been shown to exhibit binding energy differences between surface atoms and bulk atoms, thus allowing one to unambiguously distinguish between the two atomic positions. Quite clearly, surface atoms experience a potential different from the bulk due to the lower coordination number - a characteristic feature of any surface is the incomplete atomic coordination. Theoretical accounts of this phenomena are well documented in the literature for III-V and II-VI semiconductors. However, surface state energies corresponding to the equilibrium geometry of (100) and (111) surfaces of Cu-based ternary chalcopyrite semiconductors have not been calculated or experimental determined. These compounds are generating great interest for optoelectronic and photovoltaic applications, and are an isoelectronic analog of the II-VI binary compound semiconductors. Surface core-level binding energy shifts depend on the surface cohesive energies, and surface cohesive energies are related to surface structure. For ternary compound semiconductor surfaces, such as CuInSe 2 , one has the possibility of variations in surface stoichiometry. Applying standard thermodynamical calculations which consider the number of individual surface atoms and their respective chemical potentials should allow one to qualitatively determine the magnitude of surface core-level shifts and, consequently, surface state energies

  15. Positron emission tomography with ( sup 18 F)methylspiperone demonstrates D sub 2 dopamine receptor binding differences of clozapine and haloperidol

    Energy Technology Data Exchange (ETDEWEB)

    Karbe, H; Wienhard, K; Huber, M; Herholz, K; Heiss, W D [Klinik fuer Neurologie, Univ. of Koeln, Koeln (Germany); Hamacher, K; Coenen, H H; Stoecklin, G [Inst. fuer Chemie 1, Forschungszentrum Juelich Gmbh (Germany); Loevenich, A [Klinik fuer Psychiatrie, Univ. of Koeln, Koeln (Germany)

    1991-01-01

    Four schizophrenic patients were investigated with dynamic positron emission tomography (PET) using ({sup 18}F)fluorodeoxyglucose (FDG) and ({sup 18}F)methylspiperone (MSP) as tracers. Two schizophrenics were on haloperidol therapy at the time of MSP PET. The other two schizophrenics were treated with clozapine, in one of them MSP PET was carried out twice with different daily doses (100 mg and 450mg respectively). Neuroleptic serum levels were measured in all patients. Results were compared with MSP PET of two drugfree male control subjects and with a previous fluoroethylspiperone (FESP) study of normals. Three hours after tracer injection specific binding of MSP was observed in the striatum in all cases. The striatum to cerebellum ratio was used to estimate the degree of neuroleptic-caused striatal D{sub 2} dopamine receptor occupancy. In the haloperidol treated patients MSP binding was significantly decreased, whereas in the clozapine treated patients striatum to cerebellum ratio was normal. Even the increase of clozapine dose in the same patient had no influence on this ratio. Despite the smaller number of patients the study shows for the first time in humans that striatal MSP binding reflects the different D{sub 2} dopamine receptor affinities of clozapine and haloperidol. (authors).

  16. An interaction study in mammalian cells demonstrates weak binding of HSPB2 to BAG3, which is regulated by HSPB3 and abrogated by HSPB8.

    Science.gov (United States)

    Morelli, Federica F; Mediani, Laura; Heldens, Lonneke; Bertacchini, Jessika; Bigi, Ilaria; Carrà, Arianna Dorotea; Vinet, Jonathan; Carra, Serena

    2017-07-01

    The ten mammalian small heat shock proteins (sHSPs/HSPBs) show a different expression profile, although the majority of them are abundant in skeletal and cardiac muscles. HSPBs form hetero-oligomers and homo-oligomers by interacting together and complexes containing, e.g., HSPB2/HSPB3 or HSPB1/HSPB5 have been documented in mammalian cells and muscles. Moreover, HSPB8 associates with the Hsc70/Hsp70 co-chaperone BAG3, in mammalian, skeletal, and cardiac muscle cells. Interaction of HSPB8 with BAG3 regulates its stability and function. Weak association of HSPB5 and HSPB6 with BAG3 has been also reported upon overexpression in cells, supporting the idea that BAG3 might indirectly modulate the function of several HSPBs. However, it is yet unknown whether other HSPBs highly expressed in muscles such as HSPB2 and HSPB3 also bind to BAG3. Here, we report that in mammalian cells, upon overexpression, HSPB2 binds to BAG3 with an affinity weaker than HSPB8. HSPB2 competes with HSPB8 for binding to BAG3. In contrast, HSPB3 negatively regulates HSPB2 association with BAG3. In human myoblasts that express HSPB2, HSPB3, HSPB8, and BAG3, the latter interacts selectively with HSPB8. Combining these data, it supports the interpretation that HSPB8-BAG3 is the preferred interaction.

  17. Demonstration of high-performance p-type tin oxide thin-film transistors using argon-plasma surface treatments

    Science.gov (United States)

    Bae, Sang-Dae; Kwon, Soo-Hun; Jeong, Hwan-Seok; Kwon, Hyuck-In

    2017-07-01

    In this work, we investigated the effects of low-temperature argon (Ar)-plasma surface treatments on the physical and chemical structures of p-type tin oxide thin-films and the electrical performance of p-type tin oxide thin-film transistors (TFTs). From the x-ray photoelectron spectroscopy measurement, we found that SnO was the dominant phase in the deposited tin oxide thin-film, and the Ar-plasma treatment partially transformed the tin oxide phase from SnO to SnO2 by oxidation. The resistivity of the tin oxide thin-film increased with the plasma-treatment time because of the reduced hole concentration. In addition, the root-mean-square roughness of the tin oxide thin-film decreased as the plasma-treatment time increased. The p-type oxide TFT with an Ar-plasma-treated tin oxide thin-film exhibited excellent electrical performance with a high current on-off ratio (5.2 × 106) and a low off-current (1.2 × 10-12 A), which demonstrates that the low-temperature Ar-plasma treatment is a simple and effective method for improving the electrical performance of p-type tin oxide TFTs.

  18. Plasmonic Heterodimers with Binding Site-Dependent Hot Spot for Surface-Enhanced Raman Scattering.

    Science.gov (United States)

    Tian, Yuanyuan; Shuai, Zhenhua; Shen, Jingjing; Zhang, Lei; Chen, Shufen; Song, Chunyuan; Zhao, Baomin; Fan, Quli; Wang, Lianhui

    2018-05-07

    A novel plasmonic heterodimer nanostructure with a controllable self-assembled hot spot is fabricated by the conjugation of individual Au@Ag core-shell nanocubes (Au@Ag NCs) and varisized gold nanospheres (GNSs) via the biotin-streptavidin interaction from the ensemble to the single-assembly level. Due to their featured configurations, three types of heterogeneous nanostructures referred to as Vertice, Vicinity, and Middle are proposed and a single hot spot forms between the nanocube and nanosphere, which exhibits distinct diversity in surface plasmon resonance effect. Herein, the calculated surface-enhanced Raman scattering enhancement factors of the three types of heterodimers show a narrow distribution and can be tuned in orders of magnitude by controlling the size of GNSs onto individual Au@Ag NCs. Particularly, the Vertice heterodimer with unique configuration can provide extraordinary enhancement of the electric field for the single hot spot region due to the collaborative interaction of lightning rod effect and interparticle plasmon coupling effect. This established relationship between the architecture and the corresponding optical properties of the heterodimers provides the basis for creating controllable platforms which can be exploited in the applications of plasmonic devices, electronics, and biodetection. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Micrometer-sized TPM emulsion droplets with surface-mobile binding groups

    Science.gov (United States)

    van der Wel, Casper; van de Stolpe, Guido L.; Verweij, Ruben W.; Kraft, Daniela J.

    2018-03-01

    Colloids coated with lipid membranes have been widely employed for fundamental studies of lipid membrane processes, biotechnological applications such as drug delivery and biosensing, and more recently, for self-assembly. The latter has been made possible by inserting DNA oligomers with covalently linked hydrophobic anchors into the membrane. The lateral mobility of the DNA linkers on micrometer-sized droplets and solid particles has opened the door to creating structures with unprecedented structural flexibility. Here, we investigate micro-emulsions of TPM (3-(trimethoxysilyl)propyl methacrylate) as a platform for lipid monolayers and further functionalization with proteins and DNA oligonucleotides. TPM droplets can be produced with a narrow size distribution and are polymerizable, thus providing supports for model lipid membranes with controlled size and curvature. With fluorescence recovery after photobleaching, we observed that droplet-attached lipids, NeutrAvidin proteins, as well as DNA oligonucleotides all show mobility on the surface. We explored the assembly of micron-sized particles on TPM-droplets by exploiting either avidin-biotin interactions or double-stranded DNA with complementary single-stranded end groups. While the single molecules are mobile, the particles that are attached to them are not. We propose that this is caused by the heterogeneous nature of emulsified TPM, which forms an oligomer network that limits the collective motion of linkers, but allows the surface mobility of individual molecules.

  20. The Extracellular Protein Factor Epf from Streptococcus pyogenes Is a Cell Surface Adhesin That Binds to Cells through an N-terminal Domain Containing a Carbohydrate-binding Module*

    Science.gov (United States)

    Linke, Christian; Siemens, Nikolai; Oehmcke, Sonja; Radjainia, Mazdak; Law, Ruby H. P.; Whisstock, James C.; Baker, Edward N.; Kreikemeyer, Bernd

    2012-01-01

    Streptococcus pyogenes is an exclusively human pathogen. Streptococcal attachment to and entry into epithelial cells is a prerequisite for a successful infection of the human host and requires adhesins. Here, we demonstrate that the multidomain protein Epf from S. pyogenes serotype M49 is a streptococcal adhesin. An epf-deficient mutant showed significantly decreased adhesion to and internalization into human keratinocytes. Cell adhesion is mediated by the N-terminal domain of Epf (EpfN) and increased by the human plasma protein plasminogen. The crystal structure of EpfN, solved at 1.6 Å resolution, shows that it consists of two subdomains: a carbohydrate-binding module and a fibronectin type III domain. Both fold types commonly participate in ligand receptor and protein-protein interactions. EpfN is followed by 18 repeats of a domain classified as DUF1542 (domain of unknown function 1542) and a C-terminal cell wall sorting signal. The DUF1542 repeats are not involved in adhesion, but biophysical studies show they are predominantly α-helical and form a fiber-like stalk of tandem DUF1542 domains. Epf thus conforms with the widespread family of adhesins known as MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), in which a cell wall-attached stalk enables long range interactions via its adhesive N-terminal domain. PMID:22977243

  1. The extracellular protein factor Epf from Streptococcus pyogenes is a cell surface adhesin that binds to cells through an N-terminal domain containing a carbohydrate-binding module.

    Science.gov (United States)

    Linke, Christian; Siemens, Nikolai; Oehmcke, Sonja; Radjainia, Mazdak; Law, Ruby H P; Whisstock, James C; Baker, Edward N; Kreikemeyer, Bernd

    2012-11-02

    Streptococcus pyogenes is an exclusively human pathogen. Streptococcal attachment to and entry into epithelial cells is a prerequisite for a successful infection of the human host and requires adhesins. Here, we demonstrate that the multidomain protein Epf from S. pyogenes serotype M49 is a streptococcal adhesin. An epf-deficient mutant showed significantly decreased adhesion to and internalization into human keratinocytes. Cell adhesion is mediated by the N-terminal domain of Epf (EpfN) and increased by the human plasma protein plasminogen. The crystal structure of EpfN, solved at 1.6 Å resolution, shows that it consists of two subdomains: a carbohydrate-binding module and a fibronectin type III domain. Both fold types commonly participate in ligand receptor and protein-protein interactions. EpfN is followed by 18 repeats of a domain classified as DUF1542 (domain of unknown function 1542) and a C-terminal cell wall sorting signal. The DUF1542 repeats are not involved in adhesion, but biophysical studies show they are predominantly α-helical and form a fiber-like stalk of tandem DUF1542 domains. Epf thus conforms with the widespread family of adhesins known as MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), in which a cell wall-attached stalk enables long range interactions via its adhesive N-terminal domain.

  2. The Anti-sigma Factor RsiV Is a Bacterial Receptor for Lysozyme: Co-crystal Structure Determination and Demonstration That Binding of Lysozyme to RsiV Is Required for σV Activation.

    Directory of Open Access Journals (Sweden)

    Jessica L Hastie

    2016-09-01

    Full Text Available σ factors provide RNA polymerase with promoter specificity in bacteria. Some σ factors require activation in order to interact with RNA polymerase and transcribe target genes. The Extra-Cytoplasmic Function (ECF σ factor, σV, is encoded by several Gram-positive bacteria and is specifically activated by lysozyme. This activation requires the proteolytic destruction of the anti-σ factor RsiV via a process of regulated intramembrane proteolysis (RIP. In many cases proteases that cleave at site-1 are thought to directly sense a signal and initiate the RIP process. We previously suggested binding of lysozyme to RsiV initiated the proteolytic destruction of RsiV and activation of σV. Here we determined the X-ray crystal structure of the RsiV-lysozyme complex at 2.3 Å which revealed that RsiV and lysozyme make extensive contacts. We constructed RsiV mutants with altered abilities to bind lysozyme. We find that mutants that are unable to bind lysozyme block site-1 cleavage of RsiV and σV activation in response to lysozyme. Taken together these data demonstrate that RsiV is a receptor for lysozyme and binding of RsiV to lysozyme is required for σV activation. In addition, the co-structure revealed that RsiV binds to the lysozyme active site pocket. We provide evidence that in addition to acting as a sensor for the presence of lysozyme, RsiV also inhibits lysozyme activity. Thus we have demonstrated that RsiV is a protein with multiple functions. RsiV inhibits σV activity in the absence of lysozyme, RsiV binds lysozyme triggering σV activation and RsiV inhibits the enzymatic activity of lysozyme.

  3. Diverted Total Synthesis of Promysalin Analogs Demonstrates That an Iron-Binding Motif Is Responsible for Its Narrow-Spectrum Antibacterial Activity.

    Science.gov (United States)

    Steele, Andrew D; Keohane, Colleen E; Knouse, Kyle W; Rossiter, Sean E; Williams, Sierra J; Wuest, William M

    2016-05-11

    Promysalin is a species-specific Pseudomonad metabolite with unique bioactivity. To better understand the mode of action of this natural product, we synthesized 16 analogs utilizing diverted total synthesis (DTS). Our analog studies revealed that the bioactivity of promysalin is sensitive to changes within its hydrogen bond network whereby alteration has drastic biological consequences. The DTS library not only yielded three analogs that retained potency but also provided insights that resulted in the identification of a previously unknown ability of promysalin to bind iron. These findings coupled with previous observations hint at a complex multifaceted role of the natural product within the rhizosphere.

  4. Binding of Thrombin-Activated Platelets to a Fibrin Scaffold through ?IIb?3 Evokes Phosphatidylserine Exposure on Their Cell Surface

    OpenAIRE

    Brzoska, Tomasz; Suzuki, Yuko; Mogami, Hideo; Sano, Hideto; Urano, Tetsumei

    2013-01-01

    Recently, by employing intra-vital confocal microscopy, we demonstrated that platelets expose phosphatidylserine (PS) and fibrin accumulate only in the center of the thrombus but not in its periphery. To address the question how exposure of platelet anionic phospholipids is regulated within the thrombus, an in-vitro experiment using diluted platelet-rich plasma was employed, in which the fibrin network was formed in the presence of platelets, and PS exposure on the platelet surface was analyz...

  5. The meningococcal vaccine candidate neisserial surface protein A (NspA binds to factor H and enhances meningococcal resistance to complement.

    Directory of Open Access Journals (Sweden)

    Lisa A Lewis

    2010-07-01

    Full Text Available Complement forms an important arm of innate immunity against invasive meningococcal infections. Binding of the alternative complement pathway inhibitor factor H (fH to fH-binding protein (fHbp is one mechanism meningococci employ to limit complement activation on the bacterial surface. fHbp is a leading vaccine candidate against group B Neisseria meningitidis. Novel mechanisms that meningococci employ to bind fH could undermine the efficacy of fHbp-based vaccines. We observed that fHbp deletion mutants of some meningococcal strains showed residual fH binding suggesting the presence of a second receptor for fH. Ligand overlay immunoblotting using membrane fractions from one such strain showed that fH bound to a approximately 17 kD protein, identified by MALDI-TOF analysis as Neisserial surface protein A (NspA, a meningococcal vaccine candidate whose function has not been defined. Deleting nspA, in the background of fHbp deletion mutants, abrogated fH binding and mAbs against NspA blocked fH binding, confirming NspA as a fH binding molecule on intact bacteria. NspA expression levels vary among strains and expression correlated with the level of fH binding; over-expressing NspA enhanced fH binding to bacteria. Progressive truncation of the heptose (Hep I chain of lipooligosaccharide (LOS, or sialylation of lacto-N-neotetraose LOS both increased fH binding to NspA-expressing meningococci, while expression of capsule reduced fH binding to the strains tested. Similar to fHbp, binding of NspA to fH was human-specific and occurred through fH domains 6-7. Consistent with its ability to bind fH, deleting NspA increased C3 deposition and resulted in increased complement-dependent killing. Collectively, these data identify a key complement evasion mechanism with important implications for ongoing efforts to develop meningococcal vaccines that employ fHbp as one of its components.

  6. Vitamin A transport and the transmembrane pore in the cell-surface receptor for plasma retinol binding protein.

    Directory of Open Access Journals (Sweden)

    Ming Zhong

    Full Text Available Vitamin A and its derivatives (retinoids play diverse and crucial functions from embryogenesis to adulthood and are used as therapeutic agents in human medicine for eye and skin diseases, infections and cancer. Plasma retinol binding protein (RBP is the principal and specific vitamin A carrier in the blood and binds vitamin A at 1:1 ratio. STRA6 is the high-affinity membrane receptor for RBP and mediates cellular vitamin A uptake. STRA6 null mice have severely depleted vitamin A reserves for vision and consequently have vision loss, even under vitamin A sufficient conditions. STRA6 null humans have a wide range of severe pathological phenotypes in many organs including the eye, brain, heart and lung. Known membrane transport mechanisms involve transmembrane pores that regulate the transport of the substrate (e.g., the gating of ion channels. STRA6 represents a new type of membrane receptor. How this receptor interacts with its transport substrate vitamin A and the functions of its nine transmembrane domains are still completely unknown. These questions are critical to understanding the molecular basis of STRA6's activities and its regulation. We employ acute chemical modification to introduce chemical side chains to STRA6 in a site-specific manner. We found that modifications with specific chemicals at specific positions in or near the transmembrane domains of this receptor can almost completely suppress its vitamin A transport activity. These experiments provide the first evidence for the existence of a transmembrane pore, analogous to the pore of ion channels, for this new type of cell-surface receptor.

  7. Novel antibody binding determinants on the capsid surface of serotype O foot-and-mouth disease virus

    Science.gov (United States)

    Asfor, Amin S.; Upadhyaya, Sasmita; Knowles, Nick J.; King, Donald P.; Paton, David J.

    2014-01-01

    Five neutralizing antigenic sites have been described for serotype O foot-and-mouth disease viruses (FMDV) based on monoclonal antibody (mAb) escape mutant studies. However, a mutant virus selected to escape neutralization of mAb binding at all five sites was previously shown to confer complete cross-protection with the parental virus in guinea pig challenge studies, suggesting that amino acid residues outside the mAb binding sites contribute to antibody-mediated in vivo neutralization of FMDV. Comparison of the ability of bovine antisera to neutralize a panel of serotype O FMDV identified three novel putative sites at VP2-74, VP2-191 and VP3-85, where amino acid substitutions correlated with changes in sero-reactivity. The impact of these positions was tested using site-directed mutagenesis to effect substitutions at critical amino acid residues within an infectious copy of FMDV O1 Kaufbeuren (O1K). Recovered viruses containing additional mutations at VP2-74 and VP2-191 exhibited greater resistance to neutralization with both O1K guinea pig and O BFS bovine antisera than a virus that was engineered to include only mutations at the five known antigenic sites. The changes at VP2-74 and VP3-85 are adjacent to critical amino acids that define antigenic sites 2 and 4, respectively. However VP2-191 (17 Å away from VP2-72), located at the threefold axis and more distant from previously identified antigenic sites, exhibited the most profound effect. These findings extend our knowledge of the surface features of the FMDV capsid known to elicit neutralizing antibodies, and will improve our strategies for vaccine strain selection and rational vaccine design. PMID:24584474

  8. Surface binding and uptake of cadmium (Cd2+) by LLC-PK1 cells on permeable membrane supports

    International Nuclear Information System (INIS)

    Prozialeck, W.C.; Lamar, P.C.

    1993-01-01

    Recent studies have shown that Cd 2+ has relatively specific damaging effects on cell-cell junctions in the renal epithelial cell line, LLC-PK 1 . The objective of the present studies was to examine the surface binding and uptake of Cd 2+ by LLC-PK 1 cells in relation to the disruption of cell-cell junctions. LLC-PK 1 cells on Falcon Cell Culture Inserts were exposed to CdCl 2 containing trace amounts of 109 Cd 2+ from either the apical or the basolateral compartments, and the accumulation of 109 Cd 2+ was monitored for up to 8 h. The integrity of cell-cell junctions was assessed by monitoring the transepithelial electrical resistance. The results showed that the cells accumulated 3-4 times more Cd 2+ from the basolateral compartment than from the apical compartment. The accumulation of Cd 2+ from the basolateral compartment occurred in two phases: a rapid, exponential phase that occurred in 1-2 h and coincided with a decrease in transepithelial resistance, and a slower, linear phase that continued for 6-8 h. The Cd 2+ that accumulated during the rapid phase was easily removed by washing the cells in EGTA, indicating that most of it was bound to sites on the cell surface. By contrast, most of the Cd 2+ that accumulated during the slower phase could not be removed by EGTA, indicating that it had been taken up by the cells. Additional studies showed that the rapid phase of Cd 2+ accumulation was enhanced when Ca 2+ was present at low concentrations (0.1 mM), and was greatly reduced when Ca 2+ was present at high concentrations (10 mM). These results suggest that ld 2+ damages the junctions between LLC-PK 1 cells by interacting with Ca 2+ -sensitive sites on the basolateral cell surface. (orig.)

  9. An investigation of drug binding ability of a surface active ionic liquid: micellization, electrochemical, and spectroscopic studies.

    Science.gov (United States)

    Mahajan, Suruchi; Sharma, Rabia; Mahajan, Rakesh Kumar

    2012-12-18

    Keeping in view the use of surfactants in drug delivery, the interactions of surface active ionic liquids, such as 1-tetradecyl-3-methylimidazolium bromide (C(14)mimBr), with drugs, viz., dopamine hydrochloride (DH) and acetylcholine chloride (AC), have been studied, and the results are further compared with that of the structurally similar conventional cationic surfactant tetradecyltrimethylammonium bromide (TTAB). The micellization and interfacial behavior of C(14)mimBr and TTAB, in the presence of DH and AC, has been investigated from conductivity and surface tension measurements. Various micellar and adsorption characteristics for these drug-surfactant systems (DH/AC + C(14)mimBr/TTAB) have been investigated, indicating favorable interactions between them. The more detailed information regarding the nature of interactions between C(14)mimBr/TTAB and DH/AC is obtained from cyclic voltammetry (CV) and (1)H NMR measurements. CV measurements have been employed to evaluate the binding constant (K) and the Gibbs free energy change (ΔG) for these drug-surfactant complexes. These measurements indicate the existence of cation-π as well as π-π interactions between drugs and surfactants. A detailed analysis of chemical shifts of protons of drug molecules (DH and AC) in the presence of C(14)mimBr and TTAB has been done by (1)H NMR. The results obtained from (1)H NMR are in agreement with those of CV measurements. (1)H NMR studies along with the conductivity and surface tension measurements help in predicting the possible location of adsorption of these drug molecules in C(14)mimBr and TTAB micelles.

  10. Identification and Characterization of the UL37 Protein of Herpes Simplex Virus Type 1 and Demonstration that it Interacts with ICP8, the Major DNA Binding Protein of Herpes Simplex Virus

    Science.gov (United States)

    1992-10-20

    R . 1974 . Recovery of herpes simplex virus from human sacral gangl ions. N. Engl. J. Med. 291 :828-830. Baringer, J.R . 1975. Herpes simplex virus...AII’I fORCE MEDICAL C(NTEIt Title of Dissertation : "Ideatification and Characterization of the UL37 Protein of Herpes Simplex Virus Type 1 and...Demonstration that It Interacts with reps. the Major DNA Binding Protein of Herpes Simplex Virus" Name of Candidate: Lisa Shelton Doctor of

  11. Positively-charged semi-tunnel is a structural and surface characteristic of polyphosphate-binding proteins: an in-silico study.

    Directory of Open Access Journals (Sweden)

    Zheng Zachory Wei

    Full Text Available Phosphate is essential for all major life processes, especially energy metabolism and signal transduction. A linear phosphate polymer, polyphosphate (polyP, linked by high-energy phosphoanhydride bonds, can interact with various proteins, playing important roles as an energy source and regulatory factor. However, polyP-binding structures are largely unknown. Here we proposed a putative polyP binding site, a positively-charged semi-tunnel (PCST, identified by surface electrostatics analyses in polyP kinases (PPKs and many other polyP-related proteins. We found that the PCSTs in varied proteins were folded in different secondary structure compositions. Molecular docking calculations revealed a significant value for binding affinity to polyP in PCST-containing proteins. Utilizing the PCST identified in the β subunit of PPK3, we predicted the potential polyP-binding domain of PPK3. The discovery of this feature facilitates future searches for polyP-binding proteins and discovery of the mechanisms for polyP-binding activities. This should greatly enhance the understanding of the many physiological functions of protein-bound polyP and the involvement of polyP and polyP-binding proteins in various human diseases.

  12. Immunohistochemical study of Ulex europaeus agglutinin 1 (UEA-1) binding of megakaryocytes in bone marrow biopsy specimens: demonstration of heterogeneity in staining pattern reflecting the stages of differentiation.

    Science.gov (United States)

    Liu, S M; Li, C Y

    1996-01-01

    During differentiation, megakaryocytes undergo nuclear endoreplication, an increase in cell size, cytoplasmic granulation, and release of platelets. The changes in highly lobulated nuclei with varying degree of polyploidy and increasing cell size are easily recognized morphologically. However, the actual cytoplasmic changes are more difficult to perceive morphologically. With the peroxidase-antiperoxidase (PAP) method using UEA-1 as the binding protein to the alpha-L-fucose of glycoprotein synthesized by megakaryocytes, we observed significant variation in cytoplasmic staining of megakaryocytes in routinely processed bone marrow biopsy sections. A total of 3344 megakaryocytes in bone marrow sections from 10 patients with nonhematologic diseases and from 10 patients with idiopathic thrombocytopenic purpura (ITP) was studied. According to the intensity and pattern of cytoplasmic staining, we divided megakaryocytes into at least six groups: (1) low granular (LG), (2) diffuse granular (DG), (3) diffuse dense granular (DDG), (4) marginal granular (MG), (5) denuded (DMK), and (6) endomitotic (EndoM). Most of the megakaryocytes were DG (mean, 42.75% +/- 19.21%) and DDG (mean, 50.25% +/- 21.23%). In correlation with nuclear morphology and cell size, it appears that substances binding to UEA-1 are located in the paranuclear region in early megakaryocytes and produce a low granular focal staining pattern (LG cells). Next, the granules spread throughout the cytoplasm (DG cells) and increase in quantity (DDG). This is followed by migration of granules to the periphery of the cytoplasm (MG cells) and is associated with the liberation of platelets and eventual formation of DMK megakaryocytes. Endomitosis, regulated by unknown factors, occurred in the MG stage. In comparing the group with nonhematologic disease (mean DG, 35.4% +/- 18.48%; DDG, 58.4% +/- 21.8%) and the group with ITP (mean DG, 50.1% +/- 17.82%; DDG, 42.1% +/- 18.12%), we found an increasing proportion of DG

  13. Two Secondary Carbohydrate Binding Sites on the Surface of Barley alpha-Amylase 1 Have Distinct Functions and Display Synergy in Hydrolysis of Starch Granules

    DEFF Research Database (Denmark)

    Nielsen, Morten Munch; Bozonnet, Sophie; Seo, Eun-Seong

    2009-01-01

    Some polysaccharide processing enzymes possess secondary carbohydrate binding sites situated on the surface far from the active site. In barley alpha-amylase 1 (AMY1), two such sites, SBS1 and SBS2, are found on the catalytic (beta/alpha)8-barrel and the noncatalytic C-terminal domain, respective...

  14. Comparison of the binding properties of the mushroom Marasmius oreades lectin and Griffonia simplicifolia I-B isolectin to alphagalactosyl carbohydrate antigens in the surface phase

    DEFF Research Database (Denmark)

    Kirkeby, Svend; Winter, Harry C; Goldstein, Irwin J

    2004-01-01

    The binding of two alpha-galactophilic lectins, Marasmius oreades agglutinin (MOA), and Griffonia simplicifolia I isolectin B(4) (GS I-B(4)) to neoglycoproteins and natural glycoproteins were compared in a surface phase assay. Neoglycoproteins carrying various alpha-galactosylated glycans and lam...

  15. A new generation of starch products as excipient in pharmaceutical tablets .1. Preparation and binding properties of high surface area potato starch products

    NARCIS (Netherlands)

    Wierik, GHPT; ArendsScholte, AW; Eissens, AC; Lerk, CF

    1996-01-01

    A new pharmaceutical excipient with a high binding capacity was prepared from potato starch by enzymatic degradation, followed by suitable dehydration of the precipitated and filtered retrograded starch to produce high specific surface area products. Thermal dehydration methods like drying at room

  16. Borrelia burgdorferi outer surface protein C (OspC) binds complement component C4b and confers bloodstream survival.

    Science.gov (United States)

    Caine, Jennifer A; Lin, Yi-Pin; Kessler, Julie R; Sato, Hiromi; Leong, John M; Coburn, Jenifer

    2017-12-01

    Borrelia burgdorferi (Bb) is the causative agent of Lyme disease in the United States, a disease that can result in carditis, and chronic and debilitating arthritis and/or neurologic symptoms if left untreated. Bb survives in the midgut of the Ixodes scapularis tick, or within tissues of immunocompetent hosts. In the early stages of infection, the bacteria are present in the bloodstream where they must resist clearance by the innate immune system of the host. We have found a novel role for outer surface protein C (OspC) from B. burgdorferi and B. garinii in interactions with the complement component C4b and bloodstream survival in vivo. Our data show that OspC inhibits the classical and lectin complement pathways and competes with complement protein C2 for C4b binding. Resistance to complement is important for maintenance of the lifecycle of Bb, enabling survival of the pathogen within the host as well as in the midgut of a feeding tick when ospC expression is induced. © 2017 John Wiley & Sons Ltd.

  17. Independent associations of polymorphisms in vitamin D binding protein (GC) and vitamin D receptor (VDR) genes with obesity and plasma 25OHD3 levels demonstrate sex dimorphism.

    Science.gov (United States)

    Almesri, Norah; Das, Nagalla S; Ali, Muhallab E; Gumaa, Khalid; Giha, Hayder Ahmed

    2016-04-01

    We investigated a possible association between polymorphisms in vitamin D binding protein (GC) and vitamin D receptor (VDR) genes and obesity in Bahraini adults. For this purpose, 406 subjects with varying body mass indexes (BMIs) were selected. Plasma levels of 25-hydroxyvitamin D3 (25OHD3) were measured by chemiluminescence immunoassay. Six single nucleotide polymorphisms, 2 in the VDR gene (rs731236 TC and rs12721377 AG) and 4 in the GC gene (rs2282679 AC, rs4588 CA, rs7041 GT, and rs2298849 TC), were genotyped by real-time polymerase chain reaction. We found that the rs7041 minor allele (G) and rare genotype (GG) were associated with higher BMI (p = 0.007 and p = 0.012, respectively), but they did not influence 25OHD3 levels. However, the minor alleles of rs2282679 (A) and rs4588 (C) were associated with low 25OHD3 plasma levels (p = 0.039 and p = 0.021, respectively), but not with BMI. Having categorized the subjects based on their sex, we found that (i) rs7041 GG associated with high BMI in females (p = 0.003), (ii) rs4588 CC associated with high BMI in females (p = 0.034) and low 25OHD3 levels in males (p = 0.009), and (iii) rs12721377 AA associated with low 25OHD3 levels in females (p = 0.039). Notably, none of the common haplotypes (6 in the GC gene and 3 in the VDR gene) were associated with BMI. Therefore, polymorphisms in the GC (rs2282679, rs4588, rs7041) and VDR (rs12721377) genes were independently associated with obesity and 25OHD3 levels with a clear sex dimorphism.

  18. Further structural insights into the binding of complement factor H by complement regulator-acquiring surface protein 1 (CspA) of Borrelia burgdorferi

    International Nuclear Information System (INIS)

    Caesar, Joseph J. E.; Wallich, Reinhard; Kraiczy, Peter; Zipfel, Peter F.; Lea, Susan M.

    2013-01-01

    B. burgdorferi binds complement factor H using a dimeric surface protein, CspA (BbCRASP-1). Presented here is a new structure of CspA that suggests that there is a degree of flexibility between subunits which may have implications for complement regulator binding. Borrelia burgdorferi has evolved many mechanisms of evading the different immune systems across its range of reservoir hosts, including the capture and presentation of host complement regulators factor H and factor H-like protein-1 (FHL-1). Acquisition is mediated by a family of complement regulator-acquiring surface proteins (CRASPs), of which the atomic structure of CspA (BbCRASP-1) is known and shows the formation of a homodimeric species which is required for binding. Mutagenesis studies have mapped a putative factor H binding site to a cleft between the two subunits. Presented here is a new atomic structure of CspA which shows a degree of flexibility between the subunits which may be critical for factor H scavenging by increasing access to the binding interface and allows the possibility that the assembly can clamp around the bound complement regulators

  19. Mutations in the putative zinc-binding motif of UL52 demonstrate a complex interdependence between the UL5 and UL52 subunits of the human herpes simplex virus type 1 helicase/primase complex.

    Science.gov (United States)

    Chen, Yan; Carrington-Lawrence, Stacy D; Bai, Ping; Weller, Sandra K

    2005-07-01

    Herpes simplex virus type 1 (HSV-1) encodes a heterotrimeric helicase-primase (UL5/8/52) complex. UL5 contains seven motifs found in helicase superfamily 1, and UL52 contains conserved motifs found in primases. The contributions of each subunit to the biochemical activities of the complex, however, remain unclear. We have previously demonstrated that a mutation in the putative zinc finger at UL52 C terminus abrogates not only primase but also ATPase, helicase, and DNA-binding activities of a UL5/UL52 subcomplex, indicating a complex interdependence between the two subunits. To test this hypothesis and to further investigate the role of the zinc finger in the enzymatic activities of the helicase-primase, a series of mutations were constructed in this motif. They differed in their ability to complement a UL52 null virus: totally defective, partial complementation, and potentiating. In this study, four of these mutants were studied biochemically after expression and purification from insect cells infected with recombinant baculoviruses. All mutants show greatly reduced primase activity. Complementation-defective mutants exhibited severe defects in ATPase, helicase, and DNA-binding activities. Partially complementing mutants displayed intermediate levels of these activities, except that one showed a wild-type level of helicase activity. These data suggest that the UL52 zinc finger motif plays an important role in the activities of the helicase-primase complex. The observation that mutations in UL52 affected helicase, ATPase, and DNA-binding activities indicates that UL52 binding to DNA via the zinc finger may be necessary for loading UL5. Alternatively, UL5 and UL52 may share a DNA-binding interface.

  20. Residues essential for Panton-Valentine leukocidin S component binding to its cell receptor suggest both plasticity and adaptability in its interaction surface.

    Directory of Open Access Journals (Sweden)

    Benoit-Joseph Laventie

    Full Text Available Panton-Valentine leukocidin (PVL, a bicomponent staphylococcal leukotoxin, is involved in the poor prognosis of necrotizing pneumonia. The present study aimed to elucidate the binding mechanism of PVL and in particular its cell-binding domain. The class S component of PVL, LukS-PV, is known to ensure cell targeting and exhibits the highest affinity for the neutrophil membrane (Kd∼10(-10 M compared to the class F component of PVL, LukF-PV (Kd∼10(-9 M. Alanine scanning mutagenesis was used to identify the residues involved in LukS-PV binding to the neutrophil surface. Nineteen single alanine mutations were performed in the rim domain previously described as implicated in cell membrane interactions. Positions were chosen in order to replace polar or exposed charged residues and according to conservation between leukotoxin class S components. Characterization studies enabled to identify a cluster of residues essential for LukS-PV binding, localized on two loops of the rim domain. The mutations R73A, Y184A, T244A, H245A and Y250A led to dramatically reduced binding affinities for both human leukocytes and undifferentiated U937 cells expressing the C5a receptor. The three-dimensional structure of five of the mutants was determined using X-ray crystallography. Structure analysis identified residues Y184 and Y250 as crucial in providing structural flexibility in the receptor-binding domain of LukS-PV.

  1. Curved reconstructions versus three-dimensional surface rendering in the demonstration of cortical lesions in patients with extratemporal epilepsy

    NARCIS (Netherlands)

    Meiners, LC; Scheffers, JM; De Kort, GAP; Burger, H; Van Huffelen, AC; Van Rijen, PC; Van Veelen, CWM

    RATIONALE AND OBJECTIVES. TO compare the visibility and localization of extratemporal cortical lesions in extratemporal epilepsy by using curved reconstruction (CR) and three-dimensional surface rendering (3D SR) of 3D-acquired MR images and to study the degree of confidence with which localizations

  2. Ultrastructural demonstration of a glycoproteinic surface coat in allergenic pollen grains by combined cetylpyridinium chloride precipitation and silver proteinate staining.

    Science.gov (United States)

    Grote, M; Fromme, H G

    1984-01-01

    In allergenic birch pollen grains, highly watersoluble surface substances were precipitated by the cationic detergent cetylpyridinium chloride (CPC) during aqueous fixation. After processing the pollen for electron microscopy, ultrathin sections of pollen grains were subjected to the periodic acid - thiocarbohydrazide - silver proteinate (PA-TCH-SP) procedure according to Thiery (1967) for the detection of vicinal glycol groups. It was found that the material precipitated by CPC on the surface and within the exine cavities of the pollen wall strongly reacted with the PA-TCH-SP reagent thus indicating the presence of polysaccharides on the surface of birch pollen grains. In samples which had not been treated with the cationic detergent, PA-TCH-SP reactivity was reduced to thin linings on the surface and within the exine cavities. In both cases the exine proper did not stain whereas the intine showed moderate staining. Within the aperture region of the intine, PA-TCH-SP reactivity is preferably associated with fibrillar or reticular structures. The results are discussed with special reference to biochemical findings on allergenic birch pollen proteins.

  3. Identification of an estrogen receptor α non covalent ubiquitin-binding surface: role in 17β-estradiol-induced transcriptional activity.

    Science.gov (United States)

    Pesiri, Valeria; La Rosa, Piergiorgio; Stano, Pasquale; Acconcia, Filippo

    2013-06-15

    Ubiquitin (Ub)-binding domains (UBDs) located in Ub receptors decode the ubiquitination signal by non-covalently engaging the Ub modification on their binding partners and transduce the Ub signalling through Ub-based molecular interactions. In this way, inducible protein ubiquitination regulates diverse biological processes. The estrogen receptor alpha (ERα) is a ligand-activated transcription factor that mediates the pleiotropic effects of the sex hormone 17β-estradiol (E2). Fine regulation of E2 pleiotropic actions depends on E2-dependent ERα association with a plethora of binding partners and/or on the E2 modulation of receptor ubiquitination. Indeed, E2-induced ERα polyubiquitination triggers receptor degradation and transcriptional activity, and E2-dependent reduction in ERα monoubiquitination is crucial for E2 signalling. Monoubiquitinated proteins often contain UBDs, but whether non-covalent Ub-ERα binding could occur and play a role in E2-ERα signalling is unknown. Here, we report an Ub-binding surface within the ERα ligand binding domain that directs in vitro the receptor interaction with both ubiquitinated proteins and recombinant Ub chains. Mutational analysis reveals that ERα residues leucine 429 and alanine 430 are involved in Ub binding. Moreover, impairment of ERα association to ubiquitinated species strongly affects E2-induced ERα transcriptional activity. Considering the importance of UBDs in the Ub-based signalling network and the central role of different ERα binding partners in the modulation of E2-dependent effects, our discoveries provide novel insights into ERα activity that could also be relevant for ERα-dependent diseases.

  4. Surface-enhanced Raman Scattering Study of the Binding Modes of a Dibenzotetraaza[14]annulene Derivative with DNA/RNA Polynucleotides

    OpenAIRE

    Miljanić, Snežana; Dijanošić, Adriana; Kalac, Matea; Radić Stojković, Marijana; Piantanida, Ivo; Pawlica, Dariusz; Eilmes, Julita

    2012-01-01

    Binding modes of a dibenzotetraaza14annulene (DBTAA) derivative with synthetic nucleic acids were studied using surface-enhanced Raman spectroscopy (SERS). Changes in SERS intensity and appearance of new bands in spectra were attributed to different complexes formed between the DBTAA molecules and DNA/RNA polynucleotides. A decrease in intensity pointed to intercalation as the dominant binding mode of the annulene derivative with poly dGdC-poly dGdC and poly rA-poly rU, whereas new bands in...

  5. Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

    KAUST Repository

    Arenas, Jesú s; Nijland, Reindert; Rodriguez, Francisco J.; Bosma, Tom N. P.; Tommassen, Jan

    2012-01-01

    Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

  6. Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

    KAUST Repository

    Arenas, Jesús

    2012-12-04

    Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

  7. Analysis of gold(I/III)-complexes by HPLC-ICP-MS demonstrates gold(III) stability in surface waters.

    Science.gov (United States)

    Ta, Christine; Reith, Frank; Brugger, Joël; Pring, Allan; Lenehan, Claire E

    2014-05-20

    Understanding the form in which gold is transported in surface- and groundwaters underpins our understanding of gold dispersion and (bio)geochemical cycling. Yet, to date, there are no direct techniques capable of identifying the oxidation state and complexation of gold in natural waters. We present a reversed phase ion-pairing HPLC-ICP-MS method for the separation and determination of aqueous gold(III)-chloro-hydroxyl, gold(III)-bromo-hydroxyl, gold(I)-thiosulfate, and gold(I)-cyanide complexes. Detection limits for the gold species range from 0.05 to 0.30 μg L(-1). The [Au(CN)2](-) gold cyanide complex was detected in five of six waters from tailings and adjacent monitoring bores of working gold mines. Contrary to thermodynamic predictions, evidence was obtained for the existence of Au(III)-complexes in circumneutral, hypersaline waters of a natural lake overlying a gold deposit in Western Australia. This first direct evidence for the existence and stability of Au(III)-complexes in natural surface waters suggests that Au(III)-complexes may be important for the transport and biogeochemical cycling of gold in surface environments. Overall, these results show that near-μg L(-1) enrichments of Au in environmental waters result from metastable ligands (e.g., CN(-)) as well as kinetically controlled redox processes leading to the stability of highly soluble Au(III)-complexes.

  8. Importance of length and sequence order on magnesium binding to surface-bound oligonucleotides studied by second harmonic generation and atomic force microscopy.

    Science.gov (United States)

    Holland, Joseph G; Geiger, Franz M

    2012-06-07

    The binding of magnesium ions to surface-bound single-stranded oligonucleotides was studied under aqueous conditions using second harmonic generation (SHG) and atomic force microscopy (AFM). The effect of strand length on the number of Mg(II) ions bound and their free binding energy was examined for 5-, 10-, 15-, and 20-mers of adenine and guanine at pH 7, 298 K, and 10 mM NaCl. The binding free energies for adenine and guanine sequences were calculated to be -32.1(4) and -35.6(2) kJ/mol, respectively, and invariant with strand length. Furthermore, the ion density for adenine oligonucleotides did not change as strand length increased, with an average value of 2(1) ions/strand. In sharp contrast, guanine oligonucleotides displayed a linear relationship between strand length and ion density, suggesting that cooperativity is important. This data gives predictive capabilities for mixed strands of various lengths, which we exploit for 20-mers of adenines and guanines. In addition, the role sequence order plays in strands of hetero-oligonucleotides was examined for 5'-A(10)G(10)-3', 5'-(AG)(10)-3', and 5'-G(10)A(10)-3' (here the -3' end is chemically modified to bind to the surface). Although the free energy of binding is the same for these three strands (averaged to be -33.3(4) kJ/mol), the total ion density increases when several guanine residues are close to the 3' end (and thus close to the solid support substrate). To further understand these results, we analyzed the height profiles of the functionalized surfaces with tapping-mode atomic force microscopy (AFM). When comparing the average surface height profiles of the oligonucleotide surfaces pre- and post- Mg(II) binding, a positive correlation was found between ion density and the subsequent height decrease following Mg(II) binding, which we attribute to reductions in Coulomb repulsion and strand collapse once a critical number of Mg(II) ions are bound to the strand.

  9. European demonstration program on the effect-based and chemical identification and monitoring of organic pollutants in European surface waters

    NARCIS (Netherlands)

    Tousova, Zuzana; Oswald, Peter; Slobodnik, Jaroslav; Blaha, Ludek; Muz, Melis; Hu, Meng; Brack, Werner; Krauss, Martin; Di Paolo, Carolina; Tarcai, Zsolt; Seiler, Thomas Benjamin; Hollert, Henner; Koprivica, Sanja; Ahel, Marijan; Schollée, Jennifer E.; Hollender, Juliane; Suter, Marc J.F.; Hidasi, Anita O.; Schirmer, Kristin; Sonavane, Manoj; Ait-Aissa, Selim; Creusot, Nicolas; Brion, Francois; Froment, Jean; Almeida, Ana Catarina; Thomas, Kevin; Tollefsen, Knut Erik; Tufi, Sara; Ouyang, Xiyu; Leonards, Pim; Lamoree, Marja; Torrens, Victoria Osorio; Kolkman, Annemieke; Schriks, Merijn; Spirhanzlova, Petra; Tindall, Andrew; Schulze, Tobias

    2017-01-01

    Growing concern about the adverse environmental and human health effects of a wide range of micropollutants requires the development of novel tools and approaches to enable holistic monitoring of their occurrence, fate and effects in the aquatic environment. A European-wide demonstration program

  10. Interrogating the Plasmodium Sporozoite Surface: Identification of Surface-Exposed Proteins and Demonstration of Glycosylation on CSP and TRAP by Mass Spectrometry-Based Proteomics.

    Directory of Open Access Journals (Sweden)

    Kristian E Swearingen

    2016-04-01

    Full Text Available Malaria parasite infection is initiated by the mosquito-transmitted sporozoite stage, a highly motile invasive cell that targets hepatocytes in the liver for infection. A promising approach to developing a malaria vaccine is the use of proteins located on the sporozoite surface as antigens to elicit humoral immune responses that prevent the establishment of infection. Very little of the P. falciparum genome has been considered as potential vaccine targets, and candidate vaccines have been almost exclusively based on single antigens, generating the need for novel target identification. The most advanced malaria vaccine to date, RTS,S, a subunit vaccine consisting of a portion of the major surface protein circumsporozoite protein (CSP, conferred limited protection in Phase III trials, falling short of community-established vaccine efficacy goals. In striking contrast to the limited protection seen in current vaccine trials, sterilizing immunity can be achieved by immunization with radiation-attenuated sporozoites, suggesting that more potent protection may be achievable with a multivalent protein vaccine. Here, we provide the most comprehensive analysis to date of proteins located on the surface of or secreted by Plasmodium falciparum salivary gland sporozoites. We used chemical labeling to isolate surface-exposed proteins on sporozoites and identified these proteins by mass spectrometry. We validated several of these targets and also provide evidence that components of the inner membrane complex are in fact surface-exposed and accessible to antibodies in live sporozoites. Finally, our mass spectrometry data provide the first direct evidence that the Plasmodium surface proteins CSP and TRAP are glycosylated in sporozoites, a finding that could impact the selection of vaccine antigens.

  11. Real-time and label-free analysis of binding thermodynamics of carbohydrate-protein interactions on unfixed cancer cell surfaces using a QCM biosensor

    Science.gov (United States)

    Li, Xueming; Song, Siyu; Shuai, Qi; Pei, Yihan; Aastrup, Teodor; Pei, Yuxin; Pei, Zhichao

    2015-01-01

    A novel approach to the study of binding thermodynamics and kinetics of carbohydrate-protein interactions on unfixed cancer cell surfaces using a quartz crystal microbalance (QCM) biosensor was developed, in which binding events take place at the cell surface, more closely mimicking a biologically relevant environment. In this study, colon adenocarcinoma cells (KM-12) and ovary adenocarcinoma cells (SKOV-3) grew on the optimized polystyrene-coated biosensor chip without fixation. The association and dissociation between the cell surface carbohydrates and a range of lectins, including WGA, Con A, UEA-I, GS-II, PNA and SBA, were monitored in real time and without label for evaluation of cell surface glycosylation. Furthermore, the thermodynamic and kinetic parameters of the interaction between lectins and cell surface glycan were studied, providing detailed information about the interactions, such as the association rate constant, dissociation rate constant, affinity constant, as well as the changes of entropy, enthalpy and Gibbs free energy. This application provides an insight into the cell surface glycosylation and the complex molecular recognition on the intact cell surface, which may have impacts on disease diagnosis and drug discovery. PMID:26369583

  12. Cosmogenic 3He in hematite and goethite from Brazilian "canga" duricrust demonstrates the extreme stability of these surfaces

    Science.gov (United States)

    Shuster, David L.; Farley, Kenneth A.; Vasconcelos, Paulo M.; Balco, Greg; Monteiro, Hevelyn S.; Waltenberg, Kathryn; Stone, John O.

    2012-05-01

    Helium isotopes were measured in hematite and goethite samples from several lateritiric duricrusts (canga) developed on banded iron formations. These samples uniformly have high 3He concentrations which must arise from long periods of cosmic ray exposure. From coexisting phases from the Quadrilátero Ferrífero in east central Brazil, we determined the ratio of cosmogenic 3He in hematite to that of 21Ne in quartz to be 3.96 ± 0.19. Combined with best current estimates of the 21Ne production rate in quartz, this ratio implies a sea-level high latitude (SLHL) 3He production rate in hematite of 68.1 ± 8.1 atoms/g/yr; from the chemical composition we estimate the 3He production rate in goethite to be ~ 5% higher. We use these production rate estimates to interpret 3He concentrations measured in goethite and hematite from a ~ 10 m depth profile collected from a surface canga in Carajás, in the Amazon basin of Brazil. We find that the Carajás canga has experienced a very low rate of surface erosion (~ 0.16-0.54 m/Myr) over at least the last few millions of years. This iron-rich canga surface is remarkably resistant to erosion despite its location in a wet tropical environment. Details of the depth profile suggest that despite its stability, the canga has also been internally dynamic (translocation of material; solution and reprecipitation) over million-year timescales.

  13. Production of Fibronectin Binding Protein A at the surface of Lactococcus lactis increases plasmid transfer in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Daniela Pontes

    Full Text Available Lactococci are noninvasive lactic acid bacteria frequently used as protein delivery vectors and, more recently, as DNA delivery vehicles. We previously showed that Lactococcus lactis (LL expressing the Fibronectin-Binding Protein A of Staphylococcus aureus (LL-FnBPA+ showed higher internalization rates in vitro in Caco-2 cells than the native (wt lactococci and were able to deliver a eukaryotic Green Fluorescent Protein (GFP expression plasmid in 1% of human Caco-2 cells. Here, using the bovine beta-lactoglobulin (BLG, one of the major cow's milk allergen, and GFP we characterized the potential of LL-FnBPA+ as an in vivo DNA vaccine delivery vehicle. We first showed that the invasive strain LL-FnBPA+ carrying the plasmid pValac:BLG (LL-FnBPA+ BLG was more invasive than LL-BLG and showed the same invasivity as LL-FnBPA+. Then we demonstrated that the Caco-2 cells, co-incubated with LL-FnBPA+ BLG produced up to 30 times more BLG than the Caco-2 cells co-incubated with the non invasive LL-BLG. Using two different gene reporters, BLG and GFP, and two different methods of detection, EIA and fluorescence microscopy, we showed in vivo that: i in order to be effective, LL-FnBPA+ required a pre-coating with Fetal Calf Serum before oral administration; ii plasmid transfer occurred in enterocytes without regard to the strains used (invasive or not; iii the use of LL-FnBPA+ increased the number of mice producing BLG, but not the level of BLG produced. We thus confirmed the good potential of invasive recombinant lactic acid bacteria as DNA delivery vector in vivo.

  14. Structure of a Glomulin-RBX1-CUL1 complex: inhibition of a RING E3 ligase through masking of its E2-binding surface

    Science.gov (United States)

    Duda, David M.; Olszewski, Jennifer L.; Tron, Adriana E.; Hammel, Michal; Lambert, Lester J.; Waddell, M. Brett; Mittag, Tanja; DeCaprio, James A.; Schulman, Brenda A.

    2012-01-01

    Summary The ~300 human Cullin-RING ligases (CRLs) are multisubunit E3s in which a RING protein, either RBX1 or RBX2, recruits an E2 to catalyze ubiquitination. RBX1-containing CRLs also can bind Glomulin (GLMN), which binds RBX1’s RING domain, regulates the RBX1-CUL1-containing SCFFBW7 complex, and is disrupted in the disease Glomuvenous Malformation. Here we report the crystal structure of a complex between GLMN, RBX1, and a fragment of CUL1. Structural and biochemical analyses reveal that GLMN adopts a HEAT-like repeat fold that tightly binds the E2-interacting surface of RBX1, inhibiting CRL-mediated chain formation by the E2 CDC34. The structure explains the basis for GLMN’s selectivity toward RBX1 over RBX2, and how disease-associated mutations disrupt GLMN-RBX1 interactions. Our study reveals a mechanism for RING E3 ligase regulation whereby an inhibitor blocks E2 access, and raises the possibility that other E3s are likewise controlled by cellular proteins that mask E2-binding surfaces to mediate inhibition. PMID:22748924

  15. A novel RNA binding surface of the TAM domain of TIP5/BAZ2A mediates epigenetic regulation of rRNA genes.

    Science.gov (United States)

    Anosova, Irina; Melnik, Svitlana; Tripsianes, Konstantinos; Kateb, Fatiha; Grummt, Ingrid; Sattler, Michael

    2015-05-26

    The chromatin remodeling complex NoRC, comprising the subunits SNF2h and TIP5/BAZ2A, mediates heterochromatin formation at major clusters of repetitive elements, including rRNA genes, centromeres and telomeres. Association with chromatin requires the interaction of the TAM (TIP5/ARBP/MBD) domain of TIP5 with noncoding RNA, which targets NoRC to specific genomic loci. Here, we show that the NMR structure of the TAM domain of TIP5 resembles the fold of the MBD domain, found in methyl-CpG binding proteins. However, the TAM domain exhibits an extended MBD fold with unique C-terminal extensions that constitute a novel surface for RNA binding. Mutation of critical amino acids within this surface abolishes RNA binding in vitro and in vivo. Our results explain the distinct binding specificities of TAM and MBD domains to RNA and methylated DNA, respectively, and reveal structural features for the interaction of NoRC with non-coding RNA. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. Detection of ligand binding hot spots on protein surfaces via fragment-based methods: application to DJ-1 and glucocerebrosidase

    Energy Technology Data Exchange (ETDEWEB)

    Landon, Melissa R.; Lieberman, Raquel L.; Hoang, Quyen Q.; Ju, Shulin; Caaveiro, Jose M.M.; Orwig, Susan D.; Kozakov, Dima; Brenke, Ryan; Chuang, Gwo-Yu; Beglov, Dmitry; Vajda, Sandor; Petsko, Gregory A.; Ringe, Dagmar; (BU-M); (Brandeis); (GIT)

    2010-08-04

    The identification of hot spots, i.e., binding regions that contribute substantially to the free energy of ligand binding, is a critical step for structure-based drug design. Here we present the application of two fragment-based methods to the detection of hot spots for DJ-1 and glucocerebrosidase (GCase), targets for the development of therapeutics for Parkinson's and Gaucher's diseases, respectively. While the structures of these two proteins are known, binding information is lacking. In this study we employ the experimental multiple solvent crystal structures (MSCS) method and computational fragment mapping (FTMap) to identify regions suitable for the development of pharmacological chaperones for DJ-1 and GCase. Comparison of data derived via MSCS and FTMap also shows that FTMap, a computational method for the identification of fragment binding hot spots, is an accurate and robust alternative to the performance of expensive and difficult crystallographic experiments.

  17. Implications of surface charge and curvature for the binding orientation of Thermomyces lanuginosus lipase on negatively charged or zwitterionic phospholipid vesicles as studied by ESR spectroscopy

    DEFF Research Database (Denmark)

    Hedin, E.M.K.; Høyrup, Lise Pernille Kristine; Patkar, S.A.

    2005-01-01

    fluorescence quenching efficiency between each spin-label positioned on TLL, and the lipid membrane. ESR exposure and fluorescence quenching data show that TILL associates closer to the negatively charged PG surface than the zwitterionic PC surface, and binds to both POPG LUV and POPC SUV predominantly through......The triglyceride lipase (EC 3.1.1.3) Thermomyces lanuginosus lipase (TLL) binds with high affinity to unilamellar phospholipid vesicles that serve as a diluent interface for both lipase and substrate, but it displays interfacial activation on only small and negatively charged such vesicles [Cajal......) spectroscopy in combination with site-directed spin-labeling [Hedin, E. M. K., et al. (2002) Biochemistry 41, 1418514196]. In our investigation, we have studied the interfacial orientation of TLL when bound to large unilamellar vesicles (LUV) consisting of POPG, and bound to SUV consisting of 1-palmitoyl-2...

  18. Insights into the "pair of sugar tongs" surface binding site in barley alpha-amylase isozymes and crystallization of appropriate sugar tongs mutants

    DEFF Research Database (Denmark)

    Tranier, S.; Deville, K.; Robert, X.

    2005-01-01

    of an additional surface binding site called a "pair of sugar tongs" due to the sugar capturing by Tyr380 which is situated in domain C of AMYL For the first time, a biological role for the domain C was suggested as well as a hypothetical explanation of enzymatic differences between the two barley a......-amylase isozymes. However, no sugar was bound at the "sugar tongs" site in the AMY2/acarbose complex. Comparative studies of this domain on the basis of sequence, secondary structure and spatial organization allow to propose factors needed for such a site. One of the most obvious is the replacement of Ser378(AMY1......, surface plasmon resonance sugar binding experiments have proven unambiguously that this residue cannot totally explain the lack of the "pair of sugar tongs" and other tracks must be studied as, for example, the differences in orientation of Asp381 and the critical role of His395, both good candidates...

  19. Human small cell lung cancer NYH cells selected for resistance to the bisdioxopiperazine topoisomerase II catalytic inhibitor ICRF-187 demonstrate a functional R162Q mutation in the Walker A consensus ATP binding domain of the alpha isoform

    DEFF Research Database (Denmark)

    Wessel, I; Jensen, L H; Jensen, P B

    1999-01-01

    in expression of the beta isoform. Sequencing of the entire topoisomerase IIalpha cDNA from NYH/187 cells demonstrated a homozygous G-->A point mutation at nucleotide 485, leading to a R162Q conversion in the Walker A consensus ATP binding site (residues 161-165 in the alpha isoform), this being the first drug......-selected mutation described at this site. Western blotting after incubation with ICRF-187 showed no depletion of the alpha isoform in NYH/187 cells in contrast to wild-type (wt) cells, whereas equal depletion of the beta isoform was observed in the two sublines. Alkaline elution assay demonstrated a lack...... of inhibition of etoposide-induced DNA single-stranded breaks in NYH/187 cells, whereas this inhibition was readily apparent in NYH cells. Site-directed mutagenesis in human topoisomerase IIalpha introduced into a yeast Saccharomyces cerevisiae strain with a temperature-conditional yeast TOP2 mutant...

  20. TetR-dependent gene regulation in intracellular Listeria monocytogenes demonstrates the spatiotemporal surface distribution of ActA.

    Science.gov (United States)

    Schmitter, Sibylle; Fieseler, Lars; Klumpp, Jochen; Bertram, Ralph; Loessner, Martin J

    2017-08-01

    To enable specific and tightly controlled gene expression both in vitro and during the intracellular lifecycle of the pathogen Listeria monocytogenes, a TetR-dependent genetic induction system was developed. Highest concentration of cytoplasmic TetR and best repression of tetO-controlled genes was obtained by tetR expression from the synthetic promoter Pt 17 . Anhydrotetracycline (ATc) as inducer permitted concentration-dependent, fine-tuned expression of genes under control of the tetO operator and a suitable promoter. The actin-polymerizing ActA protein represents a major virulence factor of L. monocytogenes, required for actin-based motility and cell-to-cell spread in infected host cells. To be able to observe its spatial and temporal distribution on intracellular L. monocytogenes cells, conditional mutants featuring actA placed under TetR control were used to infect PtK2 epithelial cells. Following induction at different time intervals, the subsequent recruitment of actin by L. monocytogenes could be monitored. We found that cells displayed functional ActA after approximately 15 min, while formation of polarized actin tail was complete after 90-120 min. At this point, intracellular motility of the induced mutants was indistinguishable from wild-type bacteria. Interestingly, de novo ActA synthesis in intracellular Listeria also demonstrated the temporal, asymmetric redistribution of the membrane-anchored proteins from the lateral walls toward the cell poles. © 2017 John Wiley & Sons Ltd.

  1. In vivo stem cell tracking with imageable nanoparticles that bind bioorthogonal chemical receptors on the stem cell surface.

    Science.gov (United States)

    Lee, Sangmin; Yoon, Hwa In; Na, Jin Hee; Jeon, Sangmin; Lim, Seungho; Koo, Heebeom; Han, Sang-Soo; Kang, Sun-Woong; Park, Soon-Jung; Moon, Sung-Hwan; Park, Jae Hyung; Cho, Yong Woo; Kim, Byung-Soo; Kim, Sang Kyoon; Lee, Taekwan; Kim, Dongkyu; Lee, Seulki; Pomper, Martin G; Kwon, Ick Chan; Kim, Kwangmeyung

    2017-09-01

    It is urgently necessary to develop reliable non-invasive stem cell imaging technology for tracking the in vivo fate of transplanted stem cells in living subjects. Herein, we developed a simple and well controlled stem cell imaging method through a combination of metabolic glycoengineering and bioorthogonal copper-free click chemistry. Firstly, the exogenous chemical receptors containing azide (-N 3 ) groups were generated on the surfaces of stem cells through metabolic glycoengineering using metabolic precursor, tetra-acetylated N-azidoacetyl-d-mannosamine(Ac 4 ManNAz). Next, bicyclo[6.1.0]nonyne-modified glycol chitosan nanoparticles (BCN-CNPs) were prepared as imageable nanoparticles to deliver different imaging agents. Cy5.5, iron oxide nanoparticles and gold nanoparticles were conjugated or encapsulated to BCN-CNPs for optical, MR and CT imaging, respectively. These imageable nanoparticles bound chemical receptors on the Ac 4 ManNAz-treated stem cell surface specifically via bioorthogonal copper-free click chemistry. Then they were rapidly taken up by the cell membrane turn-over mechanism resulting in higher endocytic capacity compared non-specific uptake of nanoparticles. During in vivo animal test, BCN-CNP-Cy5.5-labeled stem cells could be continuously tracked by non-invasive optical imaging over 15 days. Furthermore, BCN-CNP-IRON- and BCN-CNP-GOLD-labeled stem cells could be efficiently visualized using in vivo MR and CT imaging demonstrating utility of our stem cell labeling method using chemical receptors. These results conclude that our method based on metabolic glycoengineering and bioorthogonal copper-free click chemistry can stably label stem cells with diverse imageable nanoparticles representing great potential as new stem cell imaging technology. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Structure and calcium binding activity of LipL32, the major surface antigen of pathogenic Leptospira sp

    International Nuclear Information System (INIS)

    Hauk, Pricila; Roman-Ramos, Henrique; Ho, Paulo Lee; Guzzo, Cristiane R.; Farah, Chuck S.

    2009-01-01

    Leptospirosis, caused by the spirochaete Leptospira is an important emerging infectious disease. LipL32 is the major exposed outer membrane protein found exclusively in pathogenic leptospira. It is highly immunogenic and has been shown to bind to host extracellular matrix components, including collagens, fibronectin and laminin. In this work we crystallized recombinant LipL32 protein and determined its structure to 2.25 A resolution. Initial phases were determined using the multi-wavelength anomalous dispersion technique with data collected from selenomethionine-containing crystals at the MX2 beamline at the LNLS. The LipL32 monomer is made of a jelly-roll fold core from which protrude several peripheral secondary structures. Some structural features suggested that LipL32 could bind Ca 2+ ions and indeed, spectroscopic data (circular (dichroism. intrinsic tryptophan fluorescence and extrinsic 1-amino-2-anaphthol-4-sulfonic acid fluorescence) confirmed the calcium binding properties of LipL32. (author)

  3. Reconstructing satellite images to quantify spatially explicit land surface change caused by fires and succession: A demonstration in the Yukon River Basin of interior Alaska

    Science.gov (United States)

    Huang, Shengli; Jin, Suming; Dahal, Devendra; Chen, Xuexia; Young, Claudia; Liu, Heping; Liu, Shuguang

    2013-01-01

    Land surface change caused by fires and succession is confounded by many site-specific factors and requires further study. The objective of this study was to reveal the spatially explicit land surface change by minimizing the confounding factors of weather variability, seasonal offset, topography, land cover, and drainage. In a pilot study of the Yukon River Basin of interior Alaska, we retrieved Normalized Difference Vegetation Index (NDVI), albedo, and land surface temperature (LST) from a postfire Landsat image acquired on August 5th, 2004. With a Landsat reference image acquired on June 26th, 1986, we reconstructed NDVI, albedo, and LST of 1987–2004 fire scars for August 5th, 2004, assuming that these fires had not occurred. The difference between actual postfire and assuming-no-fire scenarios depicted the fires and succession impact. Our results demonstrated the following: (1) NDVI showed an immediate decrease after burning but gradually recovered to prefire levels in the following years, in which burn severity might play an important role during this process; (2) Albedo showed an immediate decrease after burning but then recovered and became higher than prefire levels; and (3) Most fires caused surface warming, but cooler surfaces did exist; time-since-fire affected the prefire and postfire LST difference but no absolute trend could be found. Our approach provided spatially explicit land surface change rather than average condition, enabling a better understanding of fires and succession impact on ecological consequences at the pixel level.

  4. Comparison of gas-solid chromatography and MM2 force field molecular binding energies for greenhouse gases on a carbonaceous surface.

    Science.gov (United States)

    Rybolt, Thomas R; Bivona, Kevin T; Thomas, Howard E; O'Dell, Casey M

    2009-10-01

    Gas-solid chromatography was used to determine B(2s) (gas-solid virial coefficient) values for eight molecular adsorbates interacting with a carbon powder (Carbopack B, Supelco). B(2s) values were determined by multiple size variant injections within the temperature range of 313-553 K. The molecular adsorbates included: carbon dioxide (CO(2)); tetrafluoromethane (CF(4)); hexafluoroethane (C(2)F(6)); 1,1-difluoroethane (C(2)H(4)F(2)); 1-chloro-1,1-difluoroethane (C(2)H(3)ClF(2)); dichlorodifluoromethane (CCl(2)F(2)); trichlorofluoromethane (CCl(3)F); and 1,1,1-trichloroethane (C(2)H(3)Cl(3)). Two of these molecules are of special interest because they are "super greenhouse gases". The global warming potential, GWP, for CF(4) is 6500 and for C(2)F(6) is 9200 relative to the reference value of 1 for CO(2). The GWP index considers both radiative blocking and molecular lifetime. For these and other industrial greenhouse gases, adsorptive trapping on a carbonaceous solid, which depends on molecule-surface binding energy, could avoid atmospheric release. The temperature variations of the gas-solid virial coefficients in conjunction with van't Hoff plots were used to find the experimental adsorption energy or binding energy values (E(*)) for each adsorbate. A molecular mechanics based, rough-surface model was used to calculate the molecule-surface binding energy (Ecal(*)) using augmented MM2 parameters. The surface model consisted of parallel graphene layers with two separated nanostructures each containing 17 benzene rings arranged in linear strips. The separation of the parallel nanostructures had been optimized in a prior study to appropriately represent molecule-surface interactions for Carbopack B. Linear regressions of E(*) versus Ecal(*) for the current data set of eight molecules and the same surface model gave E(*)=0.926 Ecal(*) and r(2)=0.956. A combined set of the current and prior Carbopack B adsorbates studied (linear alkanes, branched alkanes, cyclic alkanes

  5. Two secondary carbohydrate binding sites on the surface of barley alpha-amylase 1 have distinct functions and display synergy in hydrolysis of starch granules.

    Science.gov (United States)

    Nielsen, Morten M; Bozonnet, Sophie; Seo, Eun-Seong; Mótyán, János A; Andersen, Joakim M; Dilokpimol, Adiphol; Abou Hachem, Maher; Gyémánt, Gyöngyi; Naested, Henrik; Kandra, Lili; Sigurskjold, Bent W; Svensson, Birte

    2009-08-18

    Some polysaccharide processing enzymes possess secondary carbohydrate binding sites situated on the surface far from the active site. In barley alpha-amylase 1 (AMY1), two such sites, SBS1 and SBS2, are found on the catalytic (beta/alpha)(8)-barrel and the noncatalytic C-terminal domain, respectively. Site-directed mutagenesis of Trp(278) and Trp(279), stacking onto adjacent ligand glucosyl residues at SBS1, and of Tyr(380) and His(395), making numerous ligand contacts at SBS2, suggested that SBS1 and SBS2 act synergistically in degradation of starch granules. While SBS1 makes the major contribution to binding and hydrolysis of starch granules, SBS2 exhibits a higher affinity for the starch mimic beta-cyclodextrin. Compared to that of wild-type AMY1, the K(d) of starch granule binding by the SBS1 W278A, W279A, and W278A/W279A mutants thus increased 15-35 times; furthermore, the k(cat)/K(m) of W278A/W279A was 2%, whereas both affinity and activity for Y380A at SBS2 were 10% of the wild-type values. Dual site double and triple SBS1/SBS2 substitutions eliminated binding to starch granules, and the k(cat)/K(m) of W278A/W279A/Y380A AMY1 was only 0.4% of the wild-type value. Surface plasmon resonance analysis of mutants showed that beta-cyclodextrin binds to SBS2 and SBS1 with K(d,1) and K(d,2) values of 0.07 and 1.40 mM, respectively. A model that accounts for the observed synergy in starch hydrolysis, where SBS1 and SBS2 bind ordered and free alpha-glucan chains, respectively, thus targeting the enzyme to single alpha-glucan chains accessible for hydrolysis, is proposed. SBS1 and SBS2 also influence the kinetics of hydrolysis for amylose and maltooligosaccharides, the degree of multiple attack on amylose, and subsite binding energies.

  6. Replacement of the V3 domain in the surface subunit of the feline immunodeficiency virus envelope glycoprotein with the equivalent region of a T cell-tropic human immunodeficiency virus type 1 results in a chimeric surface protein that efficiently binds to CXCR4.

    Science.gov (United States)

    González, Silvia A; Falcón, Juan I; Affranchino, José L

    2014-03-01

    Feline immunodeficiency virus (FIV) and the T cell-tropic strains of human immunodeficiency virus type 1 (HIV-1) share the use of the chemokine receptor CXCR4 for cell entry. To study this process further we developed a cell surface binding assay based on the expression of a soluble version of the FIV SU C-terminally tagged with the influenza virus hemagglutinin epitope (HA). The specificity of the assay was demonstrated by the following evidence: (1) the SU-HA protein bound to HeLa cells that express CXCR4 but not to MDCK cells that lack this chemokine receptor; and (2) binding of the SU-HA to HeLa cells was blocked by incubation with the CXCR4 antagonist AMD3100 as well as with the anti-CXCR4 monoclonal antibody (MAb) 12G5. Deletion of the V3 region from the FIV SU glycoprotein abolished its ability to bind CXCR4-expressing cells. Remarkably, substitution of the V3 domain of the FIV SU by the equivalent region of the HIV-1 NL4-3 isolate resulted in efficient cell surface binding of the chimeric SU protein to CXCR4. Moreover, transfection of MDCK cells with a plasmid encoding human CXCR4 allowed the association of the chimeric SU-HA glycoprotein to the transfected cells. Interestingly, while cell binding of the chimeric FIV-HIV SU was inhibited by an anti-HIV-1 V3 MAb, its association with CXCR4 was found to be resistant to AMD3100. Of note, the chimeric FIV-HIV Env glycoprotein was capable of promoting CXCR4-dependent cell-to-cell fusion.

  7. Proton and aluminum binding properties of organic acids in surface waters of the northeastern U.S.

    Science.gov (United States)

    Fakhraei, Habibollah; Driscoll, Charles T

    2015-03-03

    A variety of mathematical estimators have been used to quantify the degree of protonation of naturally occurring organic acids. These estimators range from monoprotic, diprotic, and triprotic analog models to the discrete and continuous (Gaussian) distributions of a single proton binding-dissociation. Natural water samples from two long-term monitoring programs in the northeastern U.S. were used to quantify proton- and aluminum-binding properties of naturally occurring organic matter. Water chemistry observations were clustered into 0.05 pH intervals (over 3.75-7.35 pH range) and fit to a triprotic analog model. The model optimization indicates that about 5% of dissolved organic carbon participates in ion binding, and organic acids are composed of both strong and weak acids (i.e., pKa1 = 2.54, pKa2 = 6.19, and pKa3 = 7.52 for Adirondack samples). Binding between organic acids and aluminum can substantially influence the acid behavior of dissolved organic matter and the availability of the toxic form of aluminum (i.e., inorganic monomeric aluminum).

  8. Population heterogeneity in the surface expression of Ulex europaeus I-lectin (UEA I)-binding sites in cultured malignant and transformed cells

    Energy Technology Data Exchange (ETDEWEB)

    Virtanen, I.; Lehtonen, E.; Naervaenen, O.; Leivo, I.; Lehto, V.P.

    1985-11-01

    We studied the binding of fluorochrome-coupled Ulex europaeus I-lectin (UEA-I) to cultured malignant cells: all human malignant and transformed cells and also mouse teratocarcinoma cells examined gave a homogeneous cell membrane-type of surface staining only in some of the cells. Such a population heterogeneity appeared to be independent of the cell cycle. Instead, other lectin conjugates used bound homogeneously to all cell. In permeabilized cells, a juxtanuclear reticular staining of the Golgi apparatus was seen in the UEA-I-positive cells. No staining of the pericellular matrix components, produced by malignant cells grown in serum-free culture medium, could be obtained with TRITC-UEA-I. UEA-I-lectin recognized most polypeptides from A8387 fibrosarcoma cells and HeLa cells, metabolically labelled with (/sup 3/H)fucose. Furthermore, surface labelling of these cells with the neuraminidase-galactose oxidase/sodium borohydride method disclosed that both UEA-I and Ricinus communis agglutinin I revealed the same major surface glycoproteins. Results with metabolically labelled cells showed, in addition, that UEA-I-lectin did not bind to secreted glycoproteins produced by A8387 cells and recognized by other lectins. The results indicate that transformed and malignant cells show a distinct population heterogeneity in their expression of some cell surface-associated fucosyl glycoconjugates. The results also suggest that malignant cells can glycosylate their membrane and secreted glycoproteins in a different manner.

  9. Surface Proteome Analysis of a Natural Isolate of Lactococcus lactis Reveals the Presence of Pili Able to Bind Human Intestinal Epithelial Cells*

    Science.gov (United States)

    Meyrand, Mickael; Guillot, Alain; Goin, Mélodie; Furlan, Sylviane; Armalyte, Julija; Kulakauskas, Saulius; Cortes-Perez, Naima G.; Thomas, Ginette; Chat, Sophie; Péchoux, Christine; Dupres, Vincent; Hols, Pascal; Dufrêne, Yves F.; Trugnan, Germain; Chapot-Chartier, Marie-Pierre

    2013-01-01

    Surface proteins of Gram-positive bacteria play crucial roles in bacterial adhesion to host tissues. Regarding commensal or probiotic bacteria, adhesion to intestinal mucosa may promote their persistence in the gastro-intestinal tract and their beneficial effects to the host. In this study, seven Lactococcus lactis strains exhibiting variable surface physico-chemical properties were compared for their adhesion to Caco-2 intestinal epithelial cells. In this test, only one vegetal isolate TIL448 expressed a high-adhesion phenotype. A nonadhesive derivative was obtained by plasmid curing from TIL448, indicating that the adhesion determinants were plasmid-encoded. Surface-exposed proteins in TIL448 were analyzed by a proteomic approach consisting in shaving of the bacterial surface with trypsin and analysis of the released peptides by LC-MS/MS. As the TIL448 complete genome sequence was not available, the tryptic peptides were identified by a mass matching approach against a database including all Lactococcus protein sequences and the sequences deduced from partial DNA sequences of the TIL448 plasmids. Two surface proteins, encoded by plasmids in TIL448, were identified as candidate adhesins, the first one displaying pilin characteristics and the second one containing two mucus-binding domains. Inactivation of the pilin gene abolished adhesion to Caco-2 cells whereas inactivation of the mucus-binding protein gene had no effect on adhesion. The pilin gene is located inside a cluster of four genes encoding two other pilin-like proteins and one class-C sortase. Synthesis of pili was confirmed by immunoblotting detection of high molecular weight forms of pilins associated to the cell wall as well as by electron and atomic force microscopy observations. As a conclusion, surface proteome analysis allowed us to detect pilins at the surface of L. lactis TIL448. Moreover we showed that pili appendages are formed and involved in adhesion to Caco-2 intestinal epithelial cells

  10. Structure and calcium binding activity of LipL32, the major surface antigen of pathogenic Leptospira sp

    Energy Technology Data Exchange (ETDEWEB)

    Hauk, Pricila; Roman-Ramos, Henrique; Ho, Paulo Lee [Instituto Butantan, Sao Paulo, SP (Brazil). Centro de Biotecnologia; Guzzo, Cristiane R.; Farah, Chuck S. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica. Dept. de Bioquimica

    2009-07-01

    Leptospirosis, caused by the spirochaete Leptospira is an important emerging infectious disease. LipL32 is the major exposed outer membrane protein found exclusively in pathogenic leptospira. It is highly immunogenic and has been shown to bind to host extracellular matrix components, including collagens, fibronectin and laminin. In this work we crystallized recombinant LipL32 protein and determined its structure to 2.25 A resolution. Initial phases were determined using the multi-wavelength anomalous dispersion technique with data collected from selenomethionine-containing crystals at the MX2 beamline at the LNLS. The LipL32 monomer is made of a jelly-roll fold core from which protrude several peripheral secondary structures. Some structural features suggested that LipL32 could bind Ca{sup 2+} ions and indeed, spectroscopic data (circular (dichroism. intrinsic tryptophan fluorescence and extrinsic 1-amino-2-anaphthol-4-sulfonic acid fluorescence) confirmed the calcium binding properties of LipL32. (author)

  11. Effects of Surface Charge and Functional Groups on the Adsorption and Binding Forms of Cu and Cd on Roots of indica and japonica Rice Cultivars

    Directory of Open Access Journals (Sweden)

    Zhao-Dong Liu

    2017-08-01

    Full Text Available This work was designed to understand the mechanisms of adsorption of copper (Cu and cadmium (Cd on roots of indica and japonica varieties of rice. Six varieties each of indica and japonica rice were grown in hydroponics and the chemical properties of the root surface were analyzed, including surface charges and functional groups (-COO- groups as measured by the streaming potential and attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR. Binding forms of heavy metals adsorbed on rice roots were identified using sequential extraction methods. In rice roots exposed to Cu and Cd solutions, Cu existed mainly in both exchangeable and complexed forms, whereas Cd existed mainly in the exchangeable form. The amounts of exchangeable Cu and Cd and total adsorbed metal cations on the roots of indica varieties were significantly greater than those on the roots of japonica varieties, and the higher negative charges and the larger number of functional groups on the roots of indica varieties were responsible for their higher adsorption capacity and greater binding strength for Cu and Cd. Surface charge and functional groups on roots play an important role in the adsorption of Cu and Cd on the rice roots.

  12. A Lagrangian stochastic model to demonstrate multi-scale interactions between convection and land surface heterogeneity in the atmospheric boundary layer

    Science.gov (United States)

    Parsakhoo, Zahra; Shao, Yaping

    2017-04-01

    Near-surface turbulent mixing has considerable effect on surface fluxes, cloud formation and convection in the atmospheric boundary layer (ABL). Its quantifications is however a modeling and computational challenge since the small eddies are not fully resolved in Eulerian models directly. We have developed a Lagrangian stochastic model to demonstrate multi-scale interactions between convection and land surface heterogeneity in the atmospheric boundary layer based on the Ito Stochastic Differential Equation (SDE) for air parcels (particles). Due to the complexity of the mixing in the ABL, we find that linear Ito SDE cannot represent convections properly. Three strategies have been tested to solve the problem: 1) to make the deterministic term in the Ito equation non-linear; 2) to change the random term in the Ito equation fractional, and 3) to modify the Ito equation by including Levy flights. We focus on the third strategy and interpret mixing as interaction between at least two stochastic processes with different Lagrangian time scales. The model is in progress to include the collisions among the particles with different characteristic and to apply the 3D model for real cases. One application of the model is emphasized: some land surface patterns are generated and then coupled with the Large Eddy Simulation (LES).

  13. Anti-citrullinated protein antibodies promote apoptosis of mature human Saos-2 osteoblasts via cell-surface binding to citrullinated heat shock protein 60.

    Science.gov (United States)

    Lu, Ming-Chi; Yu, Chia-Li; Yu, Hui-Chun; Huang, Hsien-Bin; Koo, Malcolm; Lai, Ning-Sheng

    2016-01-01

    We hypothesized that anti-citrullinated protein antibodies (ACPAs) react with osteoblast surface citrullinated proteins and affect cell function, leading to joint damage in patients with rheumatoid arthritis (RA). First, we purified ACPAs by cyclic citrullinated peptide (CCP)-conjugated affinity column chromatography. The cognate antigens of ACPAs on Saos-2 cells, a sarcoma osteogenic cell line generated from human osteoblasts, were probed by ACPAs, and the reactive bands were analyzed using proteomic analyses. We found that ACPAs bind to Saos-2 cell membrane, and several protein candidates, including HSP60, were identified. We then cloned and purified recombinant heat shock protein 60 (HSP60) and citrullinated HSP60 (citHSP60) and investigated the effect of ACPAs on Saos-2 cell. We confirmed that HSP60 obtained from Saos-2 cell membrane were citrullinated and reacted with ACPAs, which induces Saos-2 cells apoptosis via binding to surface-expressed citHSP60 through Toll-like receptor 4 signaling. ACPAs promoted interleukin (IL)-6 and IL-8 expression in Saos-2 cells. Finally, sera from patients with RA and healthy controls were examined for their titers of anti-HSP60 and anti-citHSP60 antibodies using an enzyme-linked immunosorbent assay. The radiographic change in patients with RA was evaluated using the Genant-modified Sharp scoring system. Patients with RA showed higher sera titers of anti-citHSP60, but not anti-HSP60, antibodies when compared with controls. In addition, the anti-citHSP60 level was positively associated with increased joint damage in patients with RA. In conclusion, Saos-2 cell apoptosis was mediated by ACPAs via binding to cell surface-expressed citHSP60 and the titer of anti-citHSP60 in patients with RA positively associated with joint damage. Copyright © 2015 Elsevier GmbH. All rights reserved.

  14. Correction to: Evaluation of cell binding to collagen and gelatin: a study of the effect of 2D and 3D architecture and surface chemistry.

    Science.gov (United States)

    Davidenko, Natalia; Schuster, Carlos F; Bax, Daniel V; Farndale, Richard W; Hamaia, Samir; Best, Serena M; Cameron, Ruth E

    2018-03-21

    The article "Evaluation of cell binding to collagen and gelatin: a study of the effect of 2D and 3D architecture and surface chemistry", written by Natalia Davidenko, Carlos F. Schuster, Daniel V. Bax, Richard W. Farndale, Samir Hamaia, Serena M. Best and Ruth E. Cameron, was originally published Online First without open access. After publication in volume 27, issue 10, page 148 it was noticed that the copyright was wrong in the PDF version of the article. The copyright of the article should read as "© The Author(s) 2016". The Open Access license terms were also missing.

  15. The Importance of Surface-Binding Site towards Starch-Adsorptivity Level in α-Amylase: A Review on Structural Point of View

    Directory of Open Access Journals (Sweden)

    Umi Baroroh

    2017-01-01

    Full Text Available Starch is a polymeric carbohydrate composed of glucose. As a source of energy, starch can be degraded by various amylolytic enzymes, including α-amylase. In a large-scale industry, starch processing cost is still expensive due to the requirement of high temperature during the gelatinization step. Therefore, α-amylase with raw starch digesting ability could decrease the energy cost by avoiding the high gelatinization temperature. It is known that the carbohydrate-binding module (CBM and the surface-binding site (SBS of α-amylase could facilitate the substrate binding to the enzyme’s active site to enhance the starch digestion. These sites are a noncatalytic module, which could interact with a lengthy substrate such as insoluble starch. The major interaction between these sites and the substrate is the CH/pi-stacking interaction with the glucose ring. Several mutation studies on the Halothermothrix orenii, SusG Bacteroides thetaiotamicron, Barley, Aspergillus niger, and Saccharomycopsis fibuligera α-amylases have revealed that the stacking interaction through the aromatic residues at the SBS is essential to the starch adsorption. In this review, the SBS in various α-amylases is also presented. Therefore, based on the structural point of view, SBS is suggested as an essential site in α-amylase to increase its catalytic activity, especially towards the insoluble starch.

  16. Ribosomal protein L5 has a highly twisted concave surface and flexible arms responsible for rRNA binding.

    OpenAIRE

    Nakashima, T; Yao, M; Kawamura, S; Iwasaki, K; Kimura, M; Tanaka, I

    2001-01-01

    Ribosomal protein L5 is a 5S rRNA binding protein in the large subunit and plays an essential role in the promotion of a particular conformation of 5S rRNA. The crystal structure of the ribosomal protein L5 from Bacillus stearothermophilus has been determined at 1.8 A resolution. The molecule consists of a five-stranded antiparallel beta-sheet and four alpha-helices, which fold in a way that is topologically similar to the ribonucleoprotein (RNP) domain. The molecular shape and electrostatic ...

  17. Surface Transient Binding-Based Fluorescence Correlation Spectroscopy (STB-FCS), a Simple and Easy-to-Implement Method to Extend the Upper Limit of the Time Window to Seconds.

    Science.gov (United States)

    Peng, Sijia; Wang, Wenjuan; Chen, Chunlai

    2018-05-10

    Fluorescence correlation spectroscopy is a powerful single-molecule tool that is able to capture kinetic processes occurring at the nanosecond time scale. However, the upper limit of its time window is restricted by the dwell time of the molecule of interest in the confocal detection volume, which is usually around submilliseconds for a freely diffusing biomolecule. Here, we present a simple and easy-to-implement method, named surface transient binding-based fluorescence correlation spectroscopy (STB-FCS), which extends the upper limit of the time window to seconds. We further demonstrated that STB-FCS enables capture of both intramolecular and intermolecular kinetic processes whose time scales cross several orders of magnitude.

  18. Helper T cell epitope-mapping reveals MHC-peptide binding affinities that correlate with T helper cell responses to pneumococcal surface protein A.

    Directory of Open Access Journals (Sweden)

    Rajesh Singh

    2010-02-01

    Full Text Available Understanding the requirements for protection against pneumococcal carriage and pneumonia will greatly benefit efforts in controlling these diseases. Several proteins and polysaccharide capsule have recently been implicated in the virulence of and protective immunity against Streptococcus pneumonia. Pneumococcal surface protein A (PspA is highly conserved among S. pneumonia strains, inhibits complement activation, binds lactoferrin, elicits protective systemic immunity against pneumococcal infection, and is necessary for full pneumococcal virulence. Identification of PspA peptides that optimally bind human leukocyte antigen (HLA would greatly contribute to global vaccine efforts, but this is hindered by the multitude of HLA polymorphisms. Here, we have used an experimental data set of 54 PspA peptides and in silico methods to predict peptide binding to HLA and murine major histocompatibility complex (MHC class II. We also characterized spleen- and cervical lymph node (CLN-derived helper T lymphocyte (HTL cytokine responses to these peptides after S. pneumonia strain EF3030-challenge in mice. Individual, yet overlapping peptides, 15 amino acids in length revealed residues 199 to 246 of PspA (PspA(199-246 consistently caused the greatest IFN-gamma, IL-2, IL-5 and proliferation as well as moderate IL-10 and IL-4 responses by ex vivo stimulated splenic and CLN CD4(+ T cells isolated from S. pneumonia strain EF3030-challeged F(1 (B6xBALB/c mice. IEDB, RANKPEP, SVMHC, MHCPred, and SYFPEITHI in silico analysis tools revealed peptides in PspA(199-246 also interact with a broad range of HLA-DR, -DQ, and -DP allelles. These data suggest that predicted MHC class II-peptide binding affinities do not always correlate with T helper (Th cytokine or proliferative responses to PspA peptides, but when used together with in vivo validation can be a useful tool to choose candidate pneumococcal HTL epitopes.

  19. Deposition of mannose-binding lectin and ficolins and activation of the lectin pathway of complement on the surface of polyurethane tubing used for cardiopulmonary bypass.

    Science.gov (United States)

    Eppa, Łukasz; Pągowska-Klimek, Izabela; Świerzko, Anna S; Moll, Maciej; Krajewski, Wojciech R; Cedzyński, Maciej

    2018-04-01

    The artificial surface used for cardiopulmonary bypass (CPB) is a crucial factor activating the complement system and thus contributing to the generation of a systemic inflammatory response. The activation of classical and alternative pathways on this artificial surface is well known. In contrast, lectin pathway (LP) activation has not been fully investigated, although noted during CPB in several studies. Moreover, we have recently proved the contribution of the LP to the generation of the systemic inflammatory response syndrome after pediatric cardiac surgery. The aim of this study was to assess LP-mediated complement activation on the surface of polyurethane CPB circuit tubing (noncoated Chalice ® ), used for CPB procedures in children with congenital heart disease. We found deposition of mannose-binding lectin, ficolin-1, -2, and -3 on the surface of unused tubing and on tubing used for CPB from a small minority of patients. Furthermore, we observed deposition of complement C4 activation products on tubing used for CPB and previously unused tubing after incubation with normal serum. The latter finding indicates LP activation in vitro on the polyurethane surface. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1202-1208, 2018. © 2017 Wiley Periodicals, Inc.

  20. Heteronuclear 2D NMR studies on an engineered insulin monomer: Assignments and characterization of the receptor-binding surface by selective 2H and 13C labeling with application to protein design

    International Nuclear Information System (INIS)

    Weiss, M.A.; Hua, Qingxin; Lynch, C.S.; Shoelson, S.E.; Frank, B.H.

    1991-01-01

    Insulin provides an important model for the application of genetic engineering to rational protein design and has been well characterized in the crystal state. However, self-association of insulin in solution has precluded complementary 2D NMR study under physiological conditions. The authors demonstrate here that such limitations may be circumvented by the use of a monomeric analogue that contains three amino acid substitutions on the protein surface (HisB10 → Asp, ProB28 → Lys, and LysB29 → Pro); this analogue (designated DKP-insulin) retains native receptor-binding potency. Comparative 1 H NMR studies of native human insulin and a series of three related analogues-(i) the singly substituted analogue [HisB10→Asp], (ii) the doubly substituted analogue [ProB28→Lys; LysB29→Pro], and (iii) DKP-insulin-demonstrate progressive reduction in concentration-dependent line-broadening in accord with the results of analytical ultracentrifugation. Extensive nonlocal interactions are observed in the NOESY spectrum of DKP-insulin, indicating that this analogue adopts a compact and stably folded structure as a monomer in overall accord with crystal models. Site-specific 2 H and 13 C isotopic labels are introduced by semisynthesis as probes for the structure and dynamics of the receptor-binding surface. These studies confirm and extend under physiological conditions the results of a previous 2D NMR analysis of native insulin in 20% acetic acid. Implications for the role of protein flexibility in receptor recognition are discussed with application to the design of novel insulin analogues

  1. Cell surface binding and uptake of arginine- and lysine-rich penetratin peptides in absence and presence of proteoglycans

    KAUST Repository

    Å mand, Helene L.; Rydberg, Hanna A.; Fornander, Louise H.; Lincoln, Per; Nordé n, Bengt; Esbjö rner, Elin K.

    2012-01-01

    Cell surface proteoglycans (PGs) appear to promote uptake of arginine-rich cell-penetrating peptides (CPPs), but their exact functions are unclear. To address if there is specificity in the interactions of arginines and PGs leading to improved

  2. Surface-bound capsular polysaccharide of type Ia group B Streptococcus mediates C1 binding and activation of the classic complement pathway

    International Nuclear Information System (INIS)

    Levy, N.J.; Kasper, D.L.

    1986-01-01

    The role of surface-bound type Ia group B Streptococcus (GBS) capsular polysaccharide in anti-body-independent binding of C1 and activation of the classic component pathway was investigated. In a radiolabeled bacterial-polymorphonuclear leukocyte (PMN) association assay, a measure of bacterial opsonization, preincubation of 3 H-type Ia GBS with purified F(ab') 2 to the organism blocked the association of the bacteria with PMN', and the inhibitory effect was dose dependent. The specificity of F(ab') 2 blocking was shown after adsorption of F(ab') 2 with type Ia polysaccharide-sensitized erythrocytes. Polysaccharide-adsorbed F(ab') 2 had a 70% decrease in ability to block the association of bacteria with PMN. Neuraminidase digestion removed 80% of the terminal sialic acid residues from the native polysaccharide. These neuraminidase-digested organisms had a 72% decrease in binding and transfer of purified C1 compared with non-enzyme-treated organisms. Type Ia capsular polysaccharide bound to sheep erythrocytes promoted classic complement pathway-mediated hemolysis of the cells. The role of C1 inhibitor (INH) in modulation of C1 activation by the organisms was investigated. The possibility existed that the C1 INH could be bound by the bacteria, allowing C1 activation to occur in the fluid phase. The inhibitor was purified from human serum, and its activity was measured before and after incubation with type Ia GBS. The organisms had no effect on C1 INH activity. Thus surface-bound capsular polysacchardie of type Ia GBS mediates C1 binding and classic pathway activation, and this does not involve the C1 INH

  3. Binding, folding and insertion of a β-hairpin peptide at a lipid bilayer surface: Influence of electrostatics and lipid tail packing.

    Science.gov (United States)

    Reid, Keon A; Davis, Caitlin M; Dyer, R Brian; Kindt, James T

    2018-03-01

    Antimicrobial peptides (AMPs) act as host defenses against microbial pathogens. Here we investigate the interactions of SVS-1 (KVKVKVKV d P l PTKVKVKVK), an engineered AMP and anti-cancer β-hairpin peptide, with lipid bilayers using spectroscopic studies and atomistic molecular dynamics simulations. In agreement with literature reports, simulation and experiment show preferential binding of SVS-1 peptides to anionic over neutral bilayers. Fluorescence and circular dichroism studies of a Trp-substituted SVS-1 analogue indicate, however, that it will bind to a zwitterionic DPPC bilayer under high-curvature conditions and folds into a hairpin. In bilayers formed from a 1:1 mixture of DPPC and anionic DPPG lipids, curvature and lipid fluidity are also observed to promote deeper insertion of the fluorescent peptide. Simulations using the CHARMM C36m force field offer complementary insight into timescales and mechanisms of folding and insertion. SVS-1 simulated at an anionic mixed POPC/POPG bilayer folded into a hairpin over a microsecond, the final stage in folding coinciding with the establishment of contact between the peptide's valine sidechains and the lipid tails through a "flip and dip" mechanism. Partial, transient folding and superficial bilayer contact are seen in simulation of the peptide at a zwitterionic POPC bilayer. Only when external surface tension is applied does the peptide establish lasting contact with the POPC bilayer. Our findings reveal the influence of disruption to lipid headgroup packing (via curvature or surface tension) on the pathway of binding and insertion, highlighting the collaborative effort of electrostatic and hydrophobic interactions on interaction of SVS-1 with lipid bilayers. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Quantum mechanics capacitance molecular mechanics modeling of core-electron binding energies of methanol and methyl nitrite on Ag(111) surface.

    Science.gov (United States)

    Löytynoja, T; Li, X; Jänkälä, K; Rinkevicius, Z; Ågren, H

    2016-07-14

    We study a newly devised quantum mechanics capacitance molecular mechanics (QMCMM) method for the calculation of core-electron binding energies in the case of molecules adsorbed on metal surfaces. This yet untested methodology is applied to systems with monolayer of methanol/methyl nitrite on an Ag(111) surface at 100 K temperature. It was found out that the studied C, N, and O 1s core-hole energies converge very slowly as a function of the radius of the metallic cluster, which was ascribed to build up of positive charge on the edge of the Ag slab. Further analysis revealed that an extrapolation process can be used to obtain binding energies that deviated less than 0.5 eV against experiments, except in the case of methanol O 1s where the difference was as large as 1.8 eV. Additional QM-cluster calculations suggest that the latter error can be connected to the lack of charge transfer over the QM-CMM boundary. Thus, the results indicate that the QMCMM and QM-cluster methods can complement each other in a holistic picture of molecule-adsorbate core-ionization studies, where all types of intermolecular interactions are considered.

  5. α-Helical element at the hormone-binding surface of the insulin receptor functions as a signaling element to activate its tyrosine kinase.

    Science.gov (United States)

    Whittaker, Jonathan; Whittaker, Linda J; Roberts, Charles T; Phillips, Nelson B; Ismail-Beigi, Faramarz; Lawrence, Michael C; Weiss, Michael A

    2012-07-10

    The primary hormone-binding surface of the insulin receptor spans one face of the N-terminal β-helix of the α-subunit (the L1 domain) and an α-helix in its C-terminal segment (αCT). Crystallographic analysis of the free ectodomain has defined a contiguous dimer-related motif in which the αCT α-helix packs against L1 β-strands 2 and 3. To relate structure to function, we exploited expanded genetic-code technology to insert photo-activatable probes at key sites in L1 and αCT. The pattern of αCT-mediated photo-cross-linking within the free and bound receptor is in accord with the crystal structure and prior mutagenesis. Surprisingly, L1 photo-probes in β-strands 2 and 3, predicted to be shielded by αCT, efficiently cross-link to insulin. Furthermore, anomalous mutations were identified on neighboring surfaces of αCT and insulin that impair hormone-dependent activation of the intracellular receptor tyrosine kinase (contained within the transmembrane β-subunit) disproportionately to their effects on insulin binding. Taken together, these results suggest that αCT, in addition to its hormone-recognition role, provides a signaling element in the mechanism of receptor activation.

  6. Proteins feel more than they see : Fine-tuning of binding affinity by properties of the non-interacting surface

    NARCIS (Netherlands)

    Kastritis, Panagiotis L.|info:eu-repo/dai/nl/315886668; Garcia Lopes Maia Rodrigues, João|info:eu-repo/dai/nl/330827391; Folkers, Gert E.|info:eu-repo/dai/nl/162277202; Boelens, Rolf|info:eu-repo/dai/nl/070151407; Bonvin, Alexandre M J J|info:eu-repo/dai/nl/113691238

    2014-01-01

    Protein-protein complexes orchestrate most cellular processes such as transcription, signal transduction and apoptosis. The factors governing their affinity remain elusive however, especially when it comes to describing dissociation rates (koff). Here we demonstrate that, next to direct

  7. Evanescent-wave cavity ring-down spectroscopy for enhanced detection of surface binding under flow injection analysis conditions

    NARCIS (Netherlands)

    Van Der Sneppen, L.; Ariese, F.; Gooijer, C.; Ubachs, W.

    2008-01-01

    In evanescent-wave cavity ring-down spectroscopy, one (or more) of the re°ections inside the cavity is a total internal re°ection (TIR) event. Only the evanescent wave associated with this TIR is being used for prob-ing the sample. This technique is therefore highly surface-speci-c and attractive

  8. Binding Interactions Between α-glucans from Lactobacillus reuteri and Milk Proteins Characterised by Surface Plasmon Resonance

    DEFF Research Database (Denmark)

    Diemer, Silja Kej; Svensson, Birte; Babol, Linnéa N.

    2012-01-01

    Interactions between milk proteins and α-glucans at pH 4.0–5.5 were investigated by use of surface plasmon resonance. The α-glucans were synthesised with glucansucrase enzymes from Lactobacillus reuteri strains ATCC-55730, 180, ML1 and 121. Variations in the molecular characteristics of the α...

  9. Binding Interactions Between alpha-glucans from Lactobacillus reuteri and Milk Proteins Characterised by Surface Plasmon Resonance

    NARCIS (Netherlands)

    Diemer, Silja K.; Svensson, Birte; Babol, Linnea N.; Cockburn, Darrell; Grijpstra, Pieter; Dijkhuizen, Lubbert; Folkenberg, Ditte M.; Garrigues, Christel; Ipsen, Richard H.

    Interactions between milk proteins and alpha-glucans at pH 4.0-5.5 were investigated by use of surface plasmon resonance. The alpha-glucans were synthesised with glucansucrase enzymes from Lactobacillus reuteri strains ATCC-55730, 180, ML1 and 121. Variations in the molecular characteristics of the

  10. Feature Binding in Zebrafish

    Directory of Open Access Journals (Sweden)

    P Neri

    2012-07-01

    Full Text Available Binding operations are primarily ascribed to cortex or similarly complex avian structures. My experiments show that the zebrafish, a lower vertebrate lacking cortex, supports visual feature binding of form and motion for the purpose of social behavior. These results challenge the notion that feature binding may require highly evolved neural structures and demonstrate that the nervous system of lower vertebrates can afford unexpectedly complex computations.

  11. Characterization of SynCAM surface trafficking using a SynCAM derived ligand with high homophilic binding affinity

    International Nuclear Information System (INIS)

    Breillat, Christelle; Thoumine, Olivier; Choquet, Daniel

    2007-01-01

    In order to better probe SynCAM function in neurons, we produced a fusion protein between the extracellular domain of SynCAM1 and the constant fragment of human IgG (SynCAM-Fc). Whether in soluble form or immobilized on latex microspheres, the chimera bound specifically to the surface of hippocampal neurons and recruited endogenous SynCAM molecules. SynCAM-Fc was also used in combination with Quantum Dots to follow the mobility of transfected SynCAM receptors at the neuronal surface. Both immobile and highly mobile SynCAM were found. Thus, SynCAM-Fc behaves as a high affinity ligand that can be used to study the function of SynCAM at the neuronal membrane

  12. Infrared Spectra and Binding Energies of Chemical Warfare Nerve Agent Simulants on the Surface of Amorphous Silica

    Science.gov (United States)

    2013-06-24

    surface that was nearly free from adsorbed water, had only trace hydrocarbon contamination, and contained a significant number of isolated SiO−H groups...are indicative of C−H stretches from a trace amount of hydrocarbon contamination within the silica sample. The three broad peaks at 1980, 1872, and...452. (13) Gay, I. D.; McFarlan, A. J.; Morrow, B. A. Trimethyl Phosphite Adsorbed on Silica - An NMR and Infrared Study. J. Phys. Chem. 1991, 95 (3

  13. Demonstration of spread-on peel-off consumer products for sampling surfaces contaminated with pesticides and chemical warfare agent signatures.

    Science.gov (United States)

    Behringer, Deborah L; Smith, Deborah L; Katona, Vanessa R; Lewis, Alan T; Hernon-Kenny, Laura A; Crenshaw, Michael D

    2014-08-01

    A terrorist attack using toxic chemicals is an international concern. The utility of rubber cement and latex body paint as spray-on/spread-on peel-off collection media for signatures attributable to pesticides and chemical warfare agents from interior building and public transportation surfaces two weeks post-deposition is demonstrated. The efficacy of these media to sample escalator handrail, stainless steel, vinyl upholstery fabric, and wood flooring is demonstrated for two pesticides and eight chemicals related to chemical warfare agents. The chemicals tested are nicotine, parathion, atropine, diisopropyl methylphosphonate, dimethyl methylphosphonate, dipinacolyl methylphosphonate, ethyl methylphosphonic acid, isopropyl methylphosphonic acid, methylphosphonic acid, and thiodiglycol. Amounts of each chemical found are generally greatest when latex body paint is used. Analytes with low volatility and containing an alkaline nitrogen or a sulfur atom (e.g., nicotine and parathion) usually are recovered to a greater extent than the neutral phosphonate diesters and acidic phosphonic acids (e.g., dimethyl methylphosphonate and ethyl methylphosphonic acid). Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  14. Demonstrating the effectiveness of body armour: a pilot prospective computerised surface wound mapping trial performed at the Role 3 hospital in Afghanistan.

    Science.gov (United States)

    Breeze, Johno; Allanson-Bailey, L S; Hepper, A E; Midwinter, M J

    2015-03-01

    Modern body armour clearly reduces injury incidence and severity, but evidence to actually objectively demonstrate this effect is scarce. Although the Joint Theatre Trauma Registry (JTTR) alone cannot relate injury pattern to body armour coverage, the addition of computerised Surface Wound Mapping (SWM) may enable this utility. Surface wound locations of all UK and NATO coalition soldiers, Afghan National Army and Police and local nationals injured by explosively propelled fragments and treated in the Role 3 UK-led Field Hospital in Camp Bastion, Afghanistan, between 8 July and 20 October 2012 were prospectively recorded. The Abbreviated Injury Scores (AIS) and relative risk of casualties sustaining injuries under a type of body armour were compared with those that did not wear that armour. Casualties wearing a combat helmet were 2.7 times less likely to sustain a fragmentation wound to the head than those that were unprotected (mean AIS of 2.9 compared with 4.1). Casualties wearing a body armour vest were 4.1 times less likely to sustain a fragmentation wound to the chest or abdomen than those that were unprotected (mean AIS of 2.9 compared with 3.9). Casualties wearing pelvic protection were 10 times less likely to sustain a fragmentation wound to the pelvis compared with those that were unprotected (mean AIS of 3.4 compared with 3.9). Computerised SWM has objectively demonstrated the ability of body armour worn on current operations in Afghanistan to reduce wound incidence and severity. We recognise this technique is limited in that it only records the surface wound location and may be specific to this conflict. However, gathering electronic SWM at the same time as recording injuries for the JTTR was simple, required little extra time and therefore we would recommend its collection during future conflicts. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  15. The neuronal Ca(2+) -binding protein 2 (NECAB2) interacts with the adenosine A(2A) receptor and modulates the cell surface expression and function of the receptor.

    Science.gov (United States)

    Canela, Laia; Luján, Rafael; Lluís, Carme; Burgueño, Javier; Mallol, Josefa; Canela, Enric I; Franco, Rafael; Ciruela, Francisco

    2007-09-01

    Heptaspanning membrane also known as G protein-coupled receptors (GPCR) do interact with a variety of intracellular proteins whose function is regulate receptor traffic and/or signaling. Using a yeast two-hybrid screen, NECAB2, a neuronal calcium binding protein, was identified as a binding partner for the adenosine A(2A) receptor (A(2A)R) interacting with its C-terminal domain. Co-localization, co-immunoprecipitation and pull-down experiments showed a close and specific interaction between A(2A)R and NECAB2 in both transfected HEK-293 cells and also in rat striatum. Immunoelectron microscopy detection of NECAB2 and A(2A)R in the rat striatopallidal structures indicated that both proteins are co-distributed in the same glutamatergic nerve terminals. The interaction of NECAB2 with A(2A)R modulated the cell surface expression, the ligand-dependent internalization and the receptor-mediated activation of the MAPK pathway. Overall, these results show that A(2A)R interacts with NECAB2 in striatal neurones co-expressing the two proteins and that the interaction is relevant for A(2A)R function.

  16. Sensitivity of Satellite-Based Skin Temperature to Different Surface Emissivity and NWP Reanalysis Sources Demonstrated Using a Single-Channel, Viewing-Angle-Corrected Retrieval Algorithm

    Science.gov (United States)

    Scarino, B. R.; Minnis, P.; Yost, C. R.; Chee, T.; Palikonda, R.

    2015-12-01

    Single-channel algorithms for satellite thermal-infrared- (TIR-) derived land and sea surface skin temperature (LST and SST) are advantageous in that they can be easily applied to a variety of satellite sensors. They can also accommodate decade-spanning instrument series, particularly for periods when split-window capabilities are not available. However, the benefit of one unified retrieval methodology for all sensors comes at the cost of critical sensitivity to surface emissivity (ɛs) and atmospheric transmittance estimation. It has been demonstrated that as little as 0.01 variance in ɛs can amount to more than a 0.5-K adjustment in retrieved LST values. Atmospheric transmittance requires calculations that employ vertical profiles of temperature and humidity from numerical weather prediction (NWP) models. Selection of a given NWP model can significantly affect LST and SST agreement relative to their respective validation sources. Thus, it is necessary to understand the accuracies of the retrievals for various NWP models to ensure the best LST/SST retrievals. The sensitivities of the single-channel retrievals to surface emittance and NWP profiles are investigated using NASA Langley historic land and ocean clear-sky skin temperature (Ts) values derived from high-resolution 11-μm TIR brightness temperature measured from geostationary satellites (GEOSat) and Advanced Very High Resolution Radiometers (AVHRR). It is shown that mean GEOSat-derived, anisotropy-corrected LST can vary by up to ±0.8 K depending on whether CERES or MODIS ɛs sources are used. Furthermore, the use of either NOAA Global Forecast System (GFS) or NASA Goddard Modern-Era Retrospective Analysis for Research and Applications (MERRA) for the radiative transfer model initial atmospheric state can account for more than 0.5-K variation in mean Ts. The results are compared to measurements from the Surface Radiation Budget Network (SURFRAD), an Atmospheric Radiation Measurement (ARM) Program ground

  17. A recyclable Au(I) catalyst for selective homocoupling of arylboronic acids: significant enhancement of nano-surface binding for stability and catalytic activity.

    Science.gov (United States)

    Zhang, Xin; Zhao, Haitao; Wang, Jianhui

    2010-08-01

    Au nanoparticles stabilized by polystyrene-co-polymethacrylic acid microspheres (PS-co-PMAA) were prepared and characterized via X-ray diffraction (XRD), and transmission electron microscope (TEM). The Au nanoparticles supported on the microspheres showed highly selective catalytic activity for homo-coupling reactions of arylboronic acids in a system of aryl-halides and arylboronic acids. X-ray photoelectron spectroscopy (XPS) spectra of the catalyst shows large amounts of Au(I) complexes band to the surface of the Au nanoparticles, which contributes to the selective homocoupling of the arylboronic acids. More importantly, this supported Au complex is a highly recyclable catalyst. The supported Au catalyst can be recycled and reused at least 6 times for a phenylboronic acid reactant, whereas the parent complex shows very low catalytic activity for this compound. The high catalytic activity of this material is attributed to: (1) the high surface to volume ratio which leads to more active sites being exposed to reactants; (2) the strong surface binding of the Au nanoparticle to the Au(I) complexes, which enhances both the stability and the catalytic activity of these complexes.

  18. Electroactive chain-like compounds constructed from trimetallic clusters and 4,4'-bipyridine spacers: one-pot synthesis, characterization and surface binding.

    Science.gov (United States)

    Abe, Masaaki; Inatomi, Atsushi; Hisaeda, Yoshio

    2011-03-14

    This paper reports the synthesis and characterization of a novel series of chain-like compounds where oxo-centered triruthenium cluster moieties are bridged by 4,4'-bipyridine (4,4'-bpy) spacers. A reaction of solvent-coordinated triruthenium "monomer" precursor [Ru(3)O(CH(3)CO(2))(6)(CO)(CH(3)OH)(2)] with a 0.1 equimolar amount of 4,4'-bpy in CH(3)OH gave mixture of chain-like compounds containing "dimers" to "tetramers" which were cleanly separated by column chromatography and characterized by spectroscopic and electrochemical methods. Cyclic voltammetry revealed that all chain-like compounds exhibit reversible and stepwise redox processes in solution with very weak intramolecular coupling between the triruthenium components across the 4,4'-bpy bridge. Photo-induced dissociation of CO from the compounds and electrode surface binding were also investigated.

  19. Role of surface layer collagen binding protein from indigenous Lactobacillus plantarum 91 in adhesion and its anti-adhesion potential against gut pathogen.

    Science.gov (United States)

    Yadav, Ashok Kumar; Tyagi, Ashish; Kaushik, Jai Kumar; Saklani, Asha Chandola; Grover, Sunita; Batish, Virender Kumar

    2013-12-14

    Human feacal isolates were ascertain as genus Lactobacillus using specific primer LbLMA1/R16-1 and further identified as Lactobacillus plantarum with species specific primers Lpl-3/Lpl-2. 25 L. plantarum strains were further assessed for hydrophobicity following the microbial adhesion to hydrocarbons (MATH) method and colonization potentials based on their adherence to immobilized human collagen type-1. Surface proteins were isolated from selected L. plantarum 91(Lp91) strain. The purified collagen binding protein (Cbp) protein was assessed for its anti-adhesion activity against enteric Escherichia coli 0157:H7 pathogen on immobilized collagen. Four L. plantarum strains displayed high degree of hydrophobicity and significant adhesion to collagen. A 72 kDa protein was purified which reduced 59.71% adhesion of E. coli 0157:H7 on immobilized collagen as compared to control well during adhesion assay. Cbp protein is the major influencing factor in inhibition of E. coli 0157:H7 adhesion with extracellular matrix (ECM) components. Hydrophobicity and adhesion potential are closely linked attributes precipitating in better colonization potential of the lactobacillus strains. Cbp is substantiated as a crucial surface protein contributing in adhesion of lactobacillus strains. The study can very well be the platform for commercialization of indigenous probiotic strain once their functional attributes are clinically explored. Copyright © 2013 Elsevier GmbH. All rights reserved.

  20. Tested Demonstrations.

    Science.gov (United States)

    Gilbert, George L.

    1983-01-01

    An apparatus is described in which effects of pressure, volume, and temperature changes on a gas can be observed simultaneously. Includes use of the apparatus in demonstrating Boyle's, Gay-Lussac's, and Charles' Laws, attractive forces, Dalton's Law of Partial pressures, and in illustrating measurable vapor pressures of liquids and some solids.…

  1. Tested Demonstrations.

    Science.gov (United States)

    Gilbert, George L., Ed.

    1987-01-01

    Describes two demonstrations to illustrate characteristics of substances. Outlines a method to detect the changes in pH levels during the electrolysis of water. Uses water pistols, one filled with methane gas and the other filled with water, to illustrate the differences in these two substances. (TW)

  2. The binding parameters of radiolabelled monoclonal F (ab')2 and Fab' fragments relative to immunoglobulin G in reactions with surface-bound antigens

    International Nuclear Information System (INIS)

    Fjeld, J.G.; Nustad, K.; Michaelsen, T.E.

    1992-01-01

    The binding parameters of iodine-125-labelled intact monoclonal immunoglobulin G (IgG), F(ab') 2 and Fab' fragments were compared. The study was carried out with the two monoclonal antibodies (MoAbs) K13 and K16 specific for human Ig light chains κ and λ, respectively. When testing the 125 I-MoAbs against monodisperse polymer particles coated with the specific antigens, the K a for the F(ab') 2 fragments were similar to that for IgG, while the K a for the Fab' fragments were reduced to 10%-20% of that for IgG. The number N of effective target sites revealed with Fab' was higher than with F(ab') and IgG, presumably because less surface area is occupied by the small Fab' molecules. The immunoreactive fraction F ranged according to IgG>F(ab') 2 >Fab'. The explanation of the moderate difference between the K a of the monoclonal Fab' and the divalent IgG and F(ab') 2 was that the divalent molecules were not divalently attached to the particles. When testing the same antibody preparations against humanlymphoma cells producing Ig with light chains κ or λ, the binding results were less reliable than when particles were utilised, presumably due to antigen shedding. Different MoAbs vary in their loss of immunoreactivity due to enzymatic degradation and the radiolabelling procedure. The preparation of the radiolabelled fragments should therefore be optimized for each MoAb, and evaluation is necessary before injection. Artificial targets with a low leakage of antigen, like the monodisperse polymer particles here applied, are recommended for the in vitro evaluation of the immunoreactivity of labelled MoAb preparations. (orig.)

  3. The SH3 domain-binding T cell tyrosyl phosphoprotein p120. Demonstration of its identity with the c-cbl protooncogene product and in vivo complexes with Fyn, Grb2, and phosphatidylinositol 3-kinase

    NARCIS (Netherlands)

    Fukazawa, T.; Reedquist, K. A.; Trub, T.; Soltoff, S.; Panchamoorthy, G.; Druker, B.; Cantley, L.; Shoelson, S. E.; Band, H.

    1995-01-01

    Previously, we have identified p120 as a Fyn/Lck SH3 and SH2 domain-binding protein that is tyrosine phosphorylated rapidly after T cell receptor triggering. Here, we used direct protein purification, amino acid sequence analysis, reactivity with antibodies, and two-dimensional gel analyses to

  4. Background electrolytes and pH effects on selenate adsorption using iron-impregnated granular activated carbon and surface binding mechanisms.

    Science.gov (United States)

    Zhang, Ning; Gang, Daniel Dianchen; McDonald, Louis; Lin, Lian-Shin

    2018-03-01

    Iron-impregnated granular activated carbon (Fe-GAC) has been shown effective for selenite adsorptive removal from aqueous solutions, but similar effectiveness was not observed with selenate. This study examined the effects of background electrolytes and pH on selenate adsorption on to Fe-GAC, and surface bindings to elucidate the selenate adsorption mechanisms. The decrease magnitude of selenate adsorption capacity under three background electrolytes followed the order: LiCl > NaCl > KCl, as ionic strength increased from 0.01 to 0.1 M. Larger adsorption capacity differences among the three electrolytes were observed under the higher ionic strengths (0.05 and 0.1 M) than those under 0.01 M. Multiplet peak fittings of high resolution X-ray photoelectron spectra for O1s and Fe2p 3/2 indicated the presence of iron (III) on adsorbent surface. pH variations during the adsorbent preparation within 3-8 in NaCl solutions did not cause appreciable changes in the iron redox state and composition. Raman spectra showed the formation of both monodentate and bidentate inner sphere complexes under pHs adsorption under alkaline conditions. Mechanisms for monodentate and bidentate formations and a stable six-member ring structure were proposed. Two strategies were recommended for modifying Fe-GAC preparation procedure to enhance the selenate adsorption: (1) mixed-metal oxide coatings to increase the point of zero charge (pH zpc ); and (2) ferrous iron coating to initially reduce selenate followed by selenite adsorption. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Localization of gonadotropin binding sites in human ovarian neoplasms

    International Nuclear Information System (INIS)

    Nakano, R.; Kitayama, S.; Yamoto, M.; Shima, K.; Ooshima, A.

    1989-01-01

    The binding of human luteinizing hormone and human follicle-stimulating hormone to ovarian tumor biopsy specimens from 29 patients was analyzed. The binding sites for human luteinizing hormone were demonstrated in one tumor of epithelial origin (mucinous cystadenoma) and in one of sex cord-stromal origin (theca cell tumor). The binding sites for human follicle-stimulating hormone were found in three tumors of epithelial origin (serous cystadenoma and mucinous cystadenoma) and in two of sex cord-stromal origin (theca cell tumor and theca-granulosa cell tumor). The surface-binding autoradiographic study revealed that the binding sites for gonadotropins were localized in the stromal tissue. The results suggest that gonadotropic hormones may play a role in the growth and differentiation of a certain type of human ovarian neoplasms

  6. [The Role of Membrane-Bound Heat Shock Proteins Hsp90 in Migration of Tumor Cells in vitro and Involvement of Cell Surface Heparan Sulfate Proteoglycans in Protein Binding to Plasma Membrane].

    Science.gov (United States)

    Snigireva, A V; Vrublevskaya, V V; Skarga, Y Y; Morenkov, O S

    2016-01-01

    Heat shock protein Hsp90, detected in the extracellular space and on the membrane of cells, plays an important role in cell motility, migration, invasion and metastasis of tumor cells. At present, the functional role and molecular mechanisms of Hsp90 binding to plasma membrane are not elucidated. Using isoform-specific antibodies against Hsp90, Hsp9α and Hsp90β, we showed that membrane-bound Hsp90α and Hsp90β play a significant role in migration of human fibrosarcoma (HT1080) and glioblastoma (A-172) cells in vitro. Disorders of sulfonation of cell heparan sulfates, cleavage of cell heparan. sulfates by heparinase I/III as well as treatment of cells with heparin lead to an abrupt reduction in the expression level of Hsp90 isoforms. Furthermore, heparin significantly inhibits tumor cell migration. The results obtained demonstrate that two isoforms of membrane-bound Hsp90 are involved in migration of tumor cells in vitro and that cell surface heparan sulfate proteoglycans play a pivotal role in the "anchoring" of Hsp90α and Hsp90β to the plasma membrane.

  7. The human VGF-derived bioactive peptide TLQP-21 binds heat shock 71 kDa protein 8 (HSPA8on the surface of SH-SY5Y cells.

    Directory of Open Access Journals (Sweden)

    Shamim Akhter

    Full Text Available VGF (non-acronymicis a secreted chromogranin/secretogranin that gives rise to a number of bioactive peptides by a complex proteolysis mechanism. VGF-derived peptides exert an extensive array of biological effects in energy metabolism, mood regulation, pain, gastric secretion function, reproduction and, perhaps, cancer. It is therefore surprising that very little is known about receptors and binding partners of VGF-derived peptides and their downstream molecular mechanisms of action. Here, using affinity chromatography and mass spectrometry-based protein identification, we have identified the heat shock cognate 71 kDa protein A8 (HSPA8as a binding partner of human TLQP-21 on the surface of human neuroblastomaSH-SY5Y cells. Binding of TLQP-21 to membrane associated HSPA8 in live SH-SY5Y cells was further supported by cross-linking to live cells. Interaction between HSPA8 and TLQP-21 was confirmed in vitro by label-free Dynamic Mass Redistribution (DMR studies. Furthermore, molecular modeling studies show that TLQP-21 can be docked into the HSPA8 peptide binding pocket. Identification of HSPA8 as a cell surface binding partner of TLQP-21 opens new avenues to explore the molecular mechanisms of its physiological actions, and of pharmacological modulation thereof.

  8. Substrate-Triggered Exosite Binding: Synergistic Dendrimer/Folic Acid Action for Achieving Specific, Tight-Binding to Folate Binding Protein.

    Science.gov (United States)

    Chen, Junjie; van Dongen, Mallory A; Merzel, Rachel L; Dougherty, Casey A; Orr, Bradford G; Kanduluru, Ananda Kumar; Low, Philip S; Marsh, E Neil G; Banaszak Holl, Mark M

    2016-03-14

    Polymer-ligand conjugates are designed to bind proteins for applications as drugs, imaging agents, and transport scaffolds. In this work, we demonstrate a folic acid (FA)-triggered exosite binding of a generation five poly(amidoamine) (G5 PAMAM) dendrimer scaffold to bovine folate binding protein (bFBP). The protein exosite is a secondary binding site on the protein surface, separate from the FA binding pocket, to which the dendrimer binds. Exosite binding is required to achieve the greatly enhanced binding constants and protein structural change observed in this study. The G5Ac-COG-FA1.0 conjugate bound tightly to bFBP, was not displaced by a 28-fold excess of FA, and quenched roughly 80% of the initial fluorescence. Two-step binding kinetics were measured using the intrinsic fluorescence of the FBP tryptophan residues to give a KD in the low nanomolar range for formation of the initial G5Ac-COG-FA1.0/FBP* complex, and a slow conversion to the tight complex formed between the dendrimer and the FBP exosite. The extent of quenching was sensitive to the choice of FA-dendrimer linker chemistry. Direct amide conjugation of FA to G5-PAMAM resulted in roughly 50% fluorescence quenching of the FBP. The G5Ac-COG-FA, which has a longer linker containing a 1,2,3-triazole ring, exhibited an ∼80% fluorescence quenching. The binding of the G5Ac-COG-FA1.0 conjugate was compared to poly(ethylene glycol) (PEG) conjugates of FA (PEGn-FA). PEG2k-FA had a binding strength similar to that of FA, whereas other PEG conjugates with higher molecular weight showed weaker binding. However, no PEG conjugates gave an increased degree of total fluorescence quenching.

  9. Hierarchy of ADAM12 binding to integrins in tumor cells

    DEFF Research Database (Denmark)

    Thodeti, Charles Kumar; Fröhlich, Camilla; Nielsen, Christian Kamp

    2005-01-01

    ADAMs (a disintegrin and metalloprotease) comprise a family of cell surface proteins with protease and cell-binding activities. Using different forms and fragments of ADAM12 as substrates in cell adhesion and spreading assays, we demonstrated that alpha9beta1 integrin is the main receptor for ADA...

  10. Protein A-containing staphylococcus aureus as an immunoglobulin-binding reagent: 1) in radioimmunoassays - 'staf-RIA' - recently also for antibiotics and microbial antigens/antibodies, and 2) in a non-radioactive surface immunoassay - 'Staph-ace ay' read by the naked eye - primarily for antibodies to antigens adsorbed to transparent surfaces

    International Nuclear Information System (INIS)

    Jonsson, S.

    1977-01-01

    This paper is intended to summarize recent developments for the use of protein A-containing staphylococci as an immunoglobulin-binding reagent in various types of radioimmunoassay and some related areas, particularly the staphylococcal surface immunoassay. The paper also presents a new process for the large scale preparation of a freeze-dried preparation of the immunoglobulin-binding, killed staphylococci, which thereby gain a much improved suspension stability. (orig.) [de

  11. Disinfectant wipes are appropriate to control microbial bioburden from surfaces: use of a new ASTM standard test protocol to demonstrate efficacy.

    Science.gov (United States)

    Sattar, S A; Bradley, C; Kibbee, R; Wesgate, R; Wilkinson, M A C; Sharpe, T; Maillard, J-Y

    2015-12-01

    The use of disinfectant pre-soaked wipes (DPW) to decontaminate high-touch environmental surfaces (HTES) by wiping is becoming increasingly widespread in the healthcare environment. However, DPW are rarely tested using conditions simulating their field use, and the label claims of environmental surface disinfectants seldom include wiping action. To evaluate the new E2967-15 standard test specific to wipes, particularly their ability to decontaminate surfaces and to transfer acquired contamination to clean surfaces. ASTM Standard E2967-15 was used by three independent laboratories to test the efficacy of five types of commercially available wipe products. All data generated were pulled together, and reproducibility and repeatability of the standard were measured. All the commercial DPW tested achieved a >4log10 (>99.99%) reduction in colony-forming units (CFU) of Staphylococcus aureus and Acinetobacter baumanii with 10s of wiping, but only one DPW containing 0.5% accelerated H2O2 prevented the transfer of bacteria to another surface. This newly introduced standard method represents a significant advance in assessing DPW for microbial decontamination of HTES, and should greatly assist research and development, and in making more relevant and reliable claims on marketed DPW. Copyright © 2015. Published by Elsevier Ltd.

  12. Binding of antibodies to the extractable nuclear antigens SS-A/Ro and SS-B/La is induced on the surface of human keratinocytes by ultraviolet light (UVL): Implications for the pathogenesis of photosensitive cutaneous lupus

    International Nuclear Information System (INIS)

    Furukawa, F.; Kashihara-Sawami, M.; Lyons, M.B.; Norris, D.A.

    1990-01-01

    Autoantibodies to the non-histone nucleoprotein antigens SS-A/Ro, SS-B/La, and RNP are highly associated with photosensitive cutaneous lupus erythematosus (LE). In order to better understand the potential mechanisms of ultraviolet (UV) light on photosensitivity in patients with cutaneous LE, we designed immunopathologic in vitro and in vivo experiments to evaluate the effects of UV on the binding of such autoantibodies to the surface of human keratinocytes, one major target of immunologic damage in photosensitive LE. Short-term 2% paraformaldehyde fixation of suspensions of cultured human keratinocytes previously incubated with monospecific antiserum probes enabled the detection of ENA expression on the cell surface by flow-cytometry analysis. UVB light (280-320 nm) induced the binding of monospecific antibody probes for SS-A/Ro and SS-B/La on keratinocytes in a dose-dependent pattern with maximal induction observed at the dose of 200 mJ/cm2 UVB. Binding of SS-A/Ro, SS-B/La, and RNP antibody was augmented strongly, but binding of anti-Sm was very weak. In contrast, UVA (320-400 nm) light had no effect on the induction of binding of these antibody probes. Identical results were seen by standard immunofluorescence techniques. Hydroxyurea-treated keratinocytes showed similar induction of those antigens by UVB irradiation, which suggested that ENA expression on cultured keratinocytes by UVB were cell-cycle independent. Tunicamycin, an inhibitor of glycosylation of proteins, reduced UVB light effect on the SS-A/Ro and SS-B/La antigen's expression. These in vitro FACS analyses revealed that ENA augmentation on the keratinocyte cell surface was dose dependent, UVB dependent, glycosylation dependent, and cell-cycle independent. In vivo ENA augmentation on the keratinocyte surface was examined in suction blister epidermal roofs

  13. Possible Overestimation of Surface Disinfection Efficiency by Assessment Methods Based on Liquid Sampling Procedures as Demonstrated by In Situ Quantification of Spore Viability ▿

    Science.gov (United States)

    Grand, I.; Bellon-Fontaine, M.-N.; Herry, J.-M.; Hilaire, D.; Moriconi, F.-X.; Naïtali, M.

    2011-01-01

    The standard test methods used to assess the efficiency of a disinfectant applied to surfaces are often based on counting the microbial survivors sampled in a liquid, but total cell removal from surfaces is seldom achieved. One might therefore wonder whether evaluations of microbial survivors in liquid-sampled cells are representative of the levels of survivors in whole populations. The present study was thus designed to determine the “damaged/undamaged” status induced by a peracetic acid disinfection for Bacillus atrophaeus spores deposited on glass coupons directly on this substrate and to compare it to the status of spores collected in liquid by a sampling procedure. The method utilized to assess the viability of both surface-associated and liquid-sampled spores included fluorescence labeling with a combination of Syto 61 and Chemchrome V6 dyes and quantifications by analyzing the images acquired by confocal laser scanning microscopy. The principal result of the study was that the viability of spores sampled in the liquid was found to be poorer than that of surface-associated spores. For example, after 2 min of peracetic acid disinfection, less than 17% ± 5% of viable cells were detected among liquid-sampled cells compared to 79% ± 5% or 47% ± 4%, respectively, when the viability was evaluated on the surface after or without the sampling procedure. Moreover, assessments of the survivors collected in the liquid phase, evaluated using the microscopic method and standard plate counts, were well correlated. Evaluations based on the determination of survivors among the liquid-sampled cells can thus overestimate the efficiency of surface disinfection procedures. PMID:21742922

  14. Possible overestimation of surface disinfection efficiency by assessment methods based on liquid sampling procedures as demonstrated by in situ quantification of spore viability.

    Science.gov (United States)

    Grand, I; Bellon-Fontaine, M-N; Herry, J-M; Hilaire, D; Moriconi, F-X; Naïtali, M

    2011-09-01

    The standard test methods used to assess the efficiency of a disinfectant applied to surfaces are often based on counting the microbial survivors sampled in a liquid, but total cell removal from surfaces is seldom achieved. One might therefore wonder whether evaluations of microbial survivors in liquid-sampled cells are representative of the levels of survivors in whole populations. The present study was thus designed to determine the "damaged/undamaged" status induced by a peracetic acid disinfection for Bacillus atrophaeus spores deposited on glass coupons directly on this substrate and to compare it to the status of spores collected in liquid by a sampling procedure. The method utilized to assess the viability of both surface-associated and liquid-sampled spores included fluorescence labeling with a combination of Syto 61 and Chemchrome V6 dyes and quantifications by analyzing the images acquired by confocal laser scanning microscopy. The principal result of the study was that the viability of spores sampled in the liquid was found to be poorer than that of surface-associated spores. For example, after 2 min of peracetic acid disinfection, less than 17% ± 5% of viable cells were detected among liquid-sampled cells compared to 79% ± 5% or 47% ± 4%, respectively, when the viability was evaluated on the surface after or without the sampling procedure. Moreover, assessments of the survivors collected in the liquid phase, evaluated using the microscopic method and standard plate counts, were well correlated. Evaluations based on the determination of survivors among the liquid-sampled cells can thus overestimate the efficiency of surface disinfection procedures.

  15. Construction of a novel selection system for endoglucanases exhibiting carbohydrate-binding modules optimized for biomass using yeast cell-surface engineering.

    Science.gov (United States)

    Nakanishi, Akihito; Bae, Jungu; Kuroda, Kouichi; Ueda, Mitsuyoshi

    2012-10-23

    To permit direct cellulose degradation and ethanol fermentation, Saccharomyces cerevisiae BY4741 (Δsed1) codisplaying 3 cellulases (Trichoderma reesei endoglucanase II [EG], T. reesei cellobiohydrolase II [CBH], and Aspergillus aculeatus β-glucosidase I [BG]) was constructed by yeast cell-surface engineering. The EG used in this study consists of a family 1 carbohydrate-binding module (CBM) and a catalytic module. A comparison with family 1 CBMs revealed conserved amino acid residues and flexible amino acid residues. The flexible amino acid residues were at positions 18, 23, 26, and 27, through which the degrading activity for various cellulose structures in each biomass may have been optimized. To select the optimal combination of CBMs of EGs, a yeast mixture with comprehensively mutated CBM was constructed. The mixture consisted of yeasts codisplaying EG with mutated CBMs, in which 4 flexible residues were comprehensively mutated, CBH, and BG. The yeast mixture was inoculated in selection medium with newspaper as the sole carbon source. The surviving yeast consisted of RTSH yeast (the mutant sequence of CBM: N18R, S23T, S26S, and T27H) and wild-type yeast (CBM was the original) in a ratio of 1:46. The mixture (1 RTSH yeast and 46 wild-type yeasts) had a fermentation activity that was 1.5-fold higher than that of wild-type yeast alone in the early phase of saccharification and fermentation, which indicates that the yeast mixture with comprehensively mutated CBM could be used to select the optimal combination of CBMs suitable for the cellulose of each biomass.

  16. Xyloglucan endotransglucosylase/hydrolases (XTHs) are inactivated by binding to glass and cellulosic surfaces, and released in active form by a heat-stable polymer from cauliflower florets.

    Science.gov (United States)

    Sharples, Sandra C; Nguyen-Phan, Tu C; Fry, Stephen C

    2017-11-01

    Xyloglucan endotransglucosylase (XET) activity, which cuts and re-joins hemicellulose chains in the plant cell wall, contributing to wall assembly and growth regulation, is the major activity of XTH proteins. During purification, XTHs often lose XET activity which, however, is restored by treatment with certain cold-water-extractable, heat-stable polymers (CHPs), e.g. from cauliflower florets. It was not known whether the XTH-activating factor (XAF) present in CHPs works by promoting (e.g. allosterically) XET activity or by re-solubilising sequestered XTH proteins. We now show that XTHs in dilute solution bind to diverse surfaces (e.g. glass and cellulose), and that CHPs can re-solubilise the bound enzyme, re-activating it. Cell walls prepared from cauliflower florets, mung bean shoots and Arabidopsis cell-suspension cultures each contained endogenous, tightly bound, inactive XTHs, which were likewise rapidly solubilised (within 0.5h) and thus activated by cauliflower XAF. We present a convenient quantitative assay for XAF acting on the native sequestered XTHs of Arabidopsis cell walls; using this assay, we show that CHPs from all plants tested possess XAF activity. The XAF activity of diverse CHPs does not correlate with their conductivity, showing that this activity is not a simple ionic effect. The XAF action of cauliflower CHPs was augmented by NaCl, although NaCl alone was much less effective than a CHP solution of similar conductivity, confirming that the cauliflower polymers did not simply exert a salt effect. We suggest that XAF is an endogenous regulator of XET action, modulating cell-wall loosening and/or assembly in vivo. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

  17. Binding of peanut lectin to germinal-centre cells: a marker for B-cell subsets of follicular lymphoma?

    OpenAIRE

    Rose, M. L.; Habeshaw, J. A.; Kennedy, R.; Sloane, J.; Wiltshaw, E.; Davies, A. J.

    1981-01-01

    The binding of horseradish-peroxidase-labelled peanut lectin (HRP-PNL) to cryostat sections of tonsil, lymphoma lymph nodes, reactive lymph nodes and miscellaneous tumours demonstrated that PNL binds selectively to lymphocytes in germinal centres. Lymph nodes from 21 patients with non-Hodgkin's lymphomas were phenotyped as cell suspensions for PNL binding, and the following surface markers: E rosetting, C3d, SIg, OK markers of T-cell subsets, Ig heavy-chain and light-chain classes. There was ...

  18. Curved reformat of the paediatric brain MRI into a 'flat-earth map' - standardised method for demonstrating cortical surface atrophy resulting from hypoxic-ischaemic encephalopathy.

    Science.gov (United States)

    Simpson, Ewan; Andronikou, Savvas; Vedajallam, Schadie; Chacko, Anith; Thai, Ngoc Jade

    2016-09-01

    Hypoxic-ischaemic encephalopathy is optimally imaged with brain MRI in the neonatal period. However neuroimaging is often also performed later in childhood (e.g., when parents seek compensation in cases of alleged birth asphyxia). We describe a standardised technique for creating two curved reconstructions of the cortical surface to show the characteristic surface changes of hypoxic-ischaemic encephalopathy in children imaged after the neonatal period. The technique was applied for 10 cases of hypoxic-ischaemic encephalopathy and also for age-matched healthy children to assess the visibility of characteristic features of hypoxic-ischaemic encephalopathy. In the abnormal brains, fissural or sulcal widening was seen in all cases and ulegyria was identifiable in 7/10. These images could be used as a visual aid for communicating MRI findings to clinicians and other interested parties.

  19. Copernicus Big Data and Google Earth Engine for Glacier Surface Velocity Field Monitoring: Feasibility Demonstration on San Rafael and San Quintin Glaciers

    Science.gov (United States)

    Di Tullio, M.; Nocchi, F.; Camplani, A.; Emanuelli, N.; Nascetti, A.; Crespi, M.

    2018-04-01

    The glaciers are a natural global resource and one of the principal climate change indicator at global and local scale, being influenced by temperature and snow precipitation changes. Among the parameters used for glacier monitoring, the surface velocity is a key element, since it is connected to glaciers changes (mass balance, hydro balance, glaciers stability, landscape erosion). The leading idea of this work is to continuously retrieve glaciers surface velocity using free ESA Sentinel-1 SAR imagery and exploiting the potentialities of the Google Earth Engine (GEE) platform. GEE has been recently released by Google as a platform for petabyte-scale scientific analysis and visualization of geospatial datasets. The algorithm of SAR off-set tracking developed at the Geodesy and Geomatics Division of the University of Rome La Sapienza has been integrated in a cloud based platform that automatically processes large stacks of Sentinel-1 data to retrieve glacier surface velocity field time series. We processed about 600 Sentinel-1 image pairs to obtain a continuous time series of velocity field measurements over 3 years from January 2015 to January 2018 for two wide glaciers located in the Northern Patagonian Ice Field (NPIF), the San Rafael and the San Quintin glaciers. Several results related to these relevant glaciers also validated with respect already available and renown software (i.e. ESA SNAP, CIAS) and with respect optical sensor measurements (i.e. LANDSAT8), highlight the potential of the Big Data analysis to automatically monitor glacier surface velocity fields at global scale, exploiting the synergy between GEE and Sentinel-1 imagery.

  20. Modeling Shear Induced Von Willebrand Factor Binding to Collagen

    Science.gov (United States)

    Dong, Chuqiao; Wei, Wei; Morabito, Michael; Webb, Edmund; Oztekin, Alparslan; Zhang, Xiaohui; Cheng, Xuanhong

    2017-11-01

    Von Willebrand factor (vWF) is a blood glycoprotein that binds with platelets and collagen on injured vessel surfaces to form clots. VWF bioactivity is shear flow induced: at low shear, binding between VWF and other biological entities is suppressed; for high shear rate conditions - as are found near arterial injury sites - VWF elongates, activating its binding with platelets and collagen. Based on parameters derived from single molecule force spectroscopy experiments, we developed a coarse-grain molecular model to simulate bond formation probability as a function of shear rate. By introducing a binding criterion that depends on the conformation of a sub-monomer molecular feature of our model, the model predicts shear-induced binding, even for conditions where binding is highly energetically favorable. We further investigate the influence of various model parameters on the ability to predict shear-induced binding (vWF length, collagen site density and distribution, binding energy landscape, and slip/catch bond length) and demonstrate parameter ranges where the model provides good agreement with existing experimental data. Our results may be important for understanding vWF activity and also for achieving targeted drug therapy via biomimetic synthetic molecules. National Science Foundation (NSF),Division of Mathematical Sciences (DMS).

  1. Focal cortical hypoperfusion in corticobasal degeneration demonstrated by three-dimensional surface display with {sup 123}I-IMP: a possible cause of apraxia

    Energy Technology Data Exchange (ETDEWEB)

    Okuda, B. [5. Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya (Japan); Tachibana, H. [5. Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya (Japan); Takeda, M. [5. Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya (Japan); Kawabata, K. [5. Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya (Japan); Sugita, M. [5. Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya (Japan); Fukuchi, M. [Department of Nuclear Medicine, Hyogo College of Medicine, Nishinomiya (Japan)

    1995-11-01

    To clarify cortical lesions responsible for apraxia in corticobasal degeneration (CBD), we reconstructed three-dimensional surface images from single-photon emission computed tomography (SPECT) data with N-isopropyl-p[I-123]-iodoamphetamine in two patients with CBD. Both had limb-kinetic apraxia (LKA) and one also had constructional apraxia (CA). Both showed asymmetrical cortical hypoperfusion in the perirolandic area. The patient with CA had unilateral hypoperfusion in the posterior parietal area. Thus, cortical hypoperfusion in the perirolandic area corresponded to LKA, and that in the posterior parietal area to CA. (orig.). With 4 figs.

  2. Focal cortical hypoperfusion in corticobasal degeneration demonstrated by three-dimensional surface display with 123I-IMP: a possible cause of apraxia

    International Nuclear Information System (INIS)

    Okuda, B.; Tachibana, H.; Takeda, M.; Kawabata, K.; Sugita, M.; Fukuchi, M.

    1995-01-01

    To clarify cortical lesions responsible for apraxia in corticobasal degeneration (CBD), we reconstructed three-dimensional surface images from single-photon emission computed tomography (SPECT) data with N-isopropyl-p[I-123]-iodoamphetamine in two patients with CBD. Both had limb-kinetic apraxia (LKA) and one also had constructional apraxia (CA). Both showed asymmetrical cortical hypoperfusion in the perirolandic area. The patient with CA had unilateral hypoperfusion in the posterior parietal area. Thus, cortical hypoperfusion in the perirolandic area corresponded to LKA, and that in the posterior parietal area to CA. (orig.). With 4 figs

  3. Binding Properties of Streptococcus gordonii SspA and SspB (Antigen I/II Family) Polypeptides Expressed on the Cell Surface of Lactococcus lactis MG1363

    OpenAIRE

    Holmes, Ann R.; Gilbert, Christophe; Wells, Jeremy M.; Jenkinson, Howard F.

    1998-01-01

    The oral bacterium Streptococcus gordonii expresses two cell wall-associated polypeptides, designated SspA (1,542 amino acid residues) and SspB (1,462 amino acid residues), that have 70% sequence identity. These polypeptides are members of the antigen I/II family of oral streptococcal adhesins and mediate the binding of streptococci to salivary glycoproteins, collagen, and other oral microorganisms such as Actinomyces naeslundii. To determine if SspA and SspB have differential binding propert...

  4. Evaluation of a solid-phase RIA technique and a solid-phase ELISA technique for demonstrating hepatitis-B surface antigen

    International Nuclear Information System (INIS)

    Vranckx, R.; Cole, J.; Peetermans, M.

    1978-01-01

    The sensitivities of a solid-phase radioimmunoassay (RIA), a solid-phase enzyme immunoassay (ELISA) and a haemagglutination test (RPHA) for the detection of the hepatitis-B surface antigen (HBsAg) were compared (1) by screening a panel of 300 sera (97 positives and 203 negatives), and (2) by titrating serial dilutions of 10 positive sera. Ninety-seven sera were positive by RIA, 95% were detected by ELISA and 81% were detected by RPHA. In the serial dilutions, the average end-points of the titrations were 0.005ng/ml for RIA, 0.01ng/ml for ELISA and 0.04 ng/ml for RPHA. It can be concluded that the sensitivity of the ELISA test is intermediate between that of the RIA and the RPHA. The ELISA and the RPHA tests seem to be a little more sensitive for the detection of subtype ay than for the detection of subtype ad. (author)

  5. Demonstration of a III-nitride vertical-cavity surface-emitting laser with a III-nitride tunnel junction intracavity contact

    International Nuclear Information System (INIS)

    Leonard, J. T.; Young, E. C.; Yonkee, B. P.; Cohen, D. A.; Margalith, T.; Speck, J. S.; DenBaars, S. P.; Nakamura, S.

    2015-01-01

    We report on a III-nitride vertical-cavity surface-emitting laser (VCSEL) with a III-nitride tunnel junction (TJ) intracavity contact. The violet nonpolar VCSEL employing the TJ is compared to an equivalent VCSEL with a tin-doped indium oxide (ITO) intracavity contact. The TJ VCSEL shows a threshold current density (J th ) of ∼3.5 kA/cm 2 , compared to the ITO VCSEL J th of 8 kA/cm 2 . The differential efficiency of the TJ VCSEL is also observed to be significantly higher than that of the ITO VCSEL, reaching a peak power of ∼550 μW, compared to ∼80 μW for the ITO VCSEL. Both VCSELs display filamentary lasing in the current aperture, which we believe to be predominantly a result of local variations in contact resistance, which may induce local variations in refractive index and free carrier absorption. Beyond the analyses of the lasing characteristics, we discuss the molecular-beam epitaxy (MBE) regrowth of the TJ, as well as its unexpected performance based on band-diagram simulations. Furthermore, we investigate the intrinsic advantages of using a TJ intracavity contact in a VCSEL using a 1D mode profile analysis to approximate the threshold modal gain and general loss contributions in the TJ and ITO VCSEL

  6. Demonstration of a III-nitride vertical-cavity surface-emitting laser with a III-nitride tunnel junction intracavity contact

    Energy Technology Data Exchange (ETDEWEB)

    Leonard, J. T., E-mail: jtleona01@gmail.com; Young, E. C.; Yonkee, B. P.; Cohen, D. A.; Margalith, T.; Speck, J. S. [Materials Department, University of California, Santa Barbara, California 93106 (United States); DenBaars, S. P.; Nakamura, S. [Materials Department, University of California, Santa Barbara, California 93106 (United States); Department of Electrical and Computer Engineering, University of California, Santa Barbara, California 93106 (United States)

    2015-08-31

    We report on a III-nitride vertical-cavity surface-emitting laser (VCSEL) with a III-nitride tunnel junction (TJ) intracavity contact. The violet nonpolar VCSEL employing the TJ is compared to an equivalent VCSEL with a tin-doped indium oxide (ITO) intracavity contact. The TJ VCSEL shows a threshold current density (J{sub th}) of ∼3.5 kA/cm{sup 2}, compared to the ITO VCSEL J{sub th} of 8 kA/cm{sup 2}. The differential efficiency of the TJ VCSEL is also observed to be significantly higher than that of the ITO VCSEL, reaching a peak power of ∼550 μW, compared to ∼80 μW for the ITO VCSEL. Both VCSELs display filamentary lasing in the current aperture, which we believe to be predominantly a result of local variations in contact resistance, which may induce local variations in refractive index and free carrier absorption. Beyond the analyses of the lasing characteristics, we discuss the molecular-beam epitaxy (MBE) regrowth of the TJ, as well as its unexpected performance based on band-diagram simulations. Furthermore, we investigate the intrinsic advantages of using a TJ intracavity contact in a VCSEL using a 1D mode profile analysis to approximate the threshold modal gain and general loss contributions in the TJ and ITO VCSEL.

  7. Enhanced binding capacity of boronate affinity adsorbent via surface modification of silica by combination of atom transfer radical polymerization and chain-end functionalization for high-efficiency enrichment of cis-diol molecules

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Wei; He, Maofang; Wang, Chaozhan; Wei, Yinmao, E-mail: ymwei@nwu.edu.cn

    2015-07-30

    Boronate affinity materials have been widely used for specific separation and preconcentration of cis-diol molecules, but most do not have sufficient capacity due to limited binding sites on the material surface. In this work, we prepared a phenylboronic acid-functionalized adsorbent with a high binding capacity via the combination of surface-initiated atom transfer radical polymerization (SI-ATRP) and chain-end functionalization. With this method, the terminal chlorides of the polymer chains were used fully, and the proposed adsorbent contains dense boronic acid polymers chain with boronic acid on the chain end. Consequently, the proposed adsorbent possesses excellent selectivity and a high binding capacity of 513.6 μmol g{sup −1} for catechol and 736.8 μmol g{sup −1} for fructose, which are much higher than those of other reported adsorbents. The dispersed solid-phase extraction (dSPE) based on the prepared adsorbent was used for extraction of three cis-diol drugs (i.e., epinephrine, isoprenaline and caffeic acid isopropyl ester) from plasma; the eluates were analyzed by HPLC-UV. The reduced amount of adsorbent (i.e., 2.0 mg) could still eliminate interferences efficiently and yielded a recovery range of 85.6–101.1% with relative standard deviations ranging from 2.5 to 9.7% (n = 5). The results indicated that the proposed strategy could serve as a promising alternative to increase the density of surface functional groups on the adsorbent; thus, the prepared adsorbent has the potential to effectively enrich cis-diol substances in real samples. - Highlights: • Boronate adsorbent is prepared via ATRP and chain-end functionalization. • The adsorbent has quite high binding capacity for cis-diols. • Binding capacity is easily manipulated by ATRP condition. • Chain-end functionalization can improve binding capacity significantly. • Reduced adsorbent is consumed in dispersed solid-phase extraction of cis-diols.

  8. Operational strategy for soil concentration predictions of strontium/yttrium-90 and cesium-137 in surface soil at the West Valley Demonstration Project site

    International Nuclear Information System (INIS)

    Myers, J.A.

    1995-01-01

    There are difficulties associated with the assessment of the interpretation of field measurements, determination of guideline protocols and control and disposal of low level radioactive contaminated soil in the environmental health physics field. Questions are raised among scientists and in public forums concerning the necessity and high costs of large area soil remediation versus the risks of low-dose radiation health effects. As a result, accurate soil activity assessments become imperative in decontamination situations. The West Valley Demonstration Project (WVDP), a US Department of Energy facility located in West Valley, New York is managed and operated by West Valley Nuclear Services Co., Inc. (WVNS). WVNS has identified contaminated on-site soil areas with a mixed variety of radionuclides (primarily fission product). Through the use of data obtained from a previous project performed during the summer of 1994 entitled ''Field Survey Correlation and Instrumentation Response for an In Situ Soil Measurement Program'' (Myers), the WVDP offers a unique research opportunity to investigate the possibility of soil concentration predictions based on exposure or count rate responses returned from a survey detector probe. In this study, correlations are developed between laboratory measured soil beta activity and survey probe response for the purposes of determining the optimal detector for field use and using these correlations to establish predictability of soil activity levels

  9. An insulin receptor mutant (Asp707 → Ala), involved in leprechaunism, is processed and transported to the cell surface but unable to bind insulin

    NARCIS (Netherlands)

    L.M. 't Hart (Leen); D. Lindhout (Dick); G.C.M. van der Zon (Gerard); H. Kayserilli (Hülya); M.Y. Apak (Memnune); W.J. Kleijer (Wim); E.R. van der Vorm (Eric); J.A. Maassen (Johannes)

    1996-01-01

    textabstractWe have identified a homozygous mutation near the carboxyl terminus of the insulin receptor (IR) α subunit from a leprechaun patient, changing Asp707 into Ala. Fibroblasts from this patient had no high affinity insulin binding sites. To examine the effect of the mutation on IR

  10. Further biochemical characterization of Mycobacterium leprae laminin-binding proteins

    Directory of Open Access Journals (Sweden)

    M.A.M. Marques

    2001-04-01

    Full Text Available It has been demonstrated that the alpha2 chain of laminin-2 present on the surface of Schwann cells is involved in the process of attachment of Mycobacterium leprae to these cells. Searching for M. leprae laminin-binding molecules, in a previous study we isolated and characterized the cationic proteins histone-like protein (Hlp and ribosomal proteins S4 and S5 as potential adhesins involved in M. leprae-Schwann cell interaction. Hlp was shown to bind alpha2-laminins and to greatly enhance the attachment of mycobacteria to ST88-14 Schwann cells. In the present study, we investigated the laminin-binding capacity of the ribosomal proteins S4 and S5. The genes coding for these proteins were PCR amplified and their recombinant products were shown to bind alpha2-laminins in overlay assays. However, when tested in ELISA-based assays and in adhesion assays with ST88-14 cells, in contrast to Hlp, S4 and S5 failed to bind laminin and act as adhesins. The laminin-binding property and adhesin capacity of two basic host-derived proteins were also tested, and only histones, but not cytochrome c, were able to increase bacterial attachment to ST88-14 cells. Our data suggest that the alanine/lysine-rich sequences shared by Hlp and eukaryotic H1 histones might be involved in the binding of these cationic proteins to laminin.

  11. The productive cellulase binding capacity of cellulosic substrates.

    Science.gov (United States)

    Karuna, Nardrapee; Jeoh, Tina

    2017-03-01

    Cellulosic biomass is the most promising feedstock for renewable biofuel production; however, the mechanisms of the heterogeneous cellulose saccharification reaction are still unsolved. As cellulases need to bind isolated molecules of cellulose at the surface of insoluble cellulose fibrils or larger aggregated cellulose structures in order to hydrolyze glycosidic bonds, the "accessibility of cellulose to cellulases" is considered to be a reaction limiting property of cellulose. We have defined the accessibility of cellulose to cellulases as the productive binding capacity of cellulose, that is, the concentration of productive binding sites on cellulose that are accessible for binding and hydrolysis by cellulases. Productive cellulase binding to cellulose results in hydrolysis and can be quantified by measuring hydrolysis rates. In this study, we measured the productive Trichoderma reesei Cel7A (TrCel7A) binding capacity of five cellulosic substrates from different sources and processing histories. Swollen filter paper and bacterial cellulose had higher productive binding capacities of ∼6 µmol/g while filter paper, microcrystalline cellulose, and algal cellulose had lower productive binding capacities of ∼3 µmol/g. Swelling and regenerating filter paper using phosphoric acid increased the initial accessibility of the reducing ends to TrCel7A from 4 to 6 µmol/g. Moreover, this increase in initial productive binding capacity accounted in large part for the difference in the overall digestibility between filter paper and swollen filter paper. We further demonstrated that an understanding of how the productive binding capacity declines over the course of the hydrolysis reaction has the potential to predict overall saccharification time courses. Biotechnol. Bioeng. 2017;114: 533-542. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  12. Phe783, Thr797, and Asp804 in transmembrane hairpin M5-M6 of Na+,K+-ATPase play a key role in ouabain binding.

    NARCIS (Netherlands)

    Qiu, L.Y.; Koenderink, J.B.; Swarts, H.G.P.; Willems, P.H.G.M.; Pont, J.J.H.H.M. de

    2003-01-01

    Ouabain is a glycoside that binds to and inhibits the action of Na+,K+-ATPase. Little is known, however, about the specific requirements of the protein surface for glycoside binding. Using chimeras of gastric H+,K+-ATPase and Na+,K+-ATPase, we demonstrated previously that the combined presence of

  13. Identification of an hexapeptide that binds to a surface pocket in cyclin A and inhibits the catalytic activity of the complex cyclin-dependent kinase 2-cyclin A.

    Science.gov (United States)

    Canela, Núria; Orzáez, Mar; Fucho, Raquel; Mateo, Francesca; Gutierrez, Ricardo; Pineda-Lucena, Antonio; Bachs, Oriol; Pérez-Payá, Enrique

    2006-11-24

    The protein-protein complexes formed between different cyclins and cyclin-dependent kinases (CDKs) are central to cell cycle regulation. These complexes represent interesting points of chemical intervention for the development of antineoplastic molecules. Here we describe the identification of an all d-amino acid hexapeptide, termed NBI1, that inhibits the kinase activity of the cyclin-dependent kinase 2 (cdk2)-cyclin A complex through selective binding to cyclin A. The mechanism of inhibition is non-competitive for ATP and non-competitive for protein substrates. In contrast to the existing CDKs peptide inhibitors, the hexapeptide NBI1 interferes with the formation of the cdk2-cyclin A complex. Furthermore, a cell-permeable derivative of NBI1 induces apoptosis and inhibits proliferation of tumor cell lines. Thus, the NBI1-binding site on cyclin A may represent a new target site for the selective inhibition of activity cdk2-cyclin A complex.

  14. Clearance and binding of radiolabeled glycoproteins by cells of the murine mononuclear phagocyte system

    International Nuclear Information System (INIS)

    Imber, M.J.

    1982-01-01

    The clearance and binding of radiolabeled lactoferrin and fast α 2 -macroglobulin were studied. Both glycoproteins cleared rapidly following intravenous injection in mice, and both bound specifically to discrete receptors on murine peritoneal macrophages. The simultaneous presence of excess, unlabeled ligands specific for receptors recognizing terminal fucose, mannose, N-acetylglucosamine or galactose residues did not inhibit the clearance or binding of either lactoferrin or fast-α 2 M. The clearance and binding of enzymatically defucosylated lactoferrin was indistinguishable from native lactoferrin, indicating that terminal α(1-3)-linked fucose on lactoferrin is not necessary for receptor recognition. The clearance and binding of two fast -α 2 M forms, α 2 M-trypsin and α 2 M-MeNH 2 cross compete with each other. Saturation binding studies indicated that the total binding of mannosyl -BSA, fusocyl-BSA, and N-acetylglucosaminyl-BSA to macrophages activated by BCG was approximately 15% of the levels observed with inflammatory macrophages elicited by thioglycollate broth. Cross-competition binding studies demonstrated a common surface receptor mediated binding of all three neoglycoprotein ligands and was identical to the receptor on mononuclear phagocytes that binds mannosyl- and N-acetylglucosaminyl-terminated glycoproteins. These results suggest that difference between discrete states of macrophage function may be correlated with selective changes in levels of the surface receptor for mannose-containing glycoproteins

  15. Crystal Structures and Binding Dynamics of Odorant-Binding Protein 3 from two aphid species Megoura viciae and Nasonovia ribisnigri.

    Science.gov (United States)

    Northey, Tom; Venthur, Herbert; De Biasio, Filomena; Chauviac, Francois-Xavier; Cole, Ambrose; Ribeiro, Karlos Antonio Lisboa; Grossi, Gerarda; Falabella, Patrizia; Field, Linda M; Keep, Nicholas H; Zhou, Jing-Jiang

    2016-04-22

    Aphids use chemical cues to locate hosts and find mates. The vetch aphid Megoura viciae feeds exclusively on the Fabaceae, whereas the currant-lettuce aphid Nasonovia ribisnigri alternates hosts between the Grossulariaceae and Asteraceae. Both species use alarm pheromones to warn of dangers. For N. ribisnigri this pheromone is a single component (E)-β-farnesene but M. viciae uses a mixture of (E)-β-farnesene, (-)-α-pinene, β-pinene, and limonene. Odorant-binding proteins (OBP) are believed to capture and transport such semiochemicals to their receptors. Here, we report the first aphid OBP crystal structures and examine their molecular interactions with the alarm pheromone components. Our study reveals some unique structural features: 1) the lack of an internal ligand binding site; 2) a striking groove in the surface of the proteins as a putative binding site; 3) the N-terminus rather than the C-terminus occupies the site closing off the conventional OBP pocket. The results from fluorescent binding assays, molecular docking and dynamics demonstrate that OBP3 from M. viciae can bind to all four alarm pheromone components and the differential ligand binding between these very similar OBP3s from the two aphid species is determined mainly by the direct π-π interactions between ligands and the aromatic residues of OBP3s in the binding pocket.

  16. Mapping of monoclonal antibody- and receptor-binding domains on human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) using a surface plasmon resonance-based biosensor.

    Science.gov (United States)

    Laricchia-Robbio, L; Liedberg, B; Platou-Vikinge, T; Rovero, P; Beffy, P; Revoltella, R P

    1996-10-01

    An automated surface plasmon resonance (SPR)-based biosensor system has been used for mapping antibody and receptor-binding regions on the recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) molecule. A rabbit antimouse IgG1-Fc antibody (RAM.Fc) was coupled to an extended carboxymethylated-hydrogel matrix attached to a gold surface in order to capture an anti-rhGM-CSF monoclonal antibody (MAb) injected over the sensing layer. rhGM-CSF was subsequently injected and allowed to bind to this antibody. Multisite binding assays were then performed, by flowing sequentially other antibodies and peptides over the surface, and the capacity of the latter to interact with the entrapped rhGM-CSF in a multimolecular complex was monitored in real time with SPR. Eleven MAb (all IgG1K), were analyzed: respectively, four antipeptide MAb raised against three distinct epitopes of the cytokine (two clones against residues 14-24, that includes part of the first alpha-helix toward the N-terminal region; one clone against peptide 30-41, an intrahelical loop; and one clone against residues 79-91, including part of the third alpha-helix) and seven antiprotein MAbs raised against the entire rhGM-CSF, whose target native epitopes are still undetermined. In addition, the binding capacity to rhGM-CSF of a synthetic peptide, corresponding to residues 238-254 of the extracellular human GM-CSF receptor alpha-chain, endowed with rhGM-CSF binding activity, was tested. The results from experiments performed with the biosensor were compared with those obtained by a sandwich enzyme-linked immunosorbent assay (ELISA), using the same reagents. The features of the biosensor technology (fully automated, measure in real time, sharpened yes/no response, less background disturbances, no need for washing step or labeling of the reagent) offered several advantages in these studies of MAb immunoreactivity and epitope mapping, giving a much better resolution and enabling more distinct

  17. Mechanistic positron emission tomography studies: demonstration of a deuterium isotope effect in the monoamine oxidase-catalyzed binding of (/sup 11/C)L-deprenyl in living baboon brain

    Energy Technology Data Exchange (ETDEWEB)

    Fowler, J.S.; Wolf, A.P.; MacGregor, R.R.; Dewey, S.L.; Logan, J.; Schlyer, D.J.; Langstrom, B.

    1988-11-01

    The application of positron emission tomography (PET) to the study of biochemical transformations in the living human and animal body requires the development of highly selective radiotracers whose concentrations in tissue provide a record of a discrete metabolic process. L-N-(11C-methyl)Deprenyl ((11C)L-deprenyl), a suicide inactivator of monoamine oxidase (MAO) type B, has been developed as a radiotracer for mapping MAO B in the living human and animal brain. In this investigation, (11C)L-deprenyl (1) and (11C)L-deprenyl-alpha, alpha-2H2 (2) have been compared in three different baboons by PET measurement of carbon-11 uptake and retention in the brain and the measurement of the amount of unchanged tracer in the arterial plasma over a 90-min time interval. For one baboon, N-(11C-methyl-2H3)L-deprenyl (3) was also studied. Kinetic parameters calculated using a three-compartment model revealed a deuterium isotope effect of 3.8 +/- 1.1. Comparison of the two tracers (1 and 2) in mouse brain demonstrated that deuterium substitution significantly reduced the amount of radioactivity bound to protein. HPLC and GLC analysis of the soluble radioactivity in mouse brain after injection of (11C)L-deprenyl showed the presence of (11C)methamphetamine as a major product along with unidentified labeled products. Sodium dodecyl sulfate-polyacrylamide electrophoresis with carbon-14-labeled L-deprenyl showed that a protein of molecular weight 58,000 was labeled. These results establish that MAO-catalyzed cleavage of the alpha carbon-hydrogen bond on the propargyl group is the rate limiting (or a major rate contributing) step in the retention of carbon-11 in brain and that the in vivo detection of labeled products in brain after the injection of (11C)L-deprenyl provides a record of MAO activity.

  18. Surface display of the receptor-binding region of the Lactobacillus brevis S-layer protein in Lactococcus lactis provides nonadhesive lactococci with the ability to adhere to intestinal epithelial cells.

    Science.gov (United States)

    Avall-Jääskeläinen, Silja; Lindholm, Agneta; Palva, Airi

    2003-04-01

    Lactobacillus brevis is a promising lactic acid bacterium for use as a probiotic dietary adjunct and a vaccine vector. The N-terminal region of the S-layer protein (SlpA) of L. brevis ATCC 8287 was recently shown to mediate adhesion to various human cell lines in vitro. In this study, a surface display cassette was constructed on the basis of this SlpA receptor-binding domain, a proteinase spacer, and an autolysin anchor. The cassette was expressed under control of the nisA promoter in Lactococcus lactis NZ9000. Western blot assay of lactococcal cell wall extracts with anti-SlpA antibodies confirmed that the SlpA adhesion domain of the fusion protein was expressed and located within the cell wall layer. Whole-cell enzyme-linked immunosorbent assay and immunofluorescence microscopy verified that the SlpA adhesion-mediating region was accessible on the lactococcal cell surface. In vitro adhesion assays with the human intestinal epithelial cell line Intestine 407 indicated that the recombinant lactococcal cells had gained an ability to adhere to Intestine 407 cells significantly greater than that of wild-type L. lactis NZ9000. Serum inhibition assay further confirmed that adhesion of recombinant lactococci to Intestine 407 cells was indeed mediated by the N terminus-encoding part of the slpA gene. The ability of the receptor-binding region of SlpA to adhere to fibronectin was also confirmed with this lactococcal surface display system. These results show that, with the aid of the receptor-binding region of the L. brevis SlpA protein, the ability to adhere to gut epithelial cells can indeed be transferred to another, nonadhesive, lactic acid bacterium.

  19. Surface Display of the Receptor-Binding Region of the Lactobacillus brevis S-Layer Protein in Lactococcus lactis Provides Nonadhesive Lactococci with the Ability To Adhere to Intestinal Epithelial Cells

    OpenAIRE

    Åvall-Jääskeläinen, Silja; Lindholm, Agneta; Palva, Airi

    2003-01-01

    Lactobacillus brevis is a promising lactic acid bacterium for use as a probiotic dietary adjunct and a vaccine vector. The N-terminal region of the S-layer protein (SlpA) of L. brevis ATCC 8287 was recently shown to mediate adhesion to various human cell lines in vitro. In this study, a surface display cassette was constructed on the basis of this SlpA receptor-binding domain, a proteinase spacer, and an autolysin anchor. The cassette was expressed under control of the nisA promoter in Lactoc...

  20. A transferrin-like GPI-linked iron-binding protein in detergent-insoluble noncaveolar microdomains at the apical surface of fetal intestinal epithelial cells

    DEFF Research Database (Denmark)

    Danielsen, E M; van Deurs, B

    1995-01-01

    of ultracryosections of mucosal tissue, the protein was localized to the apical surface of the enterocytes, whereas it was absent from the basolateral plasma membrane. Interestingly, it was mainly found in patches of flat or invaginated apical membrane domains rather than at the surface of microvilli. Caveolae were...

  1. Binding of collagens to an enterotoxigenic strain of Escherichia coli

    International Nuclear Information System (INIS)

    Visai, L.; Speziale, P.; Bozzini, S.

    1990-01-01

    An enterotoxigenic strain of Escherichia coli, B34289c, has been shown to bind the N-terminal region of fibronectin with high affinity. We now report that this strain also binds collagen. The binding of 125I-labeled type II collagen to bacteria was time dependent and reversible. Bacteria expressed a limited number of collagen receptors (2.2 x 10(4) per cell) and bound collagen with a Kd of 20 nM. All collagen types tested (I to V) as well as all tested cyanogen bromide-generated peptides [alpha 1(I)CB2, alpha 1(I)CB3, alpha 1(I)CB7, alpha 1(I)CB8, and alpha 2(I)CB4] were recognized by bacterial receptors, as demonstrated by the ability of these proteins to inhibit the binding of 125I-labeled collagen to bacteria. Of several unlabeled proteins tested in competition experiments, fibronectin and its N-terminal region strongly inhibited binding of the radiolabeled collagen to E. coli cells. Conversely, collagen competed with an 125I-labeled 28-kilodalton fibronectin fragment for bacterial binding. Collagen bound to bacteria could be displaced by excess amounts of either unlabeled fibronectin or its N-terminal fragment. Similarly, collagen could displace 125I-labeled N-terminal peptide of fibronectin bound to the bacterial cell surface. Bacteria grown at 41 degrees C or in the presence of glucose did not express collagen or fibronectin receptors. These results indicate the presence of specific binding sites for collagen on the surface of E. coli cells and furthermore that the collagen and fibronectin binding sites are located in close proximity, possibly on the same structure

  2. Electrodynamic Dust Shield Demonstrator

    Science.gov (United States)

    Stankie, Charles G.

    2013-01-01

    The objective of the project was to design and manufacture a device to demonstrate a new technology developed by NASA's Electrostatics and Surface Physics Laboratory. The technology itself is a system which uses magnetic principles to remove regolith dust from its surface. This project was to create an enclosure that will be used to demonstrate the effectiveness of the invention to The Office of the Chief Technologist. ONE of the most important challenges of space exploration is actually caused by something very small and seemingly insignificant. Dust in space, most notably on the moon and Mars, has caused many unforeseen issues. Dirt and dust on Earth, while a nuisance, can be easily cleaned and kept at bay. However, there is considerably less weathering and erosion in space. As a result, the microscopic particles are extremely rough and abrasive. They are also electrostatically charged, so they cling to everything they make contact with. This was first noted to be a major problem during the Apollo missions. Dust would stick to the spacesuits, and could not be wiped off as predicted. Dust was brought back into the spacecraft, and was even inhaled by astronauts. This is a major health hazard. Atmospheric storms and other events can also cause dust to coat surfaces of spacecraft. This can cause abrasive damage to the craft. The coating can also reduce the effectiveness of thermal insulation and solar panels.' A group of engineers at Kennedy Space Center's Electrostatics and Surface Physics Laboratory have developed a new technology, called the Electrodynamic Dust Shield, to help alleviate these problems. It is based off of the electric curtain concept developed at NASA in 1967. "The EDS is an active dust mitigation technology that uses traveling electric fields to transport electrostatically charged dust particles along surfaces. To generate the traveling electric fields, the EDS consists of a multilayer dielectric coating with an embedded thin electrode grid

  3. Demonstration of HITEX

    International Nuclear Information System (INIS)

    Morrison, H.D.; Woodall, K.B.

    1993-01-01

    A model reactor for HITEX successfully demonstrated the concept of high-temperature isotopic exchange in a closed loop simulating the conditions for fusion fuel cleanup. The catalyst of platinum on alumina pellets provided a surface area large enough to operate the reactor at 400 degrees celsius with flow rates up to 2 L/min. A 15-L tank containing a mixture of 4% CD 4 in H 2 was depleted in deuterium within 75 minutes down to 100 ppm HD above the natural concentration of HD in the make-up hydrogen stream. The application to tritium removal from tritiated impurities in a hydrogen stream will work as well or better

  4. Development of bisphenol A-removing recombinant Escherichia coli by monomeric and dimeric surface display of bisphenol A-binding peptide.

    Science.gov (United States)

    Maruthamuthu, Murali Kannan; Hong, Jiyeon; Arulsamy, Kulandaisamy; Somasundaram, Sivachandiran; Hong, SoonHo; Choe, Woo-Seok; Yoo, Ik-Keun

    2018-04-01

    Peptide-displaying Escherichia coli cells were investigated for use in adsorptive removal of bisphenol A (BPA) both in Luria-Bertani medium including BPA or ATM thermal paper eluted wastewater. Two recombinant strains were constructed with monomeric and dimeric repeats of the 7-mer BPA-binding peptide (KSLENSY), respectively. Greater than threefold increased adsorption of BPA [230.4 µmol BPA per g dry cell weight (DCW)] was found in dimeric peptide-displaying cells compared to monomeric strains (63.4 µmol per g DCW) in 15 ppm BPA solution. The selective removal of BPA from a mixture of BPA analogs (bisphenol F and bisphenol S) was verified in both monomeric and dimeric peptide-displaying cells. The binding chemistry of BPA with the peptide was assumed, based on molecular docking analysis, to be the interaction of BPA with serine and asparagine residues within the 7-mer peptide sequence. The peptide-displaying cells also functioned efficiently in thermal paper eluted wastewater containing 14.5 ppm BPA.

  5. Chemical Force Spectroscopy Evidence Supporting the Layer-by-Layer Model of Organic Matter Binding to Iron (oxy)Hydroxide Mineral Surfaces

    KAUST Repository

    Chassé , Alexander W.; Ohno, Tsutomu; Higgins, Steven R.; Amirbahman, Aria; Yildirim, Nadir; Parr, Thomas B.

    2015-01-01

    © 2015 American Chemical Society. The adsorption of dissolved organic matter (DOM) to metal (oxy)hydroxide mineral surfaces is a critical step for C sequestration in soils. Although equilibrium studies have described some of the factors controlling this process, the molecular-scale description of the adsorption process has been more limited. Chemical force spectroscopy revealed differing adhesion strengths of DOM extracted from three soils and a reference peat soil material to an iron (oxy)hydroxide mineral surface. The DOM was characterized using ultrahigh-resolution negative ion mode electrospray ionization Fourier Transform ion cyclotron resonance mass spectrometry. The results indicate that carboxyl-rich aromatic and N-containing aliphatic molecules of DOM are correlated with high adhesion forces. Increasing molecular mass was shown to decrease the adhesion force between the mineral surface and the DOM. Kendrick mass defect analysis suggests that mechanisms involving two carboxyl groups result in the most stable bond to the mineral surface. We conceptualize these results using a layer-by-layer "onion" model of organic matter stabilization on soil mineral surfaces.

  6. Chemical Force Spectroscopy Evidence Supporting the Layer-by-Layer Model of Organic Matter Binding to Iron (oxy)Hydroxide Mineral Surfaces

    KAUST Repository

    Chassé, Alexander W.

    2015-08-18

    © 2015 American Chemical Society. The adsorption of dissolved organic matter (DOM) to metal (oxy)hydroxide mineral surfaces is a critical step for C sequestration in soils. Although equilibrium studies have described some of the factors controlling this process, the molecular-scale description of the adsorption process has been more limited. Chemical force spectroscopy revealed differing adhesion strengths of DOM extracted from three soils and a reference peat soil material to an iron (oxy)hydroxide mineral surface. The DOM was characterized using ultrahigh-resolution negative ion mode electrospray ionization Fourier Transform ion cyclotron resonance mass spectrometry. The results indicate that carboxyl-rich aromatic and N-containing aliphatic molecules of DOM are correlated with high adhesion forces. Increasing molecular mass was shown to decrease the adhesion force between the mineral surface and the DOM. Kendrick mass defect analysis suggests that mechanisms involving two carboxyl groups result in the most stable bond to the mineral surface. We conceptualize these results using a layer-by-layer "onion" model of organic matter stabilization on soil mineral surfaces.

  7. Antioxidant mechanism of milk mineral-high-affinity iron binding.

    Science.gov (United States)

    Allen, K; Cornforth, D

    2007-01-01

    Milk mineral (MM), a by-product of whey processing, is an effective antioxidant in meat systems, but the antioxidant mechanism has not been established. MM has been postulated to chelate iron and prevent iron-catalysis of lipid oxidation. The objective of this research was to examine this putative mechanism. MM was compared to sodium tripolyphosphate (STPP), calcium phosphate monobasic (CPM), and calcium pyrophosphate (CPP) to determine iron-binding capacity, sample solubility, and eluate soluble phosphorus after treating samples with a ferrous chloride standard. Scanning electron microscopy with energy-dispersive X-ray analysis was used to localize minerals on iron-treated MM particle surfaces. Histochemical staining for calcium was performed on raw and cooked ground beef samples with added MM. MM bound more iron per gram (P compounds, and was much less soluble (P iron across the MM particle surface, directly demonstrating iron binding to MM particles. Unlike other common chelating agents, such as STPP and citrate, histochemical staining demonstrated that MM remained insoluble in ground beef, even after cooking. The ability of MM to bind iron and remain insoluble may enhance its antioxidant effect by removing iron ions from solution. However, MM particles must be small and well distributed in order to adequately bind iron throughout the food system.

  8. Waste and Disposal: Demonstration

    International Nuclear Information System (INIS)

    Neerdael, B.; Buyens, M.; De Bruyn, D.; Volckaert, G.

    2002-01-01

    Within the Belgian R and D programme on geological disposal, demonstration experiments have become increasingly important. In this contribution to the scientific report 2001, an overview is given of SCK-CEN's activities and achievements in the field of large-scale demonstration experiments. In 2001, main emphasis was on the PRACLAY project, which is a large-scale experiment to demonstrate the construction and the operation of a gallery for the disposal of HLW in a clay formation. The PRACLAY experiment will contribute to enhance understanding of water flow and mass transport in dense clay-based materials as well as to improve the design of the reference disposal concept. In the context of PRACLAY, a surface experiment (OPHELIE) has been developed to prepare and to complement PRACLAY-related experimental work in the HADES Underground Research Laboratory. In 2001, efforts were focussed on the operation of the OPHELIE mock-up. SCK-CEN also contributed to the SELFRAC roject which studies the self-healing of fractures in a clay formation

  9. Distinct DNA-binding surfaces in the ATPase and linker domains of MutLγ determine its substrate specificities and exert separable functions in meiotic recombination and mismatch repair.

    Directory of Open Access Journals (Sweden)

    Corentin Claeys Bouuaert

    2017-05-01

    Full Text Available Mlh1-Mlh3 (MutLγ is a mismatch repair factor with a central role in formation of meiotic crossovers, presumably through resolution of double Holliday junctions. MutLγ has DNA-binding, nuclease, and ATPase activities, but how these relate to one another and to in vivo functions are unclear. Here, we combine biochemical and genetic analyses to characterize Saccharomyces cerevisiae MutLγ. Limited proteolysis and atomic force microscopy showed that purified recombinant MutLγ undergoes ATP-driven conformational changes. In vitro, MutLγ displayed separable DNA-binding activities toward Holliday junctions (HJ and, surprisingly, single-stranded DNA (ssDNA, which was not predicted from current models. MutLγ bound DNA cooperatively, could bind multiple substrates simultaneously, and formed higher-order complexes. FeBABE hydroxyl radical footprinting indicated that the DNA-binding interfaces of MutLγ for ssDNA and HJ substrates only partially overlap. Most contacts with HJ substrates were located in the linker regions of MutLγ, whereas ssDNA contacts mapped within linker regions as well as the N-terminal ATPase domains. Using yeast genetic assays for mismatch repair and meiotic recombination, we found that mutations within different DNA-binding surfaces exert separable effects in vivo. For example, mutations within the Mlh1 linker conferred little or no meiotic phenotype but led to mismatch repair deficiency. Interestingly, mutations in the N-terminal domain of Mlh1 caused a stronger meiotic defect than mlh1Δ, suggesting that the mutant proteins retain an activity that interferes with alternative recombination pathways. Furthermore, mlh3Δ caused more chromosome missegregation than mlh1Δ, whereas mlh1Δ but not mlh3Δ partially alleviated meiotic defects of msh5Δ mutants. These findings illustrate functional differences between Mlh1 and Mlh3 during meiosis and suggest that their absence impinges on chromosome segregation not only via reduced

  10. Surfactant protein D binds to human immunodeficiency virus (HIV) envelope protein gp120 and inhibits HIV replication

    DEFF Research Database (Denmark)

    Meschi, Joseph; Crouch, Erika C; Skolnik, Paul

    2005-01-01

    The envelope protein (gp120) of human immunodeficiency virus (HIV) contains highly conserved mannosylated oligosaccharides. These glycoconjugates contribute to resistance to antibody neutralization, and binding to cell surface lectins on macrophages and dendritic cells. Mannose-binding lectin (MBL......) binds to gp120 and plays a role in defence against the virus. In this study it is demonstrated that surfactant protein D (SP-D) binds to gp120 and inhibits HIV infectivity at significantly lower concentrations than MBL. The binding of SP-D was mediated by its calcium-dependent carbohydrate......-binding activity and was dependent on glycosylation of gp120. Native dodecameric SP-D bound to HIV gp120 more strongly than native trimeric SP-D. Since one common polymorphic form of SP-D is predominantly expressed as trimers and associated with lower blood levels, these individuals may have less effective innate...

  11. The neck region of the C-type lectin DC-SIGN regulates its surface spatiotemporal organization and virus-binding capacity on antigen presenting cells

    NARCIS (Netherlands)

    Manzo, C.; Torreno-Pina, J.A.; Joosten, B.; Reinieren-Beeren, I.; Gualda, E.J.; Loza-Alvarez, P.; Figdor, Carl; Garcia Parajo, M.F.; Cambi, A.

    2012-01-01

    The C-type lectin DC-SIGN expressed on dendritic cells (DCs) facilitates capture and internalization of a plethora of different pathogens. Although it is known that DC-SIGN organizes in nanoclusters at the surface of DCs, the molecular mechanisms responsible for this well defined nanopatterning and

  12. The Neck Region of the C-type Lectin DC-SIGN Regulates Its Surface Spatiotemporal Organization and Virus-binding Capacity on Antigen-presenting Cells

    NARCIS (Netherlands)

    Manzo, C.; Torreno-Pina, J.A.; Joosten, B.; Reinieren-Beeren, I.; Gualda, E.J.; Loza-Alvarez, P.; Figdor, C.G.; Garcia-Parajo, M.F.; Cambi, A.

    2012-01-01

    The C-type lectin DC-SIGN expressed on dendritic cells (DCs) facilitates capture and internalization of a plethora of different pathogens. Although it is known that DC-SIGN organizes in nanoclusters at the surface of DCs, the molecular mechanisms responsible for this well defined nanopatterning and

  13. Origin of the Substitution Mechanism for the Binding of Organic Ligands on the Surface of CsPbBr3 Perovskite Nanocubes.

    Science.gov (United States)

    Ravi, Vikash Kumar; Santra, Pralay K; Joshi, Niharika; Chugh, Jeetender; Singh, Sachin Kumar; Rensmo, Håkan; Ghosh, Prasenjit; Nag, Angshuman

    2017-10-19

    Optoelectronic properties of CsPbBr 3 perovskite nanocubes (NCs) depend strongly on the interaction of the organic passivating molecules with the inorganic crystal. To understand this interaction, we employed a combination of synchrotron-based X-ray photoelectron spectroscopy (XPS), nuclear magnetic resonance (NMR) spectroscopy, and first-principles density functional theory (DFT)-based calculations. Variable energy XPS elucidated the internal structure of the inorganic part in a layer-by-layer fashion, whereas NMR characterized the organic ligands. Our experimental results confirm that oleylammonium ions act as capping ligands by substituting Cs + ions from the surface of CsPbBr 3 NCs. DFT calculations shows that the substitution mechanism does not require much energy for surface reconstruction and, in contrast, stabilizes the nanocrystal by the formation of three hydrogen bonds between the -NH 3 + moiety of oleylammonium and surrounding Br - on the surface of NCs. This substitution mechanism and its origin are in stark contrast to the usual adsorption of organic ligands on the surface of typical NCs.

  14. Dual Function of Novel Pollen Coat (Surface) Proteins: IgE-binding Capacity and Proteolytic Activity Disrupting the Airway Epithelial Barrier

    Science.gov (United States)

    Bashir, Mohamed Elfatih H.; Ward, Jason M.; Cummings, Matthew; Karrar, Eltayeb E.; Root, Michael; Mohamed, Abu Bekr A.; Naclerio, Robert M.; Preuss, Daphne

    2013-01-01

    Background The pollen coat is the first structure of the pollen to encounter the mucosal immune system upon inhalation. Prior characterizations of pollen allergens have focused on water-soluble, cytoplasmic proteins, but have overlooked much of the extracellular pollen coat. Due to washing with organic solvents when prepared, these pollen coat proteins are typically absent from commercial standardized allergenic extracts (i.e., “de-fatted”), and, as a result, their involvement in allergy has not been explored. Methodology/Principal Findings Using a unique approach to search for pollen allergenic proteins residing in the pollen coat, we employed transmission electron microscopy (TEM) to assess the impact of organic solvents on the structural integrity of the pollen coat. TEM results indicated that de-fatting of Cynodon dactylon (Bermuda grass) pollen (BGP) by use of organic solvents altered the structural integrity of the pollen coat. The novel IgE-binding proteins of the BGP coat include a cysteine protease (CP) and endoxylanase (EXY). The full-length cDNA that encodes the novel IgE-reactive CP was cloned from floral RNA. The EXY and CP were purified to homogeneity and tested for IgE reactivity. The CP from the BGP coat increased the permeability of human airway epithelial cells, caused a clear concentration-dependent detachment of cells, and damaged their barrier integrity. Conclusions/Significance Using an immunoproteomics approach, novel allergenic proteins of the BGP coat were identified. These proteins represent a class of novel dual-function proteins residing on the coat of the pollen grain that have IgE-binding capacity and proteolytic activity, which disrupts the integrity of the airway epithelial barrier. The identification of pollen coat allergens might explain the IgE-negative response to available skin-prick-testing proteins in patients who have positive symptoms. Further study of the role of these pollen coat proteins in allergic responses is

  15. Dual function of novel pollen coat (surface proteins: IgE-binding capacity and proteolytic activity disrupting the airway epithelial barrier.

    Directory of Open Access Journals (Sweden)

    Mohamed Elfatih H Bashir

    Full Text Available BACKGROUND: The pollen coat is the first structure of the pollen to encounter the mucosal immune system upon inhalation. Prior characterizations of pollen allergens have focused on water-soluble, cytoplasmic proteins, but have overlooked much of the extracellular pollen coat. Due to washing with organic solvents when prepared, these pollen coat proteins are typically absent from commercial standardized allergenic extracts (i.e., "de-fatted", and, as a result, their involvement in allergy has not been explored. METHODOLOGY/PRINCIPAL FINDINGS: Using a unique approach to search for pollen allergenic proteins residing in the pollen coat, we employed transmission electron microscopy (TEM to assess the impact of organic solvents on the structural integrity of the pollen coat. TEM results indicated that de-fatting of Cynodon dactylon (Bermuda grass pollen (BGP by use of organic solvents altered the structural integrity of the pollen coat. The novel IgE-binding proteins of the BGP coat include a cysteine protease (CP and endoxylanase (EXY. The full-length cDNA that encodes the novel IgE-reactive CP was cloned from floral RNA. The EXY and CP were purified to homogeneity and tested for IgE reactivity. The CP from the BGP coat increased the permeability of human airway epithelial cells, caused a clear concentration-dependent detachment of cells, and damaged their barrier integrity. CONCLUSIONS/SIGNIFICANCE: Using an immunoproteomics approach, novel allergenic proteins of the BGP coat were identified. These proteins represent a class of novel dual-function proteins residing on the coat of the pollen grain that have IgE-binding capacity and proteolytic activity, which disrupts the integrity of the airway epithelial barrier. The identification of pollen coat allergens might explain the IgE-negative response to available skin-prick-testing proteins in patients who have positive symptoms. Further study of the role of these pollen coat proteins in allergic

  16. Binding of (/sup 3/H)imipramine to human platelet membranes with compensation for saturable binding to filters and its implication for binding studies with brain membranes

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, O.M.; Wood, K.M.; Williams, D.C.

    1984-08-01

    Apparent specific binding of (/sup 3/H)imipramine to human platelet membranes at high concentrations of imipramine showed deviation from that expected of a single binding site, a result consistent with a low-affinity binding site. The deviation was due to displaceable, saturable binding to the glass fibre filters used in the assays. Imipramine, chloripramine, desipramine, and fluoxetine inhibited binding to filters whereas 5-hydroxytryptamine and ethanol were ineffective. Experimental conditions were developed that eliminated filter binding, allowing assay of high- and low-affinity binding to membranes. Failure to correct for filter binding may lead to overestimation of binding parameters, Bmax and KD for high-affinity binding to membranes, and may also be misinterpreted as indicating a low-affinity binding component in both platelet and brain membranes. Low-affinity binding (KD less than 2 microM) of imipramine to human platelet membranes was demonstrated and its significance discussed.

  17. Detection of Serum Lysophosphatidic Acids Using Affinity Binding and Surface Enhanced Laser Desorption/Ionization (SELDI) Time of Flight Mass Spectrometry

    Science.gov (United States)

    2005-04-01

    sensitivity to cisplatin (the most effective drug in ovarian cancer), and increases production and activity of multiple growth factors, proteases and...clinical grade assay. The detection approach of cocrystalizing LPA and Zn is an improvement of the approach proposed here and will be implemented in our...surface receptors for lysophospholipids belong to the G protein coupled receptor family. As over 40% of all drugs in current use target this family of

  18. Adsorption and covalent binding of fibrinogen as a method for probing the chemical composition of poly(styrene/α-tert-butoxy-ω-vinylbenzyl-polyglycidol) microsphere surfaces.

    Science.gov (United States)

    Gosecka, Monika; Chehimi, Mohamed M; Basinska, Teresa; Slomkowski, Stanislaw; Makowski, Tomasz

    2017-12-01

    We investigated the distribution of polyglycidol and polystyrene on the surface of poly(styrene/α-tert-butoxy-ω-vinylbenzyl-polyglycidol) microspheres (random distribution or segregated into hydrophilic and hydrophobic patches), using fibrinogen (Fb) as a macromolecular probe. The fibrinogen was adsorbed or covalently attached to the surface of the poly(styrene-co-α-tert-butoxy-ω-vinylbenzyl-polyglycidol) (P(S/PGLy)) microspheres. The P(S/PGLy) particles were prepared by emulsion copolymerization of styrene and α-tert-butoxy-ω-vinylbenzyl-polyglycidol (PGLy) macromonomer initiated with potassium persulfate. The polymerizations yielded P(S/PGLy) particles with various surface fractions of polyglycidol, depending on the amount of added macromonomer and the addition process. In some syntheses, the entire macromonomer amount was added once at the beginning of the polymerization, while in others, the macromonomer was added gradually after the formation of particle seeds from pure polystyrene. XPS studies revealed that the fraction of polyglycidol in the interfacial layer of the microspheres was larger when the entire amount of macromonomer was added at the beginning of the polymerization than when it was added after formation of the polystyrene seeds. Studies of fibrinogen adsorption provided the first evidence of segregation of the hydrophobic (polystyrene) and hydrophilic (polyglycidol) components at the surface of the composite P(S/PGLy) microspheres into patches. The hydrophobic patches are composed mainly of polystyrene. However, they also contain a small amount of polyglycidol chains, making the adsorption of fibrinogen weaker than the adsorption onto the pure polystyrene. Studies of covalent immobilization of fibrinogen on the microspheres via 1,3,5-trichlorotriazine confirmed these findings. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Regular square planer bis-(4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione)/copper(II) complex: Trans/cis-DFT isomerization, crystal structure, thermal, solvatochromism, hirshfeld surface and DNA-binding analysis

    Science.gov (United States)

    Hema, M. K.; Karthik, C. S.; Warad, Ismail; Lokanath, N. K.; Zarrouk, Abdelkader; Kumara, Karthik; Pampa, K. J.; Mallu, P.

    2018-04-01

    Trans-[Cu(O∩O)2] complex, O∩O = 4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione was reported with high potential toward CT-DNA binder. The solved XRD-structure of complex indicated a perfect regular square-planer geometry around the Cu(II) center. The trans/cis-DFT-isomerization calculation supported the XRD seen in reflecting the trans-isomer as the kinetic-favor isomer. The desired complex structure was also characterized by conductivity measurement, CHN-elemental analyses, MS, EDX, SEM, UV-Vis., FT-IR, HAS and TG/DTG. The Solvatochromism behavior of the complex was evaluated using four different polar solvents. MPE and Hirshfeld surface analysis (HSA) come to an agreement that fluoride and thiophene protons atoms are with suitable electro-potential environment to form non-classical H-bonds of type CThsbnd H⋯F. The DNA-binding properties were investigated by viscosity tests and spectrometric titrations, the results revealed the complex as strong calf-thymus DNA binder. High intrinsic-binding constants value ∼1.8 × 105 was collected.

  20. Benzodiazepines have high-affinity binding sites and induce melanogenesis in B16/C3 melanoma cells.

    OpenAIRE

    Matthew, E; Laskin, J D; Zimmerman, E A; Weinstein, I B; Hsu, K C; Engelhardt, D L

    1981-01-01

    We found that two markers of differentiation, tyrosinase (monophenol, dihydroxyphenylalanine:oxygen oxidoreductase, EC 1.14.18.1) activity and melanin synthesis, are induced by diazepam in B16/C3 mouse melanoma cells. We also demonstrated high-affinity binding sites for [3H]diazepam in these cells by radioreceptor assay, and we visualized binding to the cell surface by fluorescence microscopy with a benzodiazepine analog conjugated to a fluorescein-labeled protein. Our studies also showed tha...

  1. Exploring the Structure of Nitrogen-Rich Ionic Liquids and Their Binding to the Surface of Oxide-Free Boron Nanoparticles

    Science.gov (United States)

    2013-01-29

    results have to be interpreted with caution. Lovelock et al.61 have shown that clean surfaces can be obtained by sputtering for the [CnC1Im][Tf2N] IL...Licence, P. Langmuir 2006, 22, 9386−9392. (58) Hurisso, B. B.; Lovelock , K. R.; Licence, P. Phys. Chem. Chem. Phys. 2011, 13, 17737−48. (59) Zhang, H.; Cui...H. Langmuir 2009, 25, 2604−2612. (60) Zhang, Q.; Ma, X.; Liu, S.; Yang, B.; Lu, L.; He, Y.; Deng, Y. J. Mater. Chem. 2011, 21, 6864. (61) Lovelock , K

  2. Distribution of cyclophilin B-binding sites in the subsets of human peripheral blood lymphocytes.

    Science.gov (United States)

    Denys, A; Allain, F; Foxwell, B; Spik, G

    1997-08-01

    Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein, mainly associated with the secretory pathway and released in biological fluids. We have recently demonstrated that both free CyPB and CyPB-CsA complex specifically bind to peripheral blood T lymphocytes and are internalized. These results suggest that CyPB might promote the targeting of the drug into sensitive cells. Peripheral blood lymphocytes are subdivided in several populations according to their biological functions and sensitivity to CsA. We have investigated the binding of CyPB to these different subsets using a CyPB derivatized by fluorescein through its single cysteine which retains its binding properties. We have confirmed that only T cells were involved in the interaction with CyPB. The ligand binding was found to be heterogeneously distributed on the different T-cell subsets and surface-bound CyPB was mainly associated with the CD4-positive cells. No significant difference was noted between the CD45RA and CD45RO subsets, demonstrating that CyPB-binding sites were equally distributed between native and memory T cells. CD3 stimulation of T lymphocytes led to a decrease in the CyPB-binding capacity, that may be explained by a down-regulation of the CyPB-receptor expression upon T-cell activation. Finally, we demonstrated that CyPB-receptor-positive cells, isolated on CyPB sulphydryl-coupled affinity matrices, are more sensitive to CyPB-complexed CsA than mixed peripheral blood lymphocytes, suggesting that CyPB potentiates CsA activity through the binding of the complex. Taken together, our results demonstrate that CyPB-binding sites are mainly associated with resting cells of the helper T lymphocyte, and that CyPB might modulate the distribution of CsA through the drug targeting to sensitive cells.

  3. The agmatine-containing poly(amidoamine) polymer AGMA1 binds cell surface heparan sulfates and prevents attachment of mucosal human papillomaviruses.

    Science.gov (United States)

    Cagno, Valeria; Donalisio, Manuela; Bugatti, Antonella; Civra, Andrea; Cavalli, Roberta; Ranucci, Elisabetta; Ferruti, Paolo; Rusnati, Marco; Lembo, David

    2015-09-01

    The agmatine-containing poly(amidoamine) polymer AGMA1 was recently shown to inhibit the infectivity of several viruses, including human papillomavirus 16 (HPV-16), that exploit cell surface heparan sulfate proteoglycans (HSPGs) as attachment receptors. The aim of this work was to assess the antiviral activity of AGMA1 and its spectrum of activity against a panel of low-risk and high-risk HPVs and to elucidate its mechanism of action. AGMA1 was found to be a potent inhibitor of mucosal HPV types (i.e., types 16, 31, 45, and 6) in pseudovirus-based neutralization assays. The 50% inhibitory concentration was between 0.34 μg/ml and 0.73 μg/ml, and no evidence of cytotoxicity was observed. AGMA1 interacted with immobilized heparin and with cellular heparan sulfates, exerting its antiviral action by preventing virus attachment to the cell surface. The findings from this study indicate that AGMA1 is a leading candidate compound for further development as an active ingredient of a topical microbicide against HPV and other sexually transmitted viral infections. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  4. Protein sequences bound to mineral surfaces persist into deep time

    DEFF Research Database (Denmark)

    Demarchi, Beatrice; Hall, Shaun; Roncal-Herrero, Teresa

    2016-01-01

    of Laetoli (3.8 Ma) and Olduvai Gorge (1.3 Ma) in Tanzania. By tracking protein diagenesis back in time we find consistent patterns of preservation, demonstrating authenticity of the surviving sequences. Molecular dynamics simulations of struthiocalcin-1 and -2, the dominant proteins within the eggshell......, reveal that distinct domains bind to the mineral surface. It is the domain with the strongest calculated binding energy to the calcite surface that is selectively preserved. Thermal age calculations demonstrate that the Laetoli and Olduvai peptides are 50 times older than any previously authenticated...

  5. Surface-Layer (S-Layer) Proteins Sap and EA1 Govern the Binding of the S-Layer-Associated Protein BslO at the Cell Septa of Bacillus anthracis

    Science.gov (United States)

    Kern, Valerie J.; Kern, Justin W.; Theriot, Julie A.; Schneewind, Olaf

    2012-01-01

    The Gram-positive pathogen Bacillus anthracis contains 24 genes whose products harbor the structurally conserved surface-layer (S-layer) homology (SLH) domain. Proteins endowed with the SLH domain associate with the secondary cell wall polysaccharide (SCWP) following secretion. Two such proteins, Sap and EA1, have the unique ability to self-assemble into a paracrystalline layer on the surface of bacilli and form S layers. Other SLH domain proteins can also be found within the S layer and have been designated Bacillus S-layer-associated protein (BSLs). While both S-layer proteins and BSLs bind the same SCWP, their deposition on the cell surface is not random. For example, BslO is targeted to septal peptidoglycan zones, where it catalyzes the separation of daughter cells. Here we show that an insertional lesion in the sap structural gene results in elongated chains of bacilli, as observed with a bslO mutant. The chain length of the sap mutant can be reduced by the addition of purified BslO in the culture medium. This complementation in trans can be explained by an increased deposition of BslO onto the surface of sap mutant bacilli that extends beyond chain septa. Using fluorescence microscopy, we observed that the Sap S layer does not overlap the EA1 S layer and slowly yields to the EA1 S layer in a growth-phase-dependent manner. Although present all over bacilli, Sap S-layer patches are not observed at septa. Thus, we propose that the dynamic Sap/EA1 S-layer coverage of the envelope restricts the deposition of BslO to the SCWP at septal rings. PMID:22609927

  6. SP-A binding sites on bovine alveolar macrophages.

    Science.gov (United States)

    Plaga, S; Plattner, H; Schlepper-Schaefer, J

    1998-11-25

    Surfactant protein A (SP-A) binding to bovine alveolar macrophages was examined in order to characterize SP-A binding proteins on the cell surface and to isolate putative receptors from these cells that could be obtained in large amounts. Human SP-A, unlabeled or labeled with gold particles, was bound to freshly isolated macrophages and analyzed with ELISA or the transmission electron microscope. Binding of SP-A was inhibited by Ca2+ chelation, by an excess of unlabeled SP-A, or by the presence of 20 mg/ml mannan. We conclude that bovine alveolar macrophages expose binding sites for SP-A that are specific and that depend on Ca2+ and on mannose residues. For isolation of SP-A receptors with homologous SP-A as ligand we isolated SP-A from bovine lung lavage. SDS-PAGE analysis of the purified SP-A showed a protein of 32-36 kDa. Functional integrity of the protein was demonstrated. Bovine SP-A bound to Dynabeads was used to isolate SP-A binding proteins. From the fractionated and blotted proteins of the receptor preparation two proteins bound SP-A in a Ca2+-dependent manner, a 40-kDa protein showing mannose dependency and a 210-kDa protein, showing no mannose sensitivity. Copyright 1998 Academic Press.

  7. Characterization of salivary alpha-amylase binding to Streptococcus sanguis

    International Nuclear Information System (INIS)

    Scannapieco, F.A.; Bergey, E.J.; Reddy, M.S.; Levine, M.J.

    1989-01-01

    The purpose of this study was to identify the major salivary components which interact with oral bacteria and to determine the mechanism(s) responsible for their binding to the bacterial surface. Strains of Streptococcus sanguis, Streptococcus mitis, Streptococcus mutans, and Actinomyces viscosus were incubated for 2 h in freshly collected human submandibular-sublingual saliva (HSMSL) or parotid saliva (HPS), and bound salivary components were eluted with 2% sodium dodecyl sulfate. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western transfer, alpha-amylase was the prominent salivary component eluted from S. sanguis. Studies with 125 I-labeled HSMSL or 125 I-labeled HPS also demonstrated a component with an electrophoretic mobility identical to that of alpha-amylase which bound to S. sanguis. Purified alpha-amylase from human parotid saliva was radiolabeled and found to bind to strains of S. sanguis genotypes 1 and 3 and S. mitis genotype 2, but not to strains of other species of oral bacteria. Binding of [ 125 I]alpha-amylase to streptococci was saturable, calcium independent, and inhibitable by excess unlabeled alpha-amylases from a variety of sources, but not by secretory immunoglobulin A and the proline-rich glycoprotein from HPS. Reduced and alkylated alpha-amylase lost enzymatic and bacterial binding activities. Binding was inhibited by incubation with maltotriose, maltooligosaccharides, limit dextrins, and starch

  8. Decorin binds myostatin and modulates its activity to muscle cells

    International Nuclear Information System (INIS)

    Miura, Takayuki; Kishioka, Yasuhiro; Wakamatsu, Jun-ichi; Hattori, Akihito; Hennebry, Alex; Berry, Carole J.; Sharma, Mridula; Kambadur, Ravi; Nishimura, Takanori

    2006-01-01

    Myostatin, a member of TGF-β superfamily of growth factors, acts as a negative regulator of skeletal muscle mass. The mechanism whereby myostatin controls the proliferation and differentiation of myogenic cells is mostly clarified. However, the regulation of myostatin activity to myogenic cells after its secretion in the extracellular matrix (ECM) is still unknown. Decorin, a small leucine-rich proteoglycan, binds TGF-β and regulates its activity in the ECM. Thus, we hypothesized that decorin could also bind to myostatin and participate in modulation of its activity to myogenic cells. In order to test the hypothesis, we investigated the interaction between myostatin and decorin by surface plasmon assay. Decorin interacted with mature myostatin in the presence of concentrations of Zn 2+ greater than 10 μM, but not in the absence of Zn 2+ . Kinetic analysis with a 1:1 binding model resulted in dissociation constants (K D ) of 2.02 x 10 -8 M and 9.36 x 10 -9 M for decorin and the core protein of decorin, respectively. Removal of the glycosaminoglycan chain by chondroitinase ABC digestion did not affect binding, suggesting that decorin could bind to myostatin with its core protein. Furthermore, we demonstrated that immobilized decorin could rescue the inhibitory effect of myostatin on myoblast proliferation in vitro. These results suggest that decorin could trap myostatin and modulate its activity to myogenic cells in the ECM

  9. Triazatriangulene as binding group for molecular electronics

    DEFF Research Database (Denmark)

    Wei, Zhongming; Wang, Xintai; Borges, Anders

    2014-01-01

    The triazatriangulene (TATA) ring system was investigated as a binding group for tunnel junctions of molecular wires on gold surfaces. Self-assembled monolayers (SAMs) of TATA platforms with three different lengths of phenylene wires were fabricated, and their electrical conductance was recorded ...... with its high stability and directionality make this binding group very attractive for molecular electronic measurements and devices. (Figure Presented)....

  10. Site-directed antibody immobilization using a protein A-gold binding domain fusion protein for enhanced SPR immunosensing.

    Science.gov (United States)

    de Juan-Franco, Elena; Caruz, Antonio; Pedrajas, J R; Lechuga, Laura M

    2013-04-07

    We have implemented a novel strategy for the oriented immobilization of antibodies onto a gold surface based on the use of a fusion protein, the protein A-gold binding domain (PAG). PAG consists of a gold binding peptide (GBP) coupled to the immunoglobulin-binding domains of staphylococcal protein A. This fusion protein provides an easy and fast oriented immobilization of antibodies preserving its native structure, while leaving the antigen binding sites (Fab) freely exposed. Using this immobilization strategy, we have demonstrated the performance of the immunosensing of the human Growth Hormone by SPR. A limit of detection of 90 ng mL(-1) was obtained with an inter-chip variability lower than 7%. The comparison of this method with other strategies for the direct immobilization of antibodies over gold surfaces has showed the enhanced sensitivity provided by the PAG approach.

  11. IGF binding proteins.

    Science.gov (United States)

    Bach, Leon A

    2017-12-18

    Insulin-like growth factor binding proteins (IGFBPs) 1-6 bind IGFs but not insulin with high affinity. They were initially identified as serum carriers and passive inhibitors of IGF actions. However, subsequent studies showed that, although IGFBPs inhibit IGF actions in many circumstances, they may also potentiate these actions. IGFBPs are widely expressed in most tissues, and they are flexible endocrine and autocrine/paracrine regulators of IGF activity, which is essential for this important physiological system. More recently, individual IGFBPs have been shown to have IGF-independent actions. Mechanisms underlying these actions include (i) interaction with non-IGF proteins in compartments including the extracellular space and matrix, the cell surface and intracellularly; (ii) interaction with and modulation of other growth factor pathways including EGF, TGF- and VEGF; and (iii) direct or indirect transcriptional effects following nuclear entry of IGFBPs. Through these IGF-dependent and IGF-independent actions, IGFBPs modulate essential cellular processes including proliferation, survival, migration, senescence, autophagy and angiogenesis. They have been implicated in a range of disorders including malignant, metabolic, neurological and immune diseases. A more complete understanding of their cellular roles may lead to the development of novel IGFBP-based therapeutic opportunities.

  12. The involvement of coordinative interactions in the binding of dihydrolipoamide dehydrogenase to titanium dioxide-Localization of a putative binding site.

    Science.gov (United States)

    Dayan, Avraham; Babin, Gilad; Ganoth, Assaf; Kayouf, Nivin Samir; Nitoker Eliaz, Neta; Mukkala, Srijana; Tsfadia, Yossi; Fleminger, Gideon

    2017-08-01

    Titanium (Ti) and its alloys are widely used in orthodontic and orthopedic implants by virtue to their high biocompatibility, mechanical strength, and high resistance to corrosion. Biointegration of the implants with the tissue requires strong interactions, which involve biological molecules, proteins in particular, with metal oxide surfaces. An exocellular high-affinity titanium dioxide (TiO 2 )-binding protein (TiBP), purified from Rhodococcus ruber, has been previously studied in our lab. This protein was shown to be homologous with the orthologous cytoplasmic rhodococcal dihydrolipoamide dehydrogenase (rhDLDH). We have found that rhDLDH and its human homolog (hDLDH) share the TiO 2 -binding capabilities with TiBP. Intrigued by the unique TiO 2 -binding properties of hDLDH, we anticipated that it may serve as a molecular bridge between Ti-based medical structures and human tissues. The objective of the current study was to locate the region and the amino acids of the protein that mediate the protein-TiO 2 surface interaction. We demonstrated the role of acidic amino acids in the nonelectrostatic enzyme/dioxide interactions at neutral pH. The observation that the interaction of DLDH with various metal oxides is independent of their isoelectric values strengthens this notion. DLDH does not lose its enzymatic activity upon binding to TiO 2 , indicating that neither the enzyme undergoes major conformational changes nor the TiO 2 binding site is blocked. Docking predictions suggest that both rhDLDH and hDLDH bind TiO 2 through similar regions located far from the active site and the dimerization sites. The putative TiO 2 -binding regions of both the bacterial and human enzymes were found to contain a CHED (Cys, His, Glu, Asp) motif, which has been shown to participate in metal-binding sites in proteins. Copyright © 2017 John Wiley & Sons, Ltd.

  13. Biodiesel Mass Transit Demonstration

    Science.gov (United States)

    2010-04-01

    The Biodiesel Mass Transit Demonstration report is intended for mass transit decision makers and fleet managers considering biodiesel use. This is the final report for the demonstration project implemented by the National Biodiesel Board under a gran...

  14. Authoring Effective Demonstrations

    National Research Council Canada - National Science Library

    Fu, Dan; Jensen, Randy; Salas, Eduardo; Rosen, Michael A; Ramachandran, Sowmya; Upshaw, Christin L; Hinkelman, Elizabeth; Lampton, Don

    2007-01-01

    ... or human role-players for each training event. We report our ongoing efforts to (1) research the nature and purpose of demonstration, articulating guidelines for effective demonstration within a training context, and (2...

  15. Comparing Demonstratives in Kwa

    African Journals Online (AJOL)

    This paper is a comparative study of demonstrative forms in three K wa languages, ... relative distance from the deictic centre, such as English this and that, here and there. ... Mostly, the referents of demonstratives are 'activated' or at least.

  16. Polarized Light Corridor Demonstrations.

    Science.gov (United States)

    Davies, G. R.

    1990-01-01

    Eleven demonstrations of light polarization are presented. Each includes a brief description of the apparatus and the effect demonstrated. Illustrated are strain patterns, reflection, scattering, the Faraday Effect, interference, double refraction, the polarizing microscope, and optical activity. (CW)

  17. Fragment-based quantum mechanical calculation of protein-protein binding affinities.

    Science.gov (United States)

    Wang, Yaqian; Liu, Jinfeng; Li, Jinjin; He, Xiao

    2018-04-29

    The electrostatically embedded generalized molecular fractionation with conjugate caps (EE-GMFCC) method has been successfully utilized for efficient linear-scaling quantum mechanical (QM) calculation of protein energies. In this work, we applied the EE-GMFCC method for calculation of binding affinity of Endonuclease colicin-immunity protein complex. The binding free energy changes between the wild-type and mutants of the complex calculated by EE-GMFCC are in good agreement with experimental results. The correlation coefficient (R) between the predicted binding energy changes and experimental values is 0.906 at the B3LYP/6-31G*-D level, based on the snapshot whose binding affinity is closest to the average result from the molecular mechanics/Poisson-Boltzmann surface area (MM/PBSA) calculation. The inclusion of the QM effects is important for accurate prediction of protein-protein binding affinities. Moreover, the self-consistent calculation of PB solvation energy is required for accurate calculations of protein-protein binding free energies. This study demonstrates that the EE-GMFCC method is capable of providing reliable prediction of relative binding affinities for protein-protein complexes. © 2018 Wiley Periodicals, Inc. © 2018 Wiley Periodicals, Inc.

  18. Cyclophilin B binding to platelets supports calcium-dependent adhesion to collagen.

    Science.gov (United States)

    Allain, F; Durieux, S; Denys, A; Carpentier, M; Spik, G

    1999-08-01

    We have recently reported that cyclophilin B (CyPB), a secreted cyclosporine-binding protein, could bind to T lymphocytes through interactions with two types of binding sites. The first ones, referred to as type I, involve interactions with the conserved domain of CyPB and promote the endocytosis of surface-bound ligand, while the second type of binding sites, termed type II, are represented by glycosaminoglycans (GAG). Here, we further investigated the interactions of CyPB with blood cell populations. In addition to lymphocytes, CyPB was found to interact mainly with platelets. The binding is specific, with a dissociation constant (kd) of 9 +/- 3 nmol/L and the number of sites estimated at 960 +/- 60 per cell. Platelet glycosaminoglycans are not required for the interactions, but the binding is dramatically reduced by active cyclosporine derivatives. We then analyzed the biologic effects of CyPB and found a significant increase in platelet adhesion to collagen. Concurrently, CyPB initiates a transmembranous influx of Ca(2+) and induces the phosphorylation of the P-20 light chains of myosin. Taken together, the present results demonstrate for the first time that extracellular CyPB specifically interacts with platelets through a functional receptor related to the lymphocyte type I binding sites and might act by regulating the activity of a receptor-operated membrane Ca(2+) channel.

  19. Strategy Guideline: Demonstration Home

    Energy Technology Data Exchange (ETDEWEB)

    Savage, C.; Hunt, A.

    2012-12-01

    This guideline will provide a general overview of the different kinds of demonstration home projects, a basic understanding of the different roles and responsibilities involved in the successful completion of a demonstration home, and an introduction into some of the lessons learned from actual demonstration home projects. Also, this guideline will specifically look at the communication methods employed during demonstration home projects. And lastly, we will focus on how to best create a communication plan for including an energy efficient message in a demonstration home project and carry that message to successful completion.

  20. Strategy Guideline. Demonstration Home

    Energy Technology Data Exchange (ETDEWEB)

    Hunt, A.; Savage, C.

    2012-12-01

    This guideline will provide a general overview of the different kinds of demonstration home projects, a basic understanding of the different roles and responsibilities involved in the successful completion of a demonstration home, and an introduction into some of the lessons learned from actual demonstration home projects. Also, this guideline will specifically look at the communication methods employed during demonstration home projects. And lastly, we will focus on how to best create a communication plan for including an energy efficient message in a demonstration home project and carry that message to successful completion.

  1. Adaptive evolution of transcription factor binding sites

    Directory of Open Access Journals (Sweden)

    Berg Johannes

    2004-10-01

    Full Text Available Abstract Background The regulation of a gene depends on the binding of transcription factors to specific sites located in the regulatory region of the gene. The generation of these binding sites and of cooperativity between them are essential building blocks in the evolution of complex regulatory networks. We study a theoretical model for the sequence evolution of binding sites by point mutations. The approach is based on biophysical models for the binding of transcription factors to DNA. Hence we derive empirically grounded fitness landscapes, which enter a population genetics model including mutations, genetic drift, and selection. Results We show that the selection for factor binding generically leads to specific correlations between nucleotide frequencies at different positions of a binding site. We demonstrate the possibility of rapid adaptive evolution generating a new binding site for a given transcription factor by point mutations. The evolutionary time required is estimated in terms of the neutral (background mutation rate, the selection coefficient, and the effective population size. Conclusions The efficiency of binding site formation is seen to depend on two joint conditions: the binding site motif must be short enough and the promoter region must be long enough. These constraints on promoter architecture are indeed seen in eukaryotic systems. Furthermore, we analyse the adaptive evolution of genetic switches and of signal integration through binding cooperativity between different sites. Experimental tests of this picture involving the statistics of polymorphisms and phylogenies of sites are discussed.

  2. Multiple binding modes of ibuprofen in human serum albumin identified by absolute binding free energy calculations

    KAUST Repository

    Evoli, Stefania

    2016-11-10

    Human serum albumin possesses multiple binding sites and transports a wide range of ligands that include the anti-inflammatory drug ibuprofen. A complete map of the binding sites of ibuprofen in albumin is difficult to obtain in traditional experiments, because of the structural adaptability of this protein in accommodating small ligands. In this work, we provide a set of predictions covering the geometry, affinity of binding and protonation state for the pharmaceutically most active form (S-isomer) of ibuprofen to albumin, by using absolute binding free energy calculations in combination with classical molecular dynamics (MD) simulations and molecular docking. The most favorable binding modes correctly reproduce several experimentally identified binding locations, which include the two Sudlow\\'s drug sites (DS2 and DS1) and the fatty acid binding sites 6 and 2 (FA6 and FA2). Previously unknown details of the binding conformations were revealed for some of them, and formerly undetected binding modes were found in other protein sites. The calculated binding affinities exhibit trends which seem to agree with the available experimental data, and drastically degrade when the ligand is modeled in a protonated (neutral) state, indicating that ibuprofen associates with albumin preferentially in its charged form. These findings provide a detailed description of the binding of ibuprofen, help to explain a wide range of results reported in the literature in the last decades, and demonstrate the possibility of using simulation methods to predict ligand binding to albumin.

  3. Characterization of a Mutant Diphtheria Toxin that is Defective in Binding to Cell Membrane Receptors on Vero Cells

    Science.gov (United States)

    1982-08-13

    pinocytlc activity was demonstrated by the Increase in lysosomal vesicles ( acid phosphatase -positive vesicles) (4, 13). Poly-L-ornithine increased... wheat germ agglutinin and the protection was reversed by a-methly- mannoslde and N-acetylglucosamlne, respectively. These studies suggested that the...on the cell surface were involved in the initial binding of toxin to cell surface receptors. Concanavalin A and wheat germ agglutinin Inhibited the

  4. Radioimmunological demonstration of DNA specific antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Falck, P [Akademie der Wissenschaften der DDR, Berlin-Buch. Zentralinstitut fuer Isotopen- und Strahlenforschung

    1976-01-01

    Using /sup 125/I chemically labelled denatured (d) and native (n) DNA, specifically binding antibodies were demonstrated in the sera of Lupus erythemathodes patients by means of the Farr technique. (NH/sub 4/)/sub 2/SO/sub 4/ was used to separate the immunologically bound /sup 125/I-d-DNA. For /sup 125/I-n-DNA the use of a secondary antiserum for the precipitation of the primary immune complex is advantageous. The influence of antigen concentration upon the binding rate was studied. Titre determinations can be made with the proposed method.

  5. Total iron binding capacity

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/003489.htm Total iron binding capacity To use the sharing features on this page, please enable JavaScript. Total iron binding capacity (TIBC) is a blood test to ...

  6. Binding characteristics of copper and cadmium by cyanobacterium Spirulina platensis

    International Nuclear Information System (INIS)

    Fang Linchuan; Zhou Chen; Cai Peng; Chen Wenli; Rong Xingmin; Dai Ke; Liang Wei; Gu Jidong; Huang Qiaoyun

    2011-01-01

    Highlights: → The carboxyl groups play a vital role in the binding of Cu(II) and Cd(II) to S. platensis cells. → Ion exchange and complexation are the dominating mechanism for Cu(II) and Cd(II) adsorption. → XAFS analysis provided evidence for the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface. - Abstract: Cyanobacteria are promising biosorbent for heavy metals in bioremediation. Although sequestration of metals by cyanobacteria is known, the actual mechanisms and ligands involved are not very well understood. The binding characteristics of Cu(II) and Cd(II) by the cyanobacterium Spirulina platensis were investigated using a combination of chemical modifications, batch adsorption experiments, Fourier transform infrared (FTIR) spectroscopy and X-ray absorption fine structure (XAFS) spectroscopy. A significant increase in Cu(II) and Cd(II) binding was observed in the range of pH 3.5-5.0. Dramatical decrease in adsorption of Cu(II) and Cd(II) was observed after methanol esterification of the nonliving cells demonstrating that carboxyl functional groups play an important role in the binding of metals by S. platensis. The desorption rate of Cu(II) and Cd(II) from S. platensis surface was 72.7-80.7% and 53.7-58.0% by EDTA and NH 4 NO 3 , respectively, indicating that ion exchange and complexation are the dominating mechanisms for Cu(II) and Cd(II) adsorption. XAFS analysis provided further evidence on the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface.

  7. Binding characteristics of copper and cadmium by cyanobacterium Spirulina platensis

    Energy Technology Data Exchange (ETDEWEB)

    Fang Linchuan [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Zhou Chen; Cai Peng [Key Laboratory of Subtropical Agricultural Resources and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China); Chen Wenli [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Rong Xingmin; Dai Ke; Liang Wei [Key Laboratory of Subtropical Agricultural Resources and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China); Gu Jidong [Department of Ecology and Biodiversity, University of Hong Kong, Pokfulam Road, Hong Kong (Hong Kong); Huang Qiaoyun, E-mail: qyhuang@mail.hzau.edu.cn [State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070 (China); Key Laboratory of Subtropical Agricultural Resources and Environment, Ministry of Agriculture, College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070 (China)

    2011-06-15

    Highlights: {yields} The carboxyl groups play a vital role in the binding of Cu(II) and Cd(II) to S. platensis cells. {yields} Ion exchange and complexation are the dominating mechanism for Cu(II) and Cd(II) adsorption. {yields} XAFS analysis provided evidence for the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface. - Abstract: Cyanobacteria are promising biosorbent for heavy metals in bioremediation. Although sequestration of metals by cyanobacteria is known, the actual mechanisms and ligands involved are not very well understood. The binding characteristics of Cu(II) and Cd(II) by the cyanobacterium Spirulina platensis were investigated using a combination of chemical modifications, batch adsorption experiments, Fourier transform infrared (FTIR) spectroscopy and X-ray absorption fine structure (XAFS) spectroscopy. A significant increase in Cu(II) and Cd(II) binding was observed in the range of pH 3.5-5.0. Dramatical decrease in adsorption of Cu(II) and Cd(II) was observed after methanol esterification of the nonliving cells demonstrating that carboxyl functional groups play an important role in the binding of metals by S. platensis. The desorption rate of Cu(II) and Cd(II) from S. platensis surface was 72.7-80.7% and 53.7-58.0% by EDTA and NH{sub 4}NO{sub 3}, respectively, indicating that ion exchange and complexation are the dominating mechanisms for Cu(II) and Cd(II) adsorption. XAFS analysis provided further evidence on the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface.

  8. Manufacturing Demonstration Facility (MDF)

    Data.gov (United States)

    Federal Laboratory Consortium — The U.S. Department of Energy Manufacturing Demonstration Facility (MDF) at Oak Ridge National Laboratory (ORNL) provides a collaborative, shared infrastructure to...

  9. Binding characteristics of copper and cadmium by cyanobacterium Spirulina platensis.

    Science.gov (United States)

    Fang, Linchuan; Zhou, Chen; Cai, Peng; Chen, Wenli; Rong, Xingmin; Dai, Ke; Liang, Wei; Gu, Ji-Dong; Huang, Qiaoyun

    2011-06-15

    Cyanobacteria are promising biosorbent for heavy metals in bioremediation. Although sequestration of metals by cyanobacteria is known, the actual mechanisms and ligands involved are not very well understood. The binding characteristics of Cu(II) and Cd(II) by the cyanobacterium Spirulina platensis were investigated using a combination of chemical modifications, batch adsorption experiments, Fourier transform infrared (FTIR) spectroscopy and X-ray absorption fine structure (XAFS) spectroscopy. A significant increase in Cu(II) and Cd(II) binding was observed in the range of pH 3.5-5.0. Dramatical decrease in adsorption of Cu(II) and Cd(II) was observed after methanol esterification of the nonliving cells demonstrating that carboxyl functional groups play an important role in the binding of metals by S. platensis. The desorption rate of Cu(II) and Cd(II) from S. platensis surface was 72.7-80.7% and 53.7-58.0% by EDTA and NH(4)NO(3), respectively, indicating that ion exchange and complexation are the dominating mechanisms for Cu(II) and Cd(II) adsorption. XAFS analysis provided further evidence on the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface. Copyright © 2011 Elsevier B.V. All rights reserved.

  10. Understanding TR binding to pMHC complexes: how does a TR scan many pMHC complexes yet preferentially bind to one.

    Directory of Open Access Journals (Sweden)

    Javed Mohammed Khan

    Full Text Available Understanding the basis of the binding of a T cell receptor (TR to the peptide-MHC (pMHC complex is essential due to the vital role it plays in adaptive immune response. We describe the use of computed binding (free energy (BE, TR paratope, pMHC epitope, molecular surface electrostatic potential (MSEP and calculated TR docking angle (θ to analyse 61 TR/pMHC crystallographic structures to comprehend TR/pMHC interaction. In doing so, we have successfully demonstrated a novel/rational approach for θ calculation, obtained a linear correlation between BE and θ without any "codon" or amino acid preference, provided an explanation for TR ability to scan many pMHC ligands yet specifically bind one, proposed a mechanism for pMHC recognition by TR leading to T cell activation and illustrated the importance of the peptide in determining TR specificity, challenging the "germline bias" theory.

  11. Leptospiral outer membrane protein microarray, a novel approach to identification of host ligand-binding proteins.

    Science.gov (United States)

    Pinne, Marija; Matsunaga, James; Haake, David A

    2012-11-01

    Leptospirosis is a zoonosis with worldwide distribution caused by pathogenic spirochetes belonging to the genus Leptospira. The leptospiral life cycle involves transmission via freshwater and colonization of the renal tubules of their reservoir hosts. Infection requires adherence to cell surfaces and extracellular matrix components of host tissues. These host-pathogen interactions involve outer membrane proteins (OMPs) expressed on the bacterial surface. In this study, we developed an Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 OMP microarray containing all predicted lipoproteins and transmembrane OMPs. A total of 401 leptospiral genes or their fragments were transcribed and translated in vitro and printed on nitrocellulose-coated glass slides. We investigated the potential of this protein microarray to screen for interactions between leptospiral OMPs and fibronectin (Fn). This approach resulted in the identification of the recently described fibronectin-binding protein, LIC10258 (MFn8, Lsa66), and 14 novel Fn-binding proteins, denoted Microarray Fn-binding proteins (MFns). We confirmed Fn binding of purified recombinant LIC11612 (MFn1), LIC10714 (MFn2), LIC11051 (MFn6), LIC11436 (MFn7), LIC10258 (MFn8, Lsa66), and LIC10537 (MFn9) by far-Western blot assays. Moreover, we obtained specific antibodies to MFn1, MFn7, MFn8 (Lsa66), and MFn9 and demonstrated that MFn1, MFn7, and MFn9 are expressed and surface exposed under in vitro growth conditions. Further, we demonstrated that MFn1, MFn4 (LIC12631, Sph2), and MFn7 enable leptospires to bind fibronectin when expressed in the saprophyte, Leptospira biflexa. Protein microarrays are valuable tools for high-throughput identification of novel host ligand-binding proteins that have the potential to play key roles in the virulence mechanisms of pathogens.

  12. Mechanism of selective VEGF-A binding by neuropilin-1 reveals a basis for specific ligand inhibition.

    Directory of Open Access Journals (Sweden)

    Matthew W Parker

    Full Text Available Neuropilin (Nrp receptors function as essential cell surface receptors for the Vascular Endothelial Growth Factor (VEGF family of proangiogenic cytokines and the semaphorin 3 (Sema3 family of axon guidance molecules. There are two Nrp homologues, Nrp1 and Nrp2, which bind to both overlapping and distinct members of the VEGF and Sema3 family of molecules. Nrp1 specifically binds the VEGF-A(164/5 isoform, which is essential for developmental angiogenesis. We demonstrate that VEGF-A specific binding is governed by Nrp1 residues in the b1 coagulation factor domain surrounding the invariant Nrp C-terminal arginine binding pocket. Further, we show that Sema3F does not display the Nrp-specific binding to the b1 domain seen with VEGF-A. Engineered soluble Nrp receptor fragments that selectively sequester ligands from the active signaling complex are an attractive modality for selectively blocking the angiogenic and chemorepulsive functions of Nrp ligands. Utilizing the information on Nrp ligand binding specificity, we demonstrate Nrp constructs that specifically sequester Sema3 in the presence of VEGF-A. This establishes that unique mechanisms are used by Nrp receptors to mediate specific ligand binding and that these differences can be exploited to engineer soluble Nrp receptors with specificity for Sema3.

  13. Minor Coat and Heat Shock Proteins Are Involved in the Binding of Citrus Tristeza Virus to the Foregut of Its Aphid Vector, Toxoptera citricida.

    Science.gov (United States)

    Killiny, N; Harper, S J; Alfaress, S; El Mohtar, C; Dawson, W O

    2016-11-01

    Vector transmission is a critical stage in the viral life cycle, yet for most plant viruses how they interact with their vector is unknown or is explained by analogy with previously described relatives. Here we examined the mechanism underlying the transmission of citrus tristeza virus (CTV) by its aphid vector, Toxoptera citricida, with the objective of identifying what virus-encoded proteins it uses to interact with the vector. Using fluorescently labeled virions, we demonstrated that CTV binds specifically to the lining of the cibarium of the aphid. Through in vitro competitive binding assays between fluorescent virions and free viral proteins, we determined that the minor coat protein is involved in vector interaction. We also found that the presence of two heat shock-like proteins, p61 and p65, reduces virion binding in vitro Additionally, treating the dissected mouthparts with proteases did not affect the binding of CTV virions. In contrast, chitinase treatment reduced CTV binding to the foregut. Finally, competition with glucose, N-acetyl-β-d-glucosamine, chitobiose, and chitotriose reduced the binding. These findings together suggest that CTV binds to the sugar moieties of the cuticular surface of the aphid cibarium, and the binding involves the concerted activity of three virus-encoded proteins. Limited information is known about the specific interactions between citrus tristeza virus and its aphid vectors. These interactions are important for the process of successful transmission. In this study, we localized the CTV retention site as the cibarium of the aphid foregut. Moreover, we demonstrated that the nature of these interactions is protein-carbohydrate binding. The viral proteins, including the minor coat protein and two heat shock proteins, bind to sugar moieties on the surface of the foregut. These findings will help in understanding the transmission mechanism of CTV by the aphid vector and may help in developing control strategies which interfere

  14. Glycosylation at Asn91 of H1N1 haemagglutinin affects binding to glycan receptors.

    Science.gov (United States)

    Jayaraman, Akila; Koh, Xiaoying; Li, Jing; Raman, Rahul; Viswanathan, Karthik; Shriver, Zachary; Sasisekharan, Ram

    2012-06-15

    The glycoprotein HA (haemagglutinin) on the surface of influenza A virus plays a central role in recognition and binding to specific host cell-surface glycan receptors and in fusion of viral membrane to the host nuclear membrane during viral replication. Given the abundance of HA on the viral surface, this protein is also the primary target for host innate and adaptive immune responses. Although addition of glycosylation sites on HA are a part of viral evolution to evade the host immune responses, there are specific glycosylation sites that are conserved during most of the evolution of the virus. In the present study, it was demonstrated that one such conserved glycosylation site at Asn(91) in H1N1 HA critically governs the glycan receptor-binding specificity and hence would potentially impinge on the host adaptation of the virus.

  15. Binding of Intrinsic and Extrinsic Features in Working Memory

    Science.gov (United States)

    Ecker, Ullrich K. H.; Maybery, Murray; Zimmer, Hubert D.

    2013-01-01

    There is ongoing debate concerning the mechanisms of feature binding in working memory. In particular, there is controversy regarding the extent to which these binding processes are automatic. The present article demonstrates that binding mechanisms differ depending on whether the to-be-integrated features are perceived as forming a coherent…

  16. Is there a link between selectivity and binding thermodynamics profiles?

    Science.gov (United States)

    Tarcsay, Ákos; Keserű, György M

    2015-01-01

    Thermodynamics of ligand binding is influenced by the interplay between enthalpy and entropy contributions of the binding event. The impact of these binding free energy components, however, is not limited to the primary target only. Here, we investigate the relationship between binding thermodynamics and selectivity profiles by combining publicly available data from broad off-target assay profiling and the corresponding thermodynamics measurements. Our analysis indicates that compounds binding their primary targets with higher entropy contributions tend to hit more off-targets compared with those ligands that demonstrated enthalpy-driven binding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Innovative technology demonstration

    International Nuclear Information System (INIS)

    Anderson, D.B.; Luttrell, S.P.; Hartley, J.N.; Hinchee, R.

    1992-04-01

    The Innovative Technology Demonstration (ITD) program at Tinker Air Force Base (TAFB), Oklahoma City, Oklahoma, will demonstrate the overall utility and effectiveness of innovative technologies for site characterization, monitoring, and remediation of selected contaminated test sites. The current demonstration test sites include a CERCLA site on the NPL list, located under a building (Building 3001) that houses a large active industrial complex used for rebuilding military aircraft, and a site beneath and surrounding an abandoned underground tank vault used for storage of jet fuels and solvents. The site under Building 3001 (the NW Test Site) is contaminated with TCE and Cr +6 ; the site with the fuel storage vault (the SW Tanks Site) is contaminated with fuels, BTEX and TCE. These sites and others have been identified for cleanup under the Air Force's Installation Restoration Program (IRP). This document describes the demonstrations that have been conducted or are planned for the TAFB

  18. Laser Communications Relay Demonstration

    Data.gov (United States)

    National Aeronautics and Space Administration — LCRD is a minimum two year flight demonstration in geosynchronous Earth orbit to advance optical communications technology toward infusion into Deep Space and Near...

  19. Spacecraft Fire Safety Demonstration

    Data.gov (United States)

    National Aeronautics and Space Administration — The objective of the Spacecraft Fire Safety Demonstration project is to develop and conduct large-scale fire safety experiments on an International Space Station...

  20. Guiding, bending, and splitting of coupled defect surface modes in a surface-wave photonic crystal

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Zhen; Gao, Fei [Division of Physics and Applied Physics, School of Physical and Mathematical Sciences, Nanyang Technological University, Singapore, Singapore 637371 (Singapore); Zhang, Baile, E-mail: blzhang@ntu.edu.sg [Division of Physics and Applied Physics, School of Physical and Mathematical Sciences, Nanyang Technological University, Singapore, Singapore 637371 (Singapore); Centre for Disruptive Photonic Technologies, Nanyang Technological University, Singapore, Singapore 637371 (Singapore)

    2016-01-25

    We experimentally demonstrate a type of waveguiding mechanism for coupled surface-wave defect modes in a surface-wave photonic crystal. Unlike conventional spoof surface plasmon waveguides, waveguiding of coupled surface-wave defect modes is achieved through weak coupling between tightly localized defect cavities in an otherwise gapped surface-wave photonic crystal, as a classical wave analogue of tight-binding electronic wavefunctions in solid state lattices. Wave patterns associated with the high transmission of coupled defect surface modes are directly mapped with a near-field microwave scanning probe for various structures including a straight waveguide, a sharp corner, and a T-shaped splitter. These results may find use in the design of integrated surface-wave devices with suppressed crosstalk.

  1. Guiding, bending, and splitting of coupled defect surface modes in a surface-wave photonic crystal

    International Nuclear Information System (INIS)

    Gao, Zhen; Gao, Fei; Zhang, Baile

    2016-01-01

    We experimentally demonstrate a type of waveguiding mechanism for coupled surface-wave defect modes in a surface-wave photonic crystal. Unlike conventional spoof surface plasmon waveguides, waveguiding of coupled surface-wave defect modes is achieved through weak coupling between tightly localized defect cavities in an otherwise gapped surface-wave photonic crystal, as a classical wave analogue of tight-binding electronic wavefunctions in solid state lattices. Wave patterns associated with the high transmission of coupled defect surface modes are directly mapped with a near-field microwave scanning probe for various structures including a straight waveguide, a sharp corner, and a T-shaped splitter. These results may find use in the design of integrated surface-wave devices with suppressed crosstalk

  2. Education Payload Operation - Demonstrations

    Science.gov (United States)

    Keil, Matthew

    2009-01-01

    Education Payload Operation - Demonstrations (EPO-Demos) are recorded video education demonstrations performed on the International Space Station (ISS) by crewmembers using hardware already onboard the ISS. EPO-Demos are videotaped, edited, and used to enhance existing NASA education resources and programs for educators and students in grades K-12. EPO-Demos are designed to support the NASA mission to inspire the next generation of explorers.

  3. Buried Waste Integrated Demonstration

    International Nuclear Information System (INIS)

    1994-03-01

    The Buried Waste Integrated Demonstration (BWID) supports the applied research, development, demonstration, and evaluation of a suite of advanced technologies that offer promising solutions to the problems associated with the remediation of buried waste. BWID addresses the difficult remediation problems associated with DOE complex-wide buried waste, particularly transuranic (TRU) contaminated buried waste. BWID has implemented a systems approach to the development and demonstration of technologies that will characterize, retrieve, treat, and dispose of DOE buried wastes. This approach encompasses the entire remediation process from characterization to post-monitoring. The development and demonstration of the technology is predicated on how a technology fits into the total remediation process. To address all of these technological issues, BWID has enlisted scientific expertise of individuals and groups from within the DOE Complex, as well as experts from universities and private industry. The BWID mission is to support development and demonstration of a suite of technologies that, when integrated with commercially-available technologies, forms a comprehensive, remediation system for the effective and efficient remediation of buried waste throughout the DOE Complex. BWID will evaluate and validate demonstrated technologies and transfer this information and equipment to private industry to support the Office of Environmental Restoration (ER), Office of Waste Management (WM), and Office of Facility Transition (FT) remediation planning and implementation activities

  4. Small Molecule Microarrays Enable the Identification of a Selective, Quadruplex-Binding Inhibitor of MYC Expression.

    Science.gov (United States)

    Felsenstein, Kenneth M; Saunders, Lindsey B; Simmons, John K; Leon, Elena; Calabrese, David R; Zhang, Shuling; Michalowski, Aleksandra; Gareiss, Peter; Mock, Beverly A; Schneekloth, John S

    2016-01-15

    The transcription factor MYC plays a pivotal role in cancer initiation, progression, and maintenance. However, it has proven difficult to develop small molecule inhibitors of MYC. One attractive route to pharmacological inhibition of MYC has been the prevention of its expression through small molecule-mediated stabilization of the G-quadruplex (G4) present in its promoter. Although molecules that bind globally to quadruplex DNA and influence gene expression are well-known, the identification of new chemical scaffolds that selectively modulate G4-driven genes remains a challenge. Here, we report an approach for the identification of G4-binding small molecules using small molecule microarrays (SMMs). We use the SMM screening platform to identify a novel G4-binding small molecule that inhibits MYC expression in cell models, with minimal impact on the expression of other G4-associated genes. Surface plasmon resonance (SPR) and thermal melt assays demonstrated that this molecule binds reversibly to the MYC G4 with single digit micromolar affinity, and with weaker or no measurable binding to other G4s. Biochemical and cell-based assays demonstrated that the compound effectively silenced MYC transcription and translation via a G4-dependent mechanism of action. The compound induced G1 arrest and was selectively toxic to MYC-driven cancer cell lines containing the G4 in the promoter but had minimal effects in peripheral blood mononucleocytes or a cell line lacking the G4 in its MYC promoter. As a measure of selectivity, gene expression analysis and qPCR experiments demonstrated that MYC and several MYC target genes were downregulated upon treatment with this compound, while the expression of several other G4-driven genes was not affected. In addition to providing a novel chemical scaffold that modulates MYC expression through G4 binding, this work suggests that the SMM screening approach may be broadly useful as an approach for the identification of new G4-binding small

  5. Binding of 3H-iloprost to rat gastric mucosa: a pitfall in performing radioligand binding assays

    International Nuclear Information System (INIS)

    Beinborn, M.; Kromer, W.; Staar, U.; Sewing, K.F.

    1985-01-01

    Binding of 3 H-iloprost was studied in a 20,000 x g sediment of the rat gastric mucosa. When pH in both test tubes for total and non-specific binding was kept identical, no displaceable binding of iloprost could be detected. When no care was taken to keep the pH identical in corresponding test tubes of the binding assay, changes in pH simulated specific and displaceable binding of iloprost. Therefore it is concluded that - in contrast to earlier reports - it is not possible to demonstrate specific iloprost binding using the given method

  6. Site-directed mutagenesis of HIV-1 vpu gene demonstrates two clusters of replication-defective mutants with distinct ability to down-modulate cell surface CD4 and tetherin

    Directory of Open Access Journals (Sweden)

    Masako Nomaguchi

    2010-11-01

    Full Text Available HIV-1 Vpu acts positively on viral infectivity by mediating CD4 degradation in endoplasmic reticulum and enhances virion release by counteracting a virion release restriction factor, tetherin. In order to define the impact of Vpu activity on HIV-1 replication, we have generated a series of site-specific proviral vpu mutants. Of fifteen mutants examined, seven exhibited a replication-defect similar to that of a vpu-deletion mutant in a lymphocyte cell line H9. These mutations clustered in narrow regions within transmembrane domain (TMD and cytoplasmic domain (CTD. Replication-defective mutants displayed the reduced ability to enhance virion release from a monolayer cell line HEp2 without exception. Upon transfection with Vpu expression vectors, neither TMD mutants nor CTD mutants blocked CD4 expression at the cell surface in another monolayer cell line MAGI. While TMD mutants were unable to down-modulate cell surface tetherin in HEp2 cells, CTD mutants did quite efficiently. Confocal microscopy analysis revealed the difference of intracellular localization between TMD and CTD mutants. In total, replication capability of HIV-1 carrying vpu mutations correlates well with the ability of Vpu to enhance virion release and to impede the cell surface expression of CD4 but not with the ability to down-modulate cell surface tetherin. Our results here suggest that efficient viral replication requires not only down-regulation of cell surface tetherin but also its degradation.

  7. Learning From Demonstration?

    DEFF Research Database (Denmark)

    Koch, Christian; Bertelsen, Niels Haldor

    2014-01-01

    Demonstration projects are often used in the building sector to provide a basis for using new processes and/or products. The climate change agenda implies that construction is not only required to deliver value for the customer, cost reductions and efficiency but also sustainable buildings....... This paper reports on an early demonstration project, the Building of a passive house dormitory in the Central Region of Denmark in 2006-2009. The project was supposed to deliver value, lean design, prefabrication, quality in sustainability, certification according to German standards for passive houses......, and micro combined heat and power using hydrogen. Using sociological and business economic theories of innovation, the paper discusses how early movers of innovation tend to obtain only partial success when demonstrating their products and often feel obstructed by minor details. The empirical work...

  8. Solar renovation demonstration projects

    Energy Technology Data Exchange (ETDEWEB)

    Bruun Joergensen, O [ed.

    1998-10-01

    In the framework of the IEA SHC Programme, a Task on building renovation was initiated, `Task 20, Solar Energy in Building Renovation`. In a part of the task, Subtask C `Design of Solar Renovation Projects`, different solar renovation demonstration projects were developed. The objective of Subtask C was to demonstrate the application of advanced solar renovation concepts on real buildings. This report documents 16 different solar renovation demonstration projects including the design processes of the projects. The projects include the renovation of houses, schools, laboratories, and factories. Several solar techniques were used: building integrated solar collectors, glazed balconies, ventilated solar walls, transparent insulation, second skin facades, daylight elements and photovoltaic systems. These techniques are used in several simple as well as more complex system designs. (au)

  9. Biodenitrification demonstration test report

    International Nuclear Information System (INIS)

    Benear, A.K.; Murray, S.J.; Lahoda, E.J.; Leslie, J.W.; Patton, J.B.; Menako, C.R.

    1987-08-01

    A two-column biodenitrification (BDN) facility was constructed at the Feed Materials Production Center (FMPC) in 1985 and 1986 to test the feasibility of biological treatment for industrial nitrate-bearing waste water generated at FMPC. This demonstration facility comprises one-half of the proposed four-column production facility. A demonstration test was conducted over a four month period in 1987. The results indicate the proposed BDN production facility can process FMPC industrial wastewater in a continuous manner while maintaining an effluent that will consistently meet the proposed NPDES limits for combined nitrate nitrogen (NO 3 -N) and nitrite nitrogen (NO 2 -N). The proposed NPDES limits are 62 kg/day average and 124 kg/day maximum. These limits were proportioned to determine that the two-column demonstration facility should meet the limits of 31 kg/day average and 62 kg/day maximum

  10. Distributed picture compilation demonstration

    Science.gov (United States)

    Alexander, Richard; Anderson, John; Leal, Jeff; Mullin, David; Nicholson, David; Watson, Graham

    2004-08-01

    A physical demonstration of distributed surveillance and tracking is described. The demonstration environment is an outdoor car park overlooked by a system of four rooftop cameras. The cameras extract moving objects from the scene, and these objects are tracked in a decentralized way, over a real communication network, using the information form of the standard Kalman filter. Each node therefore has timely access to the complete global picture and because there is no single point of failure in the system, it is robust. The demonstration system and its main components are described here, with an emphasis on some of the lessons we have learned as a result of applying a corpus of distributed data fusion theory and algorithms in practice. Initial results are presented and future plans to scale up the network are also outlined.

  11. Photovoltaic demonstration projects

    Energy Technology Data Exchange (ETDEWEB)

    Gillett, W B; Hacker, R J; Kaut, W [eds.

    1991-01-01

    This book, the proceedings of the fourth PV-Contractors' Meeting organized by the Commission of the European Communities, Directorate-General for Energy, held at Brussels on 21 and 22 November 1989, provides an overview of the photovoltaic demonstration projects which have been supported in the framework of the Energy Demonstration Program since 1983. It includes reports by each of the contractors who submitted proposals in 1983, 1984, 1985 and 1986, describing progress with their projects. Summaries of the discussions held at the meeting, which included contractors whose projects were submitted in 1987, are also presented. The different technologies which are being demonstrated concern the modules, the cabling of the array, structure design, storage strategy and power conditioning. The various applications include desalination, communications, dairy farms, water pumping, and warning systems. Papers have been processed separately for inclusion on the data base.

  12. Electric vehicle demonstration

    Energy Technology Data Exchange (ETDEWEB)

    Ouellet, M. [National Centre for Advanced Transportation, Saint-Jerome, PQ (Canada)

    2010-07-01

    The desirable characteristics of Canadian projects that demonstrate vehicle use in real-world operation and the appropriate mechanism to collect and disseminate the monitoring data were discussed in this presentation. The scope of the project was on passenger cars and light duty trucks operating in plug-in electric vehicle (PHEV) or battery electric vehicle modes. The presentation also discussed the funding, stakeholders involved, Canadian travel pattern analysis, regulatory framework, current and recent electric vehicle demonstration projects, and project guidelines. It was concluded that some demonstration project activities may have been duplicated as communication between the proponents was insufficient. It was recommended that data monitoring using automatic data logging with minimum reliance on logbooks and other user entry should be emphasized. figs.

  13. Sequence similarity between the erythrocyte binding domain of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals a functional heparin binding motif involved in binding to the Duffy antigen receptor for chemokines

    Directory of Open Access Journals (Sweden)

    Bolton Michael J

    2011-11-01

    Full Text Available Abstract Background The HIV surface glycoprotein gp120 (SU, gp120 and the Plasmodium vivax Duffy binding protein (PvDBP bind to chemokine receptors during infection and have a site of amino acid sequence similarity in their binding domains that often includes a heparin binding motif (HBM. Infection by either pathogen has been found to be inhibited by polyanions. Results Specific polyanions that inhibit HIV infection and bind to the V3 loop of X4 strains also inhibited DBP-mediated infection of erythrocytes and DBP binding to the Duffy Antigen Receptor for Chemokines (DARC. A peptide including the HBM of PvDBP had similar affinity for heparin as RANTES and V3 loop peptides, and could be specifically inhibited from heparin binding by the same polyanions that inhibit DBP binding to DARC. However, some V3 peptides can competitively inhibit RANTES binding to heparin, but not the PvDBP HBM peptide. Three other members of the DBP family have an HBM sequence that is necessary for erythrocyte binding, however only the protein which binds to DARC, the P. knowlesi alpha protein, is inhibited by heparin from binding to erythrocytes. Heparitinase digestion does not affect the binding of DBP to erythrocytes. Conclusion The HBMs of DBPs that bind to DARC have similar heparin binding affinities as some V3 loop peptides and chemokines, are responsible for specific sulfated polysaccharide inhibition of parasite binding and invasion of red blood cells, and are more likely to bind to negative charges on the receptor than cell surface glycosaminoglycans.

  14. Innovative technology demonstrations

    International Nuclear Information System (INIS)

    Anderson, D.B.; Luttrell, S.P.; Hartley, J.N.

    1992-08-01

    Environmental Management Operations (EMO) is conducting an Innovative Technology Demonstration Program for Tinker Air Force Base (TAFB). Several innovative technologies are being demonstrated to address specific problems associated with remediating two contaminated test sites at the base. Cone penetrometer testing (CPT) is a form of testing that can rapidly characterize a site. This technology was selected to evaluate its applicability in the tight clay soils and consolidated sandstone sediments found at TAFB. Directionally drilled horizontal wells was selected as a method that may be effective in accessing contamination beneath Building 3001 without disrupting the mission of the building, and in enhancing the extraction of contamination both in ground water and in soil. A soil gas extraction (SGE) demonstration, also known as soil vapor extraction, will evaluate the effectiveness of SGE in remediating fuels and TCE contamination contained in the tight clay soil formations surrounding the abandoned underground fuel storage vault located at the SW Tanks Site. In situ sensors have recently received much acclaim as a technology that can be effective in remediating hazardous waste sites. Sensors can be useful for determining real-time, in situ contaminant concentrations during the remediation process for performance monitoring and in providing feedback for controlling the remediation process. Following the SGE demonstration, the SGE system and SW Tanks test site will be modified to demonstrate bioremediation as an effective means of degrading the remaining contaminants in situ. The bioremediation demonstration will evaluate a bioventing process in which the naturally occurring consortium of soil bacteria will be stimulated to aerobically degrade soil contaminants, including fuel and TCE, in situ

  15. Innovative technology demonstrations

    International Nuclear Information System (INIS)

    Anderson, D.B.; Hartley, J.N.; Luttrell, S.P.

    1992-04-01

    Currently, several innovative technologies are being demonstrated at Tinker Air Force Base (TAFB) to address specific problems associated with remediating two contaminated test sites at the base. Cone penetrometer testing (CPT) is a form of testing that can rapidly characterize a site. This technology was selected to evaluate its applicability in the tight clay soils and consolidated sandstone sediments found at TAFB. Directionally drilled horizontal wells have been successfully installed at the US Department of Energy's (DOE) Savannah River Site to test new methods of in situ remediation of soils and ground water. This emerging technology was selected as a method that may be effective in accessing contamination beneath Building 3001 without disrupting the mission of the building, and in enhancing the extraction of contamination both in ground water and in soil. A soil gas extraction (SGE) demonstration, also known as soil vapor extraction, will evaluate the effectiveness of SGE in remediating fuels and TCE contamination contained in the tight clay soil formations surrounding the abandoned underground fuel storage vault located at the SW Tanks Site. In situ sensors have recently received much acclaim as a technology that can be effective in remediating hazardous waste sites. Sensors can be useful for determining real-time, in situ contaminant concentrations during the remediation process for performance monitoring and in providing feedback for controlling the remediation process. A demonstration of two in situ sensor systems capable of providing real-time data on contamination levels will be conducted and evaluated concurrently with the SGE demonstration activities. Following the SGE demonstration, the SGE system and SW Tanks test site will be modified to demonstrate bioremediation as an effective means of degrading the remaining contaminants in situ

  16. Gigashot Optical Laser Demonstrator

    Energy Technology Data Exchange (ETDEWEB)

    Deri, R. J. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2015-10-13

    The Gigashot Optical Laser Demonstrator (GOLD) project has demonstrated a novel optical amplifier for high energy pulsed lasers operating at high repetition rates. The amplifier stores enough pump energy to support >10 J of laser output, and employs conduction cooling for thermal management to avoid the need for expensive and bulky high-pressure helium subsystems. A prototype amplifier was fabricated, pumped with diode light at 885 nm, and characterized. Experimental results show that the amplifier provides sufficient small-signal gain and sufficiently low wavefront and birefringence impairments to prove useful in laser systems, at repetition rates up to 60 Hz.

  17. Photovoltaic demonstration projects 2

    Energy Technology Data Exchange (ETDEWEB)

    Gillett, W B; Hacker, R J [Halcrow (William) and Partners, Swindon (UK); Kaut, W [eds.

    1989-01-01

    This book, the proceedings of the third Photovoltaic Contractors' Meeting organised by the Commission of the European Communities, Directorate-General for Energy provides an overview of the photovoltaic demonstration projects which have been supported by the Energy Directorate of the Commission of the European Communities since 1983. It includes reports by each of the contractors who submitted proposals in 1983, 1984 and 1985, describing progress with their projects. The different technologies which are being demonstrated concern the modules, the cabling of the array, structure design, storage strategy and power conditioning. The various applications include powering of houses, villages, recreation centres, water desalination, communications, dairy farms, water pumping and warning systems. (author).

  18. Fundamental considerations in ski binding analysis.

    Science.gov (United States)

    Mote, C D; Hull, M L

    1976-01-01

    1. The static adjustment of a ski binding by hand or by available machines is only an adjustment and is neither a static nor a dynamic evaluation of the binding design. Bindings of different design with identical static adjustments will perform differently in environments in which the forces are static or dynamic. 2. The concept of binding release force is a useful measure of binding adjustment, but it is inappropriate as a criterion for binding evaluation. First, it does not direct attention toward the injury causing mechanism, strain, or displacement in the leg. Second, it is only part of the evaluation in dynamic problems. 3. The binding release decision in present bindings is displacement controlled. The relative displacement of the boot and ski is the system variable. For any specified relative displacement the binding force can be any of an infinite number of possibilities determined by the loading path. 4. The response of the leg-ski system to external impulses applied to the ski is independent of the boot-ski relative motion as long as the boot recenters quickly in the binding. Response is dependent upon the external impulse plus system inertia, damping and stiffness. 5. When tested under half sinusoidal forces applied to a test ski, all bindings will demonstrate static and impulse loading regions. In the static region the force drives the binding to a relative release displacement. In the impulse region the initial velocity of the ski drives the binding to a release displacement. 6. The transition between the static and impulse loading regions is determined by the binding's capacity to store and dissipate energy along the principal loading path. Increased energy capacity necessitates larger external impulses to produce release. 7. In all bindings examined to date, the transmitted leg displacement or strain at release under static loading exceeds leg strain under dynamic or impact loading. Because static loading is responsible for many injuries, a skier

  19. Novel heparan sulfate-binding peptides for blocking herpesvirus entry.

    Directory of Open Access Journals (Sweden)

    Pranay Dogra

    Full Text Available Human cytomegalovirus (HCMV infection can lead to congenital hearing loss and mental retardation. Upon immune suppression, reactivation of latent HCMV or primary infection increases morbidity in cancer, transplantation, and late stage AIDS patients. Current treatments include nucleoside analogues, which have significant toxicities limiting their usefulness. In this study we screened a panel of synthetic heparin-binding peptides for their ability to prevent CMV infection in vitro. A peptide designated, p5+14 exhibited ~ 90% reduction in murine CMV (MCMV infection. Because negatively charged, cell-surface heparan sulfate proteoglycans (HSPGs, serve as the attachment receptor during the adsorption phase of the CMV infection cycle, we hypothesized that p5+14 effectively competes for CMV adsorption to the cell surface resulting in the reduction in infection. Positively charged Lys residues were required for peptide binding to cell-surface HSPGs and reducing viral infection. We show that this inhibition was not due to a direct neutralizing effect on the virus itself and that the peptide blocked adsorption of the virus. The peptide also inhibited infection of other herpesviruses: HCMV and herpes simplex virus 1 and 2 in vitro, demonstrating it has broad-spectrum antiviral activity. Therefore, this peptide may offer an adjunct therapy for the treatment of herpes viral infections and other viruses that use HSPGs for entry.

  20. Inseparable Phone Books Demonstration

    Science.gov (United States)

    Balta, Nuri; Çetin, Ali

    2017-01-01

    This study is aimed at first introducing a well-known discrepant event; inseparable phone books and second, turning it into an experiment for high school or middle school students. This discrepant event could be used especially to indicate how friction force can be effective in producing an unexpected result. Demonstration, discussion, explanation…

  1. PHARUS ASAR demonstrator

    NARCIS (Netherlands)

    Smith, A.J.E.; Bree, R.J.P. van; Calkoen, C.J.; Dekker, R.J.; Otten, M.P.G.; Rossum, W.L. van

    2001-01-01

    PHARUS is a polarimetric phased array C-band Synthetic Aperture Radar (SAR), designed and built for airborne use. Advanced SAR (ASAR) data in image and alternating polarization mode have been simulated with PHARUS to demonstrate the use of Envisat for a number of typical SAR applications that are

  2. Demonstrating the Gas Laws.

    Science.gov (United States)

    Holko, David A.

    1982-01-01

    Presents a complete computer program demonstrating the relationship between volume/pressure for Boyle's Law, volume/temperature for Charles' Law, and volume/moles of gas for Avagadro's Law. The programing reinforces students' application of gas laws and equates a simulated moving piston to theoretical values derived using the ideal gas law.…

  3. Astronomy LITE Demonstrations

    Science.gov (United States)

    Brecher, Kenneth

    2006-12-01

    Project LITE (Light Inquiry Through Experiments) is a materials, software, and curriculum development project. It focuses on light, optics, color and visual perception. According to two recent surveys of college astronomy faculty members, these are among the topics most often included in the large introductory astronomy courses. The project has aimed largely at the design and implementation of hands-on experiences for students. However, it has also included the development of lecture demonstrations that employ novel light sources and materials. In this presentation, we will show some of our new lecture demonstrations concerning geometrical and physical optics, fluorescence, phosphorescence and polarization. We have developed over 200 Flash and Java applets that can be used either by teachers in lecture settings or by students at home. They are all posted on the web at http://lite.bu.edu. For either purpose they can be downloaded directly to the user's computer or run off line. In lecture demonstrations, some of these applets can be used to control the light emitted by video projectors to produce physical effects in materials (e.g. fluorescence). Other applets can be used, for example, to demonstrate that the human percept of color does not have a simple relationship with the physical frequency of the stimulating source of light. Project LITE is supported by Grant #DUE-0125992 from the NSF Division of Undergraduate Education.

  4. A Magnetic Circuit Demonstration.

    Science.gov (United States)

    Vanderkooy, John; Lowe, June

    1995-01-01

    Presents a demonstration designed to illustrate Faraday's, Ampere's, and Lenz's laws and to reinforce the concepts through the analysis of a two-loop magnetic circuit. Can be made dramatic and challenging for sophisticated students but is suitable for an introductory course in electricity and magnetism. (JRH)

  5. Surface chemistry of "unprotected" nanoparticles

    DEFF Research Database (Denmark)

    Schrader, Imke; Warneke, Jonas; Neumann, Sarah

    2015-01-01

    The preparation of colloidal nanoparticles in alkaline ethylene glycol is a powerful approach for the preparation of model catalysts and ligand-functionalized nanoparticles. For these systems the term "unprotected" nanoparticles has been established because no strongly binding stabilizers...... study. "Unprotected" Pt and Ru nanoparticles were characterized by NMR spectroscopy, which does not evidence the presence of any C-H containing species bound to the particle surface. Instead, the colloids were found to be covered by CO, as demonstrated by IR spectroscopy. However, analysis...

  6. Identification of residues on human receptor DPP4 critical for MERS-CoV binding and entry

    Energy Technology Data Exchange (ETDEWEB)

    Song, Wenfei [Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084 (China); Wang, Ying [Comprehensive AIDS Research Center, Research Center for Public Health, School of Medicine, Tsinghua University, Beijing 100084 (China); Wang, Nianshuang; Wang, Dongli [Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084 (China); Guo, Jianying; Fu, Lili [Comprehensive AIDS Research Center, Research Center for Public Health, School of Medicine, Tsinghua University, Beijing 100084 (China); Shi, Xuanling, E-mail: shixuanlingsk@tsinghua.edu.cn [Comprehensive AIDS Research Center, Research Center for Public Health, School of Medicine, Tsinghua University, Beijing 100084 (China)

    2014-12-15

    Middle East respiratory syndrome coronavirus (MERS-CoV) infects host cells through binding the receptor binding domain (RBD) on its spike glycoprotein to human receptor dipeptidyl peptidase 4 (hDPP4). Here, we report identification of critical residues on hDPP4 for RBD binding and virus entry through analysis of a panel of hDPP4 mutants. Based on the RBD–hDPP4 crystal structure we reported, the mutated residues were located at the interface between RBD and hDPP4, which potentially changed the polarity, hydrophobic or hydrophilic properties of hDPP4, thereby interfering or disrupting their interaction with RBD. Using surface plasmon resonance (SPR) binding analysis and pseudovirus infection assay, we showed that several residues in hDPP4–RBD binding interface were important on hDPP4–RBD binding and viral entry. These results provide atomic insights into the features of interactions between hDPP4 and MERS-CoV RBD, and also provide potential explanation for cellular and species tropism of MERS-CoV infection. - Highlights: • It has been demonstrated that MERS-CoV infects host cells through binding its envelope spike (S) glycoprotein to the host cellular receptor dipeptidyl peptidase 4 (DPP4). • To identify the critical residues on hDPP4 for RBD binding and virus entry, we constructed a panel of hDPP4 mutants based on structure-guided mutagenesis. • Using surface plasmon resonance (SPR) binding analysis and pseudovirus infection assay, we showed that several residues on hDPP4 had significant impacts on virus/receptor interactions and viral entry. • Our study has provided new insights into the features of interactions between hDPP4 and MERS-CoV RBD, and provides potential explanation for cellular and species tropism of MERS-CoV infection.

  7. Binding of CFA/I Pili of Enterotoxigenic Escherichia coli to Asialo-GM1 Is Mediated by the Minor Pilin CfaE.

    Science.gov (United States)

    Madhavan, T P Vipin; Riches, James D; Scanlon, Martin J; Ulett, Glen C; Sakellaris, Harry

    2016-05-01

    CFA/I pili are representatives of a large family of related pili that mediate the adherence of enterotoxigenic Escherichia coli to intestinal epithelial cells. They are assembled via the alternate chaperone-usher pathway and consist of two subunits, CfaB, which makes up the pilus shaft and a single pilus tip-associated subunit, CfaE. The current model of pilus-mediated adherence proposes that CFA/I has two distinct binding activities; the CfaE subunit is responsible for binding to receptors of unknown structure on erythrocyte and intestinal epithelial cell surfaces, while CfaB binds to various glycosphingolipids, including asialo-GM1. In this report, we present two independent lines of evidence that, contrary to the existing model, CfaB does not bind to asialo-GM1 independently of CfaE. Neither purified CfaB subunits nor CfaB assembled into pili bind to asialo-GM1. Instead, we demonstrate that binding activity toward asialo-GM1 resides in CfaE and this is essential for pilus binding to Caco-2 intestinal epithelial cells. We conclude that the binding activities of CFA/I pili for asialo-GM1, erythrocytes, and intestinal cells are inseparable, require the same amino acid residues in CfaE, and therefore depend on the same or very similar binding mechanisms. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  8. Remote monitoring demonstration

    International Nuclear Information System (INIS)

    Caskey, Susan; Olsen, John

    2006-01-01

    The recently upgraded remote monitoring system at the Joyo Experimental Reactor uses a DCM-14 camera module and GEMINI software. The final data is compatible both with the IAEA-approved GARS review software and the ALIS software that was used for this demonstration. Features of the remote monitoring upgrade emphasized compatibility with IAEA practice. This presentation gives particular attention to the selection process for meeting network security considerations at the O'arai site. The Joyo system is different from the NNCA's ACPF system, in that it emphasizes use of IAEA standard camera technology and data acquisition and transmission software. In the demonstration itself, a temporary virtual private network (VPN) between the meeting room and the server at Sandia in Albuquerque allowed attendees to observe data stored from routine transmissions from the Joyo Fresh Fuel Storage to Sandia. Image files from a fuel movement earlier in the month showed Joyo workers and IAEA inspectors carrying out a transfer. (author)

  9. Commercial incineration demonstration

    International Nuclear Information System (INIS)

    Borduin, L.C.; Neuls, A.S.

    1981-01-01

    Low-level radioactive wastes (LLW) generated by nuclear utilities presently are shipped to commercial burial grounds for disposal. Substantially increasing shipping and disposal charges have sparked renewed industry interest in incineration and other advanced volume reduction techniques as potential cost-saving measures. Repeated inquiries from industry sources regarding LLW applicability of the Los Alamos controlled-air incineration (CAI) design led DOE to initiate this commercial demonstration program in FY-1980. The selected program approach to achieving CAI demonstration at a utility site is a DOE sponsored joint effort involving Los Alamos, a nuclear utility, and a liaison subcontractor. Required development tasks and responsibilities of the particpants are described. Target date for project completion is the end of FY-1985

  10. Photovoltaic demonstration projects

    Energy Technology Data Exchange (ETDEWEB)

    Kaut, W [Commission of the European Communities, Brussels (Belgium); Gillett, W B; Hacker, R J [Halcrow Gilbert Associates Ltd., Swindon (GB)

    1992-12-31

    This publication, comprising the proceedings of the fifth contractor`s meeting organized by the Commission of the European Communities, Directorate-General for Energy, provides an overview of the photovoltaic demonstration projects which have been supported in the framework of the energy demonstration programme since 1983. It includes reports by each of the contractors who submitted proposals in 1987 and 1988, describing progress within their projects. Projects accepted from earlier calls for proposals and not yet completed were reviewed by a rapporteur and are discussed in the summary section. The results of the performance monitoring of all projects and the lessons drawn from the practical experience of the projects are also presented in the summaries and conclusions. Contractors whose projects were submitted in 1989 were also present at the meeting and contributed to the reported discussions. This proceeding is divided into four sessions (General, Housing, technical presentations, other applications) and 24 papers are offered.

  11. AVNG system demonstration

    Energy Technology Data Exchange (ETDEWEB)

    Thron, Jonathan Louis [Los Alamos National Laboratory; Mac Arthur, Duncan W [Los Alamos National Laboratory; Kondratov, Sergey [VNIIEF; Livke, Alexander [VNIIEF; Razinkov, Sergey [VNIIEF

    2010-01-01

    An attribute measurement system (AMS) measures a number of unclassified attributes of potentially classified material. By only displaying these unclassified results as red or green lights, the AMS protects potentially classified information while still generating confidence in the measurement result. The AVNG implementation that we describe is an AMS built by RFNC - VNIIEF in Sarov, Russia. To provide additional confidence, the AVNG was designed with two modes of operation. In the secure mode, potentially classified measurements can be made with only the simple red light/green light display. In the open mode, known unclassified material can be measured with complete display of the information collected from the radiation detectors. The AVNG demonstration, which occurred in Sarov, Russia in June 2009 for a joint US/Russian audience, included exercising both modes of AVNG operation using a number of multi-kg plutonium sources. In addition to describing the demonstration, we will show photographs and/or video taken of AVNG operation.

  12. Antares: preliminary demonstrator results

    International Nuclear Information System (INIS)

    Kouchner, A.

    2000-05-01

    The ANTARES collaboration is building an undersea neutrino telescope off Toulon (Mediterranean sea) with effective area ∼ 0.1 km 2 . An extensive study of the site properties has been achieved together with software analysis in order to optimize the performance of the detector. Results are summarized here. An instrumented line, linked to shore for first time via an electro-optical cable, has been immersed late 1999. The preliminary results of this demonstrator line are reported. (author)

  13. The Majorana Demonstrator

    Energy Technology Data Exchange (ETDEWEB)

    Aguayo, Estanislao; Fast, James E.; Hoppe, Eric W.; Keillor, Martin E.; Kephart, Jeremy D.; Kouzes, Richard T.; LaFerriere, Brian D.; Merriman, Jason H.; Orrell, John L.; Overman, Nicole R.; Avignone, Frank T.; Back, Henning O.; Combs, Dustin C.; Leviner, L.; Young, A.; Barabash, Alexander S.; Konovalov, S.; Vanyushin, I.; Yumatov, Vladimir; Bergevin, M.; Chan, Yuen-Dat; Detwiler, Jason A.; Loach, J. C.; Martin, R. D.; Poon, Alan; Prior, Gersende; Vetter, Kai; Bertrand, F.; Cooper, R. J.; Radford, D. C.; Varner, R. L.; Yu, Chang-Hong; Boswell, M.; Elliott, S.; Gehman, Victor M.; Hime, Andrew; Kidd, M. F.; LaRoque, B. H.; Rielage, Keith; Ronquest, M. C.; Steele, David; Brudanin, V.; Egorov, Viatcheslav; Gusey, K.; Kochetov, Oleg; Shirchenko, M.; Timkin, V.; Yakushev, E.; Busch, Matthew; Esterline, James H.; Tornow, Werner; Christofferson, Cabot-Ann; Horton, Mark; Howard, S.; Sobolev, V.; Collar, J. I.; Fields, N.; Creswick, R.; Doe, Peter J.; Johnson, R. A.; Knecht, A.; Leon, Jonathan D.; Marino, Michael G.; Miller, M. L.; Robertson, R. G. H.; Schubert, Alexis G.; Wolfe, B. A.; Efremenko, Yuri; Ejiri, H.; Hazama, R.; Nomachi, Masaharu; Shima, T.; Finnerty, P.; Fraenkle, Florian; Giovanetti, G. K.; Green, M.; Henning, Reyco; Howe, M. A.; MacMullin, S.; Phillips, D.; Snavely, Kyle J.; Strain, J.; Vorren, Kris R.; Guiseppe, Vincente; Keller, C.; Mei, Dong-Ming; Perumpilly, Gopakumar; Thomas, K.; Zhang, C.; Hallin, A. L.; Keeter, K.; Mizouni, Leila; Wilkerson, J. F.

    2011-09-03

    A brief review of the history and neutrino physics of double beta decay is given. A description of the MAJORANA DEMONSTRATOR research and development program, including background reduction techniques, is presented in some detail. The application of point contact (PC) detectors to the experiment is discussed, including the effectiveness of pulse shape analysis. The predicted sensitivity of a PC detector array enriched to 86% to 76Ge is given.

  14. IGCC technology and demonstration

    Energy Technology Data Exchange (ETDEWEB)

    Palonen, J [A. Ahlstrom Corporation, Karhula (Finland). Hans Ahlstrom Lab.; Lundqvist, R G [A. Ahlstrom Corporation, Helsinki (Finland); Staahl, K [Sydkraft AB, Malmoe (Sweden)

    1997-12-31

    Future energy production will be performed by advanced technologies that are more efficient, more environmentally friendly and less expensive than current technologies. Integrated gasification combined cycle (IGCC) power plants have been proposed as one of these systems. Utilising biofuels in future energy production will also be emphasised since this lowers substantially carbon dioxide emissions into the atmosphere due to the fact that biomass is a renewable form of energy. Combining advanced technology and biomass utilisation is for this reason something that should and will be encouraged. A. Ahlstrom Corporation of Finland and Sydkraft AB of Sweden have as one part of company strategies adopted this approach for the future. The companies have joined their resources in developing a biomass-based IGCC system with the gasification part based on pressurised circulating fluidized-bed technology. With this kind of technology electrical efficiency can be substantially increased compared to conventional power plants. As a first concrete step, a decision has been made to build a demonstration plant. This plant, located in Vaernamo, Sweden, has already been built and is now in commissioning and demonstration stage. The system comprises a fuel drying plant, a pressurised CFB gasifier with gas cooling and cleaning, a gas turbine, a waste heat recovery unit and a steam turbine. The plant is the first in the world where the integration of a pressurised gasifier with a gas turbine will be realised utilising a low calorific gas produced from biomass. The capacity of the Vaernamo plant is 6 MW of electricity and 9 MW of district heating. Technology development is in progress for design of plants of sizes from 20 to 120 MWe. The paper describes the Bioflow IGCC system, the Vaernamo demonstration plant and experiences from the commissioning and demonstration stages. (orig.)

  15. IGCC technology and demonstration

    Energy Technology Data Exchange (ETDEWEB)

    Palonen, J. [A. Ahlstrom Corporation, Karhula (Finland). Hans Ahlstrom Lab.; Lundqvist, R.G. [A. Ahlstrom Corporation, Helsinki (Finland); Staahl, K. [Sydkraft AB, Malmoe (Sweden)

    1996-12-31

    Future energy production will be performed by advanced technologies that are more efficient, more environmentally friendly and less expensive than current technologies. Integrated gasification combined cycle (IGCC) power plants have been proposed as one of these systems. Utilising biofuels in future energy production will also be emphasised since this lowers substantially carbon dioxide emissions into the atmosphere due to the fact that biomass is a renewable form of energy. Combining advanced technology and biomass utilisation is for this reason something that should and will be encouraged. A. Ahlstrom Corporation of Finland and Sydkraft AB of Sweden have as one part of company strategies adopted this approach for the future. The companies have joined their resources in developing a biomass-based IGCC system with the gasification part based on pressurised circulating fluidized-bed technology. With this kind of technology electrical efficiency can be substantially increased compared to conventional power plants. As a first concrete step, a decision has been made to build a demonstration plant. This plant, located in Vaernamo, Sweden, has already been built and is now in commissioning and demonstration stage. The system comprises a fuel drying plant, a pressurised CFB gasifier with gas cooling and cleaning, a gas turbine, a waste heat recovery unit and a steam turbine. The plant is the first in the world where the integration of a pressurised gasifier with a gas turbine will be realised utilising a low calorific gas produced from biomass. The capacity of the Vaernamo plant is 6 MW of electricity and 9 MW of district heating. Technology development is in progress for design of plants of sizes from 20 to 120 MWe. The paper describes the Bioflow IGCC system, the Vaernamo demonstration plant and experiences from the commissioning and demonstration stages. (orig.)

  16. Curved reformat of the paediatric brain MRI into a 'flat-earth map' - standardised method for demonstrating cortical surface atrophy resulting from hypoxic-ischaemic encephalopathy