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Sample records for active dimeric form

  1. Structure, Aggregation, and Activity of a Covalent Insulin Dimer Formed During Storage of Neutral Formulation of Human Insulin.

    Science.gov (United States)

    Hjorth, Christian Fogt; Norrman, Mathias; Wahlund, Per-Olof; Benie, Andrew J; Petersen, Bent O; Jessen, Christian M; Pedersen, Thomas Å; Vestergaard, Kirsten; Steensgaard, Dorte B; Pedersen, Jan Skov; Naver, Helle; Hubálek, František; Poulsen, Christian; Otzen, Daniel

    2016-04-01

    A specific covalently linked dimeric species of insulin high molecular weight products (HMWPs), formed during prolonged incubation of a neutral pharmaceutical formulation of human insulin, were characterized in terms of tertiary structure, self-association, biological activity, and fibrillation properties. The dimer was formed by a covalent link between A21Asn and B29Lys. It was analyzed using static and dynamic light scattering and small-angle X-ray scattering to evaluate its self-association behavior. The tertiary structure was obtained using nuclear magnetic resonance and X-ray crystallography. The biological activity of HMWP was determined using 2 in vitro assays, and its influence on fibrillation was investigated using Thioflavin T assays. The dimer's tertiary structure was nearly identical to that of the noncovalent insulin dimer, and it was able to form hexamers in the presence of zinc. The dimer exhibited reduced propensity for self-association in the absence of zinc but significantly postponed the onset of fibrillation in insulin formulations. Consistent with its dimeric state, the tested species of HMWP showed little to no biological activity in the used assays. This study is the first detailed characterization of a specific type of human insulin HMWP formed during storage of a marketed pharmaceutical formulation. These results indicate that this specific type of HMWP is unlikely to antagonize the physical stability of the formulation, as HMWP retained a tertiary structure similar to the noncovalent dimer and participated in hexamer assembly in the presence of zinc. In addition, increasing amounts of HMWP reduce the rate of insulin fibrillation.

  2. Structural and biochemical studies on Vibrio cholerae Hsp31 reveals a novel dimeric form and Glutathione-independent Glyoxalase activity

    Science.gov (United States)

    Dey, Sanjay

    2017-01-01

    Vibrio cholerae experiences a highly hostile environment at human intestine which triggers the induction of various heat shock genes. The hchA gene product of V. cholerae O395, referred to a hypothetical intracellular protease/amidase VcHsp31, is one such stress-inducible homodimeric protein. Our current study demonstrates that VcHsp31 is endowed with molecular chaperone, amidopeptidase and robust methylglyoxalase activities. Through site directed mutagenesis coupled with biochemical assays on VcHsp31, we have confirmed the role of residues in the vicinity of the active site towards amidopeptidase and methylglyoxalase activities. VcHsp31 suppresses the aggregation of insulin in vitro in a dose dependent manner. Through crystal structures of VcHsp31 and its mutants, grown at various temperatures, we demonstrate that VcHsp31 acquires two (Type-I and Type-II) dimeric forms. Type-I dimer is similar to EcHsp31 where two VcHsp31 monomers associate in eclipsed manner through several intersubunit hydrogen bonds involving their P-domains. Type-II dimer is a novel dimeric organization, where some of the intersubunit hydrogen bonds are abrogated and each monomer swings out in the opposite directions centering at their P-domains, like twisting of wet cloth. Normal mode analysis (NMA) of Type-I dimer shows similar movement of the individual monomers. Upon swinging, a dimeric surface of ~400Å2, mostly hydrophobic in nature, is uncovered which might bind partially unfolded protein substrates. We propose that, in solution, VcHsp31 remains as an equilibrium mixture of both the dimers. With increase in temperature, transformation to Type-II form having more exposed hydrophobic surface, occurs progressively accounting for the temperature dependent increase of chaperone activity of VcHsp31. PMID:28235098

  3. Mechanism of FGF receptor dimerization and activation

    Science.gov (United States)

    Sarabipour, Sarvenaz; Hristova, Kalina

    2016-01-01

    Fibroblast growth factors (fgfs) are widely believed to activate their receptors by mediating receptor dimerization. Here we show, however, that the FGF receptors form dimers in the absence of ligand, and that these unliganded dimers are phosphorylated. We further show that ligand binding triggers structural changes in the FGFR dimers, which increase FGFR phosphorylation. The observed effects due to the ligands fgf1 and fgf2 are very different. The fgf2-bound dimer structure ensures the smallest separation between the transmembrane (TM) domains and the highest possible phosphorylation, a conclusion that is supported by a strong correlation between TM helix separation in the dimer and kinase phosphorylation. The pathogenic A391E mutation in FGFR3 TM domain emulates the action of fgf2, trapping the FGFR3 dimer in its most active state. This study establishes the existence of multiple active ligand-bound states, and uncovers a novel molecular mechanism through which FGFR-linked pathologies can arise.

  4. Structure of the Mature Streptococcal Cysteine Protease Exotoxin mSpeB in Its Active Dimeric Form

    DEFF Research Database (Denmark)

    Olsen, Johan G; Dagil, Robert; Niclasen, Louise Meinert

    2009-01-01

    Invasive infections of Streptococcus pyogenes are dependent on the cysteine protease streptococcal pyrogenic exotoxin B. Previous structures of the enzyme have not disclosed the proper active-site configuration. Here, the crystal structure of the mature enzyme is presented to 1.55 A, disclosing....... Based on the present structure, the active site of clan CA cysteine proteases is expanded and a detailed mechanism of the deacylation mechanism is proposed. The results may have applications for the development of protease inhibitors specific to bacterial cysteine proteases....... a homodimer. A serine from one subunit inserts into the active site of the other to donate to the oxyanion hole and coordinates the ligand proximal to the active-site cysteine. Dimerization is unique to the mature form and is clearly a prerequisite for catalysis. The present structure supports a tripartite...

  5. Antibody-mediated activation of a defective beta-D-galactosidase: dimeric form of the activatable mutant enzyme.

    Science.gov (United States)

    Conway de Macario, E; Ellis, J; Guzman, R; Rotman, B

    1978-02-01

    Sedimentation analyses of AMEF, an activatable mutant beta-D-galactosidase (beta-D-galactoside galactohydrolase, EC 3.2.1.23), and the products of its reaction with Fab fragments of activating antibody show that this enzyme exists mainly as 10S dimers. Activation of AMEF by purified antibody resulted in formation of 16S tetramers. A unifying hypothesis postulating a dimer--tetramer equilibrium accounts for this observation as the counterpart of inactivation, which was shown to involve the breakdown of tetramers into inactive subunits [Roth, R. A. & Rotman, B. (1975) Biochem. Biophys. Res. Commun. 67, 1382--1390]. Conditions are described under which AMEF loses the specific antigenic determinant(s) responsible for binding activating antibody, allowing its subsequent use as an absorption to obtain immunologically purified activating antibody,

  6. Production of recombinant orange-spotted grouper (Epinephelus coioides) follicle-stimulating hormone (FSH) in single-chain form and dimer form by Pichia pastoris and their biological activities.

    Science.gov (United States)

    Chen, Jun; Zhang, Yanhong; Tang, Zhiguo; Mao, Jiewei; Kuang, Zhonglei; Qin, Chaobin; Li, Wensheng

    2012-09-01

    FSH is a key regulator of steroidogenesis and gonadal growth in teleosts. However, function of FSH is elusive in grouper due to the lack of purified and native FSH. In the present study, we reported production of bioactive orange-spotted grouper (Epinephelus coioides) FSH in dimer form and single-chain form by Pichia pastoris. Dimer form of recombinant grouper FSH (rgFSHba) was accomplished by co-expressing mature FSHb-subunit and a-subunit genes. Fusion of mature FSHb-subunit and a-subunit genes together linking with a polypeptide (4×(Gly-Ser)-Gly-Thr) gene generated single-chain form of recombinant grouper FSH (rgFSHb-a). Recombinant grouper common α-subunit (rgCga) and FSHb-subunit (rgFSHb) were also separately produced. Recombinant proteins were verified by Western blot and mass spectrometry assays, and characterized by deglycosylation analysis. Deglycosylation assay suggested that glycosylation of recombinant FSH mainly occurred on common a-subunit. Bioactivities of recombinant proteins were initially evaluated by activating grouper FSH receptor, and further demonstrated by incubating ovarian fragments of adult grouper and intraperitoneal injection in juvenile female grouper. Two forms of recombinant FSH presented similar biological activities of activating FSH receptor and stimulating in vitro testosterone (T) and estradiol-17β (E2) secretion, though the dimer form functioned slightly weaker than the single-chain form. However, injections of rgFSHb-a or rgFSHba could significantly increase serum T and E2 levels, induce early ovarian development, reduce hypothalamic gnrh1 mRNA level, and increase hypothalamic cyp19a1b mRNA level. Data in this study suggested that recombinant gonadotropin could be produced in dimer form or single-chain form by P. pastoris, and FSH could regulate steroidogenesis and early ovarian development in juvenile grouper.

  7. Synthetic Covalently Linked Dimeric Form of H2 Relaxin Retains Native RXFP1 Activity and Has Improved In Vitro Serum Stability

    Directory of Open Access Journals (Sweden)

    Vinojini B. Nair

    2015-01-01

    Full Text Available Human (H2 relaxin is a two-chain peptide member of the insulin superfamily and possesses potent pleiotropic roles including regulation of connective tissue remodeling and systemic and renal vasodilation. These effects are mediated through interaction with its cognate G-protein-coupled receptor, RXFP1. H2 relaxin recently passed Phase III clinical trials for the treatment of congestive heart failure. However, its in vivo half-life is short due to its susceptibility to proteolytic degradation and renal clearance. To increase its residence time, a covalent dimer of H2 relaxin was designed and assembled through solid phase synthesis of the two chains, including a judiciously monoalkyne sited B-chain, followed by their combination through regioselective disulfide bond formation. Use of a bisazido PEG7 linker and “click” chemistry afforded a dimeric H2 relaxin with its active site structurally unhindered. The resulting peptide possessed a similar secondary structure to the native monomeric H2 relaxin and bound to and activated RXFP1 equally well. It had fewer propensities to activate RXFP2, the receptor for the related insulin-like peptide 3. In human serum, the dimer had a modestly increased half-life compared to the monomeric H2 relaxin suggesting that additional oligomerization may be a viable strategy for producing longer acting variants of H2 relaxin.

  8. Plasmon excitations in the dimers formed by atom chains

    Science.gov (United States)

    Xue, Hong-jie; Hao, Da-peng; Zhang, Ming; Wang, Xiao-mei

    2017-02-01

    Based on the linear response theory in the random-phase approximation and the free-electron gas model, we study the plasmon excitations in the dimers formed by atom chains. With the help of energy absorption spectrum and charge distribution, the evolutions of longitudinal and transverse plasmon, and the effect of the system parameters such as size, atomic separation and electron filling on plasmon are obtained. In addition, the dipole, quadrupole, end and central plasmon are observed.

  9. Overexpression and characterization of dimeric and tetrameric forms of recombinant serine hydroxymethyltransferase from Bacillus stearothermophilus

    Indian Academy of Sciences (India)

    Venkatakrishna R Jala; V Prakash; N Appaji Rao; H S Savithri

    2002-06-01

    Serine hydroxymethyltransferase (SHMT), a pyridoxal-5′-phosphate (PLP) dependent enzyme catalyzes the interconversion of L-Ser and Gly using tetrahydrofolate as a substrate. The gene encoding for SHMT was amplified by PCR from genomic DNA of Bacillus stearothermophilus and the PCR product was cloned and overexpressed in Escherichia coli. The purified recombinant enzyme was isolated as a mixture of dimer (90%) and tetramer (10%). This is the first report demonstrating the existence of SHMT as a dimer and tetramer in the same organism. The specific activities at 37°C of the dimeric and tetrameric forms were 6.7 U/mg and 4.1 U/mg, respectively. The purified dimer was extremely thermostable with a m of 85°C in the presence of PLP and L-Ser. The temperature optimum of the dimer was 80°C with a specific activity of 32.4 U/mg at this temperature. The enzyme catalyzed tetrahydrofolate-independent reactions at a slower rate compared to the tetrahydrofolate-dependent retro-aldol cleavage of L-Ser. The interaction with substrates and their analogues indicated that the orientation of PLP ring of B. stearothermophilus SHMT was probably different from sheep liver cytosolic recombinant SHMT (scSHMT).

  10. Structural characterization suggests models for monomeric and dimeric forms of full-length ezrin.

    Science.gov (United States)

    Phang, Juanita M; Harrop, Stephen J; Duff, Anthony P; Sokolova, Anna V; Crossett, Ben; Walsh, James C; Beckham, Simone A; Nguyen, Cuong D; Davies, Roberta B; Glöckner, Carina; Bromley, Elizabeth H C; Wilk, Krystyna E; Curmi, Paul M G

    2016-09-15

    Ezrin is a member of the ERM (ezrin-radixin-moesin) family of proteins that have been conserved through metazoan evolution. These proteins have dormant and active forms, where the latter links the actin cytoskeleton to membranes. ERM proteins have three domains: an N-terminal FERM [band Four-point-one (4.1) ERM] domain comprising three subdomains (F1, F2, and F3); a helical domain; and a C-terminal actin-binding domain. In the dormant form, FERM and C-terminal domains form a stable complex. We have determined crystal structures of the active FERM domain and the dormant FERM:C-terminal domain complex of human ezrin. We observe a bistable array of phenylalanine residues in the core of subdomain F3 that is mobile in the active form and locked in the dormant form. As subdomain F3 is pivotal in binding membrane proteins and phospholipids, these transitions may facilitate activation and signaling. Full-length ezrin forms stable monomers and dimers. We used small-angle X-ray scattering to determine the solution structures of these species. As expected, the monomer shows a globular domain with a protruding helical coiled coil. The dimer shows an elongated dumbbell structure that is twice as long as the monomer. By aligning ERM sequences spanning metazoan evolution, we show that the central helical region is conserved, preserving the heptad repeat. Using this, we have built a dimer model where each monomer forms half of an elongated antiparallel coiled coil with domain-swapped FERM:C-terminal domain complexes at each end. The model suggests that ERM dimers may bind to actin in a parallel fashion.

  11. [Antioxidant and antibacterial activities of dimeric phenol compounds].

    Science.gov (United States)

    Ogata, Masahiro

    2008-08-01

    We studied the antioxidant and antibacterial activities of monomeric and dimeric phenol compounds. Dimeric compounds had higher antioxidant activities than monomeric compounds. Electron spin resonance spin-trapping experiments showed that phenol compounds with an allyl substituent on their aromatic rings directly scavenged superoxide, and that only eugenol trapped hydroxyl radicals. We developed a generation system of the hydroxyl radical without using any metals by adding L-DOPA and DMPO to PBS or MiliQ water in vitro. We found that eugenol trapped hydroxyl radicals directly and is metabolized to a dimer. On the other hand, dipropofol, a dimer of propofol, has strong antibacterial activity against Gram-positive bacteria. However, it lacks solubility in water and this property is assumed to limit its efficacy. We tried to improve the solubility and found a new solubilization method of dipropofol in water with the addition of a monosaccharide or ascorbic acid.

  12. Amphiphile dependency of the monomeric and dimeric forms of acetylcholinesterase from human erythrocyte membrane.

    Science.gov (United States)

    Ott, P; Brodbeck, U

    1984-08-08

    Human erythrocyte membrane-bound acetylcholinesterase was converted to a monomeric species by treatment of ghosts with 2-mercaptoethanol and iodoacetic acid. After solubilization with Triton X-100, the reduced and alkylated enzyme was partially purified by affinity chromatography and separated from residual dimeric enzyme by sucrose density gradient centrifugation in a zonal rotor. Monomeric and dimeric acetylcholinesterase showed full enzymatic activity in presence of Triton X-100 whereas in the absence of detergent, activity was decreased to approx. 20% and 15%, respectively. Preformed egg phosphatidylcholine vesicles fully sustained activity of the monomeric species whereas the dimer was only 80% active. The results suggest that a dimeric structure is not required for manifestation of amphiphile dependency of membrane-bound acetylcholinesterase from human erythrocytes. Furthermore, monomeric enzyme appears to be more easily inserted into phospholipid bilayers than the dimeric species.

  13. Monoubiquitination and activity of the paracaspase MALT1 requires glutamate 549 in the dimerization interface.

    Directory of Open Access Journals (Sweden)

    Katrin Cabalzar

    Full Text Available The mucosa-associated lymphoid tissue protein-1 (MALT1, also known as paracaspase is a protease whose activity is essential for the activation of lymphocytes and the growth of cells derived from human diffuse large B-cell lymphomas of the activated B-cell subtype (ABC DLBCL. Crystallographic approaches have shown that MALT1 can form dimers via its protease domain, but why dimerization is relevant for the biological activity of MALT1 remains largely unknown. Using a molecular modeling approach, we predicted Glu 549 (E549 to be localized within the MALT1 dimer interface and thus potentially relevant. Experimental mutation of this residue into alanine (E549A led to a complete impairment of MALT1 proteolytic activity. This correlated with an impaired capacity of the mutant to form dimers of the protease domain in vitro, and a reduced capacity to promote NF-κB activation and transcription of the growth-promoting cytokine interleukin-2 in antigen receptor-stimulated lymphocytes. Moreover, this mutant could not rescue the growth of ABC DLBCL cell lines upon MALT1 silencing. Interestingly, the MALT1 mutant E549A was unable to undergo monoubiquitination, which we identified previously as a critical step in MALT1 activation. Collectively, these findings suggest a model in which E549 at the dimerization interface is required for the formation of the enzymatically active, monoubiquitinated form of MALT1.

  14. Monoubiquitination and activity of the paracaspase MALT1 requires glutamate 549 in the dimerization interface.

    Science.gov (United States)

    Cabalzar, Katrin; Pelzer, Christiane; Wolf, Annette; Lenz, Georg; Iwaszkiewicz, Justyna; Zoete, Vincent; Hailfinger, Stephan; Thome, Margot

    2013-01-01

    The mucosa-associated lymphoid tissue protein-1 (MALT1, also known as paracaspase) is a protease whose activity is essential for the activation of lymphocytes and the growth of cells derived from human diffuse large B-cell lymphomas of the activated B-cell subtype (ABC DLBCL). Crystallographic approaches have shown that MALT1 can form dimers via its protease domain, but why dimerization is relevant for the biological activity of MALT1 remains largely unknown. Using a molecular modeling approach, we predicted Glu 549 (E549) to be localized within the MALT1 dimer interface and thus potentially relevant. Experimental mutation of this residue into alanine (E549A) led to a complete impairment of MALT1 proteolytic activity. This correlated with an impaired capacity of the mutant to form dimers of the protease domain in vitro, and a reduced capacity to promote NF-κB activation and transcription of the growth-promoting cytokine interleukin-2 in antigen receptor-stimulated lymphocytes. Moreover, this mutant could not rescue the growth of ABC DLBCL cell lines upon MALT1 silencing. Interestingly, the MALT1 mutant E549A was unable to undergo monoubiquitination, which we identified previously as a critical step in MALT1 activation. Collectively, these findings suggest a model in which E549 at the dimerization interface is required for the formation of the enzymatically active, monoubiquitinated form of MALT1.

  15. Structure of the dimeric form of CTP synthase from Sulfolobus solfataricus

    DEFF Research Database (Denmark)

    Lauritsen, Iben; Willemoës, Martin; Jensen, Kaj Frank;

    2011-01-01

    CTP synthase catalyzes the last committed step in de novo pyrimidine-nucleotide biosynthesis. Active CTP synthase is a tetrameric enzyme composed of a dimer of dimers. The tetramer is favoured in the presence of the substrate nucleotides ATP and UTP; when saturated with nucleotide, the tetramer c...

  16. Dimerization of tetherin is not essential for its antiviral activity against Lassa and Marburg viruses.

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    Toshie Sakuma

    Full Text Available Tetherin (also known as BST2, CD317 or HM1.24 has recently been reported to inhibit a wide range of viruses. However, the antiviral mechanism of action of tetherin has not been determined. Both ends of the tetherin molecule are associated with the plasma membrane and it forms a homodimer. Therefore, a model in which progeny virions are retained on the cell surface by dimer formation between tetherin molecules on the viral envelope and plasma membrane has been proposed as the antiviral mechanism of action of this molecule. To investigate this possibility, we examined the correlation between dimerization and antiviral activity of tetherin in Lassa and Marburg virus-like particle production systems using tetherin mutants deficient in dimer formation. However, the tetherin mutant with complete loss of dimerization activity still showed apparent antiviral activity, indicating that dimerization of tetherin is not essential for its antiviral activity. This suggests that tetherin retains progeny virions on the cell surface by a mechanism other than dimerization.

  17. Antiviral activity of α-helical stapled peptides designed from the HIV-1 capsid dimerization domain

    Directory of Open Access Journals (Sweden)

    Cowburn David

    2011-05-01

    Full Text Available Abstract Background The C-terminal domain (CTD of HIV-1 capsid (CA, like full-length CA, forms dimers in solution and CTD dimerization is a major driving force in Gag assembly and maturation. Mutations of the residues at the CTD dimer interface impair virus assembly and render the virus non-infectious. Therefore, the CTD represents a potential target for designing anti-HIV-1 drugs. Results Due to the pivotal role of the dimer interface, we reasoned that peptides from the α-helical region of the dimer interface might be effective as decoys to prevent CTD dimer formation. However, these small peptides do not have any structure in solution and they do not penetrate cells. Therefore, we used the hydrocarbon stapling technique to stabilize the α-helical structure and confirmed by confocal microscopy that this modification also made these peptides cell-penetrating. We also confirmed by using isothermal titration calorimetry (ITC, sedimentation equilibrium and NMR that these peptides indeed disrupt dimer formation. In in vitro assembly assays, the peptides inhibited mature-like virus particle formation and specifically inhibited HIV-1 production in cell-based assays. These peptides also showed potent antiviral activity against a large panel of laboratory-adapted and primary isolates, including viral strains resistant to inhibitors of reverse transcriptase and protease. Conclusions These preliminary data serve as the foundation for designing small, stable, α-helical peptides and small-molecule inhibitors targeted against the CTD dimer interface. The observation that relatively weak CA binders, such as NYAD-201 and NYAD-202, showed specificity and are able to disrupt the CTD dimer is encouraging for further exploration of a much broader class of antiviral compounds targeting CA. We cannot exclude the possibility that the CA-based peptides described here could elicit additional effects on virus replication not directly linked to their ability to bind

  18. Dimerization in Highly Concentrated Solutions of Phosphoimidazolide Activated Monomucleotides

    Science.gov (United States)

    Kanavarioti, Anastassia

    1997-08-01

    Phosphoimidazolide activated ribomononucleotides (*pN) are useful substrates for the non-enzymatic synthesis of polynucleotides. However, dilute neutral aqueous solutions of *pN typically yield small amounts of dimers and traces of polymers; most of *pN hydrolyzes to yield nucleoside 5'-monophosphate. Here we report the self-condensation of nucleoside 5'-phosphate 2-methylimidazolide (2-MeImpN with N = cytidine, uridine or guanosine) in the presence of Mg2+ in concentrated solutions, such as might have been found in an evaporating lagoon on prebiotic Earth. The product distribution indicates that oligomerization is favored at the expense of hydrolysis. At 1.0 M, 2-MeImpU and 2-MeImpC produce about 65% of oligomers including 4% of the 3',5'-linked dimer. Examination of the product distribution of the three isomeric dimers in a self-condensation allows identification of reaction pathways that lead to dimer formation. Condensations in a concentrated mixture of all three nucleotides (U,C,G mixtures) is made possible by the enhanced solubility of 2-MeImpG in such mixtures. Although percent yield of internucleotide linked dimers is enhanced as a function of initial monomer concentration, pyrophosphate dimer yields remain practically unchanged at about 20% for 2-MeImpU, 16% for 2-MeImpC and 25% of the total pyrophosphate in the U,C,G mixtures. The efficiency by which oligomers are produced in these concentrated solutions makes the evaporating lagoon scenario a potentially interesting medium for the prebiotic synthesis of dimers and short RNAs.

  19. Dimer-tetramer transition controls RUNX1/ETO leukemogenic activity.

    Science.gov (United States)

    Wichmann, Christian; Becker, Yvonne; Chen-Wichmann, Linping; Vogel, Vitali; Vojtkova, Anna; Herglotz, Julia; Moore, Sandra; Koch, Joachim; Lausen, Jörn; Mäntele, Werner; Gohlke, Holger; Grez, Manuel

    2010-07-29

    RUNX1/ETO, the fusion protein resulting from the chromosomal translocation t(8;21), is one of the most frequent translocation products in acute myeloid leukemia. Several in vitro and in vivo studies have shown that the homo-tetramerization domain of ETO, the nervy homology region 2 (NHR2), is essential for RUNX1/ETO oncogenic activity. We analyzed the energetic contribution of individual amino acids within the NHR2 to RUNX1/ETO dimer-tetramer transition and found a clustered area of 5 distinct amino acids with strong contribution to the stability of tetramers. Substitution of these amino acids abolishes tetramer formation without affecting dimer formation. Similar to RUNX1/ETO monomers, dimers failed to bind efficiently to DNA and to alter expression of RUNX1-dependent genes. RUNX1/ETO dimers do not block myeloid differentiation, are unable to enhance the self-renewal capacity of hematopoietic progenitors, and fail to induce leukemia in a murine transplantation model. Our data reveal the existence of an essential structural motif (hot spot) at the NHR2 dimer-tetramer interface, suitable for a molecular intervention in t(8;21) leukemias.

  20. Human cystatin C forms an inactive dimer during intracellular trafficking in transfected CHO cells

    DEFF Research Database (Denmark)

    Merz, G S; Benedikz, Eirikur; Schwenk, V

    1997-01-01

    To define the cellular processing of human cystatin C as well as to lay the groundwork for investigating its contribution to lcelandic Hereditary Cerebral Hemorrhage with Amyloidosis (HCHWA-I), we have characterized the trafficking, secretion, and extracellular fate of human cystatin C...... that the cystatin C dimer, formed during intracellular trafficking, is converted to monomer at or before secretion. Cells in which exit from the endoplasmic reticulum (ER) was blocked with brefeldin A contained the 33 kDa species, indicating that cystatin C dimerization occurs in the ER. After removal of brefeldin......, presumably as a consequence of the low pH of late endosome/lysosomes. As a dimer, cystatin C would be prevented from inhibiting the lysosomal cysteine proteases. These results reveal a novel mechanism, transient dimerization, by which cystatin C is inactivated during the early part of its trafficking through...

  1. Structural and Vibrational Study on Monomer and Dimer Forms and Water Clusters of Acetazolamide

    Directory of Open Access Journals (Sweden)

    Aysen E. Ozel

    2013-01-01

    Full Text Available Experimental IR and Raman spectra of solid acetazolamide have been analysed by computing the molecular structures and vibrational spectra of monomer and dimer forms and water clusters of acetazolamide. The possible stable conformers of free acetazolamide molecule in the ground state were obtained by scanning the potential energy surface through the dihedral angles, D1 (1S-2C-6S-9N, D2 (4N-5C-12N-14C, and D3 (5C-12N-14C-16C. The final geometry parameters for the obtained stable conformers were determined by means of geometry optimization, carried out at DFT/B3LYP/6-31G++(d,p theory level. Afterwards the possible dimer forms of the molecule and acetazolamide-H2O clusters were formed and their energetically preferred conformations were investigated using the same method and the same level of theory. The effect of BSSE on the structure and energy of acetazolamide dimer has been investigated. The assignment of the vibrational modes was performed based on the potential energy distribution of the vibrational modes, calculated by using GAR2PED program. The experimental vibrational wavenumbers of solid acetazolamide are found to be in better agreement with the calculated wavenumbers of dimer form of acetazolamide than those of its monomeric form. NBO analysis has been performed on both monomer and dimer geometries.

  2. The dimeric form of HLA-G molecule is associated with the response of early rheumatoid arthritis (ERA) patients to methotrexate.

    Science.gov (United States)

    Rizzo, Roberta; Farina, Ilaria; Bortolotti, Daria; Galuppi, Elisa; Padovan, Melissa; Di Luca, Dario; Govoni, Marcello

    2017-03-01

    A growing body of evidence indicates a possible involvement of HLA (human leukocyte antigen)-G antigens in rheumatoid arthritis (RA), mainly in the HLA-G dimeric isoform, the most active HLA-G form with the strongest immunosuppression, that showed an excellent anti-inflammatory effect in collagen-induced arthritis model mice. However, the relevance of HLA-G dimers in RA response to methotrexate (MTX) treatment is still unknown. We analyzed the HLA-G dimers' amount in plasma samples from early rheumatoid arthritis (ERA) patients before MTX therapy and evaluated the role of these molecules as biomarker of the different response to the treatment. Plasma sHLA-G levels were detected by ELISA, and HLA-G dimeric and monomeric forms were revealed by Western blot in 12 MTX responder (reaching DAS28 remission G levels and the 78 kDa HLA-G dimeric form. Unresponsive ERA patients were characterized by lower plasma sHLA-G levels, and only one patient presented the 78 kDa HLA-G dimeric form (DAS28 5.1). Our preliminary results support the hypothesis that in ERA patients, sHLA-G and, in particular, the presence of the dimeric form in plasma samples before MTX therapy could be an a priori biomarker for the response to MTX treatment.

  3. Formation of the Ras dimer is essential for Raf-1 activation.

    Science.gov (United States)

    Inouye, K; Mizutani, S; Koide, H; Kaziro, Y

    2000-02-11

    Although it is well established that Ras requires membrane localization for activation of its target molecule, Raf-1, the reason for this requirement is not fully understood. In this study, we found that modified Ras, which is purified from Sf9 cells, could activate Raf-1 in a cell-free system, when incorporated into liposome. Using a bifunctional cross-linker and a protein-fragmentation complementation assay, we detected dimer formation of Ras in the liposome and in the intact cells, respectively. These results suggest that dimerization of Ras in the lipid membrane is essential for activation of Raf-1. To support this, we found that, when fused to glutathione S-transferase (GST), unprocessed Ras expressed in Escherichia coli could bypass the requirement for liposome. A Ras-dependent Raf-1 activator, which we previously reported (Mizutani, S., Koide, H., and Kaziro, Y. (1998) Oncogene 16, 2781-2786), was still required for Raf-1 activation by GST-Ras. Furthermore, an enforced dimerization of unmodified oncogenic Ras mutant in human embryonic kidney (HEK) 293 cells, using a portion of gyrase B or estrogen receptor, also resulted in activation of Raf-1. From these results, we conclude that membrane localization allows Ras to form a dimer, which is essential, although not sufficient, for Raf-1 activation.

  4. Dimerization via tandem leucine zippers is essential for the activation of the mitogen-activated protein kinase kinase kinase, MLK-3.

    Science.gov (United States)

    Leung, I W; Lassam, N

    1998-12-04

    Mixed lineage kinase-3 (MLK-3) is a mitogen-activated kinase kinase kinase that mediates stress-activating protein kinase (SAPK)/c-Jun NH2-terminal kinase activation. MLK-3 and other MLK family kinases are characterized by the presence of multiple protein-protein interaction domains including a tandem leucine/isoleucine zipper (LZs) motif. Leucine zippers are known to mediate protein dimerization raising the possibility that the tandem leucine/isoleucine zippers may function as a dimerization motif of MLK-3. Using both co-immunoprecipitation and nonreducing SDS-polyacrylamide gel electrophoresis, we demonstrated that MLK-3 forms disulfide bridged homo-dimers and that the LZs motif is sufficient for MLK-3 homodimerization. We next asked whether MLK-3 utilizes a dimerization-based activation mechanism analogous to that of receptor tyrosine kinases. We found that dimerization via the LZs motif is a prerequisite for MLK-3 autophosphorylation. We then demonstrated that co-expression of Cdc42 lead to a substantial increase in MLK-3 dimerization, indicating that binding by this GTPase may induce MLK-3 dimerization. Moreover, the LZs minus form of MLK-3 failed to activate the downstream target SAPK, and expression of a MLK-3 LZs polypeptide was found to block SAPK activation by wild type MLK-3. Taken together, these findings indicate that dimerization plays a pivotal role in MLK-3 activation.

  5. Dimerization of translationally controlled tumor protein is essential for its cytokine-like activity.

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    Miyoung Kim

    Full Text Available BACKGROUND: Translationally Controlled Tumor Protein (TCTP found in nasal lavage fluids of allergic patients was named IgE-dependent histamine-releasing factor (HRF. Human recombinant HRF (HrHRF has been recently reported to be much less effective than HRF produced from activated mononuclear cells (HRFmn. METHODS AND FINDINGS: We found that only NH(2-terminal truncated, but not C-terminal truncated, TCTP shows cytokine releasing activity compared to full-length TCTP. Interestingly, only NH(2-terminal truncated TCTP, unlike full-length TCTP, forms dimers through intermolecular disulfide bonds. We tested the activity of dimerized full-length TCTP generated by fusing it to rabbit Fc region. The untruncated-full length protein (Fc-HrTCTP was more active than HrTCTP in BEAS-2B cells, suggesting that dimerization of TCTP, rather than truncation, is essential for the activation of TCTP in allergic responses. We used confocal microscopy to evaluate the affinity of TCTPs to its putative receptor. We detected stronger fluorescence in the plasma membrane of BEAS-2B cells incubated with Del-N11TCTP than those incubated with rat recombinant TCTP (RrTCTP. Allergenic activity of Del-N11TCTP prompted us to see whether the NH(2-terminal truncated TCTP can induce allergic airway inflammation in vivo. While RrTCTP had no influence on airway inflammation, Del-N11TCTP increased goblet cell hyperplasia in both lung and rhinal cavity. The dimerized protein was found in sera from allergic patients, and bronchoalveolar lavage fluids from airway inflamed mice. CONCLUSIONS: Dimerization of TCTP seems to be essential for its cytokine-like activity. Our study has potential to enhance the understanding of pathogenesis of allergic disease and provide a target for allergic drug development.

  6. Novel dimeric β-helical model of an ice nucleation protein with bridged active sites

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    Walker Virginia K

    2011-09-01

    Full Text Available Abstract Background Ice nucleation proteins (INPs allow water to freeze at high subzero temperatures. Due to their large size (>120 kDa, membrane association, and tendency to aggregate, an experimentally-determined tertiary structure of an INP has yet to be reported. How they function at the molecular level therefore remains unknown. Results Here we have predicted a novel β-helical fold for the INP produced by the bacterium Pseudomonas borealis. The protein uses internal serine and glutamine ladders for stabilization and is predicted to dimerize via the burying of a solvent-exposed tyrosine ladder to make an intimate hydrophobic contact along the dimerization interface. The manner in which PbINP dimerizes also allows for its multimerization, which could explain the aggregation-dependence of INP activity. Both sides of the PbINP structure have tandem arrays of amino acids that can organize waters into the ice-like clathrate structures seen on antifreeze proteins. Conclusions Dimerization dramatically increases the 'ice-active' surface area of the protein by doubling its width, increasing its length, and presenting identical ice-forming surfaces on both sides of the protein. We suggest that this allows sufficient anchored clathrate waters to align on the INP surface to nucleate freezing. As PbINP is highly similar to all known bacterial INPs, we predict its fold and mechanism of action will apply to these other INPs.

  7. Apolipoprotein-E forms dimers in human frontal cortex and hippocampus

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    Halliday Glenda M

    2010-02-01

    Full Text Available Abstract Background Apolipoprotein-E (apoE plays important roles in neurobiology and the apoE4 isoform increases risk for Alzheimer's disease (AD. ApoE3 and apoE2 are known to form disulphide-linked dimers in plasma and cerebrospinal fluid whereas apoE4 cannot form these dimers as it lacks a cysteine residue. Previous in vitro research indicates dimerisation of apoE3 has a significant impact on its functions related to cholesterol homeostasis and amyloid-beta peptide degradation. The possible occurrence of apoE dimers in cortical tissues has not been examined and was therefore assessed. Human frontal cortex and hippocampus from control and AD post-mortem samples were homogenised and analysed for apoE by western blotting under both reducing and non-reducing conditions. Results In apoE3 homozygous samples, ~12% of apoE was present as a homodimer and ~2% was detected as a 43 kDa heterodimer. The level of dimerisation was not significantly different when control and AD samples were compared. As expected, these dimerised forms of apoE were not detected in apoE4 homozygous samples but were detected in apoE3/4 heterozygotes at a level approximately 60% lower than seen in the apoE3 homozygous samples. Similar apoE3 dimers were also detected in lysates of SK-N-SH neuroblastoma cells and in freshly prepared rabbit brain homogenates. The addition of the thiol trapping agent, iodoacetamide, to block reactive thiols during both human and rabbit brain sample homogenisation and processing did not reduce the amount of apoE homodimer recovered. These data indicate that the apoE dimers we detected in the human brain are not likely to be post-mortem artefacts. Conclusion The identification of disulphide-linked apoE dimers in human cortical and hippocampal tissues represents a distinct structural difference between the apoE3 and apoE4 isoforms that may have functional consequences.

  8. Monomeric banana lectin at acidic pH overrules conformational stability of its native dimeric form.

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    Javed M Khan

    Full Text Available Banana lectin (BL is a homodimeric protein categorized among jacalin-related family of lectins. The effect of acidic pH was examined on conformational stability of BL by using circular dichroism, intrinsic fluorescence, 1-anilino-8-napthalene sulfonate (ANS binding, size exclusion chromatography (SEC and dynamic light scattering (DLS. During acid denaturation of BL, the monomerization of native dimeric protein was found at pH 2.0. The elution profile from SEC showed two different peaks (59.65 ml & 87.98 ml at pH 2.0 while single peak (61.45 ml at pH 7.4. The hydrodynamic radii (R h of native BL was 2.9 nm while at pH 2.0 two species were found with R h of 1.7 and 3.7 nm. Furthermore at, pH 2.0 the secondary structures of BL remained unaltered while tertiary structure was significantly disrupted with the exposure of hydrophobic clusters confirming the existence of molten globule like state. The unfolding of BL with different subunit status was further evaluated by urea and temperature mediated denaturation to check their stability. As inferred from high Cm and ΔG values, the monomeric form of BL offers more resistance towards chemical denaturation than the native dimeric form. Besides, dimeric BL exhibited a Tm of 77°C while no loss in secondary structures was observed in monomers even up to 95°C. To the best of our knowledge, this is the first report on monomeric subunit of lectins showing more stability against denaturants than its native dimeric state.

  9. Inhibition of RAF Isoforms and Active Dimers by LY3009120 Leads to Anti-tumor Activities in RAS or BRAF Mutant Cancers.

    Science.gov (United States)

    Peng, Sheng-Bin; Henry, James R; Kaufman, Michael D; Lu, Wei-Ping; Smith, Bryan D; Vogeti, Subha; Rutkoski, Thomas J; Wise, Scott; Chun, Lawrence; Zhang, Youyan; Van Horn, Robert D; Yin, Tinggui; Zhang, Xiaoyi; Yadav, Vipin; Chen, Shih-Hsun; Gong, Xueqian; Ma, Xiwen; Webster, Yue; Buchanan, Sean; Mochalkin, Igor; Huber, Lysiane; Kays, Lisa; Donoho, Gregory P; Walgren, Jennie; McCann, Denis; Patel, Phenil; Conti, Ilaria; Plowman, Gregory D; Starling, James J; Flynn, Daniel L

    2015-09-14

    LY3009120 is a pan-RAF and RAF dimer inhibitor that inhibits all RAF isoforms and occupies both protomers in RAF dimers. Biochemical and cellular analyses revealed that LY3009120 inhibits ARAF, BRAF, and CRAF isoforms with similar affinity, while vemurafenib or dabrafenib have little or modest CRAF activity compared to their BRAF activities. LY3009120 induces BRAF-CRAF dimerization but inhibits the phosphorylation of downstream MEK and ERK, suggesting that it effectively inhibits the kinase activity of BRAF-CRAF heterodimers. Further analyses demonstrated that LY3009120 also inhibits various forms of RAF dimers including BRAF or CRAF homodimers. Due to these unique properties, LY3009120 demonstrates minimal paradoxical activation, inhibits MEK1/2 phosphorylation, and exhibits anti-tumor activities across multiple models carrying KRAS, NRAS, or BRAF mutation.

  10. Form, function and functionality of two dimeric toluene-2,4-diisocyanate polymorphs.

    Science.gov (United States)

    Vella-Zarb, Liana; Dinnebier, Robert E

    2012-04-01

    2,4-Dioxo-1,3-diazetidine-1,3-bis(methyl-m-phenylene) diisocyanate (dimerized toluene-2,4-diisocyanate, TDI) is one of the most widely used aromatic diisocyanates in the polymer industry, and it crystallizes in at least two polymorphic forms (form A and form B) depending on reaction conditions. The crystal structures of the two forms were determined from high-resolution laboratory X-ray powder diffraction data using simulated annealing and Rietveld refinement. In spite of a marked structural similarity between them, significant discrepancies in the physical properties of the two forms prompted analysis of their partitioned energy terms in an effort to better our understanding of the driving force behind such differences in behaviour.

  11. Synthesis and antiviral activity of new dimeric inhibitors against HIV-1

    DEFF Research Database (Denmark)

    Danel, Krzysztof; Larsen, Louise M.; Pedersen, Erik Bjerreg.;

    2008-01-01

    by Sonogashira reaction, ‘click' chemistry or Pd-catalyzed oxidative coupling. The iodo precursor 5 turned out as a potent compound against wild type and mutated HIV-1 virus. All dimeric compounds showed lower activity against HIV-1 than MKC-442, except the asymmetric dimer of AZT and 1a which showed an activity...

  12. Packing interface energetics in different crystal forms of the λ Cro dimer.

    Science.gov (United States)

    Ahlstrom, Logan S; Miyashita, Osamu

    2014-07-01

    Variation among crystal structures of the λ Cro dimer highlights conformational flexibility. The structures range from a wild type closed to a mutant fully open conformation, but it is unclear if each represents a stable solution state or if one may be the result of crystal packing. Here we use molecular dynamics (MD) simulation to investigate the energetics of crystal packing interfaces and the influence of site-directed mutagenesis on them in order to examine the effect of crystal packing on wild type and mutant Cro dimer conformation. Replica exchange MD of mutant Cro in solution shows that the observed conformational differences between the wild type and mutant protein are not the direct consequence of mutation. Instead, simulation of Cro in different crystal environments reveals that mutation affects the stability of crystal forms. Molecular Mechanics Poisson-Boltzmann Surface Area binding energy calculations reveal the detailed energetics of packing interfaces. Packing interfaces can have diverse properties in strength, energetic components, and some are stronger than the biological dimer interface. Further analysis shows that mutation can strengthen packing interfaces by as much as ∼5 kcal/mol in either crystal environment. Thus, in the case of Cro, mutation provides an additional energetic contribution during crystal formation that may stabilize a fully open higher energy state. Moreover, the effect of mutation in the lattice can extend to packing interfaces not involving mutation sites. Our results provide insight into possible models for the effect of crystallization on Cro conformational dynamics and emphasize careful consideration of protein crystal structures.

  13. Active control of nano dimers response using piezoelectric effect

    Science.gov (United States)

    Mekkawy, Ahmed A.; Ali, Tamer A.; Badawi, Ashraf H.

    2016-09-01

    Nano devices can be used as building blocks for Internet of Nano-Things network devices, such as sensors/actuators, transceivers, and routers. Although nano particles response can be engineered to fit in different regimes, for such a nano particle to be used as an active nano device, its properties should be dynamically controlled. This controllability is a challenge, and there are many proposed techniques to tune nanoparticle response on the spot through a sort of control signal, wither that signal is optical (for all-optical systems) or electronic (for opto-electronic systems). This will allow the use of nano particles as nano-switches or as dynamic sensors that can pick different frequencies depending on surrounding conditions or depending on a smart decisions. In this work, we propose a piezoelectric substrate as an active control mediator to control plasmonic gaps in nano dimers. This method allows for integrating nano devices with regular electronics while communicating control signals to nano devices through applying electric signals to a piezoelectric material, in order to control the gaps between nano particles in a nano cluster. We do a full numerical study to the system, analyzing the piezoelectric control resolution (minimum gap change step) and its effect on a nanodimer response as a nanoantenna. This analysis considers the dielectric functions of materials within the visible frequencies range. The effects of different parameters, such as the piezoelectric geometrical structure and materials, on the gap control resolution and the operating frequency are studied.

  14. The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA.

    Science.gov (United States)

    Booth, David S; Cheng, Yifan; Frankel, Alan D

    2014-12-08

    The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-Ran(GTP) nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

  15. The EBNA-2 N-Terminal Transactivation Domain Folds into a Dimeric Structure Required for Target Gene Activation.

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    Anders Friberg

    2015-05-01

    Full Text Available Epstein-Barr virus (EBV is a γ-herpesvirus that may cause infectious mononucleosis in young adults. In addition, epidemiological and molecular evidence links EBV to the pathogenesis of lymphoid and epithelial malignancies. EBV has the unique ability to transform resting B cells into permanently proliferating, latently infected lymphoblastoid cell lines. Epstein-Barr virus nuclear antigen 2 (EBNA-2 is a key regulator of viral and cellular gene expression for this transformation process. The N-terminal region of EBNA-2 comprising residues 1-58 appears to mediate multiple molecular functions including self-association and transactivation. However, it remains to be determined if the N-terminus of EBNA-2 directly provides these functions or if these activities merely depend on the dimerization involving the N-terminal domain. To address this issue, we determined the three-dimensional structure of the EBNA-2 N-terminal dimerization (END domain by heteronuclear NMR-spectroscopy. The END domain monomer comprises a small fold of four β-strands and an α-helix which form a parallel dimer by interaction of two β-strands from each protomer. A structure-guided mutational analysis showed that hydrophobic residues in the dimer interface are required for self-association in vitro. Importantly, these interface mutants also displayed severely impaired self-association and transactivation in vivo. Moreover, mutations of solvent-exposed residues or deletion of the α-helix do not impair dimerization but strongly affect the functional activity, suggesting that the EBNA-2 dimer presents a surface that mediates functionally important intra- and/or intermolecular interactions. Our study shows that the END domain is a novel dimerization fold that is essential for functional activity. Since this specific fold is a unique feature of EBNA-2 it might provide a novel target for anti-viral therapeutics.

  16. Dimerization of inositol monophosphatase Mycobacterium tuberculosis SuhB is not constitutive, but induced by binding of the activator Mg2+

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    Nigou Jérôme

    2007-08-01

    Full Text Available Abstract Background The cell wall of Mycobacterium tuberculosis contains a wide range of phosphatidyl inositol-based glycolipids that play critical structural roles and, in part, govern pathogen-host interactions. Synthesis of phosphatidyl inositol is dependent on free myo-inositol, generated through dephosphorylation of myo-inositol-1-phosphate by inositol monophosphatase (IMPase. Human IMPase, the putative target of lithium therapy, has been studied extensively, but the function of four IMPase-like genes in M. tuberculosis is unclear. Results We determined the crystal structure, to 2.6 Å resolution, of the IMPase M. tuberculosis SuhB in the apo form, and analysed self-assembly by analytical ultracentrifugation. Contrary to the paradigm of constitutive dimerization of IMPases, SuhB is predominantly monomeric in the absence of the physiological activator Mg2+, in spite of a conserved fold and apparent dimerization in the crystal. However, Mg2+ concentrations that result in enzymatic activation of SuhB decisively promote dimerization, with the inhibitor Li+ amplifying the effect of Mg2+, but failing to induce dimerization on its own. Conclusion The correlation of Mg2+-driven enzymatic activity with dimerization suggests that catalytic activity is linked to the dimer form. Current models of lithium inhibition of IMPases posit that Li+ competes for one of three catalytic Mg2+ sites in the active site, stabilized by a mobile loop at the dimer interface. Our data suggest that Mg2+/Li+-induced ordering of this loop may promote dimerization by expanding the dimer interface of SuhB. The dynamic nature of the monomer-dimer equilibrium may also explain the extended concentration range over which Mg2+ maintains SuhB activity.

  17. Properties and metathesis activity of monomeric and dimeric Mo centres variously located on γ-alumina A DFT study

    Science.gov (United States)

    Handzlik, Jarosław

    2007-05-01

    Ethene metathesis proceeding on monomeric and dimeric Mo species on the (1 0 0) and (1 1 0) γ-alumina is investigated by density functional theory, applying the cluster approach. The calculated vibrational frequencies of the surface OH groups are assigned to the experimental IR bands. It is shown that both monomeric and dimeric Mo forms can be the active sites of olefin metathesis. Metathesis activity and stability of the Mo-methylidene centres depend on their location on alumina. The differences in the sites reactivity are explained on the basis of their geometrical and electronic structure parameters. For the monomeric centres, isomerisation of the trigonal bipyramidal intermediate to the stable square pyramidal molybdacyclobutane is kinetically favoured over the cycloreversal step. The situation is opposite in the case of the dimeric species.

  18. Structure of 1,5-Anhydro-D-Fructose: X-ray Analysis of Crystalline Acetylated Dimeric Forms

    DEFF Research Database (Denmark)

    Lundt, Inge; Andersen, Søren Møller; Marcussen, Jan

    1998-01-01

    Acetylation of 1,5-anhydro-D-fructose under acidic conditions gave two crystalline acetylated dimeric forms, which by X-ray analysis were shown to be diastereomeric spiroketals formed between C-2 and C-2´/C-3´. The structures of the compounds differed only at the configuration at C-2. Acetylation...

  19. Influence of linker length and composition on enzymatic activity and ribosomal binding of neomycin dimers.

    Science.gov (United States)

    Watkins, Derrick; Kumar, Sunil; Green, Keith D; Arya, Dev P; Garneau-Tsodikova, Sylvie

    2015-07-01

    The human and bacterial A site rRNA binding as well as the aminoglycoside-modifying enzyme (AME) activity against a series of neomycin B (NEO) dimers is presented. The data indicate that by simple modifications of linker length and composition, substantial differences in rRNA selectivity and AME activity can be obtained. We tested five different AMEs with dimeric NEO dimers that were tethered via triazole, urea, and thiourea linkages. We show that triazole-linked dimers were the worst substrates for most AMEs, with those containing the longer linkers showing the largest decrease in activity. Thiourea-linked dimers that showed a decrease in activity by AMEs also showed increased bacterial A site binding, with one compound (compound 14) even showing substantially reduced human A site binding. The urea-linked dimers showed a substantial decrease in activity by AMEs when a conformationally restrictive phenyl linker was introduced. The information learned herein advances our understanding of the importance of the linker length and composition for the generation of dimeric aminoglycoside antibiotics capable of avoiding the action of AMEs and selective binding to the bacterial rRNA over binding to the human rRNA.

  20. Euglena gracilis ascorbate peroxidase forms an intramolecular dimeric structure: its unique molecular characterization.

    Science.gov (United States)

    Ishikawa, Takahiro; Tajima, Naoko; Nishikawa, Hitoshi; Gao, Yongshun; Rapolu, Madhusudhan; Shibata, Hitoshi; Sawa, Yoshihiro; Shigeoka, Shigeru

    2010-02-09

    Euglena gracilis lacks a catalase and contains a single APX (ascorbate peroxidase) and enzymes related to the redox cycle of ascorbate in the cytosol. In the present study, a full-length cDNA clone encoding the Euglena APX was isolated and found to contain an open reading frame encoding a protein of 649 amino acids with a calculated molecular mass of 70.5 kDa. Interestingly, the enzyme consisted of two entirely homologous catalytic domains, designated APX-N and APX-C, and an 102 amino acid extension in the N-terminal region, which had a typical class II signal proposed for plastid targeting in Euglena. A computer-assisted analysis indicated a novel protein structure with an intramolecular dimeric structure. The analysis of cell fractionation showed that the APX protein is distributed in the cytosol, but not the plastids, suggesting that Euglena APX becomes mature in the cytosol after processing of the precursor. The kinetics of the recombinant mature FL (full-length)-APX and the APX-N and APX-C domains with ascorbate and H2O2 were almost the same as that of the native enzyme. However, the substrate specificity of the mature FL-APX and the native enzyme was different from that of APX-N and APX-C. The mature FL-APX, but not the truncated forms, could reduce alkyl hydroperoxides, suggesting that the dimeric structure is correlated with substrate recognition. In Euglena cells transfected with double-stranded RNA, the silencing of APX expression resulted in a significant increase in the cellular level of H2O2, indicating the physiological importance of APX to the metabolism of H2O2.

  1. Tor forms a dimer through an N-terminal helical solenoid with a complex topology.

    Science.gov (United States)

    Baretić, Domagoj; Berndt, Alex; Ohashi, Yohei; Johnson, Christopher M; Williams, Roger L

    2016-04-13

    The target of rapamycin (Tor) is a Ser/Thr protein kinase that regulates a range of anabolic and catabolic processes. Tor is present in two complexes, TORC1 and TORC2, in which the Tor-Lst8 heterodimer forms a common sub-complex. We have determined the cryo-electron microscopy (EM) structure of Tor bound to Lst8. Two Tor-Lst8 heterodimers assemble further into a dyad-symmetry dimer mediated by Tor-Tor interactions. The first 1,300 residues of Tor form a HEAT repeat-containing α-solenoid with four distinct segments: a highly curved 800-residue N-terminal 'spiral', followed by a 400-residue low-curvature 'bridge' and an extended 'railing' running along the bridge leading to the 'cap' that links to FAT region. This complex topology was verified by domain insertions and offers a new interpretation of the mTORC1 structure. The spiral of one TOR interacts with the bridge of another, which together form a joint platform for the Regulatory Associated Protein of TOR (RAPTOR) regulatory subunit.

  2. Tor forms a dimer through an N-terminal helical solenoid with a complex topology

    Science.gov (United States)

    Baretić, Domagoj; Berndt, Alex; Ohashi, Yohei; Johnson, Christopher M.; Williams, Roger L.

    2016-04-01

    The target of rapamycin (Tor) is a Ser/Thr protein kinase that regulates a range of anabolic and catabolic processes. Tor is present in two complexes, TORC1 and TORC2, in which the Tor-Lst8 heterodimer forms a common sub-complex. We have determined the cryo-electron microscopy (EM) structure of Tor bound to Lst8. Two Tor-Lst8 heterodimers assemble further into a dyad-symmetry dimer mediated by Tor-Tor interactions. The first 1,300 residues of Tor form a HEAT repeat-containing α-solenoid with four distinct segments: a highly curved 800-residue N-terminal 'spiral', followed by a 400-residue low-curvature 'bridge' and an extended `railing' running along the bridge leading to the 'cap' that links to FAT region. This complex topology was verified by domain insertions and offers a new interpretation of the mTORC1 structure. The spiral of one TOR interacts with the bridge of another, which together form a joint platform for the Regulatory Associated Protein of TOR (RAPTOR) regulatory subunit.

  3. Dimer-dependent intrinsic/basal activity of the class B G protein-coupled receptor PAC1 promotes cellular anti-apoptotic activity through Wnt/β-catenin pathways that are associated with dimer endocytosis.

    Directory of Open Access Journals (Sweden)

    Rongjie Yu

    Full Text Available The high expression of PACAP (pituitary adenylate cyclase-activating polypeptide-preferring receptor PAC1 is associated with nerve injury and tumors. Our previous report (Yu R, et al. PLoS One 2012; 7: e51811 confirmed the dimerization of PAC1 and found that the M-PAC1 mutation in the N-terminal first Cys/Ala lost the ability to form dimers. In this study, Chinese hamster ovary (CHO-K1 cells overexpressing wild-type PAC1 (PAC1-CHO had significantly higher anti-apoptotic activities against serum withdrawal-induced apoptosis associated with a lower caspase 3 activity and a higher Bcl-2 level in a ligand-independent manner than those of CHO cells overexpressing the mutant M-PAC1 (M-PAC1-CHO. PAC1-CHO had significantly higher β-catenin, cyclin D1 and c-myc levels corresponding to the Wnt/β-catenin signal than did M-PAC1-CHO. In addition, the Wnt/β-catenin pathway inhibitor XAV939 significantly inhibited the anti-apoptotic activities of PAC1-CHO. Top-flash assays demonstrated that PAC1-CHO had a significantly stronger Wnt/β-catenin signal than did M-PAC1-CHO. Acetylcysteine (NAC as an inhibitor of the dimerization of PAC1 inhibited the anti-apoptotic activities that were endowed by PAC1 and decreased the Wnt/β-catenin signal in Top-flash assays. In the PAC1 Tet (tetracycline-on inducible gene expression system by doxycycline (Dox, higher expression levels of PAC1 resulted in higher anti-apoptotic activities that were associated with a stronger Wnt/β-catenin signal. A similar correlation was also found with the down-regulation of PAC1 in the Neuro2a neuroblastoma cell. BiFC combined with fluorescence confocal imaging indicated that during serum-withdrawal-induced apoptosis, PAC1 dimers displayed significant endocytosis. These findings indicate that PAC1 has ligand-independent and dimer-dependent intrinsic/basal activity, conferring cells with anti-apoptotic activities against serum withdrawal, which is involved in the Wnt/β-catenin signal and

  4. The crystal structure of human IRE1 luminal domain reveals a conserved dimerization interface required for activation of the unfolded protein response

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jiahai; Liu, Chuan Yin; Back, Sung Hoon; Clark, Robert L.; Peisach, Daniel; Xu, Zhaohui; Kaufman, Randal J. (Michigan)

    2010-03-08

    The unfolded protein response (UPR) is an evolutionarily conserved mechanism by which all eukaryotic cells adapt to the accumulation of unfolded proteins in the endoplasmic reticulum (ER). Inositol-requiring kinase 1 (IRE1) and PKR-related ER kinase (PERK) are two type I transmembrane ER-localized protein kinase receptors that signal the UPR through a process that involves homodimerization and autophosphorylation. To elucidate the molecular basis of the ER transmembrane signaling event, we determined the x-ray crystal structure of the luminal domain of human IRE1{alpha}. The monomer of the luminal domain comprises a unique fold of a triangular assembly of {beta}-sheet clusters. Structural analysis identified an extensive dimerization interface stabilized by hydrogen bonds and hydrophobic interactions. Dimerization creates an MHC-like groove at the interface. However, because this groove is too narrow for peptide binding and the purified luminal domain forms high-affinity dimers in vitro, peptide binding to this groove is not required for dimerization. Consistent with our structural observations, mutations that disrupt the dimerization interface produced IRE1{alpha} molecules that failed to either dimerize or activate the UPR upon ER stress. In addition, mutations in a structurally homologous region within PERK also prevented dimerization. Our structural, biochemical, and functional studies in vivo altogether demonstrate that IRE1 and PERK have conserved a common molecular interface necessary and sufficient for dimerization and UPR signaling.

  5. ERIS, an endoplasmic reticulum IFN stimulator, activates innate immune signaling through dimerization.

    Science.gov (United States)

    Sun, Wenxiang; Li, Yang; Chen, Lu; Chen, Huihui; You, Fuping; Zhou, Xiang; Zhou, Yi; Zhai, Zhonghe; Chen, Danying; Jiang, Zhengfan

    2009-05-26

    We report here the identification and characterization of a protein, ERIS, an endoplasmic reticulum (ER) IFN stimulator, which is a strong type I IFN stimulator and plays a pivotal role in response to both non-self-cytosolic RNA and dsDNA. ERIS (also known as STING or MITA) resided exclusively on ER membrane. The ER retention/retrieval sequence RIR was found to be critical to retain the protein on ER membrane and to maintain its integrity. ERIS was dimerized on innate immune challenges. Coumermycin-induced ERIS dimerization led to strong and fast IFN induction, suggesting that dimerization of ERIS was critical for self-activation and subsequent downstream signaling.

  6. Crystal Structure of PKG I:cGMP Complex Reveals a cGMP-Mediated Dimeric Interface that Facilitates cGMP-Induced Activation

    KAUST Repository

    Kim, Jeong Joo

    2016-04-09

    Cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG) is a key regulator of smooth muscle and vascular tone and represents an important drug target for treating hypertensive diseases and erectile dysfunction. Despite its importance, its activation mechanism is not fully understood. To understand the activation mechanism, we determined a 2.5 Å crystal structure of the PKG I regulatory (R) domain bound with cGMP, which represents the activated state. Although we used a monomeric domain for crystallization, the structure reveals that two R domains form a symmetric dimer where the cGMP bound at high-affinity pockets provide critical dimeric contacts. Small-angle X-ray scattering and mutagenesis support this dimer model, suggesting that the dimer interface modulates kinase activation. Finally, structural comparison with the homologous cyclic AMP-dependent protein kinase reveals that PKG is drastically different from protein kinase A in its active conformation, suggesting a novel activation mechanism for PKG. Kim et al. obtain the first crystal structure of the PKG I R domain bound with cGMP representing its activated state. It reveals a symmetric R dimer where cGMP molecules provide dimeric contacts. This R-R interaction prevents the high-affinity inhibitory interaction between R-C domain and sustains activation. © 2016 Elsevier Ltd.

  7. Structural features of piperazinyl-linked ciprofloxacin dimers required for activity against drug-resistant strains of Staphylococcus aureus.

    Science.gov (United States)

    Kerns, Robert J; Rybak, Michael J; Kaatz, Glenn W; Vaka, Flamur; Cha, Raymond; Grucz, Richard G; Diwadkar, Veena U

    2003-07-07

    We previously demonstrated that piperazinyl-linked fluoroquinolone dimers possess potent antibacterial activity against drug-resistant strains of Staphylococcus aureus. In this study, we report the preparation and evaluation of a series of incomplete dimers toward ascertaining structural features of piperazinyl-linked ciprofloxacin dimers that render these agents refractory to fluoroquinolone-resistance mechanisms in Staphylococcus aureus.

  8. An artemisinin-derived dimer has highly potent anti-cytomegalovirus (CMV and anti-cancer activities.

    Directory of Open Access Journals (Sweden)

    Ran He

    Full Text Available We recently reported that two artemisinin-derived dimers (dimer primary alcohol 606 and dimer sulfone 4-carbamate 832-4 are significantly more potent in inhibiting human cytomegalovirus (CMV replication than artemisinin-derived monomers. In our continued evaluation of the activities of artemisinins in CMV inhibition, twelve artemisinin-derived dimers and five artemisinin-derived monomers were used. Dimers as a group were found to be potent inhibitors of CMV replication. Comparison of CMV inhibition and the slope parameter of dimers and monomers suggest that dimers are distinct in their anti-CMV activities. A deoxy dimer (574, lacking the endoperoxide bridge, did not have any effect on CMV replication, suggesting a role for the endoperoxide bridge in CMV inhibition. Differences in anti-CMV activity were observed among three structural analogs of dimer sulfone 4-carbamate 832-4 indicating that the exact placement and oxidation state of the sulfur atom may contribute to its anti-CMV activity. Of all tested dimers, artemisinin-derived diphenyl phosphate dimer 838 proved to be the most potent inhibitor of CMV replication, with a selectivity index of approximately 1500, compared to our previously reported dimer sulfone 4-carbamate 832-4 with a selectivity index of about 900. Diphenyl phosphate dimer 838 was highly active against a Ganciclovir-resistant CMV strain and was also the most active dimer in inhibition of cancer cell growth. Thus, diphenyl phosphate dimer 838 may represent a lead for development of a highly potent and safe anti-CMV compound.

  9. Bowman-Birk protease inhibitor from the seeds of Vigna unguiculata forms a highly stable dimeric structure.

    Science.gov (United States)

    Rao, K N; Suresh, C G

    2007-10-01

    Different protease inhibitors including Bowman-Birk type (BBI) have been reported from the seeds of Vigna unguiculata. Protease isoinhibitors of double-headed Bowman-Birk type from the seeds of Vigna unguiculata have been purified and characterized. The BBI from Vigna unguiculata (Vu-BBI) has been found to undergo self-association to form very stable dimers and more complex oligomers, by size-exclusion chromatography and SDS-PAGE in the presence of urea. Many BBIs have been reported to undergo self-association to form homodimers or more complex oligomers in solution. Only one dimeric crystal structure of a BBI (pea-BBI) is reported to date. We report the three-dimensional structure of a Vu-BBI determined at 2.5 A resolution. Although, the inhibitor has a monomer fold similar to that found in other known structures of Bowman-Birk protease inhibitors, its quaternary structure is different from that commonly observed in this family. The structural elements responsible for the stability of monomer molecule and dimeric association are discussed. The Vu-BBI may use dimeric or higher quaternary association to maintain the physiological state and to execute its biological function.

  10. Expanded activity of dimer nucleases by combining ZFN and TALEN for genome editing.

    Science.gov (United States)

    Yan, Wei; Smith, Cory; Cheng, Linzhao

    2013-01-01

    Our ability to precisely and efficiently edit mammalian and plant genomes has been significantly improved in recent years, partially due to increasing use of designer nucleases that recognize a pre-determined DNA sequence, make a specific DNA double-strand break, and stimulate gene targeting. A pair of zinc finger nucleases (ZFNs) or transcription activator-like effector nucleases (TALENs) that recognize two adjacent unique DNA sequences dimerize through the fused FokI nuclease domain and cut in the middle of target DNA sequences. We report here that increasing the length of recognition DNA sequences by TALENs or ZFNs does not necessarily translate to a higher efficiency or specificity. We also discover that one subunit of ZFNs and one subunit of TALENs can form a pair of hybrid nucleases with expanded specificity at two diverse targets, and stimulate gene targeting in multiple cell types including human induced pluripotent stem (iPS) cells with improved efficiency.

  11. Model for growth hormone receptor activation based on subunit rotation within a receptor dimer

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Richard J.; Adams, Julian J.; Pelekanos, Rebecca A.; Wan, Yu; McKinstry, William J.; Palethorpe, Kathryn; Seeber, Ruth M.; Monks, Thea A.; Eidne, Karin A.; Parker, Michael W.; Waters, Michael J. (UWA); (St. Vincent); (Queensland)

    2010-07-13

    Growth hormone is believed to activate the growth hormone receptor (GHR) by dimerizing two identical receptor subunits, leading to activation of JAK2 kinase associated with the cytoplasmic domain. However, we have reported previously that dimerization alone is insufficient to activate full-length GHR. By comparing the crystal structure of the liganded and unliganded human GHR extracellular domain, we show here that there is no substantial change in its conformation on ligand binding. However, the receptor can be activated by rotation without ligand by inserting a defined number of alanine residues within the transmembrane domain. Fluorescence resonance energy transfer (FRET), bioluminescence resonance energy transfer (BRET) and coimmunoprecipitation studies suggest that receptor subunits undergo specific transmembrane interactions independent of hormone binding. We propose an activation mechanism involving a relative rotation of subunits within a dimeric receptor as a result of asymmetric placement of the receptor-binding sites on the ligand.

  12. Modulation of Bacillus thuringiensis Phosphatidylinositol-Specific Phospholipase C Activity by Mutations in the Putative Dimerization Interface

    Energy Technology Data Exchange (ETDEWEB)

    Shi, X.; Shao, C; Zhang, X; Zambonelli, C; Redfield, A; Head, J; Seaton, B; Roberts, M

    2009-01-01

    Cleavage of phosphatidylinositol (PI) to inositol 1,2-(cyclic)-phosphate (cIP) and cIP hydrolysis to inositol 1-phosphate by Bacillus thuringiensis phosphatidylinositol-specific phospholipase C are activated by the enzyme binding to phosphatidylcholine (PC) surfaces. Part of this reflects improved binding of the protein to interfaces. However, crystallographic analysis of an interfacially impaired phosphatidylinositol-specific phospholipase (W47A/W242A) suggested protein dimerization might occur on the membrane. In the W47A/W242A dimer, four tyrosine residues from one monomer interact with the same tyrosine cluster of the other, forming a tight dimer interface close to the membrane binding regions. We have constructed mutant proteins in which two or more of these tyrosine residues have been replaced with serine. Phospholipid binding and enzymatic activity of these mutants have been examined to assess the importance of these residues to enzyme function. Replacing two tyrosines had small effects on enzyme activity. However, removal of three or four tyrosine residues weakened PC binding and reduced PI cleavage by the enzyme as well as PC activation of cIP hydrolysis. Crystal structures of Y247S/Y251S in the absence and presence of myo-inositol as well as Y246S/Y247S/Y248S/Y251S indicate that both mutant proteins crystallized as monomers, were very similar to one another, and had no change in the active site region. Kinetic assays, lipid binding, and structural results indicate that either (i) a specific PC binding site, critical for vesicle activities and cIP activation, has been impaired, or (ii) the reduced dimerization potential for Y246S/Y247S/Y248S and Y246S/Y247S/Y248S/Y251S is responsible for their reduced catalytic activity in all assay systems.

  13. Lipophilic vancomycin aglycon dimer with high activity against vancomycin-resistant bacteria.

    Science.gov (United States)

    Yarlagadda, Venkateswarlu; Sarkar, Paramita; Manjunath, Goutham B; Haldar, Jayanta

    2015-12-01

    Antibiotic-resistant superbugs such as vancomycin-resistant Enterococci (VRE) and Staphylococci have become a major global health hazard. To address this issue, we synthesized vancomycin aglycon dimers to systematically probe the impact of a linker on biological activity. A dimer having a pendant lipophilic moiety in the linker showed ∼300-fold more activity than vancomycin against VRE. The high activity of the compound is attributed to its enhanced binding affinity to target peptides which resulted in improved peptidoglycan (cell wall) biosynthesis inhibition. Therefore, our studies suggest that these compounds, prepared by using facile synthetic methodology, can be used to combat vancomycin-resistant bacterial infections.

  14. Catalytic activity of titania zirconia mixed oxide catalyst for dimerization eugenol

    Science.gov (United States)

    Tursiloadi, S.; Kristiani, A.; Jenie, S. N. Aisyiyah; Laksmono, J. A.

    2017-01-01

    Clove oil has been found to possess antibacterial, antifungal, antiviral, antitumor, antioxidant and insecticidal properties. The major compound of clove oil is eugenol about 49-87%. Eugenol as phenolic compounds exhibits antioxidant and antimicrobial activities. The derivative compound of eugenol, dieugenol, show antioxidant potency better than parent eugenol. A series of TiO2-ZrO2 mixed oxides (TZ) with various titanium contents from 0 to 100wt%, prepared by using sol gel method were tested their catalytic activity for dimerization eugenol, Their catalytic activity show that these catalysts resulted a low yield of dimer eugenol, dieugenol, about 2-9 % and the purity is more than 50%.

  15. Potent immunosuppressive principles, dimeric sesquiterpene thioalkaloids, isolated from nupharis rhizoma, the rhizoma of Nuphar pumilum (nymphaeaceae): structure-requirement of nuphar-alkaloid for immunosuppressive activity.

    Science.gov (United States)

    Yamahara, J; Shimoda, H; Matsuda, H; Yoshikawa, M

    1996-09-01

    Potent immunosuppressants, the dimeric sesquiterpene thioalkaloids, 6-hydroxythiobinupharidine (2), 6,6'-dihydroxythiobinupharidine (3), 6-hydroxythionuphlutine B (5) and 6'-hydroxythionuphlutine B (6), were isolated from a natural medicine, Nupharis Rhizoma, the rhizoma of Nuphar pumilum (TIMM.) DC., through bioassay-guided separation together with five quinolizidine alkaloids (8, 9, 10, 11, 12). Dimeric sesquiterpene thioalkaloids (2, 3, 5, 6) were found to significantly inhibit anti-sheep erythrocyte plaque forming cell formation in mice spleen cells at 10(-6) M concentration. At this concentration, 2, 5 and 6 were found to exhibit no cytotoxicity to mice spleen cells, and 3 also showed only a little cytotoxicity. In addition, the inhibitory activity of several Nuphar alkaloids, dimeric sesquiterpene thioalkaloids (1, 4, 7, 8), and monomeric sesquiterpene alkaloids (9, 10, 11, 12) on anti-sheep erythrocyte plaque forming cell formation was examined and some structural requirement of Nuphar alkaloid for immunosuppressive activity was determined.

  16. Dimerization Domain of Retinal Membrane Guanylyl Cyclase 1 (RetGC1) Is an Essential Part of Guanylyl Cyclase-activating Protein (GCAP) Binding Interface.

    Science.gov (United States)

    Peshenko, Igor V; Olshevskaya, Elena V; Dizhoor, Alexander M

    2015-08-01

    The photoreceptor-specific proteins guanylyl cyclase-activating proteins (GCAPs) bind and regulate retinal membrane guanylyl cyclase 1 (RetGC1) but not natriuretic peptide receptor A (NPRA). Study of RetGC1 regulation in vitro and its association with fluorescently tagged GCAP in transfected cells showed that R822P substitution in the cyclase dimerization domain causing congenital early onset blindness disrupted RetGC1 ability to bind GCAP but did not eliminate its affinity for another photoreceptor-specific protein, retinal degeneration 3 (RD3). Likewise, the presence of the NPRA dimerization domain in RetGC1/NPRA chimera specifically disabled binding of GCAPs but not of RD3. In subsequent mapping using hybrid dimerization domains in RetGC1/NPRA chimera, multiple RetGC1-specific residues contributed to GCAP binding by the cyclase, but the region around Met(823) was the most crucial. Either positively or negatively charged residues in that position completely blocked GCAP1 and GCAP2 but not RD3 binding similarly to the disease-causing mutation in the neighboring Arg(822). The specificity of GCAP binding imparted by RetGC1 dimerization domain was not directly related to promoting dimerization of the cyclase. The probability of coiled coil dimer formation computed for RetGC1/NPRA chimeras, even those incapable of binding GCAP, remained high, and functional complementation tests showed that the RetGC1 active site, which requires dimerization of the cyclase, was formed even when Met(823) or Arg(822) was mutated. These results directly demonstrate that the interface for GCAP binding on RetGC1 requires not only the kinase homology region but also directly involves the dimerization domain and especially its portion containing Arg(822) and Met(823).

  17. [Activity of the inositol-containing phospholipid dimer analogues against human immunodeficiency virus].

    Science.gov (United States)

    Baranova, E O; Shastina, N S; Lobach, O A; Chataeva, M S; Nosik, D N; Shvets, V I

    2014-01-01

    For the purpose of finding effective inhibitors of virus adsorption the series of inositol-containing phospholipid dimer analogues were previously synthesized. In the present work, the antiretroviral activity of these compounds against HIV-1 was demonstrated on the model of cells infected with the virus. The highest effect was found in the case of dimer poliol 5, EC50 (50%-effective concentration) was 3.9 microg/ml. The development of new polyanionic compounds, which can interfere with early steps of the virus life cycle, is a promising addition to the antiretroviral therapy based on the virus enzyme inhibitors.

  18. Dimerization of DOCK2 is essential for DOCK2-mediated Rac activation and lymphocyte migration.

    Directory of Open Access Journals (Sweden)

    Masao Terasawa

    Full Text Available The migratory properties of lymphocytes depend on DOCK2, an atypical Rac activator predominantly expressed in hematopoietic cells. Although DOCK2 does not contain the Dbl homology domain typically found in guanine nucleotide exchange factors (GEFs, DOCK2 mediates the GTP-GDP exchange reaction for Rac via its DOCK homology region (DHR-2 (also known as CZH2 or Docker domain. DOCK2 DHR-2 domain is composed of three lobes, and Rac binding site and catalytic center are generated entirely from lobes B and C. On the other hand, lobe A has been implicated in dimer formation, yet its physiological significance remains unknown. Here, we report that lobe A-mediated DOCK2 dimerization is crucial for Rac activation and lymphocyte migration. We found that unlike wild-type DOCK2, DOCK2 mutant lacking lobe A failed to restore motility and polarity when expressed in thymoma cells and primary T cells lacking endogenous expression of DOCK2. Similar results were obtained with the DOCK2 point mutant having a defect in dimerization. Deletion of lobe A from the DHR-2 domain did not affect Rac GEF activity in vitro. However, fluorescence resonance energy transfer analyses revealed that lobe A is required for DOCK2 to activate Rac effectively during cell migration. Our results thus indicate that DOCK2 dimerization is functionally important under the physiological condition where only limited amounts of DOCK2 and Rac are localized to the plasma membrane.

  19. Dimer monomer transition and dimer re-formation play important role for ATM cellular function during DNA repair.

    Science.gov (United States)

    Du, Fengxia; Zhang, Minjie; Li, Xiaohua; Yang, Caiyun; Meng, Hao; Wang, Dong; Chang, Shuang; Xu, Ye; Price, Brendan; Sun, Yingli

    2014-10-03

    The ATM protein kinase, is a serine/threonine protein kinase that is recruited and activated by DNA double-strand breaks, mediates responses to ionizing radiation in mammalian cells. Here we show that ATM is held inactive in unirradiated cells as a dimer and phosphorylates the opposite strand of the dimer in response to DNA damage. Cellular irradiation induces rapid intermolecular autophosphorylation of serine 1981 that causes dimer dissociation and initiates cellular ATM kinase activity. ATM cannot phosphorylate the substrates when it could not undergo dimer monomer transition. After DNA repair, the active monomer will undergo dephosphorylation to form dimer again and dephosphorylation is critical for dimer re-formation. Our work reveals novel function of ATM dimer monomer transition and explains why ATM dimer monomer transition plays such important role for ATM cellular activity during DNA repair.

  20. Optical tuning of near and far fields form hybrid dimer nanoantennas via laser-induced melting

    Science.gov (United States)

    Kolodny, Stanislav A.; Sun, Yali; Zuev, Dmitry A.; Makarov, Sergey V.; Krasnok, Alexander E.; Belov, Pavel A.

    2016-08-01

    Hybrid nanophotonics based on metal-dielectric nanostructures unifies the advantages of plasmonics and all-dielectric nanophotonics providing strong localization of light, magnetic optical response and specifically designed scattering properties. Here, we propose a new method for optical properties tuning of hybrid dimer nanoantenas via laser-induced melting at the nanoscale. We demonstrate numerically that near- and farfield properties of a hybrid nanoantenna dramatically changes with fs-laser modification of Au particle. The results lay the groundwork for the fine-tuning of hybrid nanoantennas and can be applied for effective light manipulation at the nanoscale, as well as biomedical and energy applications.

  1. The four canonical tpr subunits of human APC/C form related homo-dimeric structures and stack in parallel to form a TPR suprahelix.

    Science.gov (United States)

    Zhang, Ziguo; Chang, Leifu; Yang, Jing; Conin, Nora; Kulkarni, Kiran; Barford, David

    2013-11-15

    The anaphase-promoting complex or cyclosome (APC/C) is a large E3 RING-cullin ubiquitin ligase composed of between 14 and 15 individual proteins. A striking feature of the APC/C is that only four proteins are involved in directly recognizing target proteins and catalyzing the assembly of a polyubiquitin chain. All other subunits, which account for >80% of the mass of the APC/C, provide scaffolding functions. A major proportion of these scaffolding subunits are structurally related. In metazoans, there are four canonical tetratricopeptide repeat (TPR) proteins that form homo-dimers (Apc3/Cdc27, Apc6/Cdc16, Apc7 and Apc8/Cdc23). Here, we describe the crystal structure of the N-terminal homo-dimerization domain of Schizosaccharomyces pombe Cdc23 (Cdc23(Nterm)). Cdc23(Nterm) is composed of seven contiguous TPR motifs that self-associate through a related mechanism to those of Cdc16 and Cdc27. Using the Cdc23(Nterm) structure, we generated a model of full-length Cdc23. The resultant "V"-shaped molecule docks into the Cdc23-assigned density of the human APC/C structure determined using negative stain electron microscopy (EM). Based on sequence conservation, we propose that Apc7 forms a homo-dimeric structure equivalent to those of Cdc16, Cdc23 and Cdc27. The model is consistent with the Apc7-assigned density of the human APC/C EM structure. The four canonical homo-dimeric TPR proteins of human APC/C stack in parallel on one side of the complex. Remarkably, the uniform relative packing of neighboring TPR proteins generates a novel left-handed suprahelical TPR assembly. This finding has implications for understanding the assembly of other TPR-containing multimeric complexes.

  2. Towards squalamine mimics: synthesis and antibacterial activities of head-to-tail dimeric sterol-polyamine conjugates.

    Science.gov (United States)

    Chen, Wen-Hua; Wennersten, Christine; Moellering, Robert C; Regen, Steven L

    2013-03-01

    Four dimeric sterol-polyamine conjugates have been synthesized from the homo- and hetero-connection of monomeric sterol-polyamine analogs in a head-to-tail manner. These dimeric conjugates show strong antibacterial activity against a broad spectrum of Gram-positive bacteria, whereas their corresponding activities against Gram-negative bacteria are relatively moderate. Though no significant difference was observed in the activities of these conjugates, cholic acid-containing dimeric conjugates generally exhibit higher activities than the corresponding deoxycholic acid-derived analogs. This is in contrast to the finding that a monomeric deoxycholic acid-spermine conjugate was more active than the corresponding cholic acid-derived analog.

  3. G domain dimerization controls dynamin's assembly-stimulated GTPase activity

    Energy Technology Data Exchange (ETDEWEB)

    Chappie, Joshua S.; Acharya, Sharmistha; Leonard, Marilyn; Schmid, Sandra L.; Dyda, Fred (NIH); (Scripps)

    2010-06-14

    Dynamin is an atypical GTPase that catalyses membrane fission during clathrin-mediated endocytosis. The mechanisms of dynamin's basal and assembly-stimulated GTP hydrolysis are unknown, though both are indirectly influenced by the GTPase effector domain (GED). Here we present the 2.0 {angstrom} resolution crystal structure of a human dynamin 1-derived minimal GTPase-GED fusion protein, which was dimeric in the presence of the transition state mimic GDP.AlF{sub 4}{sup -}. The structure reveals dynamin's catalytic machinery and explains how assembly-stimulated GTP hydrolysis is achieved through G domain dimerization. A sodium ion present in the active site suggests that dynamin uses a cation to compensate for the developing negative charge in the transition state in the absence of an arginine finger. Structural comparison to the rat dynamin G domain reveals key conformational changes that promote G domain dimerization and stimulated hydrolysis. The structure of the GTPase-GED fusion protein dimer provides insight into the mechanisms underlying dynamin-catalysed membrane fission.

  4. Crystal Structure of PKG I:cGMP Complex Reveals a cGMP-Mediated Dimeric Interface that Facilitates cGMP-Induced Activation.

    Science.gov (United States)

    Kim, Jeong Joo; Lorenz, Robin; Arold, Stefan T; Reger, Albert S; Sankaran, Banumathi; Casteel, Darren E; Herberg, Friedrich W; Kim, Choel

    2016-05-03

    Cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG) is a key regulator of smooth muscle and vascular tone and represents an important drug target for treating hypertensive diseases and erectile dysfunction. Despite its importance, its activation mechanism is not fully understood. To understand the activation mechanism, we determined a 2.5 Å crystal structure of the PKG I regulatory (R) domain bound with cGMP, which represents the activated state. Although we used a monomeric domain for crystallization, the structure reveals that two R domains form a symmetric dimer where the cGMP bound at high-affinity pockets provide critical dimeric contacts. Small-angle X-ray scattering and mutagenesis support this dimer model, suggesting that the dimer interface modulates kinase activation. Finally, structural comparison with the homologous cyclic AMP-dependent protein kinase reveals that PKG is drastically different from protein kinase A in its active conformation, suggesting a novel activation mechanism for PKG.

  5. Dimerization by a cytokine receptor is necessary for constitutive activation of JAK2V617F.

    Science.gov (United States)

    Lu, Xiaohui; Huang, Lily Jun-Shen; Lodish, Harvey F

    2008-02-29

    The majority of the BCR-ABL-negative myeloproliferative disorders express the mutant JAK2, JAK2V617F. Previously we showed that constitutive activation of this oncogenic JAK2 mutant in Ba/F3 or 32D cells requires coexpression of a cognate homodimeric cytokine receptor, such as the EpoR. However, overexpression of JAK2V617F in Ba/F3 cells renders them cytokine-independent for growth in the absence of an exogenous cytokine receptor. Here, we demonstrated that JAK2V617F domains required for receptor association are essential for cytokine-independent growth by overexpressed JAK2V617F, suggesting JAK2V617F is binding to an unknown endogenous cytokine receptor(s) for its activation. We further showed that disruption of EpoR dimerization by coexpressing a truncated EpoR disrupted JAK2V617F-mediated transformation, indicating that EpoR dimerization plays an essential role in the activation of JAK2V617F. Interestingly, coexpression of JAK2V617F with EpoR mutants that retain JAK2 binding but are defective in mediating Epo-dependent JAK2 activation due to mutations in a conserved juxtamembrane motif does lead to cytokine-independent activation of JAK2V617F. Overall, these findings confirm that JAK2V617F requires binding to a dimerized cytokine receptor for its activation, and that the key EpoR juxtamembrane regulatory motif essential for Epo-dependent JAK2 activation is not essential for the activation of JAK2V617F. The structure of the activated JAK2V617F is thus likely to be different from that of the activated wild-type JAK2, raising the possibility of developing a specifically targeted therapy for myeloproliferative disorders.

  6. Identification and super-resolution imaging of ligand-activated receptor dimers in live cells

    CERN Document Server

    Winckler, Pascale; Giannone, Gregory; De Giorgi, Francesca; Ichas, François; Sibarita, Jean-Baptiste; Lounis, Brahim; Cognet, Laurent

    2013-01-01

    Molecular interactions are key to many chemical and biological processes like protein function. In many signaling processes they occur in sub-cellular areas displaying nanoscale organizations and involving molecular assemblies. The nanometric dimensions and the dynamic nature of the interactions make their investigations complex in live cells. While super-resolution fluorescence microscopies offer live-cell molecular imaging with sub-wavelength resolutions, they lack specificity for distinguishing interacting molecule populations. Here we combine super-resolution microscopy and single-molecule F\\"orster Resonance Energy Transfer (FRET) to identify dimers of receptors induced by ligand binding and provide super-resolved images of their membrane distribution in live cells. By developing a two-color universal-Point-Accumulation-In-the-Nanoscale-Topography (uPAINT) method, dimers of epidermal growth factor receptors (EGFR) activated by EGF are studied at ultra-high densities, revealing preferential cell-edge sub-...

  7. Dimerization of complement factor H-related proteins modulates complement activation in vivo.

    Science.gov (United States)

    Goicoechea de Jorge, Elena; Caesar, Joseph J E; Malik, Talat H; Patel, Mitali; Colledge, Matthew; Johnson, Steven; Hakobyan, Svetlana; Morgan, B Paul; Harris, Claire L; Pickering, Matthew C; Lea, Susan M

    2013-03-19

    The complement system is a key component regulation influences susceptibility to age-related macular degeneration, meningitis, and kidney disease. Variation includes genomic rearrangements within the complement factor H-related (CFHR) locus. Elucidating the mechanism underlying these associations has been hindered by the lack of understanding of the biological role of CFHR proteins. Here we present unique structural data demonstrating that three of the CFHR proteins contain a shared dimerization motif and that this hitherto unrecognized structural property enables formation of both homodimers and heterodimers. Dimerization confers avidity for tissue-bound complement fragments and enables these proteins to efficiently compete with the physiological complement inhibitor, complement factor H (CFH), for ligand binding. Our data demonstrate that these CFHR proteins function as competitive antagonists of CFH to modulate complement activation in vivo and explain why variation in the CFHRs predisposes to disease.

  8. Molecular recognition: monomer of the yeast transcriptional activator GCN4 recognizes its dimer DNA binding target sites specifically

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    It is widely believed that dimerization is a requirement for the yeast transcriptional activator GCN4 to recognize its specific DNA target sites. We used the basic region (226-252) of the yeast transcriptional activator GCN4, as both a monomeric peptide and a disulfide-linked dimer to investigate the interaction of the peptides with the DNA target sites AP-1 and CRE. CD and ITC experiments indicate that although the monomeric peptide GCN4-M has a weaker affinity with the DNA relative to the disulfide-linked dimer peptide GCN4-D, it recognizes AP-1 and CRE target sites specifically.

  9. Dimeric Matrine-Type Alkaloids from the Roots of Sophora flavescens and Their Anti-Hepatitis B Virus Activities.

    Science.gov (United States)

    Zhang, Yu-Bo; Zhan, Li-Qin; Li, Guo-Qiang; Wang, Feng; Wang, Ying; Li, Yao-Lan; Ye, Wen-Cai; Wang, Guo-Cai

    2016-08-01

    Six unusual matrine-type alkaloid dimers, flavesines A-F (1-6, respectively), together with three proposed biosynthetic intermediates (7-9) were isolated from the roots of Sophora flavescens. Compounds 1-5 were the first natural matrine-type alkaloid dimers, and compound 6 represented an unprecedented dimerization pattern constructed by matrine and (-)-cytisine. Their structures were elucidated by NMR, MS, single-crystal X-ray diffraction, and a chemical method. The hypothetical biogenetic pathways of 1-6 were also proposed. Compounds 1-9 exhibited inhibitory activities against hepatitis B virus.

  10. Mechanistic basis of Nek7 activation through Nek9 binding and induced dimerization

    Science.gov (United States)

    Haq, Tamanna; Richards, Mark W.; Burgess, Selena G.; Gallego, Pablo; Yeoh, Sharon; O'Regan, Laura; Reverter, David; Roig, Joan; Fry, Andrew M.; Bayliss, Richard

    2015-11-01

    Mitotic spindle assembly requires the regulated activities of protein kinases such as Nek7 and Nek9. Nek7 is autoinhibited by the protrusion of Tyr97 into the active site and activated by the Nek9 non-catalytic C-terminal domain (CTD). CTD binding apparently releases autoinhibition because mutation of Tyr97 to phenylalanine increases Nek7 activity independently of Nek9. Here we find that self-association of the Nek9-CTD is needed for Nek7 activation. We map the minimal Nek7 binding region of Nek9 to residues 810-828. A crystal structure of Nek7Y97F bound to Nek9810-828 reveals a binding site on the C-lobe of the Nek7 kinase domain. Nek7Y97F crystallizes as a back-to-back dimer between kinase domain N-lobes, in which the specific contacts within the interface are coupled to the conformation of residue 97. Hence, we propose that the Nek9-CTD activates Nek7 through promoting back-to-back dimerization that releases the autoinhibitory tyrosine residue, a mechanism conserved in unrelated kinase families.

  11. Photoinduced formation of an azobenzene-based CD-active supramolecular cyclic dimer.

    Science.gov (United States)

    Sogawa, Hiromitsu; Terada, Kayo; Miyagi, Yu; Shiotsuki, Masashi; Inai, Yoshihito; Masuda, Toshio; Sanda, Fumio

    2015-04-27

    A series of new photo-responsive amino acid-derived azobenzenedicarboxylic acid derivatives (S)-1 a-e were synthesized. Compound (S)-1 a in the trans form exhibited no circular dichroism (CD) signal in DMF under ambient conditions, whereas intense Cotton effects were observed upon UV irradiation, indicating the formation of a chiral supramolecular structure in the cis form. The CD signals disappeared when trifluoroacetic acid (TFA) was added to the solution. The ester counterpart [(S)-1 a'] showed no CD signal. Hydrogen bonding between the carboxy groups seemed necessary for constructing the supramolecular structure. The kinetic studies of cis to trans isomerization of (S)-1 a demonstrated that the formation of a chiral supramolecule enhances the stability of the cis-azobenzene structure. The ESI mass spectrum of stilbenedicarboxylic acid (S)-4, an analogue of (S)-1 b, confirmed the formation of a dimer. A theoretical CD study revealed that (S)-1 a in the cis form should be present as a cyclic chiral dimer.

  12. The Amino Acid Specificity for Activation of Phenylalanine Hydroxylase Matches the Specificity for Stabilization of Regulatory Domain Dimers.

    Science.gov (United States)

    Zhang, Shengnan; Hinck, Andrew P; Fitzpatrick, Paul F

    2015-08-25

    Liver phenylalanine hydroxylase is allosterically activated by phenylalanine. The structural changes that accompany activation have not been identified, but recent studies of the effects of phenylalanine on the isolated regulatory domain of the enzyme support a model in which phenylalanine binding promotes regulatory domain dimerization. Such a model predicts that compounds that stabilize the regulatory domain dimer will also activate the enzyme. Nuclear magnetic resonance spectroscopy and analytical ultracentrifugation were used to determine the ability of different amino acids and phenylalanine analogues to stabilize the regulatory domain dimer. The abilities of these compounds to activate the enzyme were analyzed by measuring their effects on the fluorescence change that accompanies activation and on the activity directly. At concentrations of 10-50 mM, d-phenylalanine, l-methionine, l-norleucine, and (S)-2-amino-3-phenyl-1-propanol were able to activate the enzyme to the same extent as 1 mM l-phenylalanine. Lower levels of activation were seen with l-4-aminophenylalanine, l-leucine, l-isoleucine, and 3-phenylpropionate. The ability of these compounds to stabilize the regulatory domain dimer agreed with their ability to activate the enzyme. These results support a model in which allosteric activation of phenylalanine hydroxylase is linked to dimerization of regulatory domains.

  13. The polar domain of the b subunit of Escherichia coli F1F0-ATPase forms an elongated dimer that interacts with the F1 sector.

    Science.gov (United States)

    Dunn, S D

    1992-04-15

    A soluble form of the b subunit of the F0 sector of the F1F0-ATPase of Escherichia coli has been produced, purified, and characterized. In this form of the protein, designated bsol, residues 25-146 (the carboxyl terminus) of b have been fused to an amino-terminal octapeptide extension derived from the vector pUC8. The inferred subunit molecular weight of bsol is 15,459. bsol protein was expressed in E. coli as a soluble cytoplasmic protein and was readily purified to homogeneity by conventional methods. The molecular weight of bsol, determined by sedimentation equilibrium, was 31,200, indicating that the protein is dimeric. Chemical cross-linking studies supported this conclusion. However, bsol sedimented with a coefficient of just 1.8 S and behaved on size exclusion chromatography with an apparent molecular weight of 80,000-85,000. These results indicate that the protein exists in solution as a highly elongated dimer. The circular dichroism spectrum indicated that bsol is highly alpha-helical. Binding of bsol to F1-ATPase was directly demonstrated by size exclusion chromatography. bsol also inhibited the binding of F1-ATPase to F1-depleted membrane vesicles, as measured by reconstitution of energy-dependent quinacrine fluorescence quenching. This result implies that bsol and F0 compete for binding to the same site on F1. The apparently normal interaction of bsol with F1-ATPase strongly suggests that the recombinant protein assumes the correct structure. No substantial effects of bsol on the ATPase activity of purified F1 were observed.

  14. A new flavan-3-ol dimer from Ficus spragueana leaves and its cytotoxic activity

    Directory of Open Access Journals (Sweden)

    Ehab A Ragab

    2013-01-01

    Full Text Available Background: Isolation and structure elucidation of flavan-3-ol constituents from the leaves of Ficus spragueana and their cytotoxic activity. Materials and Methods: Different open silica gel column chromatographic techniques with different solvent systems were used for the separation of the constituents of the ethyl acetate-soluble fraction of the alcoholic extract of Ficus spragueana leaves. The structures of these compounds were assigned on the basis of spectroscopic analyses and comparison with literature data. MTT colorimetric assay method (Viability assay was used for the evaluation of cytotoxic activity of compound 1 against human breast cancer (MCF-7 and human liver cancer (HepG2 cell lines. Results: The isolation of one flavan-3-ol dimer and was identified as (--afzelechin-(4α→8-epicatechin 1, and two flavan-3-ol monomers and were identified as (--epiafzelechin 2 and (--epicatechin 3. Compound 1 was relatively inactive against human breast cancer (MCF-7 cell line at the tested concentrations as compared with the standard. However, at a concentration (50 ΅g it was found to give inhibition upon the proliferation of examined human liver (HepG2 tumor cell line. Conclusions: Compound 1 is a new flavan-3-ol dimer and it showed a potent cytotoxic activity against human liver (HepG2 tumor cell line.

  15. Dimerization and enzymatic activity of fungal 17β-hydroxysteroid dehydrogenase from the short-chain dehydrogenase/reductase superfamily

    Directory of Open Access Journals (Sweden)

    Kristan Katja

    2005-12-01

    Full Text Available Abstract Background 17β-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17β-HSDcl is a member of the short-chain dehydrogenase/reductase (SDR superfamily. SDR proteins usually function as dimers or tetramers and 17β-HSDcl is also a homodimer under native conditions. Results We have investigated here which secondary structure elements are involved in the dimerization of 17β-HSDcl and examined the importance of dimerization for the enzyme activity. Sequence similarity with trihydroxynaphthalene reductase from Magnaporthe grisea indicated that Arg129 and His111 from the αE-helices interact with the Asp121, Glu117 and Asp187 residues from the αE and αF-helices of the neighbouring subunit. The Arg129Asp and His111Leu mutations both rendered 17β-HSDcl monomeric, while the mutant 17β-HSDcl-His111Ala was dimeric. Circular dichroism spectroscopy analysis confirmed the conservation of the secondary structure in both monomers. The three mutant proteins all bound coenzyme, as shown by fluorescence quenching in the presence of NADP+, but both monomers showed no enzymatic activity. Conclusion We have shown by site-directed mutagenesis and structure/function analysis that 17β-HSDcl dimerization involves the αE and αF helices of both subunits. Neighbouring subunits are connected through hydrophobic interactions, H-bonds and salt bridges involving amino acid residues His111 and Arg129. Since the substitutions of these two amino acid residues lead to inactive monomers with conserved secondary structure, we suggest dimerization is a prerequisite for catalysis. A detailed understanding of this dimerization could lead to the development of compounds that will specifically prevent dimerization, thereby serving as a new type of inhibitor.

  16. Correlating in Vitro and in Vivo Activities of Light-Inducible Dimers: A Cellular Optogenetics Guide.

    Science.gov (United States)

    Hallett, Ryan A; Zimmerman, Seth P; Yumerefendi, Hayretin; Bear, James E; Kuhlman, Brian

    2016-01-15

    Light-inducible dimers are powerful tools for cellular optogenetics, as they can be used to control the localization and activity of proteins with high spatial and temporal resolution. Despite the generality of the approach, application of light-inducible dimers is not always straightforward, as it is frequently necessary to test alternative dimer systems and fusion strategies before the desired biological activity is achieved. This process is further hindered by an incomplete understanding of the biophysical/biochemical mechanisms by which available dimers behave and how this correlates to in vivo function. To better inform the engineering process, we examined the biophysical and biochemical properties of three blue-light-inducible dimer variants (cryptochrome2 (CRY2)/CIB1, iLID/SspB, and LOVpep/ePDZb) and correlated these characteristics to in vivo colocalization and functional assays. We find that the switches vary dramatically in their dark and lit state binding affinities and that these affinities correlate with activity changes in a variety of in vivo assays, including transcription control, intracellular localization studies, and control of GTPase signaling. Additionally, for CRY2, we observe that light-induced changes in homo-oligomerization can have significant effects on activity that are sensitive to alternative fusion strategies.

  17. Subunit b-dimer of the Escherichia coli ATP synthase can form left-handed coiled-coils.

    Science.gov (United States)

    Wise, John G; Vogel, Pia D

    2008-06-01

    One remaining challenge to our understanding of the ATP synthase concerns the dimeric coiled-coil stator subunit b of bacterial synthases. The subunit b-dimer has been implicated in important protein interactions that appear necessary for energy conservation and that may be instrumental in energy conservation during rotary catalysis by the synthase. Understanding the stator structure and its interactions with the rest of the enzyme is crucial to the understanding of the overall catalytic mechanism. Controversy exists on whether subunit b adopts a classic left-handed or a presumed right-handed dimeric coiled-coil and whether or not staggered pairing between nonhomologous residues in the homodimer is required for intersubunit packing. In this study we generated molecular models of the Escherichia coli subunit b-dimer that were based on the well-established heptad-repeat packing exhibited by left-handed, dimeric coiled-coils by employing simulated annealing protocols with structural restraints collected from known structures. In addition, we attempted to create hypothetical right-handed coiled-coil models and left- and right-handed models with staggered packing in the coiled-coil domains. Our analyses suggest that the available structural and biochemical evidence for subunit b can be accommodated by classic left-handed, dimeric coiled-coil quaternary structures.

  18. Inhibition of aberrant complement activation by a dimer of acetylsalicylic acid.

    Science.gov (United States)

    Lee, Moonhee; Wathier, Matthew; Love, Jennifer A; McGeer, Edith; McGeer, Patrick L

    2015-10-01

    We here report synthesis for the first time of the acetyl salicylic acid dimer 5,5'-methylenebis(2-acetoxybenzoic acid) (DAS). DAS inhibits aberrant complement activation by selectively blocking factor D of the alternative complement pathway and C9 of the membrane attack complex. We have previously identified aurin tricarboxylic and its oligomers as promising agents in this regard. DAS is much more potent, inhibiting erythrocyte hemolysis by complement-activated serum with an IC50 in the 100-170 nanomolar range. There are numerous conditions where self-damage from the complement system has been implicated in the pathology, including such chronic degenerative diseases of aging as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and age-related macular degeneration. Consequently, there is a high priority for the discovery and development of agents that can successfully treat such conditions. DAS holds considerable promise for being such an agent.

  19. Dnmt3b Methylates DNA by a Noncooperative Mechanism, and Its Activity Is Unaffected by Manipulations at the Predicted Dimer Interface.

    Science.gov (United States)

    Norvil, Allison B; Petell, Christopher J; Alabdi, Lama; Wu, Lanchen; Rossie, Sandra; Gowher, Humaira

    2016-11-04

    The catalytic domains of the de novo DNA methyltransferases Dnmt3a-C and Dnmt3b-C are highly homologous. However, their unique biochemical properties could potentially contribute to differences in the substrate preferences or biological functions of these enzymes. Dnmt3a-C forms tetramers through interactions at the dimer interface, which also promote multimerization on DNA and cooperativity. Similar to the case for processive enzymes, cooperativity allows Dnmt3a-C to methylate multiple sites on the same DNA molecule; however, it is unclear whether Dnmt3b-C methylates DNA by a cooperative or processive mechanism. The importance of the tetramer structure and cooperative mechanism is emphasized by the observation that the R882H mutation in the dimer interface of DNMT3A is highly prevalent in acute myeloid leukemia and leads to a substantial loss of its activity. Under conditions that distinguish between cooperativity and processivity, we show that in contrast to that of Dnmt3a-C, the activity of Dnmt3b-C is not cooperative and confirm the processivity of Dnmt3b-C and the full length Dnmt3b enzyme. Whereas the R878H mutation (mouse homologue of R882H) led to the loss of cooperativity of Dnmt3a-C, the activity and processivity of the analogous Dnmt3b-C R829H variant were comparable to those of the wild-type enzyme. Additionally, buffer acidification that attenuates the dimer interface interactions of Dnmt3a-C had no effect on Dnmt3b-C activity. Taken together, these results demonstrate an important mechanistic difference between Dnmt3b and Dnmt3a and suggest that interactions at the dimer interface may play a limited role in regulating Dnmt3b-C activity. These new insights have potential implications for the distinct biological roles of Dnmt3a and Dnmt3b.

  20. Characterization of microtubule-binding and dimerization activity of Giardia lamblia end-binding 1 protein.

    Science.gov (United States)

    Kim, Juri; Nagami, Sara; Lee, Kyu-Ho; Park, Soon-Jung

    2014-01-01

    End-binding 1 (EB1) proteins are evolutionarily conserved components of microtubule (MT) plus-end tracking protein that regulate MT dynamics. Giardia lamblia, with two nuclei and cytoskeletal structures, requires accurate MT distribution for division. In this study, we show that a single EB1 homolog gene of G. lamblia regulates MT dynamics in mitosis. The haemagglutinin-tagged G. lamblia EB1 (GlEB1) localizes to the nuclear envelopes and median bodies, and is transiently present in mitotic spindles of dividing cells. Knockdown of GlEB1 expression using the morpholinos-based anti-EB1 oligonucleotides, resulted in a significant defect in mitosis of Giardia trophozoites. The MT-binding assays using recombinant GlEB1 (rGlEB1) proteins demonstrated that rGlEB1102-238, but not rGlEB11-184, maintains an MT-binding ability comparable with that of the full length protein, rGlEB11-238. Size exclusion chromatography showed that rGlEB1 is present as a dimer formed by its C-terminal domain and a disulfide bond. In vitro-mutagenesis of GlEB1 indicated that an intermolecular disulfide bond is made between cysteine #13 of the two monomers. Complementation assay using the BIM1 knockout mutant yeast, the yeast homolog of mammalian EB1, indicated that expression of the C13S mutant GlEB1 protein cannot rescue the mitotic defect of the BIM1 mutant yeast. These results suggest that dimerization of GlEB1 via the 13th cysteine residues plays a role during mitosis in Giardia.

  1. Structural basis for morpheein-type allosteric regulation of Escherichia coli glucosamine-6-phosphate synthase: equilibrium between inactive hexamer and active dimer.

    Science.gov (United States)

    Mouilleron, Stéphane; Badet-Denisot, Marie-Ange; Pecqueur, Ludovic; Madiona, Karine; Assrir, Nadine; Badet, Bernard; Golinelli-Pimpaneau, Béatrice

    2012-10-01

    The amino-terminal cysteine of glucosamine-6-phosphate synthase (GlmS) acts as a nucleophile to release and transfer ammonia from glutamine to fructose 6-phosphate through a channel. The crystal structure of the C1A mutant of Escherichia coli GlmS, solved at 2.5 Å resolution, is organized as a hexamer, where the glutaminase domains adopt an inactive conformation. Although the wild-type enzyme is active as a dimer, size exclusion chromatography, dynamic and quasi-elastic light scattering, native polyacrylamide gel electrophoresis, and ultracentrifugation data show that the dimer is in equilibrium with a hexameric state, in vitro and in cellulo. The previously determined structures of the wild-type enzyme, alone or in complex with glucosamine 6-phosphate, are also consistent with a hexameric assembly that is catalytically inactive because the ammonia channel is not formed. The shift of the equilibrium toward the hexameric form in the presence of cyclic glucosamine 6-phosphate, together with the decrease of the specific activity with increasing enzyme concentration, strongly supports product inhibition through hexamer stabilization. Altogether, our data allow us to propose a morpheein model, in which the active dimer can rearrange into a transiently stable form, which has the propensity to form an inactive hexamer. This would account for a physiologically relevant allosteric regulation of E. coli GlmS. Finally, in addition to cyclic glucose 6-phosphate bound at the active site, the hexameric organization of E. coli GlmS enables the binding of another linear sugar molecule. Targeting this sugar-binding site to stabilize the inactive hexameric state is therefore suggested for the development of specific antibacterial inhibitors.

  2. Structure of cyanase reveals that a novel dimeric and decameric arrangement of subunits is required for formation of the enzyme active site.

    Energy Technology Data Exchange (ETDEWEB)

    Walsh, M. A.; Otwinowski, Z.; Perrakis, A.; Anderson, P. M.; Joachimiak, A.; Biosciences Division; Univ. of Texas; European Molecular Biology Lab.; Univ. of Minnesota; Northwestern Univ.

    2000-01-01

    Cyanase is an enzyme found in bacteria and plants that catalyzes the reaction of cyanate with bicarbonate to produce ammonia and carbon dioxide. In Escherichia coli, cyanase is induced from the cyn operon in response to extracellular cyanate. The enzyme is functionally active as a homodecamer of 17 kDa subunits, and displays half-site binding of substrates or substrate analogs. The enzyme shows no significant amino acid sequence homology with other proteins. We have determined the crystal structure of cyanase at 1.65 {angstrom} resolution using the multiwavelength anomalous diffraction (MAD) method. Cyanase crystals are triclinic and contain one homodecamer in the asymmetric unit. Selenomethionine-labeled protein offers 40 selenium atoms for use in phasing. Structures of cyanase with bound chloride or oxalate anions, inhibitors of the enzyme, allowed identification of the active site. The cyanase monomer is composed of two domains. The N-terminal domain shows structural similarity to the DNA-binding {alpha}-helix bundle motif. The C-terminal domain has an 'open fold' with no structural homology to other proteins. The subunits of cyanase are arranged in a novel manner both at the dimer and decamer level. The dimer structure reveals the C-terminal domains to be intertwined, and the decamer is formed by a pentamer of these dimers. The active site of the enzyme is located between dimers and is comprised of residues from four adjacent subunits of the homodecamer. The structural data allow a conceivable reaction mechanism to be proposed.

  3. Intermolecular Interactions in the TMEM16A Dimer Controlling Channel Activity

    Science.gov (United States)

    Scudieri, Paolo; Musante, Ilaria; Gianotti, Ambra; Moran, Oscar; Galietta, Luis J. V.

    2016-12-01

    TMEM16A and TMEM16B are plasma membrane proteins with Ca2+-dependent Cl‑ channel function. By replacing the carboxy-terminus of TMEM16A with the equivalent region of TMEM16B, we obtained channels with potentiation of channel activity. Progressive shortening of the chimeric region restricted the “activating domain” to a short sequence close to the last transmembrane domain and led to TMEM16A channels with high activity at very low intracellular Ca2+ concentrations. To elucidate the molecular mechanism underlying this effect, we carried out experiments based on double chimeras, Forster resonance energy transfer, and intermolecular cross-linking. We also modeled TMEM16A structure using the Nectria haematococca TMEM16 protein as template. Our results indicate that the enhanced activity in chimeric channels is due to altered interaction between the carboxy-terminus and the first intracellular loop in the TMEM16A homo-dimer. Mimicking this perturbation with a small molecule could be the basis for a pharmacological stimulation of TMEM16A-dependent Cl‑ transport.

  4. Structure of the dimeric N-glycosylated form of fungal β-N-acetylhexosaminidase revealed by computer modeling, vibrational spectroscopy, and biochemical studies

    Directory of Open Access Journals (Sweden)

    Sklenář Jan

    2007-05-01

    Full Text Available Abstract Background Fungal β-N-acetylhexosaminidases catalyze the hydrolysis of chitobiose into its constituent monosaccharides. These enzymes are physiologically important during the life cycle of the fungus for the formation of septa, germ tubes and fruit-bodies. Crystal structures are known for two monomeric bacterial enzymes and the dimeric human lysosomal β-N-acetylhexosaminidase. The fungal β-N-acetylhexosaminidases are robust enzymes commonly used in chemoenzymatic syntheses of oligosaccharides. The enzyme from Aspergillus oryzae was purified and its sequence was determined. Results The complete primary structure of the fungal β-N-acetylhexosaminidase from Aspergillus oryzae CCF1066 was used to construct molecular models of the catalytic subunit of the enzyme, the enzyme dimer, and the N-glycosylated dimer. Experimental data were obtained from infrared and Raman spectroscopy, and biochemical studies of the native and deglycosylated enzyme, and are in good agreement with the models. Enzyme deglycosylated under native conditions displays identical kinetic parameters but is significantly less stable in acidic conditions, consistent with model predictions. The molecular model of the deglycosylated enzyme was solvated and a molecular dynamics simulation was run over 20 ns. The molecular model is able to bind the natural substrate – chitobiose with a stable value of binding energy during the molecular dynamics simulation. Conclusion Whereas the intracellular bacterial β-N-acetylhexosaminidases are monomeric, the extracellular secreted enzymes of fungi and humans occur as dimers. Dimerization of the fungal β-N-acetylhexosaminidase appears to be a reversible process that is strictly pH dependent. Oligosaccharide moieties may also participate in the dimerization process that might represent a unique feature of the exclusively extracellular enzymes. Deglycosylation had only limited effect on enzyme activity, but it significantly affected

  5. Adlayers of C60-C60 and C60-C70 fullerene dimers formed on au(111) in benzene solutions studied by STM and LEED.

    Science.gov (United States)

    Matsumoto, Masashi; Inukai, Junji; Tsutsumi, Eishi; Yoshimoto, Soichiro; Itaya, Kingo; Ito, Osamu; Fujiwara, Koichi; Murata, Michihisa; Murata, Yasujiro; Komatsu, Koichi

    2004-02-17

    Scanning tunneling microscopy (STM) and low-energy electron diffraction were used to reveal the structures of ordered adlayers of [2+2]-type C60-C60 fullerene dimer (C120) and C60-C70 cross-dimer (C130) formed on Au(111) by immersingit in abenzene solution containing C120 or C130 molecules. High-resolution STM images clearly showed the packing arrangements and the electronic structures of C120 and C130 on the Au(111) surface in ultrahigh vacuum. The (2 square root3 x 4square root3)R30 degrees, (2square root3 x 5square root3)R30 degrees, and (7 x 7) structures were found for the C120 adlayer on the Au(111) surface, whereas C130 molecules were closely packed on the surface. Each C60 or C70 monomer cage was discerned in the STM image of a C130 molecule.

  6. Collisional properties of weakly bound heteronuclear dimers

    NARCIS (Netherlands)

    Marcelis, B.; Kokkelmans, S.J.J.M.F.; Shlyapnikov, G.V.; Petrov, D.S.

    2008-01-01

    We consider collisional properties of weakly bound heteronuclear molecules (dimers) formed in a two-species mixture of atoms with a large mass difference. We focus on dimers containing light fermionic atoms as they manifest collisional stability due to an effective dimer-dimer repulsion originating

  7. Reversible dissociation of active octamer of cyanase to inactive dimer promoted by alteration of the sulfhydryl group.

    Science.gov (United States)

    Anderson, P M; Johnson, W V; Korte, J J; Xiong, X F; Sung, Y C; Fuchs, J A

    1988-04-25

    Cyanase is an inducible enzyme in Escherichia coli that catalyzes the reaction of cyanate with bicarbonate resulting in the decomposition of cyanate to ammonia and bicarbonate. In this study, the role of the single sulfhydryl group in each of the eight identical subunits of cyanase was investigated. Tetranitromethane, methyl methanethiosulfonate, N-ethylmaleimide, and Hg2+ all reacted with the sulfhydryl group to give derivatives which had reduced activities and which dissociated reversibly to inactive dimer. Association of inactive dimer to active octamer was facilitated by the presence of azide (cyanate analog) and bicarbonate, increased temperature and enzyme concentration, and presence of phosphate. Nitration of tyrosine residues by tetranitromethane occurred only in the absence of azide and bicarbonate, suggesting that at least some of the tyrosine residues become exposed when octamer dissociates to dimer. Site-directed mutagenesis was used to prepare a mutant enzyme in which serine was substituted for cysteine. The mutant enzyme was catalytically active and had properties very similar to native enzyme, except that it was less stable to treatment with urea and to high temperatures. These results establish that in native cyanase the sulfhydryl group per se is not required for catalytic activity, but it may play a role in stabilizing octameric structure, and that octameric structure is required for catalytic activity.

  8. Growth hormone (GH)-independent dimerization of GH receptor by a leucine zipper results in constitutive activation

    DEFF Research Database (Denmark)

    Behncken, S N; Billestrup, Nils; Brown, R;

    2000-01-01

    Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers of the gro......Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers...... proliferation after interleukin 3 withdrawal at a rate equal to maximally stimulated wild type GHR-expressing cells. Activation of STAT 5b was also observed in Fos-Jun-GHR-expressing cells at a level equal to that in chronically GH-treated GHR-expressing cells. Thus, forced dimerization of the transmembrane...... and cytoplasmic domains of the GHR in the absence of the extracellular domain can lead to the constitutive activation of known GH signaling end points, supporting the view that proximity of Janus kinase 2 (JAK2) kinases is the essential element in signaling. Such constitutively active GH receptors may have...

  9. Quinine dimers are potent inhibitors of the Plasmodium falciparum chloroquine resistance transporter and are active against quinoline-resistant P. falciparum.

    Science.gov (United States)

    Hrycyna, Christine A; Summers, Robert L; Lehane, Adele M; Pires, Marcos M; Namanja, Hilda; Bohn, Kelsey; Kuriakose, Jerrin; Ferdig, Michael; Henrich, Philipp P; Fidock, David A; Kirk, Kiaran; Chmielewski, Jean; Martin, Rowena E

    2014-03-21

    Chloroquine (CQ) resistance in the human malaria parasite Plasmodium falciparum is primarily conferred by mutations in the "chloroquine resistance transporter" (PfCRT). The resistance-conferring form of PfCRT (PfCRT(CQR)) mediates CQ resistance by effluxing the drug from the parasite's digestive vacuole, the acidic compartment in which CQ exerts its antiplasmodial effect. PfCRT(CQR) can also decrease the parasite's susceptibility to other quinoline drugs, including the current antimalarials quinine and amodiaquine. Here we describe interactions between PfCRT(CQR) and a series of dimeric quinine molecules using a Xenopus laevis oocyte system for the heterologous expression of PfCRT and using an assay that detects the drug-associated efflux of H(+) ions from the digestive vacuole in parasites that harbor different forms of PfCRT. The antiplasmodial activities of dimers 1 and 6 were also examined in vitro (against drug-sensitive and drug-resistant strains of P. falciparum) and in vivo (against drug-sensitive P. berghei). Our data reveal that the quinine dimers are the most potent inhibitors of PfCRT(CQR) reported to date. Furthermore, the lead compounds (1 and 6) were not effluxed by PfCRT(CQR) from the digestive vacuole but instead accumulated to very high levels within this organelle. Both 1 and 6 exhibited in vitro antiplasmodial activities that were inversely correlated with CQ. Moreover, the additional parasiticidal effect exerted by 1 and 6 in the drug-resistant parasites was attributable, at least in part, to their ability to inhibit PfCRT(CQR). This highlights the potential for devising new antimalarial therapies that exploit inherent weaknesses in a key resistance mechanism of P. falciparum.

  10. Dehydrodiisoeugenol, an isoeugenol dimer, inhibits lipopolysaccharide-stimulated nuclear factor kappa B activation and cyclooxygenase-2 expression in macrophages.

    Science.gov (United States)

    Murakami, Yukio; Shoji, Masao; Hirata, Atsushi; Tanaka, Shoji; Yokoe, Ichiro; Fujisawa, Seiichiro

    2005-02-15

    o-Methoxyphenols such as eugenol and isoeugenol exhibit anti-oxidant and anti-inflammatory activities, but at higher concentrations act as oxidants and potent allergens. We recently demonstrated the eugenol dimer bis-eugenol to be an efficient inhibitor of lipopolysaccharide (LPS)-induced inflammatory cytokine expression in macrophages without cytotoxicity. This result suggested that dimer compound of o-methoxyphenols may possess anti-inflammatory activity. Thus, we further synthesized dehydrodiisoeugenol and alpha-diisoeugenol from isoeugenols, and investigated whether these dimers could inhibit LPS-stimulated nuclear factor kappa B (NF-kappaB) activation and cyclooxygenase (COX)-2 gene expression, both of which are closely involved in inflammation and mutagenesis. The expression of the COX-2 gene was strongly inhibited by dehydrodiisoeugenol in RAW264.7 murine macrophages stimulated with LPS. In contrast, isoeugenol and alpha-diisoeugenol did not inhibit it. Dehydrodiisoeugenol also significantly inhibited LPS-stimulated phosphorylation-dependent proteolysis of inhibitor kappaB-alpha and transcriptional activity of NF-kappaB in the cells. These findings suggest that dehydrodiisoeugenol acts as a potent anti-inflammatory agent.

  11. Light-induced activation of class II cyclobutane pyrimidine dimer photolyases.

    Science.gov (United States)

    Okafuji, Asako; Biskup, Till; Hitomi, Kenichi; Getzoff, Elizabeth D; Kaiser, Gebhard; Batschauer, Alfred; Bacher, Adelbert; Hidema, Jun; Teranishi, Mika; Yamamoto, Kazuo; Schleicher, Erik; Weber, Stefan

    2010-05-04

    Light-induced activation of class II cyclobutane pyrimidine dimer (CPD) photolyases of Arabidopsis thaliana and Oryza sativa has been examined by UV/Vis and pulsed Davies-type electron-nuclear double resonance (ENDOR) spectroscopy, and the results compared with structure-known class I enzymes, CPD photolyase and (6-4) photolyase. By ENDOR spectroscopy, the local environment of the flavin adenine dinucleotide (FAD) cofactor is probed by virtue of proton hyperfine couplings that report on the electron-spin density at the positions of magnetic nuclei. Despite the amino-acid sequence dissimilarity as compared to class I enzymes, the results indicate similar binding motifs for FAD in the class II photolyases. Furthermore, the photoreduction kinetics starting from the FAD cofactor in the fully oxidized redox state, FAD(ox), have been probed by UV/Vis spectroscopy. In Escherichia coli (class I) CPD photolyase, light-induced generation of FADH from FAD(ox), and subsequently FADH(-) from FADH, proceeds in a step-wise fashion via a chain of tryptophan residues. These tryptophans are well conserved among the sequences and within all known structures of class I photolyases, but completely lacking from the equivalent positions of class II photolyase sequences. Nevertheless, class II photolyases show photoreduction kinetics similar to those of the class I enzymes. We propose that a different, but also effective, electron-transfer cascade is conserved among the class II photolyases. The existence of such electron transfer pathways is supported by the observation that the catalytically active fully reduced flavin state obtained by photoreduction is maintained even under oxidative conditions in all three classes of enzymes studied in this contribution.

  12. Dimerization of endogenous MT1-MMP is a regulatory step in the activation of the 72-kDa gelatinase MMP-2 on fibroblasts and fibrosarcoma cells

    DEFF Research Database (Denmark)

    Ingvarsen, S.; Madsen, D.H.; Hillig, T.

    2008-01-01

    with monovalent Fab fragments. Since only a negligible level of proMMP-2 activation was obtained with MT1-MMP-expressing cells in the absence of dimerization, our results identify the dimerization event as a critical level of proteolytic cascade regulation Udgivelsesdato: 2008/7...... by a monoclonal antibody that binds specifically to MT1-MMP as shown by immunofluorescence experiments. The antibody has no effect on the catalytic activity. The effect on proMMP-2 activation involves MT1-MMP dimerization because it requires the divalent monoclonal antibody, with no effect obtained...

  13. Cold inactivation and dissociation into dimers of Escherichia coli tryptophanase and its W330F mutant form.

    Science.gov (United States)

    Erez, T; Gdalevsky GYa; Torchinsky, Y M; Phillips, R S; Parola, A H

    1998-05-19

    The kinetics and mechanism of reversible cold inactivation of the tetrameric enzyme tryptophanase have been studied. Cold inactivation is shown to occur slowly in the presence of K+ ions and much faster in their absence. The W330F mutant tryptophanase undergoes rapid cold inactivation even in the presence of K+ ions. In all cases the inactivation is accompanied by a decrease of the coenzyme 420-nm CD and absorption peaks and a shift of the latter peak to shorter wavelengths. The spectral changes and the NaBH4 test indicate that cooling of tryptophanase leads to breaking of the internal aldimine bond and release of the coenzyme. HPLC analysis showed that the ensuing apoenzyme dissociates into dimers. The dissociation depends on the nature and concentration of anions in the buffer solution. It readily occurs at low protein concentrations in the presence of salting-in anions Cl-, NO3- and I-, whereas salting-out anions, especially HPO4(2-), hinder the dissociation. K+ ions do not influence the dissociation of the apoenzyme, but partially protect holotryptophanase from cold inactivation. Thus, the two processes, cold inactivation of tryptophanase and dissociation of its apoform into dimers exhibit different dependencies on K+ ions and anions.

  14. Heparin modulates the mitogenic activity of fibroblast growth factor by inducing dimerization of its receptor. a 3D view by using NMR.

    Science.gov (United States)

    Nieto, Lidia; Canales, Ángeles; Fernández, Israel S; Santillana, Elena; González-Corrochano, Rocío; Redondo-Horcajo, Mariano; Cañada, F Javier; Nieto, Pedro; Martín-Lomas, Manuel; Giménez-Gallego, Guillermo; Jiménez-Barbero, Jesús

    2013-09-23

    In vitro mitogenesis assays have shown that sulfated glycosaminoglycans (GAGs; heparin and heparan sulfate) cause an enhancement of the mitogenic activity of fibroblast growth factors (FGFs). Herein, we report that the simultaneous presence of FGF and the GAG is not an essential requisite for this event to take place. Indeed, preincubation with heparin (just before FGF addition) of cells lacking heparan sulfate produced an enhancing effect equivalent to that observed when the GAG and the protein are simultaneously added. A first structural characterization of this effect by analytical ultracentrifugation of a soluble preparation of the heparin-binding domain of fibroblast growth factor receptor 2 (FGFR2) and a low molecular weight (3 kDa) heparin showed that the GAG induces dimerization of FGFR2. To derive a high resolution structural picture of this molecular recognition process, the interactions of a soluble heparin-binding domain of FGFR2 with two different homogeneous, synthetic, and mitogenically active sulfated GAGs were analyzed by NMR spectroscopy. These studies, assisted by docking protocols and molecular dynamics simulations, have demonstrated that the interactions of these GAGs with the soluble heparin-binding domain of FGFR induces formation of an FGFR dimer; its architecture is equivalent to that in one of the two distinct crystallographic structures of FGFR in complex with both heparin and FGF1. This preformation of the FGFR dimer (with similar topology to that of the signaling complex) should favor incorporation of the FGF component to form the final assemblage of the signaling complex, without major entropy penalty. This cascade of events is probably at the heart of the observed activating effect of heparin in FGF-driven mitogenesis.

  15. Dimerization of a flocculent protein from Moringa oleifera: experimental evidence and in silico interpretation.

    Science.gov (United States)

    Pavankumar, Asalapuram R; Kayathri, Rajarathinam; Murugan, Natarajan A; Zhang, Qiong; Srivastava, Vaibhav; Okoli, Chuka; Bulone, Vincent; Rajarao, Gunaratna K; Ågren, Hans

    2014-01-01

    Many proteins exist in dimeric and other oligomeric forms to gain stability and functional advantages. In this study, the dimerization property of a coagulant protein (MO2.1) from Moringa oleifera seeds was addressed through laboratory experiments, protein-protein docking studies and binding free energy calculations. The structure of MO2.1 was predicted by homology modelling, while binding free energy and residues-distance profile analyses provided insight into the energetics and structural factors for dimer formation. Since the coagulation activities of the monomeric and dimeric forms of MO2.1 were comparable, it was concluded that oligomerization does not affect the biological activity of the protein.

  16. Circular dimers of lambda DNA in infected, nonlysogenic Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Freifelder, D.; Baran, N.; Folkmanis, A.; Freifelder, D.L.R.

    1977-09-01

    Covalently closed circular dimerss of phage lambda DNA have been found in Escherichia coli infected with lambda. These dimers can be formed by either the lambda Red or Int systems, by a nonrecombinational replicative mechanism requiring the activity of the lambda O and P genes or by joining of the cohesive ends. Dimers mediated by the E. coli Rec system have not been observed. Those formed by the Int system often result from recombination between different DNA molecules; however, the Red-mediated dimer may be a result of replicative extension of a single DNA molecule. Trimers have also been observed but studied only briefly.

  17. Universality in bosonic dimer-dimer scattering

    Energy Technology Data Exchange (ETDEWEB)

    Deltuva, A. [Centro de Fisica Nuclear, Universidade de Lisboa, P-1649-003 Lisboa (Portugal)

    2011-08-15

    Bosonic dimer-dimer scattering is studied near the unitary limit using momentum-space equations for the four-particle transition operators. The impact of the Efimov effect on the dimer-dimer scattering observables is explored, and a number of universal relations is established with high accuracy. The rate for the creation of Efimov trimers via dimer-dimer collisions is calculated.

  18. Structure of a Light-Activated LOV Protein Dimer That Regulates Transcription

    Energy Technology Data Exchange (ETDEWEB)

    Vaidya, Anand T.; Chen, Chen-Hui; Dunlap, Jay C.; Loros, Jennifer J.; Crane, Brian R. (Dartmouth-MED); (Cornell)

    2012-10-25

    Light, oxygen, or voltage (LOV) protein domains are present in many signaling proteins in bacteria, archaea, protists, plants, and fungi. The LOV protein VIVID (VVD) of the filamentous fungus Neurospora crassa enables the organism to adapt to constant or increasing amounts of light and facilitates proper entrainment of circadian rhythms. Here, we determined the crystal structure of the fully light-adapted VVD dimer and reveal the mechanism by which light-driven conformational change alters the oligomeric state of the protein. Light-induced formation of a cysteinyl-flavin adduct generated a new hydrogen bond network that released the amino (N) terminus from the protein core and restructured an acceptor pocket for binding of the N terminus on the opposite subunit of the dimer. Substitution of residues critical for the switch between the monomeric and the dimeric states of the protein had profound effects on light adaptation in Neurospora. The mechanism of dimerization of VVD provides molecular details that explain how members of a large family of photoreceptors convert light responses to alterations in protein-protein interactions.

  19. Dimerization of matrix metalloproteinase-2 (MMP-2): functional implication in MMP-2 activation.

    Science.gov (United States)

    Koo, Bon-Hun; Kim, Yeon Hyang; Han, Jung Ho; Kim, Doo-Sik

    2012-06-29

    Matrix metalloproteinase-2 (MMP-2) functions in diverse biological processes through the degradation of extracellular and non-extracellular matrix molecules. Because of its potential for tissue damage, there are several ways to regulate MMP-2 activity, including gene expression, compartmentalization, zymogen activation, and enzyme inactivation by extracellular inhibitors. Enzyme regulation through zymogen activation is important for the regulation of MMP-2 activity. In our previous studies, we showed that thrombin directly cleaved the propeptide of MMP-2 at specific sites for enzyme activation. We also demonstrated that heparan sulfate was required for thrombin-mediated activation of pro-MMP-2 by binding to thrombin, presumably through conformational changes at the active site of the enzyme. This suggests a regulatory mechanism for thrombin-mediated activation of pro-MMP-2. In this study, we found that MMP-2 formed a reduction-sensitive homodimer in a controlled manner and that Ca(2+) ion was essential for homodimerization of MMP-2. Homodimerization was not associated with protein kinase C-mediated phosphorylation of MMP-2. MMP-2 formed a homodimer through an intermolecular disulfide bond between Cys(102) and the neighboring Cys(102). Homodimerization of MMP-2 enhanced thrombin-mediated activation of pro-MMP-2. Moreover, the MMP-2 homodimer could cleave a small peptide substrate without removal of the propeptide. Taken together, our experimental data suggest a novel regulatory mechanism for pro-MMP-2 activation that is modulated through homodimerization of MMP-2.

  20. Studies on synthesis and activation mechanism of mitomycin dimers connected by 1,2-dithiolane and diol linkers.

    Science.gov (United States)

    Kim, Hyoung Rae; Kim, Jae Jin; Park, Jung Jae; Lee, Sang Hyup

    2012-10-01

    We report the synthetic and mechanistic studies on a new cyclic disulfide mitomycin dimer, 7-N,7'-N'-(1″,2″-dithiolanyl-3″,5″-dimethylenyl)bismitomycin C (8), and a diol mitomycin dimer, 7-N,7'-N'-(2″,4″-dihydroxy-1″,5″-pentanediyl)bismitomycin C (9). Mitomycin 8 is a dimer connected by a 1,2-dithiolane (a five-membered cyclic disulfide) linker, and was specifically designed to undergo nucleophilic activation and double DNA alkylations leading to efficient production of DNA interstrand cross-link (DNA ISC) adducts. Disulfide cleavage in 8 would generate two thiol groups that could serve as probes to activate two mitomycin rings. At first, the target mitomycin 8 was synthesized using mitomycin A (1) and the key intermediate, cyclic disulfide (10), which was prepared through a seven-step synthetic sequence. Diol mitomycin 9 was also synthesized from 1 and diamine salt 13. Next, kinetic studies using solvolysis reaction revealed that the activation rates of 8 were much higher than those of 9 and mitomycin C (2) under nucleophilic conditions provided by Et(3)P presumably due to the presence of a cyclic disulfide unit in 8. These findings led us to propose a nucleophilic activation pathway for 8. Then, DNA ISC experiments further revealed that the levels of DNA ISC caused by 8 in the presence of Et(3)P were much higher (97%) than those by 9 (5%) and 2 (4%). More importantly, mitomycin 8 underwent much faster activation and produced slightly higher levels of DNA ISC than the previously reported mitomycins 5-7. Overall, we concluded that 8 was highly efficient for both nucleophilic activation and corresponding DNA ISC formation, and that this differentiation came from the crucial function of the cyclic disulfide unit in 8.

  1. Crystalline products isolated from solutions with commercially available 2,3-bis(2-pyridyl)pyrazine (dpp) as reactant: Detection of a dimerized form of dpp

    Science.gov (United States)

    Grove, Hilde; Frøystein, Nils Åge; Sæthre, Leif J.; Sletten, Jorunn

    2006-12-01

    From reaction mixtures of commercially available 2,3-bis(2-pyridyl)pyrazine (dpp) and perchloric acid four different solid products have been isolated and structurally characterized by X-ray crystallography; (dppH 2)(ClO 4) 2·3H 2O ( 1), (dppH 2)(ClO 4) 2 ( 2), (ddppH 2)(ClO 4) 2 ( 3) and (dppH)(ClO 4) ( 4) (dppH is monoprotonated dpp, dppH 2 is the diprotonated dication of dpp, ddppH 2 is the dication 5,5'-bis(2,3-bis(2-pyridyl)pyrazinium), i.e. dimerized dppH). In 1 and 2 the nitrogen atom in both of the pyridyl rings in dpp is protonated. Hydrogen bonding and packing arrangements differ in the two compounds. In 3 a protonated and dimerized form of dpp is found; two 2,3-bis(2-pyridyl)pyrazinium units are connected by a C(sp 2) sbnd C(sp 2) bond in the 5-positions of the pyrazine rings, one pyridyl ring in each dpp moiety being protonated at the nitrogen. Strong intra-cation N sbnd H⋯N bonds between pyridyl rings are present. In 4 the monomeric dppH moieties display the same type of intramolecular hydrogen bonds as found in 3. Upon reacting dpp with copper(II) salts and oxalate, a dinuclear copper (II) complex, [Cu 2(ddpp)(ox) 2(H 2O) 2] (ddpp = 5,5'-bis(2,3-bis(2-pyridyl)pyrazine)), as well as mononuclear [Cu(dpp)(ox)(H 2O)] have been obtained in the same reaction mixtures. In one of the isolated crystalline products a dimer of dpp is found, as described in the case of compound 3, but no protonation has occurred. The crystal structure determination reveals a dinuclear complex with the neutral ddpp (dimerized dpp) as bridging ligand. Oxalate occurs as a bidentate, terminal ligand; water completes the copper coordination sphere. The dinuclear complex cocrystallizes with a mononuclear Cu-dpp-ox complex; in the mixed crystal two mononuclear units replace one dinuclear unit in 25% of the unit cells, thus the average formula is [Cu 2(ddpp)(ox) 2(H 2O) 2] 0.75[{Cu(dpp)(ox)(H 2O)} 2] 0.25·8H 2O ( 5). The monomeric species also crystallizes in the form of compound

  2. Effect of rubidium and cesium ions on the dimeric quaduplex formed by the Oxytricha nova telomeric repeat oligonucleotide d(GGGGTTTTGGGG).

    Science.gov (United States)

    Marincola, Flaminia Cesare; Virno, Ada; Randazzo, Antonio; Lai, Adolfo

    2007-01-01

    The DNA sequence d(GGGGTTTTGGGG) consists of 1.5 units of the repeat in telomeres of Oxytricha nova. It has been shown by NMR and x-ray crystallographic analysis that it is capable to form a dimeric quadruplex structure and that a variety of cations, namely K(+), Na(+), and NH(4)(+), are able to interact with this complex with different affinity, leading to complexes characterized by different local conformations. Thus, in order to improve the knowledge of this kind of molecule, and in particular to provide further insight into the role of monovalent cations in the G-quadruplex folding and conformation, we have investigated by (1)H-NMR the effect of the addition of Rb(+) and Cs(+) to the quadruplex formed by the oligonucleotide d(GGGGTTTTGGGG).

  3. Analysis of the DNA-binding and dimerization activities of Neurospora crassa transcription factor NUC-1.

    Science.gov (United States)

    Peleg, Y; Metzenberg, R L

    1994-12-01

    NUC-1, a positive regulatory protein of Neurospora crassa, controls the expression of several unlinked target genes involved in phosphorus acquisition. The carboxy-terminal end of the NUC-1 protein has sequence similarity to the helix-loop-helix family of transcription factors. Bacterially expressed and in vitro-synthesized proteins, which consist of the carboxy-terminal portion of NUC-1, bind specifically to upstream sequences of two of its target genes, pho2+ and pho-4+. These upstream sequences contain the core sequence, CACGTG, a target for many helix-loop-helix proteins. A large loop region (47 amino acids) separates the helix I and helix II domains. Mutations and deletion within the loop region did not interfere with the in vitro or in vivo functions of the protein. Immediately carboxy-proximal to the helix II domain, the NUC-1 protein contains an atypical zipper domain which is essential for function. This domain consists of a heptad repeat of alanine and methionine rather than leucine residues. Analysis of mutant NUC-1 proteins suggests that the helix II and the zipper domains are essential for the protein dimerization, whereas the basic and the helix I domains are involved in DNA binding. The helix I domain, even though likely to participate in dimer formation while NUC-1 is bound to DNA, is not essential for in vitro dimerization.

  4. Circular dimers of a lambda DNA in infected, nonlysogenic Escherichia coli.

    Science.gov (United States)

    Freifelder, D; Baran, N; Folkmanis, A; Freifelder, D L

    1977-09-01

    Covalently closed circular dimers of phage lambda DNA have been found in Escherichia coli infected with lambda. These dimers can be formed by either the lambda Red or Int systems, by a nonrecombinational replicative mechanism requiring the activity of the lambda O and P genes or by joining of the cohesive ends. Dimers mediated by the E. coli Rec system have not been observed. Those formed by the Int system often result from recombination between different DNA molecules; however, the Red-mediated dimers may be a result of replicative extension of a single DNA molecule. Trimers have also been observed but studied only briefly.

  5. Fibroblast growth factor 2 dimer with superagonist in vitro activity improves granulation tissue formation during wound healing.

    Science.gov (United States)

    Decker, Caitlin G; Wang, Yu; Paluck, Samantha J; Shen, Lu; Loo, Joseph A; Levine, Alex J; Miller, Lloyd S; Maynard, Heather D

    2016-03-01

    Site-specific chemical dimerization of fibroblast growth factor 2 (FGF2) with the optimal linker length resulted in a FGF2 homodimer with improved granulation tissue formation and blood vessel formation at exceptionally low concentrations. Homodimers of FGF2 were synthesized through site-specific linkages to both ends of different molecular weight poly(ethylene glycols) (PEGs). The optimal linker length was determined by screening dimer-induced metabolic activity of human dermal fibroblasts and found to be that closest to the inter-cysteine distance, 70 Å, corresponding to 2 kDa PEG. A straightforward analysis of the kinetics of second ligand binding as a function of tether length showed that, as the polymerization index (the number of monomer repeat units in the polymer, N) of the tether decreases, the mean time for second ligand capture decreases as ∼N(3/2), leading to an enhancement of the number of doubly bound ligands in steady-state for a given (tethered) ligand concentration. FGF2-PEG2k-FGF2 induced greater fibroblast metabolic activity than FGF2 alone, all other dimers, and all monoconjugates, at each concentration tested, with the greatest difference observed at low (0.1 ng/mL) concentration. FGF2-PEG2k-FGF2 further exhibited superior activity compared to FGF2 for both metabolic activity and migration in human umbilical vein endothelial cells, as well as improved angiogenesis in a coculture model in vitro. Efficacy in an in vivo wound healing model was assessed in diabetic mice. FGF2-PEG2k-FGF2 increased granulation tissue and blood vessel density in the wound bed compared to FGF2. The results suggest that this rationally designed construct may be useful for improving the fibroblast matrix formation and angiogenesis in chronic wound healing.

  6. Ni-, Pd-, or Pt-catalyzed ethylene dimerization: a mechanistic description of the catalytic cycle and the active species.

    Science.gov (United States)

    Roy, Dipankar; Sunoj, Raghavan B

    2010-03-07

    Two key mechanistic possibilities for group 10 transition metal [M(eta(3)-allyl)(PMe(3))](+) catalyzed (where M = Ni(II), Pd(II) and Pt(II)) ethylene dimerization are investigated using density functional theory methods. The nature of the potential active catalysts in these pathways is analyzed to gain improved insights into the mechanism of ethylene dimerization to butene. The catalytic cycle is identified as involving typical elementary steps in transition metal-catalyzed C-C bond formation reactions, such as oxidative insertion as well as beta-H elimination. The computed kinetic and thermodynamic features indicate that a commonly proposed metal hydride species (L(n)M-H) is less likely to act as the active species as compared to a metal-ethyl species (L(n)M-CH(2)CH(3)). Of the two key pathways considered, the active species is predicted to be a metal hydride in pathway-1, whereas a metal alkyl complex serves as the active catalyst in pathway-2. A metal-mediated hydride shift from a growing metal alkyl chain to the ethylene molecule, bound to the metal in an eta(2) fashion, is predicted to be the preferred route for the generation of the active species. Among the intermediates involved in the catalytic cycle, metal alkyls with a bound olefin are identified as thermodynamically stable for all three metal ions. In general, the Ni-catalyzed pathways are found to be energetically more favorable than those associated with Pd and Pt catalysts.

  7. Structure–Activity Relationship of Oligomeric Flavan-3-ols: Importance of the Upper-Unit B-ring Hydroxyl Groups in the Dimeric Structure for Strong Activities

    Directory of Open Access Journals (Sweden)

    Yoshitomo Hamada

    2015-10-01

    Full Text Available Proanthocyanidins, which are composed of oligomeric flavan-3-ol units, are contained in various foodstuffs (e.g., fruits, vegetables, and drinks and are strongly biologically active compounds. We investigated which element of the proanthocyanidin structure is primarily responsible for this functionality. In this study, we elucidate the importance of the upper-unit of 4–8 condensed dimeric flavan-3-ols for antimicrobial activity against Saccharomyces cerevisiae (S. cerevisiae and cervical epithelioid carcinoma cell line HeLa S3 proliferation inhibitory activity. To clarify the important constituent unit of proanthocyanidin, we synthesized four dimeric compounds, (−-epigallocatechin-[4,8]-(+-catechin, (−-epigallocatechin-[4,8]-(−-epigallocatechin, (−-epigallocatechin-[4,8]-(−-epigallocatechin-3-O-gallate, and (+-catechin-[4,8]-(−-epigallocatechin and performed structure–activity relationship (SAR studies. In addition to antimicrobial activity against S. cerevisiae and proliferation inhibitory activity on HeLa S3 cells, the correlation of 2,2-diphenyl-l-picrylhydrazyl radical scavenging activity with the number of phenolic hydroxyl groups was low. On the basis of the results of our SAR studies, we concluded that B-ring hydroxyl groups of the upper-unit of the dimer are crucially important for strong and effective activity.

  8. Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity.

    Science.gov (United States)

    Lim, Seng Koon; Sandén, Camilla; Selegård, Robert; Liedberg, Bo; Aili, Daniel

    2016-01-01

    Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

  9. Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity

    Science.gov (United States)

    Lim, Seng Koon; Sandén, Camilla; Selegård, Robert; Liedberg, Bo; Aili, Daniel

    2016-02-01

    Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

  10. Functional and evolutionary analyses of Helicobacter pylori HP0231 (DsbK) protein with strong oxidative and chaperone activity characterized by a highly diverged dimerization domain.

    Science.gov (United States)

    Bocian-Ostrzycka, Katarzyna M; Łasica, Anna M; Dunin-Horkawicz, Stanisław; Grzeszczuk, Magdalena J; Drabik, Karolina; Dobosz, Aneta M; Godlewska, Renata; Nowak, Elżbieta; Collet, Jean-Francois; Jagusztyn-Krynicka, Elżbieta K

    2015-01-01

    Helicobacter pylori does not encode the classical DsbA/DsbB oxidoreductases that are crucial for oxidative folding of extracytoplasmic proteins. Instead, this microorganism encodes an untypical two proteins playing a role in disulfide bond formation - periplasmic HP0231, which structure resembles that of EcDsbC/DsbG, and its redox partner, a membrane protein HpDsbI (HP0595) with a β-propeller structure. The aim of presented work was to assess relations between HP0231 structure and function. We showed that HP0231 is most closely related evolutionarily to the catalytic domain of DsbG, even though it possesses a catalytic motif typical for canonical DsbA proteins. Similarly, the highly diverged N-terminal dimerization domain is homologous to the dimerization domain of DsbG. To better understand the functioning of this atypical oxidoreductase, we examined its activity using in vivo and in vitro experiments. We found that HP0231 exhibits oxidizing and chaperone activities but no isomerizing activity, even though H. pylori does not contain a classical DsbC. We also show that HP0231 is not involved in the introduction of disulfide bonds into HcpC (Helicobacter cysteine-rich protein C), a protein involved in the modulation of the H. pylori interaction with its host. Additionally, we also constructed a truncated version of HP0231 lacking the dimerization domain, denoted HP0231m, and showed that it acts in Escherichia coli cells in a DsbB-dependent manner. In contrast, HP0231m and classical monomeric EcDsbA (E. coli DsbA protein) were both unable to complement the lack of HP0231 in H. pylori cells, though they exist in oxidized forms. HP0231m is inactive in the insulin reduction assay and possesses high chaperone activity, in contrast to EcDsbA. In conclusion, HP0231 combines oxidative functions characteristic of DsbA proteins and chaperone activity characteristic of DsbC/DsbG, and it lacks isomerization activity.

  11. Functional and evolutionary analyses of Helicobacter pylori HP0231 (DsbK protein with strong oxidative and chaperone activity characterized by a highly diverged dimerization domain

    Directory of Open Access Journals (Sweden)

    Katarzyna Marta Bocian-Ostrzycka

    2015-10-01

    Full Text Available Helicobacter pylori does not encode the classical DsbA/DsbB oxidoreductases that are crucial for oxidative folding of extracytoplasmic proteins. Instead, this microorganism encodes an untypical two proteins playing a role in disulfide bond formation – periplasmic HP0231, which structure resembles that of EcDsbC/DsbG, and its redox partner, a membrane protein HpDsbI (HP0595 with a -propeller structure. The aim of presented work was to assess relations between HP0231 structure and function.We showed that HP0231 is most closely related evolutionarily to the catalytic domain of DsbG, even though it possesses a catalytic motif typical for canonical DsbA proteins. Similarly, the highly diverged N-terminal dimerization domain is homologous to the dimerization domain of DsbG. To better understand the functioning of this atypical oxidoreductase, we examined its activity using in vivo and in vitro experiments. We found that HP0231 exhibits oxidizing and chaperone activities but no isomerizing activity, even though H. pylori does not contain a classical DsbC. We also show that HP0231 is not involved in the introduction of disulfide bonds into HcpC (Helicobacter cysteine-rich protein C, a protein involved in the modulation of the H. pylori interaction with its host. Additionally, we also constructed a truncated version of HP0231 lacking the dimerization domain, denoted HP0231m, and showed that it acts in E. coli cells in a DsbB-dependent manner. In contrast, HP0231m and classical monomeric EcDsbA (Escherichia coli DsbA protein were both unable to complement the lack of HP0231 in H. pylori cells, though they exist in oxidized forms. HP0231m is inactive in the insulin reduction assay and possesses high chaperone activity, in contrast to EcDsbA. In conclusion, HP0231 combines oxidative functions characteristic of DsbA proteins and chaperone activity characteristic of DsbC/DsbG, and it lacks isomerization activity.

  12. ERIS, an endoplasmic reticulum IFN stimulator, activates innate immune signaling through dimerization

    OpenAIRE

    2009-01-01

    We report here the identification and characterization of a protein, ERIS, an endoplasmic reticulum (ER) IFN stimulator, which is a strong type I IFN stimulator and plays a pivotal role in response to both non–self-cytosolic RNA and dsDNA. ERIS (also known as STING or MITA) resided exclusively on ER membrane. The ER retention/retrieval sequence RIR was found to be critical to retain the protein on ER membrane and to maintain its integrity. ERIS was dimerized on innate immune challenges. Coume...

  13. The solution configurations of inactive and activated DntR have implications for the sliding dimer mechanism of LysR transcription factors.

    Science.gov (United States)

    Lerche, Michael; Dian, Cyril; Round, Adam; Lönneborg, Rosa; Brzezinski, Peter; Leonard, Gordon A

    2016-01-28

    LysR Type Transcriptional Regulators (LTTRs) regulate basic metabolic pathways or virulence gene expression in prokaryotes. Evidence suggests that the activation of LTTRs involves a conformational change from an inactive compact apo- configuration that represses transcription to an active, expanded holo- form that promotes it. However, no LTTR has yet been observed to adopt both configurations. Here, we report the results of structural studies of various forms of the LTTR DntR. Crystal structures of apo-DntR and of a partially autoinducing mutant H169T-DntR suggest that active and inactive DntR maintain a compact homotetrameric configuration. However, Small Angle X-ray Scattering (SAXS) studies on solutions of apo-, H169T- and inducer-bound holo-DntR indicate a different behaviour, suggesting that while apo-DntR maintains a compact configuration in solution both H169T- and holo-DntR adopt an expanded conformation. Models of the SAXS-obtained solution conformations of apo- and holo-DntR homotetramers in complex with promoter-operator region DNA are consistent with previous observations of a shifting of LTTR DNA binding sites upon activation and a consequent relaxation in the bend of the promoter-operator region DNA. Our results thus provide clear evidence at the molecular level which strongly supports the 'sliding dimer' hypothesis concerning LTTR activation mechanisms.

  14. Relationship between structure and P-glycoprotein inhibitory activity of dimeric peptides related to the Dmt-Tic pharmacophore.

    Science.gov (United States)

    Ambo, Akihiro; Ohkatsu, Hiromichi; Minamizawa, Motoko; Watanabe, Hideko; Sugawara, Shigeki; Nitta, Kazuo; Tsuda, Yuko; Okada, Yoshio; Sasaki, Yusuke

    2012-03-15

    To develop novel inhibitors of P-glycoprotein (P-gp), dimeric peptides related to an opioid peptide containing the Dmt-Tic pharmacophore were synthesized and their P-gp inhibitory activities were analyzed. Of the 30 analogs synthesized, N(α),N(ε)-[(CH(3))(2)Mle-Tic](2)Lys-NH(2) and its D-Lys analog were found to exhibit potent P-gp inhibitory activity, twice that of verapamil, in doxorubicin-resistant K562 cells. Structure-activity studies indicated that the correct hydrophobicity and spacer length between two aromatic rings are important structural elements in this series of analogs for inhibition of P-gp.

  15. DNA melting properties of the dityrosine cross-linked dimer of Ribonuclease A.

    Science.gov (United States)

    Dinda, Amit Kumar; Chattaraj, Saparya; Ghosh, Sudeshna; Tripathy, Debi Ranjan; Dasgupta, Swagata

    2016-09-01

    Several DNA binding proteins exist in dimeric form when bound with DNA to be able to exhibit various biological processes such as DNA repair, DNA replication and gene expression. Various dimeric forms of Ribonuclease A (RNase A) and other members of the ribonuclease A superfamily are endowed with a multitude of biological activities such as antitumor and antiviral activity. In the present study, we have compared the DNA binding properties between the RNase A monomer and the dityrosine (DT) cross-linked RNase A dimer, and checked the inhibitory effect of DNA on the ribonucleolytic activity of the dimeric protein. An agarose gel based assay shows that like the monomer, the dimer also binds with DNA. The number of nucleotides bound per monomer unit of the dimer is higher than the number of nucleotides that bind with the each monomer. From fluorescence measurements, the association constant (Ka) values for complexation of the monomer and the dimer with ct-DNA are (4.95±0.45)×10(4)M(-1) and (1.29±0.05)×10(6)M(-1) respectively. Binding constant (Kb) values for the binding of the monomer and the dimer with ct-DNA were determined using UV-vis spectroscopy and were found to be (4.96±1.67)×10(4)M(-1) and (4.32±0.31)×10(5)M(-1) respectively. Circular dichroism studies shows that the dimer possesses significant effect on DNA conformation. The melting profile for the ct-DNA-dimer indicated that the melting temperature (Tm) for the ct-DNA-dimer complex is lower compared to the ct-DNA-monomer complex. The ribonucleolytic activity of the dimer, like the monomer, diminishes upon binding with DNA.

  16. In vitro release and antibacterial activity of poly(oleic/linoleic acid dimer:sebacic acid)-gentamicin

    Institute of Scientific and Technical Information of China (English)

    YANGXiu-Fen; ZHOUZhi-Bin; 等

    2003-01-01

    AIM:To investigate whether poly(oleic/linoleic acid dimer:sebacic acid)-getamicin[Poly(OAD/LOAD:SA)-gentamicin]delivery system was useful to treat chronic osteomyelitis.METHODS:Drug delivery system consisted of gentamicin sufate dispersed in a copolymer containing oleic/linoleic acid dimer(OAD/LOAD)and sebacic acid(SA)in a 1:1 weight ration.The gentamicin releast from[Poly(OAD/LOAD:SA)-gentamicin]was tested in water 0.9% saline,and phosphate buffer 0.1mol/L,RESULTS:The gentamicin concentration peak was found on d2,then slowly decreased.considerable amout of gentamicin was still released on d 50.From d 2 o d 50,the gentamicin concentration in the releasing fluids was from 59 to 42128-fold and 1.8 to 1314-fold of the MIC for Staphylococcus aureus and Escherichia coli,respectively.Staphylococcus aureus and Escherichia coli were strongly inhibited by the releasing fluids for 50d.The gentamicin release and anti-bacterial activity in the three media were similar.only in 0.1mol/L phosphate buffer,from d 2 to 14 it was lower.CONCLUSION:Poly(OAD/LOAD:SA)-gentamicin was useful to treat chronic osteomyelitis.

  17. Crystal Structure of a Novel Dimeric Form of NS5A Domain I Protein from Hepatitis C Virus

    Energy Technology Data Exchange (ETDEWEB)

    Love, Robert A.; Brodsky, Oleg; Hickey, Michael J.; Wells, Peter A.; Cronin, Ciarán N.; Pfizer

    2009-07-10

    A new protein expression vector design utilizing an N-terminal six-histidine tag and tobacco etch virus protease cleavage site upstream of the hepatitis C virus NS5A sequence has resulted in a more straightforward purification method and improved yields of purified NS5A domain I protein. High-resolution diffracting crystals of NS5A domain I (amino acids 33 to 202) [NS5A(33-202)] were obtained by using detergent additive crystallization screens, leading to the structure of a homodimer which is organized differently from that published previously (T. L. Tellinghuisen, J. Marcotrigiano, and C. M. Rice, Nature 435:374-379, 2005) yet is consistent with a membrane association model for NS5A. The monomer-monomer interface of NS5A(33-202) features an extensive buried surface area involving the most-highly conserved face of each monomer. The two alternate structural forms of domain I now available may be indicative of the multiple roles emerging for NS5A in viral RNA replication and viral particle assembly.

  18. Doubly chloro bridged dimeric copper(II) complex: magneto-structural correlation and anticancer activity.

    Science.gov (United States)

    Sikdar, Yeasin; Modak, Ritwik; Bose, Dipayan; Banerjee, Saswati; Bieńko, Dariusz; Zierkiewicz, Wiktor; Bieńko, Alina; Das Saha, Krishna; Goswami, Sanchita

    2015-05-21

    We have synthesized and structurally characterized a new doubly chloro bridged dimeric copper(II) complex, [Cu2(μ-Cl)2(HL)2Cl2] (1) based on a Schiff base ligand, 5-[(pyridin-2-ylmethylene)-amino]-pentan-1-ol). Single crystal X-ray diffraction shows the presence of dinuclear copper(II) centres in a square pyramidal geometry linked by obtuse double chloro bridge. The magnetic study illustrated that weak antiferromagnetic interactions (J = -0.47 cm(-1)) prevail in complex 1 which is well supported by magneto-structural correlation. This compound adds to the library of doubly chloro bridged copper(ii) complexes in the regime of spin state cross over. DFT calculations have been conducted within a broken-symmetry (BS) framework to investigate the exchange interaction further which depicts that the approximate spin projection technique yields the best corroboration of the experimental J value. Spin density plots show the presence of an ∼0.52e charge residing on the copper atom along with a substantial charge on bridging and peripheral chlorine atoms. The potential of complex1 to act as an anticancer agent is thoroughly examined on a series of liver cancer cell lines and screening shows the HepG2 cell line exhibits maximum cytotoxicity by phosphatidyl serine exposure in the outer cell membrane associated with ROS generation and mitochondrial depolarization with increasing time in the in vitro model system.

  19. Determination of Activity Coefficients of di-(2-ethylhexyl) Phosphoric Acid Dimer in Select Organic Solvents Using Vapor Phase Osmometry

    Energy Technology Data Exchange (ETDEWEB)

    Michael F. Gray; Peter Zalupski; Mikael Nilsson

    2013-08-01

    Effective models for solvent extraction require accurate characterization of the nonideality effects for each component, including the extractants. In this study, the nonideal behavior of the industrial extractant di(2-ethylhexyl) phosphoric acid has been investigated using vapor pressure osmometry (VPO). From the osmometry data, activity coefficients for the HDEHP dimer were obtained based on a formulation of the regular solution theory of Scatchard and Hildebrand, and the Margules two- and three-suffix equations. The results show similarity with a slope-analysis based relation from previous literature, although important differences are highlighted. The work points towards VPO as a useful technique for this type of study, but care must be taken with the choice of standard and method of analysis.

  20. Studies on activation mechanism of a mitomycin dimer, 7-N,7'-N'-(1″,2″-dithiepanyl-3″,7″-dimethylenyl)bismitomycin C.

    Science.gov (United States)

    Kim, Jae Jin; Kim, Hyoung Rae; Lee, Sang Hyup

    2012-09-01

    We report the studies on nucleophilic activation and DNA alkylation of a cyclic disulfide mitomycin dimer, 7-N,7'-N'-(1″,2″-dithiepanyl-3″,7″-dimethylenyl)bismitomycin C (6) along with a diol mitomycin dimer, 7-N,7'-N'-(2″,6″-dihydroxy-1″,7″-heptanediyl)bismitomycin C (7). We wished to see if disulfide mitomycin 6 undergoes efficient nucleophilic activation and corresponding formation of DNA interstrand cross-link (DNA ISC) products compared to diol mitomycin 7. Mitomycin 6 is a dimer connected by a seven-membered cyclic disulfide (a 1,2-dithiepane) linker, and mitomycin 7 is also a dimer containing 2,6-dihydroxyheptane linker that was employed as a reference one to identify the effect of disulfide unit in 6. Through kinetic studies using solvolysis reaction, we found that 6 underwent much faster nucleophilic activation by Et3P compared to 7, and that the enhanced activation rates were induced by the disulfide unit in 6. These findings led us to propose a nucleophilic activation mechanism for 6. We further demonstrated that 6 produced much higher levels of DNA ISC (86%) by the action of Et3P compared with 7 (5%) and 1 (4%). Therefore, we have concluded that 6 was highly efficient for nucleophilic activation and DNA ISC formation due to the key role of cyclic disulfide unit in 6.

  1. Cooperative binding mode of the inhibitors of R6K replication, pi dimers.

    Science.gov (United States)

    Bowers, Lisa M; Filutowicz, Marcin

    2008-03-28

    The replication initiator protein, pi, plays an essential role in the initiation of plasmid R6K replication. Both monomers and dimers of pi bind to iterons in the gamma origin of plasmid R6K, yet monomers facilitate open complex formation, while dimers, the predominant form in the cell, do not. Consequently, pi monomers activate replication, while pi dimers inhibit replication. Recently, it was shown that the monomeric form of pi binds multiple tandem iterons in a strongly cooperative fashion, which might explain how monomers outcompete dimers for replication initiation when plasmid copy number and pi supply are low. Here, we examine cooperative binding of pi dimers and explore the role that these interactions may have in the inactivation of gamma origin. To examine pi dimer/iteron interactions in the absence of competing pi monomer/iteron interactions using wild-type pi, constructs were made with key base changes to each iteron that eliminate pi monomer binding yet have no impact on pi dimer binding. Our results indicate that, in the absence of pi monomers, pi dimers bind with greater cooperativity to alternate iterons than to adjacent iterons, thus preferentially leaving intervening iterons unbound and the origin unsaturated. We discuss new insights into plasmid replication control by pi dimers.

  2. The PH Domain of PDK1 Exhibits a Novel, Phospho-Regulated Monomer-Dimer Equilibrium With Important Implications for Kinase Domain Activation: Single Molecule and Ensemble Studies†

    Science.gov (United States)

    Ziemba, Brian P.; Pilling, Carissa; Calleja, Véronique; Larijani, Banafshé; Falke, Joseph J.

    2013-01-01

    Phosphoinositide-Dependent Kinase-1 (PDK1) is an essential master kinase recruited to the plasma membrane by the binding of its C-terminal PH domain to the signaling lipid phosphatidylinositol-3,4-5-trisphosphate (PIP3). Membrane binding leads to PDK1 phospho-activation, but despite the central role of PDK1 in signaling and cancer biology this activation mechanism remains poorly understood. PDK1 has been shown to exist as a dimer in cells, and one crystal structure of its isolated PH domain exhibits a putative dimer interface. It has been proposed that phosphorylation of PH domain residue T513 (or the phospho-mimetic T513E mutation) may regulate a novel PH domain dimer-monomer equilibrium, thereby converting an inactive PDK1 dimer to an active monomer. However, the oligomeric state(s) of the PH domain on the membrane have not yet been determined, nor whether a negative charge at position 513 is sufficient to regulate its oligomeric state. The present study investigates the binding of purified WT and T513E PDK1 PH domains to lipid bilayers containing the PIP3 target lipid, using both single molecule and ensemble measurements. Single molecule analysis of the brightness of fluorescent PH domain shows that the PIP3-bound WT PH domain on membranes is predominantly dimeric, while the PIP3-bound T513E PH domain is monomeric, demonstrating that negative charge at the T513 position is sufficient to dissociate the PH domain dimer and is thus likely to play a central role in PDK1 monomerization and activation. Single molecule analysis of 2-D diffusion of PH domain-PIP3 complexes reveals that the dimeric WT PH domain diffuses at the same rate a single lipid molecule, indicating that only one of its two PIP3 binding sites is occupied and there is little protein penetration into the bilayer as observed for other PH domains. The 2-D diffusion of T513E PH domain is slower, suggesting the negative charge disrupts local structure in a way that enables greater protein insertion into

  3. Structure of a dimeric crenarchaeal Cas6 enzyme with an atypical active site for CRISPR RNA processing

    Science.gov (United States)

    Reeks, Judith; Sokolowski, Richard D.; Graham, Shirley; Liu, Huanting; Naismith, James H.; White, Malcolm F.

    2013-01-01

    The competition between viruses and hosts is played out in all branches of life. Many prokaryotes have an adaptive immune system termed ‘CRISPR’ (clustered regularly interspaced short palindromic repeats) which is based on the capture of short pieces of viral DNA. The captured DNA is integrated into the genomic DNA of the organism flanked by direct repeats, transcribed and processed to generate crRNA (CRISPR RNA) that is loaded into a variety of effector complexes. These complexes carry out sequence-specific detection and destruction of invading mobile genetic elements. In the present paper, we report the structure and activity of a Cas6 (CRISPR-associated 6) enzyme (Sso1437) from Sulfolobus solfataricus responsible for the generation of unit-length crRNA species. The crystal structure reveals an unusual dimeric organization that is important for the enzyme's activity. In addition, the active site lacks the canonical catalytic histidine residue that has been viewed as an essential feature of the Cas6 family. Although several residues contribute towards catalysis, none is absolutely essential. Coupled with the very low catalytic rate constants of the Cas6 family and the plasticity of the active site, this suggests that the crRNA recognition and chaperone-like activities of the Cas6 family should be considered as equal to or even more important than their role as traditional enzymes. PMID:23527601

  4. Effects of Dimerization of Serratia marcescens Endonuclease on Water Dynamics.

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Chuanying; Beck, Brian W.; Krause, Kurt; Weksberg, Tiffany E.; Pettitt, Bernard M.

    2007-02-15

    The research described in this product was performed in part in the Environmental Molecular Sciences Laboratory, a national scientific user facility sponsored by the Department of Energy's Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory. The dynamics and structure of Serratia marcescens endonuclease and its neighboring solvent are investigated by molecular dynamics (MD). Comparisons are made with structural and biochemical experiments. The dimer form is physiologic and functions more processively than the monomer. We previously found a channel formed by connected clusters of waters from the active site to the dimer interface. Here, we show that dimerization clearly changes correlations in the water structure and dynamics in the active site not seen in the monomer. Our results indicate that water at the active sites of the dimer is less affected compared with bulk solvent than in the monomer where it has much slower characteristic relaxation times. Given that water is a required participant in the reaction, this gives a clear advantage to dimerization in the absence of an apparent ability to use both active sites simultaneously.

  5. Laforin, a dual specificity phosphatase involved in Lafora disease, is present mainly as monomeric form with full phosphatase activity.

    Directory of Open Access Journals (Sweden)

    Vikas V Dukhande

    Full Text Available Lafora Disease (LD is a fatal neurodegenerative epileptic disorder that presents as a neurological deterioration with the accumulation of insoluble, intracellular, hyperphosphorylated carbohydrates called Lafora bodies (LBs. LD is caused by mutations in either the gene encoding laforin or malin. Laforin contains a dual specificity phosphatase domain and a carbohydrate-binding module, and is a member of the recently described family of glucan phosphatases. In the current study, we investigated the functional and physiological relevance of laforin dimerization. We purified recombinant human laforin and subjected the monomer and dimer fractions to denaturing gel electrophoresis, mass spectrometry, phosphatase assays, protein-protein interaction assays, and glucan binding assays. Our results demonstrate that laforin prevalently exists as a monomer with a small dimer fraction both in vitro and in vivo. Of mechanistic importance, laforin monomer and dimer possess equal phosphatase activity, and they both associate with malin and bind glucans to a similar extent. However, we found differences between the two states' ability to interact simultaneously with malin and carbohydrates. Furthermore, we tested other members of the glucan phosphatase family. Cumulatively, our data suggest that laforin monomer is the dominant form of the protein and that it contains phosphatase activity.

  6. Purification of a dimeric arginine deiminase from Enterococcus faecium GR7 and study of its anti-cancerous activity.

    Science.gov (United States)

    Kaur, Baljinder; Kaur, Rajinder

    2016-09-01

    The arginine deiminase (ADI, E.C 3.5.3.6) - a key enzyme of ADI pathway of Enterococcus faecium GR7 was purified to homogeneity. A sequential purification strategy involving ammonium sulfate fractionation, molecular sieve followed by Sephadex G-100 gel filtration was applied to the crude culture filtrate to obtain a pure enzyme preparation. The enzyme was purified with a fold of 16.92 and showed a final specific activity of 76.65IU/mg with a 49.17% yield. The dimeric ADI has a molecular mass of about 94,364.929Da, and comprises of hetrodimers of 49.1kDa and 46.5kDa as determined by MALDI-TOF and PAGE analysis. To assess anti-cancerous activity of ADI by MTT assay was carried out against cancer cell lines (MCF-7, Sp2/0-Ag14 and Hep-G2). Purified ADI exhibited the most profound antiproliferative activity against Hep-G2 cells; with half-maximal inhibitory concentration (IC50) of 1.95μg/ml. Purified ADI from E. faecium GR7 was observed to induce apoptosis in the Hep-G2 cells by DNA fragmentation assay. Our findings suggest the possibility of a future use of ADI from E. faecium GR7 as a potential anticancer drug.

  7. Circumnuclear Star Forming Activity in NGC 3982

    Institute of Scientific and Technical Information of China (English)

    Shui-Nai Zhang; Qiu-Sheng Gu; Yi-Peng Wang

    2008-01-01

    We present a study of the nearby Seyfert galaxy NGC 3982 using optical,infrared and X-ray data acquired by SDSS,Spitzer and Chandra.Our main results are as follows:(1) A simple stellar population synthesis on the nuclear and circumnuclear SDSS spectra gives unambiguous evidence of young stellar components in both the nuclear and circumnuclear regions.(2) The Spitzer Infrared Spectrograph (IRS) spectrum of the central region (~3") shows a power-law continuum,a silicate emission feature at 9.7 μm,and significant PAH emission features at 7.7,8.6,11.3 and 12.7/zm,suggesting the coexistence of AGN and starburst activities in the central region of NGC 3982.(3) We estimate the star formation rate (SFR) of the circumnuclear (~5"-20") region from the Ha luminosity to be for the active nucleus of NGC 3982 from radio to X-ray,and obtain a bolometric luminosity of Lbol=4.5×1042 erg s-1,corresponding to an Eddington ratio (Lbol/LEdd) of 0.014.The HST image of NGC 3982 shows a nuclear mini-spiral between the circumnuclear starforming region and the nucleus,which could be the channel through which gas is transported to the supermassive black hole from the circumnuclear star-forming region.

  8. Structural Basis for Enhanced HIV-1 Neutralization by a Dimeric Immunoglobulin G Form of the Glycan-Recognizing Antibody 2G12

    Directory of Open Access Journals (Sweden)

    Yunji Wu

    2013-12-01

    Full Text Available The human immunoglobulin G (IgG 2G12 recognizes high-mannose carbohydrates on the HIV type 1 (HIV-1 envelope glycoprotein gp120. Its two antigen-binding fragments (Fabs are intramolecularly domain exchanged, resulting in a rigid (Fab2 unit including a third antigen-binding interface not found in antibodies with flexible Fab arms. We determined crystal structures of dimeric 2G12 IgG created by intermolecular domain exchange, which exhibits increased breadth and >50-fold increased neutralization potency compared with monomeric 2G12. The four Fab and two fragment crystalline (Fc regions of dimeric 2G12 were localized at low resolution in two independent structures, revealing IgG dimers with two (Fab2 arms analogous to the Fabs of conventional monomeric IgGs. Structures revealed three conformationally distinct dimers, demonstrating flexibility of the (Fab2-Fc connections that was confirmed by electron microscopy, small-angle X-ray scattering, and binding studies. We conclude that intermolecular domain exchange, flexibility, and bivalent binding to allow avidity effects are responsible for the increased potency and breadth of dimeric 2G12.

  9. Fatty acids modulate Toll-like receptor 4 activation through regulation of receptor dimerization and recruitment into lipid rafts in a reactive oxygen species-dependent manner.

    Science.gov (United States)

    Wong, Scott W; Kwon, Myung-Ja; Choi, Augustine M K; Kim, Hong-Pyo; Nakahira, Kiichi; Hwang, Daniel H

    2009-10-02

    The saturated fatty acids acylated on Lipid A of lipopolysaccharide (LPS) or bacterial lipoproteins play critical roles in ligand recognition and receptor activation for Toll-like Receptor 4 (TLR4) and TLR2. The results from our previous studies demonstrated that saturated and polyunsaturated fatty acids reciprocally modulate the activation of TLR4. However, the underlying mechanism has not been understood. Here, we report for the first time that the saturated fatty acid lauric acid induced dimerization and recruitment of TLR4 into lipid rafts, however, dimerization was not observed in non-lipid raft fractions. Similarly, LPS and lauric acid enhanced the association of TLR4 with MD-2 and downstream adaptor molecules, TRIF and MyD88, into lipid rafts leading to the activation of downstream signaling pathways and target gene expression. However, docosahexaenoic acid (DHA), an n-3 polyunsaturated fatty acid, inhibited LPS- or lauric acid-induced dimerization and recruitment of TLR4 into lipid raft fractions. Together, these results demonstrate that lauric acid and DHA reciprocally modulate TLR4 activation by regulation of the dimerization and recruitment of TLR4 into lipid rafts. In addition, we showed that TLR4 recruitment to lipid rafts and dimerization were coupled events mediated at least in part by NADPH oxidase-dependent reactive oxygen species generation. These results provide a new insight in understanding the mechanism by which fatty acids differentially modulate TLR4-mediated signaling pathway and consequent inflammatory responses which are implicated in the development and progression of many chronic diseases.

  10. Molecular structure investigation of neutral, dimer and anion forms of 3,4-pyridinedicarboxylic acid: A combined experimental and theoretical study

    Science.gov (United States)

    Karabacak, Mehmet; Bilgili, Sibel; Atac, Ahmet

    2015-01-01

    In this study, the structural and vibrational analysis of 3,4-pyridinedicarboxylic acid (3,4-PDCA) are presented using experimental techniques as FT-IR, FT-Raman, NMR, UV and quantum chemical calculations. FT-IR and FT-Raman spectra of 3,4-pyridinedicarboxylic acid in the solid phase are recorded in the region 4000-400 cm-1 and 4000-50 cm-1, respectively. The geometrical parameters and energies of all different and possible monomer, dimer, anion-1 and anion-2 conformers of 3,4-PDCA are obtained from Density Functional Theory (DFT) with B3LYP/6-311++G(d,p) basis set. There are sixteen conformers (C1sbnd C16) for this molecule (neutral form). The most stable conformer of 3,4-PDCA is the C1 conformer. The complete assignments are performed on the basis of the total energy distribution (TED) of the vibrational modes calculated with scaled quantum mechanics (SQM) method. 1H and 13C NMR spectra are recorded and the chemical shifts are calculated by using DFT/B3LYP methods with 6-311++G(d,p) basis set. The UV absorption spectrum of the studied compound is recorded in the range of 200-400 nm by dissolved in ethanol. The optimized geometric parameters were compared with experimental data via the X-ray results derived from complexes of this molecule. In addition these, molecular electrostatic potential (MEP), thermodynamic and electronic properties, HOMO-LUMO energies and Mulliken atomic charges, are performed.

  11. Tetramerization reinforces the dimer interface of MnSOD.

    Directory of Open Access Journals (Sweden)

    Yuewei Sheng

    Full Text Available Two yeast manganese superoxide dismutases (MnSOD, one from Saccharomyces cerevisiae mitochondria (ScMnSOD and the other from Candida albicans cytosol (CaMnSODc, have most biochemical and biophysical properties in common, yet ScMnSOD is a tetramer and CaMnSODc is a dimer or "loose tetramer" in solution. Although CaMnSODc was found to crystallize as a tetramer, there is no indication from the solution properties that the functionality of CaMnSODc in vivo depends upon the formation of the tetrameric structure. To elucidate further the functional significance of MnSOD quaternary structure, wild-type and mutant forms of ScMnSOD (K182R, A183P mutant and CaMnSODc (K184R, L185P mutant with the substitutions at dimer interfaces were analyzed with respect to their oligomeric states and resistance to pH, heat, and denaturant. Dimeric CaMnSODc was found to be significantly more subject to thermal or denaturant-induced unfolding than tetrameric ScMnSOD. The residue substitutions at dimer interfaces caused dimeric CaMnSODc but not tetrameric ScMnSOD to dissociate into monomers. We conclude that the tetrameric assembly strongly reinforces the dimer interface, which is critical for MnSOD activity.

  12. 75 FR 26782 - Agency Information Collection Activities: Form I-864, Form I-864A, Form I-864EZ, and Form I-864W...

    Science.gov (United States)

    2010-05-12

    ... Agency Information Collection Activities: Form I-864, Form I- 864A, Form I-864EZ, and Form I-864W... Collection Under Review; Form I- 864, Affidavit of Support Under Section 213A of the Act; Form I-864A, Contract Between Sponsor and Household Member, Form I-864EZ, Affidavit of Support Under Section 213A of...

  13. Elements located upstream and downstream of the major splice donor site influence the ability of HIV-2 leader RNA to dimerize in vitro.

    Science.gov (United States)

    Lanchy, Jean-Marc; Rentz, Casey A; Ivanovitch, John D; Lodmell, J Stephen

    2003-03-11

    An essential step in the replication cycle of all retroviruses is the dimerization of genomic RNA prior to or during budding and maturation of the viral particle. In HIV-1, a 5' leader region site termed stem-loop 1 (SL1) promotes RNA dimerization in vitro and influences dimerization in vivo. In HIV-2, two sequences promote dimerization of RNA fragments in vitro: the 5'-end of the primer-binding site (PBS) and a stem-loop region homologous to the HIV-1 SL1 sequence. Because HIV-2 RNA constructs of different lengths use these two dimerization signals disproportionately, we hypothesized that other sequences could modulate their relative utilization. Here, we characterized the influence of sequences upstream and downstream of the major splice donor site on the formation of HIV-2 RNA dimers in vitro using a variety of RNA constructs and dimerization and electrophoresis protocols. We first assayed the formation of loose or tight dimers for 1-444 and 1-561 model RNAs. Although both RNAs could form PBS-dependent loose dimers, the 1-561 RNA was unable to make SL1-dependent tight dimers. Using RNAs truncated at their 5'- and/or 3'-ends and by making compensatory base substitutions, we found that two elements interfere with the formation of SL1-dependent tight dimers. The cores of these elements are located at nucleotides 189-196 and 543-550. Our results suggest that base pairing between these sequences prevents the formation of SL1-dependent tight dimers, probably by sequestering SL1 in a stable intramolecular arrangement. Moreover, we found that nucleotides downstream of SL1 decreased the rate of tight dimerization. Interestingly, dimerization at 37 degrees C in the presence of nucleocapsid protein increased the yield of SL1-mediated tight dimerization in vitro, even in the presence of the two interfering elements, suggesting a relationship between the nucleocapsid protein and activation of the SL1 dimerization signal in vivo.

  14. Novel covalently linked insulin dimer engineered to investigate the function of insulin dimerization

    DEFF Research Database (Denmark)

    Vinther, Tine N.; Norrman, Mathias; Strauss, Holger M.;

    2012-01-01

    and insulin stability and function, we engineered a covalently linked insulin dimer in which two monomers were linked by a disulfide bond. The structure of this covalent dimer was identical to the self-association dimer of human insulin. Importantly, this covalent dimer was capable of further oligomerization...... to form the structural equivalent of the classical hexamer. The covalently linked dimer neither bound to the insulin receptor, nor induced a metabolic response in vitro. However, it was extremely thermodynamically stable and did not form amyloid fibrils when subjected to mechanical stress, underlining......An ingenious system evolved to facilitate insulin binding to the insulin receptor as a monomer and at the same time ensure sufficient stability of insulin during storage. Insulin dimer is the cornerstone of this system. Insulin dimer is relatively weak, which ensures dissociation into monomers...

  15. Products and mechanism of acene dimerization. A computational study.

    Science.gov (United States)

    Zade, Sanjio S; Zamoshchik, Natalia; Reddy, A Ravikumar; Fridman-Marueli, Galit; Sheberla, Dennis; Bendikov, Michael

    2011-07-20

    The high reactivity of acenes can reduce their potential applications in the field of molecular electronics. Although pentacene is an important material for use in organic field-effect transistors because of its high charge mobility, its reactivity is a major disadvantage hindering the development of pentacene applications. In this study, several reaction pathways for the thermal dimerization of acenes were considered computationally. The formation of acene dimers via a central benzene ring and the formation of acene-based polymers were found to be the preferred pathways, depending on the length of the monomer. Interestingly, starting from hexacene, acene dimers are thermodynamically disfavored products, and the reaction pathway is predicted to proceed instead via a double cycloaddition reaction (polymerization) to yield acene-based polymers. A concerted asynchronous reaction mechanism was found for benzene and naphthalene dimerization, while a stepwise biradical mechanism was predicted for the dimerization of anthracene, pentacene, and heptacene. The biradical mechanism for dimerization of anthracene and pentacene proceeds via syn or anti transition states and biradical minima through stepwise biradical pathways, while dimerization of heptacene proceeds via asynchronous ring closure of the complex formed by two heptacene molecules. The activation barriers for thermal dimerization decrease rapidly with increasing acene chain length and are calculated (at M06-2X/6-31G(d)+ZPVE) to be 77.9, 57.1, 33.3, -0.3, and -12.1 kcal/mol vs two isolated acene molecules for benzene, naphthalene, anthracene, pentacene, and heptacene, respectively. If activation energy is calculated vs the initially formed complex of two acene molecules, then the calculated barriers are 80.5, 63.2, 43.7, 16.7, and 12.3 kcal/mol. Dimerization is exothermic from anthracene onward, but it is endothermic at the terminal rings, even for heptacene. Phenyl substitution at the most reactive meso

  16. Solubility parameter and activity coefficient of HDEHP dimer in select organic diluents by vapor pressure osmometry

    Energy Technology Data Exchange (ETDEWEB)

    Gray, M.; Nilsson, M. [University of California Irvine, 916 Engineering Tower, UC Irvine, Irvine, CA 92697-2575 (United States); Zalupski, P. [Idaho National Laboratory, 2525 Fremont Avenue, Idaho Falls, ID 83415 (United States)

    2013-07-01

    A thorough understanding of the non-ideal behavior of the chemical components utilized in solvent extraction contributes to the success of any large-scale spent nuclear fuel treatment. To address this, our current work uses vapor pressure osmometry to characterize the non-ideal behavior of the solvent extraction agent di-(2-ethylhexyl) phosphoric acid (HDEHP), a common extractant in proposed separation schemes. Solubility parameters were fit to data on HDEHP at four temperatures using models based on Scatchard Hildebrand regular solution theory with Flory Huggins entropic corrections. The results are comparable but not identical to the activity coefficients from prior slope analysis in the literature. (authors)

  17. RecFOR is not required for pneumococcal transformation but together with XerS for resolution of chromosome dimers frequently formed in the process.

    Science.gov (United States)

    Johnston, Calum; Mortier-Barrière, Isabelle; Granadel, Chantal; Polard, Patrice; Martin, Bernard; Claverys, Jean-Pierre

    2015-01-01

    Homologous recombination (HR) is required for both genome maintenance and generation of diversity in eukaryotes and prokaryotes. This process initiates from single-stranded (ss) DNA and is driven by a universal recombinase, which promotes strand exchange between homologous sequences. The bacterial recombinase, RecA, is loaded onto ssDNA by recombinase loaders, RecBCD and RecFOR for genome maintenance. DprA was recently proposed as a third loader dedicated to genetic transformation. Here we assessed the role of RecFOR in transformation of the human pathogen Streptococcus pneumoniae. We firstly established that RecFOR proteins are not required for plasmid transformation, strongly suggesting that DprA ensures annealing of plasmid single-strands internalized in the process. We then observed no reduction in chromosomal transformation using a PCR fragment as donor, contrasting with the 10,000-fold drop in dprA- cells and demonstrating that RecFOR play no role in transformation. However, a ∼1.45-fold drop in transformation was observed with total chromosomal DNA in recFOR mutants. To account for this limited deficit, we hypothesized that transformation with chromosomal DNA stimulated unexpectedly high frequency (>30% of cells) formation of chromosome dimers as an intermediate in the generation of tandem duplications, and that RecFOR were crucial for dimer resolution. We validated this hypothesis, showing that the site-specific recombinase XerS was also crucial for dimer resolution. An even higher frequency of dimer formation (>80% of cells) was promoted by interspecies transformation with Streptococcus mitis chromosomal DNA, which contains numerous inversions compared to pneumococcal chromosome, each potentially promoting dimerization. In the absence of RecFOR and XerS, dimers persist, as confirmed by DAPI staining, and can limit the efficiency of transformation, since resulting in loss of transformant chromosome. These findings strengthen the view that different HR

  18. RecFOR is not required for pneumococcal transformation but together with XerS for resolution of chromosome dimers frequently formed in the process.

    Directory of Open Access Journals (Sweden)

    Calum Johnston

    2015-01-01

    Full Text Available Homologous recombination (HR is required for both genome maintenance and generation of diversity in eukaryotes and prokaryotes. This process initiates from single-stranded (ss DNA and is driven by a universal recombinase, which promotes strand exchange between homologous sequences. The bacterial recombinase, RecA, is loaded onto ssDNA by recombinase loaders, RecBCD and RecFOR for genome maintenance. DprA was recently proposed as a third loader dedicated to genetic transformation. Here we assessed the role of RecFOR in transformation of the human pathogen Streptococcus pneumoniae. We firstly established that RecFOR proteins are not required for plasmid transformation, strongly suggesting that DprA ensures annealing of plasmid single-strands internalized in the process. We then observed no reduction in chromosomal transformation using a PCR fragment as donor, contrasting with the 10,000-fold drop in dprA- cells and demonstrating that RecFOR play no role in transformation. However, a ∼1.45-fold drop in transformation was observed with total chromosomal DNA in recFOR mutants. To account for this limited deficit, we hypothesized that transformation with chromosomal DNA stimulated unexpectedly high frequency (>30% of cells formation of chromosome dimers as an intermediate in the generation of tandem duplications, and that RecFOR were crucial for dimer resolution. We validated this hypothesis, showing that the site-specific recombinase XerS was also crucial for dimer resolution. An even higher frequency of dimer formation (>80% of cells was promoted by interspecies transformation with Streptococcus mitis chromosomal DNA, which contains numerous inversions compared to pneumococcal chromosome, each potentially promoting dimerization. In the absence of RecFOR and XerS, dimers persist, as confirmed by DAPI staining, and can limit the efficiency of transformation, since resulting in loss of transformant chromosome. These findings strengthen the view that

  19. Structural insights into lipid-dependent reversible dimerization of human GLTP

    Energy Technology Data Exchange (ETDEWEB)

    Samygina, Valeria R.; Ochoa-Lizarralde, Borja [CIC bioGUNE, Technology Park of Bizkaia, 48160 Derio (Spain); Popov, Alexander N. [European Synchrotron Radiation Facility, 38043 Grenoble (France); Cabo-Bilbao, Aintzane; Goni-de-Cerio, Felipe [CIC bioGUNE, Technology Park of Bizkaia, 48160 Derio (Spain); Molotkovsky, Julian G. [Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, RAS, Moscow 117997 (Russian Federation); Patel, Dinshaw J. [Memorial Sloan–Kettering Cancer Center, New York, NY 10021 (United States); Brown, Rhoderick E., E-mail: reb@umn.edu [University of Minnesota, Austin, MN 55912 (United States); Malinina, Lucy, E-mail: reb@umn.edu [CIC bioGUNE, Technology Park of Bizkaia, 48160 Derio (Spain)

    2013-04-01

    It is shown that dimerization is promoted by glycolipid binding to human GLTP. The importance of dimer flexibility in wild-type protein is manifested by point mutation that ‘locks’ the dimer while diversifying ligand/protein adaptations. Human glycolipid transfer protein (hsGLTP) forms the prototypical GLTP fold and is characterized by a broad transfer selectivity for glycosphingolipids (GSLs). The GLTP mutation D48V near the ‘portal entrance’ of the glycolipid binding site has recently been shown to enhance selectivity for sulfatides (SFs) containing a long acyl chain. Here, nine novel crystal structures of hsGLTP and the SF-selective mutant complexed with short-acyl-chain monoSF and diSF in different crystal forms are reported in order to elucidate the potential functional roles of lipid-mediated homodimerization. In all crystal forms, the hsGLTP–SF complexes displayed homodimeric structures supported by similarly organized intermolecular interactions. The dimerization interface always involved the lipid sphingosine chain, the protein C-terminus (C-end) and α-helices 6 and 2, but the D48V mutant displayed a ‘locked’ dimer conformation compared with the hinge-like flexibility of wild-type dimers. Differences in contact angles, areas and residues at the dimer interfaces in the ‘flexible’ and ‘locked’ dimers revealed a potentially important role of the dimeric structure in the C-end conformation of hsGLTP and in the precise positioning of the key residue of the glycolipid recognition centre, His140. ΔY207 and ΔC-end deletion mutants, in which the C-end is shifted or truncated, showed an almost complete loss of transfer activity. The new structural insights suggest that ligand-dependent reversible dimerization plays a role in the function of human GLTP.

  20. Active compounds release from semisolid dosage forms.

    Science.gov (United States)

    Olejnik, Anna; Goscianska, Joanna; Nowak, Izabela

    2012-11-01

    The aim of this paper is to review all the aspects of the in vitro release testing (IVRT) from semisolid dosage forms. Although none of the official dissolution methods has been specified for use with semisolid dosage forms, their utility for assessing release rates of drugs from semisolid dosage forms has become a topic of considerable interest. One can expect to overcome such complexity in the future, when the official "Topical and Transdermal Drug Products-Product Performance Tests" will be published in an issue of the Pharmacopeial Forum. Many factors such as type of the dissolution medium, membrane, temperature, and speed have an influence on the mechanism and kinetics of the release testing from gels, creams, and ointments; therefore, those parameters have been widely discussed.

  1. Structural insights into the IgE mediated responses induced by the allergens Hev b 8 and Zea m 12 in their dimeric forms

    Science.gov (United States)

    Mares-Mejía, Israel; Martínez-Caballero, Siseth; Garay-Canales, Claudia; Cano-Sánchez, Patricia; Torres-Larios, Alfredo; Lara-González, Samuel; Ortega, Enrique; Rodríguez-Romero, Adela

    2016-01-01

    Oligomerization of allergens plays an important role in IgE-mediated reactions, as effective crosslinking of IgE- FcεRI complexes on the cell membrane is dependent on the number of exposed B-cell epitopes in a single allergen molecule or on the occurrence of identical epitopes in a symmetrical arrangement. Few studies have attempted to experimentally demonstrate the connection between allergen dimerization and the ability to trigger allergic reactions. Here we studied plant allergenic profilins rHev b 8 (rubber tree) and rZea m 12 (maize) because they represent an important example of cross-reactivity in the latex-pollen-food syndrome. Both allergens in their monomeric and dimeric states were isolated and characterized by exclusion chromatography and mass spectrometry and were used in immunological in vitro experiments. Their crystal structures were solved, and for Hev b 8 a disulfide-linked homodimer was found. Comparing the structures we established that the longest loop is relevant for recognition by IgE antibodies, whereas the conserved regions are important for cross-reactivity. We produced a novel monoclonal murine IgE (mAb 2F5), specific for rHev b 8, which was useful to provide evidence that profilin dimerization considerably increases the IgE-mediated degranulation in rat basophilic leukemia cells. PMID:27586352

  2. Inducible removal of UV-induced pyrimidine dimers from transcriptionally active and inactive genes of Saccharomyces cerevisiae.

    Science.gov (United States)

    Waters, R; Zhang, R; Jones, N J

    1993-05-01

    The prior UV irradiation of alpha haploid Saccharomyces cerevisiae with a UV dose of 25 J/m2 substantially increases the repairability of damage subsequently induced by a UV dose of 70 J/m2 given 1 h after the first irradiation. This enhancement of repair is seen at both the MAT alpha and HML alpha loci, which are, respectively, transcriptionally active and inactive in alpha haploid cells. The presence in the medium of the protein synthesis inhibitor, cycloheximide in the period between the two irradiations eliminated this effect. Enhanced repair still occurred if cycloheximide was present only after the final UV irradiation. This indicated that the first result is not due to cycloheximide merely blocking the synthesis of repair enzymes associated with a hypothetical rapid turnover of such molecules. The enhanced repairability is not the result of changes in chromatin accessibility without protein synthesis, merely caused by the repair of the damage induced by the prior irradiation. The data clearly show that a UV-inducible removal of pyrimidine dimers has occurred which involves the synthesis of new proteins. The genes known to possess inducible promoters, and which are involved in excision are RAD2, RAD7, RAD16 and RAD23. Studies with the rad7 and rad16 mutants which are defective in the ability to repair HML alpha and proficient in the repair of MAT alpha showed that in rad7, preirradiation enhanced the repair at MAT alpha, whereas in rad16 this increased repair of MAT alpha was absent. The preirradiation did not modify the inability to repair HML alpha in either strain. Thus RAD16 has a role in this inducible repair.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Novel covalently linked insulin dimer engineered to investigate the function of insulin dimerization

    DEFF Research Database (Denmark)

    Vinther, Tine N.; Norrman, Mathias; Strauss, Holger M.;

    2012-01-01

    An ingenious system evolved to facilitate insulin binding to the insulin receptor as a monomer and at the same time ensure sufficient stability of insulin during storage. Insulin dimer is the cornerstone of this system. Insulin dimer is relatively weak, which ensures dissociation into monomers...... in the circulation, and it is stabilized by hexamer formation in the presence of zinc ions during storage in the pancreatic ß-cell. Due to the transient nature of insulin dimer, direct investigation of this important form is inherently difficult. To address the relationship between insulin oligomerization...... and insulin stability and function, we engineered a covalently linked insulin dimer in which two monomers were linked by a disulfide bond. The structure of this covalent dimer was identical to the self-association dimer of human insulin. Importantly, this covalent dimer was capable of further oligomerization...

  4. Dimerization of norbornene on zeolite catalysts

    Institute of Scientific and Technical Information of China (English)

    N. G. Grigor’eva; S. V. Bubennov; L. M. Khalilov; B. I. Kutepov

    2015-01-01

    The high activity and selectivity of H‐Beta and H‐ZSM‐12 zeolites in the dimerization of norbornene was established. The norbornene conversion reached 100%in chlorinated paraffin and argon gas medium, with a selectivity of dimer formation of 88%–98%. Four stereo‐isomers of the bis‐2,2’‐norbornylidene structure were identified in the dimer fraction, with the (Z)‐anti‐bis‐2,2’‐norbornylidene prevailing over the others.

  5. Taxation and forms of organizing business activities

    Directory of Open Access Journals (Sweden)

    Đinđić Srđan

    2013-01-01

    Full Text Available This paper takes sample tax regimes and tendencies from the developed countries in the EU-15 and the USA, and uses them to analyse the influence of taxation on the choice of organizational form of profit-oriented entities in Serbia. In order to understand how the procedure of taxation affects the sphere of business decision-making it is necessary to focus on the tax status of business losses and valorization and the effects of the double taxation of dividends. The rule of successive deduction of losses ensures the fiscally transparent entity receives a tax saving in the form of a reduction of the present value of the total paid tax. Meanwhile the corporation is handicapped because it postpones loss deductions, that is, it postpones tax saving, which directly influences the level of the present value of saved tax. The global trend of gradually moving from the classical system towards shareholder relief provision, above all in the form of a reduced withholding tax rate on dividends, has two opposing features: it simplifies the tax procedure while neglecting the distributional aims (consequences of taxation. The analysis of a particular practical example from the Serbian tax context enables us to draw a conclusion in relation to the relative taxes paid by entrepreneurs versus enterprises. The developed countries favour fiscally transparent entities, whereas Serbia allocates tax privileges to enterprises.

  6. A redox-dependent dimerization switch regulates activity and tolerance for reactive oxygen species of barley seed glutathione peroxidase

    DEFF Research Database (Denmark)

    Navrot, Nicolas; Skjoldager, Nicklas; Bunkenborg, Jakob;

    2015-01-01

    -butyl hydroperoxide, but is more sensitive to inactivation by hydrogen peroxide. Treatment of the monomer with hydrogen peroxide results in dimer formation. This observed new behavior of a plant glutathione peroxidase suggests a mechanism involving a switch from a highly catalytically competent monomer to a less...

  7. The crystal structures of the calcium-bound con-G and con-T[K7gamma] dimeric peptides demonstrate a metal-dependent helix-forming motif.

    Science.gov (United States)

    Cnudde, Sara E; Prorok, Mary; Dai, Qiuyun; Castellino, Francis J; Geiger, James H

    2007-02-14

    Short peptides that have the ability to form stable alpha-helices in solution are rare, and a number of strategies have been used to produce them, including the use of metal chelation to stabilize folding of the backbone. However, no example exists of a structurally well-defined helix stabilized exclusively through metal ion chelation. Conantokins (con)-G and -T are short peptides that are potent antagonists of N-methyl-D-aspartate receptor channels. While con-G exhibits no helicity alone, it undergoes a structural transition to a helical conformation in the presence of a variety of multivalent cations, especially Mg2+ and Ca2+. This complexation also results in antiparallel dimerization of two peptide helices in the presence of Ca2+, but not Mg2+. A con-T variant, con-T[K7gamma], displays very similar behavior. We have solved the crystal structures of both Ca2+/con-G and Ca2+/con-T [K7gamma] at atomic resolution. These structures clearly show the nature of the metal-dependent dimerization and helix formation and surprisingly also show that the con-G dimer interface is completely different from the con-T[K7gamma] interface, even though the metal chelation is similar in the two peptides. This represents a new paradigm in helix stabilization completely independent of the hydrophobic effect, which we define as the "metallo-zipper."

  8. 76 FR 41279 - Agency Information Collection Activities; Form I-864, Form I-864A, Form I-864EZ, and Form I-864W...

    Science.gov (United States)

    2011-07-13

    ... SECURITY U.S. Citizenship and Immigration Services Agency Information Collection Activities; Form I-864, Form I- 864A, Form I-864EZ, and Form I-864W; Extension of an Existing Information Collection; Comment Request ACTION: 30-Day Notice of Information Collection Under Review: Form I- 864, Affidavit of...

  9. 75 FR 51093 - Agency Information Collection Activities: Form I-864, Form I-864A, Form I-864EZ, and Form I-864W...

    Science.gov (United States)

    2010-08-18

    ... SECURITY U.S. Citizenship and Immigration Services Agency Information Collection Activities: Form I-864, Form I- 864A, Form I-864EZ, and Form I-864W; Extension of a Currently Approved Information Collection; Comment Request ACTION: 30-Day Notice of Information Collection Under Review: Form I- 864, Affidavit...

  10. Monitoring of receptor dimerization using plasmonic coupling of gold nanoparticles.

    Science.gov (United States)

    Crow, Matthew J; Seekell, Kevin; Ostrander, Julie H; Wax, Adam

    2011-11-22

    The dimerization of receptors on the cell membrane is an important step in the activation of cell signaling pathways. Several methods exist for observing receptor dimerization, including coimmunoprecipitation, chemical cross-linking, and fluorescence resonance energy transfer (FRET). These techniques are limited in that only FRET is appropriate for live cells, but even that method suffers from photobleaching and bleed-through effects. In this study, we implement an alternative method for the targeting of HER-2 homodimer formation based on the plasmonic coupling of gold nanoparticles functionalized with HER-2 Ab. In the presented studies, SK-BR-3 cells, known to overexpress HER-2, are labeled with these nanoparticles and receptor colocalization is observed using plasmonic coupling. HER-2 targeted nanoparticles bound to these cells exhibit a peak resonance that is significantly red-shifted relative to those bound to similar receptors on A549 cells, which have significantly lower levels of HER-2 expression. This significant red shift indicates plasmonic coupling is occurring and points to a new avenue for assessing dimerization by monitoring their colocalization. To determine that dimerization is occurring, the refractive index of the nanoenvironment of the labels is assessed using a theoretical analysis based on the Mie coated sphere model. The results indicate scattering by single, isolated nanoparticles for the low HER-2 expressing A549 cell line, but the scattering observed for the HER-2 overexpressing SK-BR-3 cell line may only be explained by plasmonic-coupling of proximal nanoparticle pairs. To validate the conformation of nanoparticles bound to HER-2 receptors undergoing dimerization, discrete dipole approximation (DDA) models are used to assess spectra of scattering by coupled nanoparticles. Comparison of the experimental results with theoretical models indicates that NP dimers are formed for the labeling of SK-BR-3 cells, suggesting that receptor

  11. Atomic resolution crystal structure of VcLMWPTP-1 from Vibrio cholerae O395: Insights into a novel mode of dimerization in the low molecular weight protein tyrosine phosphatase family

    Energy Technology Data Exchange (ETDEWEB)

    Nath, Seema; Banerjee, Ramanuj; Sen, Udayaditya, E-mail: udayaditya.sen@saha.ac.in

    2014-07-18

    Highlights: • VcLMWPTP-1 forms dimer in solution. • The dimer is catalytically active unlike other reported dimeric LMWPTPs. • The formation of extended dimeric surface excludes the active site pocket. • The surface bears closer resemblance to eukaryotic LMWPTPs. - Abstract: Low molecular weight protein tyrosine phosphatase (LMWPTP) is a group of phosphotyrosine phosphatase ubiquitously found in a wide range of organisms ranging from bacteria to mammals. Dimerization in the LMWPTP family has been reported earlier which follows a common mechanism involving active site residues leading to an enzymatically inactive species. Here we report a novel form of dimerization in a LMWPTP from Vibrio cholera 0395 (VcLMWPTP-1). Studies in solution reveal the existence of the dimer in solution while kinetic study depicts the active form of the enzyme. This indicates that the mode of dimerization in VcLMWPTP-1 is different from others where active site residues are not involved in the process. A high resolution (1.45 Å) crystal structure of VcLMWPTP-1 confirms a different mode of dimerization where the active site is catalytically accessible as evident by a tightly bound substrate mimicking ligand, MOPS at the active site pocket. Although being a member of a prokaryotic protein family, VcLMWPTP-1 structure resembles very closely to LMWPTP from a eukaryote, Entamoeba histolytica. It also delineates the diverse surface properties around the active site of the enzyme.

  12. The pore-forming toxin proaerolysin is activated by furin.

    Science.gov (United States)

    Abrami, L; Fivaz, M; Decroly, E; Seidah, N G; Jean, F; Thomas, G; Leppla, S H; Buckley, J T; van der Goot, F G

    1998-12-04

    Aerolysin is secreted as an inactive dimeric precursor by the bacterium Aeromonas hydrophila. Proteolytic cleavage within a mobile loop near the C terminus of the protoxin is required for oligomerization and channel formation. This loop contains the sequence KVRRAR432, which should be recognized by mammalian proprotein convertases such as furin, PACE4, and PC5/6A. Here we show that these three proteases cleave proaerolysin after Arg-432 in vitro, yielding active toxin. We also investigated the potential role of these enzymes in the in vivo activation of the protoxin. We found that Chinese hamster ovary cells were able to convert the protoxin to aerolysin in the absence of exogenous proteases and that activation did not require internalization of the toxin. The furin inhibitor alpha1-antitrypsin Portland reduced the rate of proaerolysin activation in vivo, and proaerolysin processing was even further reduced in furin-deficient FD11 Chinese hamster ovary cells. The cells were also less sensitive to proaerolysin than wild type cells; however, transient transfection of FD11 cells with the cDNA encoding furin conferred normal sensitivity to the protoxin. Together these findings argue that furin catalyzes the cell-surface activation of proaerolysin in vivo.

  13. Dimerization of the glucan phosphatase laforin requires the participation of cysteine 329.

    Directory of Open Access Journals (Sweden)

    Pablo Sánchez-Martín

    Full Text Available Laforin, encoded by a gene that is mutated in Lafora Disease (LD, OMIM 254780, is a modular protein composed of a carbohydrate-binding module and a dual-specificity phosphatase domain. Laforin is the founding member of the glucan-phosphatase family and regulates the levels of phosphate present in glycogen. Multiple reports have described the capability of laforin to form dimers, although the function of these dimers and their relationship with LD remains unclear. Recent evidence suggests that laforin dimerization depends on redox conditions, suggesting that disulfide bonds are involved in laforin dimerization. Using site-directed mutagenesis we constructed laforin mutants in which individual cysteine residues were replaced by serine and then tested the ability of each protein to dimerize using recombinant protein as well as a mammalian cell culture assay. Laforin-Cys329Ser was the only Cys/Ser mutant unable to form dimers in both assays. We also generated a laforin truncation lacking the last three amino acids, laforin-Cys329X, and this truncation also failed to dimerize. Interestingly, laforin-Cys329Ser and laforin-Cys329X were able to bind glucans, and maintained wild type phosphatase activity against both exogenous and biologically relevant substrates. Furthermore, laforin-Cys329Ser was fully capable of participating in the ubiquitination process driven by a laforin-malin complex. These results suggest that dimerization is not required for laforin phosphatase activity, glucan binding, or for the formation of a functional laforin-malin complex. Cumulatively, these results suggest that cysteine 329 is specifically involved in the dimerization process of laforin. Therefore, the C329S mutant constitutes a valuable tool to analyze the physiological implications of laforin's oligomerization.

  14. A Nove Asymmetric ent—Kauranoid Dimer from Isodon enanderianus

    Institute of Scientific and Technical Information of China (English)

    纳智; 黎胜红; 等

    2002-01-01

    Further investigation on the aerial parts of Isodon enanderianus afforded a novel asymmetric ent-kauranoid dimer,enanderi-nanin J(1).The structure of the dimer was elucidated by means of spectroscopic methods (including 2D NMR tecniques ),Enanderinanin J was a dimer of xerophilusin A and probably formed by [4+2] cycloaddition.

  15. A Novel Asymmetric ent-Kauranoid Dimer from Isodon enanderianus

    Institute of Scientific and Technical Information of China (English)

    NA,Zhi(纳智); LI,Sheng-Hong(黎胜红); XIANG,Wei(项伟); ZHAO,Ai-Hua(赵爱华); LI,Chao-Ming(李朝明); SUN,Han-Dong(孙汉董)

    2002-01-01

    Further investigation on the aerial parts of Isodon enanderianus afforded a novel asymmetric ent-kauranoid dimer, enanuderinaninJ (1). The structure of the dimer was elucidated by means of spectroscopic methods (including 2D NMR techniques ). Enanderinanin J was a dimer of xerophilusin A and probably formed by [ 4 + 2] cycloaddition.

  16. Role of Rydberg states in the photostability of heterocyclic dimers: the case of pyrazole dimer.

    Science.gov (United States)

    Zilberg, Shmuel; Haas, Yehuda

    2012-11-26

    A new route for the nonradiative decay of photoexcited, H-bonded, nitrogen-containing, heterocyclic dimers is offered and exemplified by a study of the pyrazole dimer. In some of these systems the N(3s) Rydberg state is the lowest excited singlet state. This state is formed by direct light absorption or by nonradiative transition from the allowed ππ* state. An isomer of this Rydberg state is formed by H atom transfer to the other component of the dimer. The newly formed H-bonded radical pair is composed of two radicals (a H-adduct of pyrazole, a heterocyclic analogue of the NH(4) radical) and the pyrazolium π-radical. It is calculated to have a shallow local minimum and is the lowest point on the PES of the H-pyrazole/pyrazolium radical pair. This species can cross back to the ground state of the original dimer through a relatively small energy gap and compete with the H-atom loss channel, known for the monomer. In both Rydberg dimers, an electron occupies a Rydberg orbital centered mostly on one of the two components of the dimer. This Rydberg Center Shift (RCS) mechanism, proposed earlier (Zilberg, S.; Kahan, A.; Haas, Y. Phys. Chem. Chem. Phys. 2012, 14, 8836), leads to deactivation of the electronically excited dimer while keeping it intact. It, thus, may explain the high photostability of the pyrazole dimer as well as other heterocyclic dimers.

  17. Dimerization interface of 3-hydroxyacyl-CoA dehydrogenase tunes the formation of its catalytic intermediate.

    Directory of Open Access Journals (Sweden)

    Yingzhi Xu

    Full Text Available 3-Hydroxyacyl-CoA dehydrogenase (HAD, EC 1.1.1.35 is a homodimeric enzyme localized in the mitochondrial matrix, which catalyzes the third step in fatty acid β-oxidation. The crystal structures of human HAD and subsequent complexes with cofactor/substrate enabled better understanding of HAD catalytic mechanism. However, numerous human diseases were found related to mutations at HAD dimerization interface that is away from the catalytic pocket. The role of HAD dimerization in its catalytic activity needs to be elucidated. Here, we solved the crystal structure of Caenorhabditis elegans HAD (cHAD that is highly conserved to human HAD. Even though the cHAD mutants (R204A, Y209A and R204A/Y209A with attenuated interactions on the dimerization interface still maintain a dimerization form, their enzymatic activities significantly decrease compared to that of the wild type. Such reduced activities are in consistency with the reduced ratios of the catalytic intermediate formation. Further molecular dynamics simulations results reveal that the alteration of the dimerization interface will increase the fluctuation of a distal region (a.a. 60-80 that plays an important role in the substrate binding. The increased fluctuation decreases the stability of the catalytic intermediate formation, and therefore the enzymatic activity is attenuated. Our study reveals the molecular mechanism about the essential role of the HAD dimerization interface in its catalytic activity via allosteric effects.

  18. Formation and dimerization of the phosphodiesterase active site of the Pseudomonas aeruginosa MorA, a bi-functional c-di-GMP regulator.

    Science.gov (United States)

    Phippen, Curtis William; Mikolajek, Halina; Schlaefli, Henry George; Keevil, Charles William; Webb, Jeremy Stephen; Tews, Ivo

    2014-12-20

    Diguanylate cyclases (DGC) and phosphodiesterases (PDE), respectively synthesise and hydrolyse the secondary messenger cyclic dimeric GMP (c-di-GMP), and both activities are often found in a single protein. Intracellular c-di-GMP levels in turn regulate bacterial motility, virulence and biofilm formation. We report the first structure of a tandem DGC-PDE fragment, in which the catalytic domains are shown to be active. Two phosphodiesterase states are distinguished by active site formation. The structures, in the presence or absence of c-di-GMP, suggest that dimerisation and binding pocket formation are linked, with dimerisation being required for catalytic activity. An understanding of PDE activation is important, as biofilm dispersal via c-di-GMP hydrolysis has therapeutic effects on chronic infections.

  19. Trans-dimerization of JAM-A regulates Rap2 and is mediated by a domain that is distinct from the cis-dimerization interface.

    Science.gov (United States)

    Monteiro, Ana C; Luissint, Anny-Claude; Sumagin, Ronen; Lai, Caroline; Vielmuth, Franziska; Wolf, Mattie F; Laur, Oskar; Reiss, Kerstin; Spindler, Volker; Stehle, Thilo; Dermody, Terence S; Nusrat, Asma; Parkos, Charles A

    2014-05-01

    Junctional adhesion molecule-A (JAM-A) is a tight junction-associated signaling protein that regulates epithelial cell proliferation, migration, and barrier function. JAM-A dimerization on a common cell surface (in cis) has been shown to regulate cell migration, and evidence suggests that JAM-A may form homodimers between cells (in trans). Indeed, transfection experiments revealed accumulation of JAM-A at sites between transfected cells, which was lost in cells expressing cis- or predicted trans-dimerization null mutants. Of importance, microspheres coated with JAM-A containing alanine substitutions to residues 43NNP45 (NNP-JAM-A) within the predicted trans-dimerization site did not aggregate. In contrast, beads coated with cis-null JAM-A demonstrated enhanced clustering similar to that observed with wild-type (WT) JAM-A. In addition, atomic force microscopy revealed decreased association forces in NNP-JAM-A compared with WT and cis-null JAM-A. Assessment of effects of JAM-A dimerization on cell signaling revealed that expression of trans- but not cis-null JAM-A mutants decreased Rap2 activity. Furthermore, confluent cells, which enable trans-dimerization, had enhanced Rap2 activity. Taken together, these results suggest that trans-dimerization of JAM-A occurs at a unique site and with different affinity compared with dimerization in cis. Trans-dimerization of JAM-A may thus act as a barrier-inducing molecular switch that is activated when cells become confluent.

  20. Activity of the HMGB1-Derived Immunostimulatory Peptide Hp91 Resides in the Helical C-terminal Portion and is Enhanced by Dimerization

    Science.gov (United States)

    Saenz, R.; Messmer, B.; Futalan, D.; Tor, Y.; Larsson, M.; Daniels, G.; Esener, S.; Messmer, D.

    2013-01-01

    We have previously shown that an 18 amino acid long peptide, named Hp91, whose sequence corresponds to a region within the endogenous protein HMGB1, activates dendritic cells (DCs) and acts as adjuvant in vivo by potentiating Th1-type antigen-specific immune responses. We analyzed the structure-function relationship of the Hp91 peptide to investigate the amino acids and structure responsible for immune responses. We found that the cysteine at position 16 of Hp91 enabled formation of reversible peptide dimmers, monomer and dimmer were compared for DC binding and activation. Stable monomers and dimers were generated using a maleimide conjugation reaction. The dimer showed enhanced ability to bind to and activate DCs. Furthermore, the C-terminal 9 amino acids of Hp91, named UC1018 were sufficient for DC binding and Circular dichroism showed that UC1018 assumes an alpha-helical structure. The ninemer peptide UC1018 induced more potent antigen-specific CTL responses in vivo as compared to Hp91 and it protected mice from tumor development when used in a prophylactic vaccine setting. We have identified a short alpha helical peptide that acts as potent adjuvant inducing protective immune responses in vivo. PMID:24172222

  1. SGN-CD33A: a novel CD33-targeting antibody-drug conjugate using a pyrrolobenzodiazepine dimer is active in models of drug-resistant AML.

    Science.gov (United States)

    Kung Sutherland, May S; Walter, Roland B; Jeffrey, Scott C; Burke, Patrick J; Yu, Changpu; Kostner, Heather; Stone, Ivan; Ryan, Maureen C; Sussman, Django; Lyon, Robert P; Zeng, Weiping; Harrington, Kimberly H; Klussman, Kerry; Westendorf, Lori; Meyer, David; Bernstein, Irwin D; Senter, Peter D; Benjamin, Dennis R; Drachman, Jonathan G; McEarchern, Julie A

    2013-08-22

    Outcomes in acute myeloid leukemia (AML) remain unsatisfactory, and novel treatments are urgently needed. One strategy explores antibodies and their drug conjugates, particularly those targeting CD33. Emerging data with gemtuzumab ozogamicin (GO) demonstrate target validity and activity in some patients with AML, but efficacy is limited by heterogeneous drug conjugation, linker instability, and a high incidence of multidrug resistance. We describe here the development of SGN-CD33A, a humanized anti-CD33 antibody with engineered cysteines conjugated to a highly potent, synthetic DNA cross-linking pyrrolobenzodiazepine dimer via a protease-cleavable linker. The use of engineered cysteine residues at the sites of drug linker attachment results in a drug loading of approximately 2 pyrrolobenzodiazepine dimers per antibody. In preclinical testing, SGN-CD33A is more potent than GO against a panel of AML cell lines and primary AML cells in vitro and in xenotransplantation studies in mice. Unlike GO, antileukemic activity is observed with SGN-CD33A in AML models with the multidrug-resistant phenotype. Mechanistic studies indicate that the cytotoxic effects of SGN-CD33A involve DNA damage with ensuing cell cycle arrest and apoptotic cell death. Together, these data suggest that SGN-CD33A has CD33-directed antitumor activity and support clinical testing of this novel therapeutic in patients with AML.

  2. NMR detection of intermolecular interaction sites in the dimeric 5'-leader of the HIV-1 genome.

    Science.gov (United States)

    Keane, Sarah C; Van, Verna; Frank, Heather M; Sciandra, Carly A; McCowin, Sayo; Santos, Justin; Heng, Xiao; Summers, Michael F

    2016-11-15

    HIV type-1 (HIV-1) contains a pseudodiploid RNA genome that is selected for packaging and maintained in virions as a noncovalently linked dimer. Genome dimerization is mediated by conserved elements within the 5'-leader of the RNA, including a palindromic dimer initiation signal (DIS) that has been proposed to form kissing hairpin and/or extended duplex intermolecular contacts. Here, we have applied a (2)H-edited NMR approach to directly probe for intermolecular interactions in the full-length, dimeric HIV-1 5'-leader (688 nucleotides; 230 kDa). The interface is extensive and includes DIS:DIS base pairing in an extended duplex state as well as intermolecular pairing between elements of the upstream Unique-5' (U5) sequence and those near the gag start site (AUG). Other pseudopalindromic regions of the leader, including the transcription activation (TAR), polyadenylation (PolyA), and primer binding (PBS) elements, do not participate in intermolecular base pairing. Using a (2)H-edited one-dimensional NMR approach, we also show that the extended interface structure forms on a time scale similar to that of overall RNA dimerization. Our studies indicate that a kissing dimer-mediated structure, if formed, exists only transiently and readily converts to the extended interface structure, even in the absence of the HIV-1 nucleocapsid protein or other RNA chaperones.

  3. HLA-G Dimers in the Prolongation of Kidney Allograft Survival

    Directory of Open Access Journals (Sweden)

    Maureen Ezeakile

    2014-01-01

    Full Text Available Human leukocyte antigen-G (HLA-G contributes to acceptance of allografts in solid organ/tissue transplantation. Most studies have determined that soluble HLA-G isoforms are systematically detected in serum/plasma of transplanted patients with significantly fewer episodes of acute and/or chronic rejection of allogeneic tissue/organ. Current models of the interactions of HLA-G and its specific receptors explain it as functioning in a monomeric form. However, in recent years, new data has revealed the ability of HLA-G to form disulfide-linked dimeric complexes with high preferential binding and functional activities. Limited data are available on the role of soluble HLA-G dimers in clinical pathological conditions. We describe here the presence of soluble HLA-G dimers in kidney transplant patients. Our study showed that a high level of HLA-G dimers in plasma and increased expression of the membrane-bound form of HLA-G on monocytes are associated with prolongation of kidney allograft survival. We also determined that the presence of soluble HLA-G dimers links to the lower levels of proinflammatory cytokines, suggesting a potential role of HLA-G dimers in controlling the accompanying inflammatory state.

  4. Monomeric 14-3-3ζ Has a Chaperone-Like Activity and Is Stabilized by Phosphorylated HspB6

    OpenAIRE

    Sluchanko, Nikolai N.; Artemova, Natalya V.; Sudnitsyna, Maria V.; Safenkova, Irina V.; Antson, Alfred A.; Levitsky, Dmitrii I.; Gusev, Nikolai B.

    2012-01-01

    Members of the 14-3-3 eukaryotic protein family predominantly function as dimers. The dimeric form can be converted into monomers upon phosphorylation of Ser58 located at the subunit interface. Monomers are less stable than dimers and have been considered to be either less active or even inactive during binding and regulation of phosphorylated client proteins. However, like dimers, monomers contain the phosphoserine-binding site and therefore can retain some functions of the dimeric 14-3-3. F...

  5. Calix[4]arene supported clusters: a dimer of [Mn(III)Mn(II)] dimers

    DEFF Research Database (Denmark)

    Taylor, Stephanie M; McIntosh, Ruaraidh D; Beavers, Christine M;

    2011-01-01

    Phosphinate ligands allow for the transformation of a calix[4]arene supported [Mn(III)(2)Mn(II)(2)] tetramer cluster motif into an unusual [Mn(III)Mn(II)](2) dimer of dimers; the clusters self-assemble in the crystal to form bi-layer arrays reminiscent of the typical packing of calixarene solvates....

  6. Water Aerobics as a Form of Health Activities

    Directory of Open Access Journals (Sweden)

    Anna S. Batrak

    2013-09-01

    Full Text Available The offered literature review considers water aerobics as a form of health activities. Water aerobics is wide spread and popular, especially among women, because it is also the form of adaptive and health activities. It enlarges general physiological effect of physical exercises on the human body. Regular exercises improve physical fitness and physical development, health, mood, sleep, intensify activities and working efficiency.

  7. Design of dimerization inhibitors of HIV-1 aspartic proteinase: A computer-based combinatorial approach

    Science.gov (United States)

    Caflisch, Amedeo; Schramm, Hans J.; Karplus, Martin

    2000-02-01

    Inhibition of dimerization to the active form of the HIV-1 aspartic proteinase (HIV-1 PR) may be a way to decrease the probability of escape mutations for this viral protein. The Multiple Copy Simultaneous Search (MCSS) methodology was used to generate functionality maps for the dimerization interface of HIV-1 PR. The positions of the MCSS minima of 19 organic fragments, once postprocessed to take into account solvation effects, are in good agreement with experimental data on peptides that bind to the interface. The MCSS minima combined with an approach for computational combinatorial ligand design yielded a set of modified HIV-1 PR C-terminal peptides that are similar to known nanomolar inhibitors of HIV-1 PR dimerization. A number of N-substituted 2,5-diketopiperazines are predicted to be potential dimerization inhibitors of HIV-1 PR.

  8. Hetero-metallic trigonal cage-shaped dimeric Ni3K core complex of L-proline ligand: Synthesis, structural, electrochemical and DNA binding and cleavage activities

    Indian Academy of Sciences (India)

    S Nagasubramanian; A Jayamani; V Thamilarasan; G Aravindan; V Ganesan; N Sengottuvelan

    2014-05-01

    Hetero-metallic trigonal cage-shaped dimeric Ni3K core complex of L-proline ligand has been synthesized and characterized. Single crystal X-ray diffraction analysis showed that the hetero-metallic Ni(II)-K(I) complex has a dimeric structure with nine coordinated potassium atoms and six coordinated nickel atoms. The cyclic voltammograms of the complex exhibited two successive quasireversible reduction waves at ($E^{1}_{pc} = −1.02$ V and $E^{2}_{pc} = −1.33$ V) and two successive irreversible oxidation waves ($E^{1}_{pa} = 0.95$ V and $E^{2}_{pa} = 1.45$ V) versus Ag/AgCl in DMF solution. Interaction of the complex with calf-thymus DNA (CT DNA) has been studied using spectroscopic techniques. The complex is an avid DNA binder with a binding constant of 3.6 × 108 M-1. The complex showed efficient oxidative cleavage of supercoiled pBR322 DNA in the presence of the reducing agent hydrogen peroxide involving hydroxyl radical (°OH) species. As evidenced from the control experiment, DNA cleavage in the presence of °OH radical was inhibited by quenchers, viz. DMSO and KI. The complex showed in vitro antimicrobial activity against four bacteria and two fungi and the activity is greater than that of the free ligand.

  9. Novel covalently linked insulin dimer engineered to investigate the function of insulin dimerization.

    Directory of Open Access Journals (Sweden)

    Tine N Vinther

    Full Text Available An ingenious system evolved to facilitate insulin binding to the insulin receptor as a monomer and at the same time ensure sufficient stability of insulin during storage. Insulin dimer is the cornerstone of this system. Insulin dimer is relatively weak, which ensures dissociation into monomers in the circulation, and it is stabilized by hexamer formation in the presence of zinc ions during storage in the pancreatic β-cell. Due to the transient nature of insulin dimer, direct investigation of this important form is inherently difficult. To address the relationship between insulin oligomerization and insulin stability and function, we engineered a covalently linked insulin dimer in which two monomers were linked by a disulfide bond. The structure of this covalent dimer was identical to the self-association dimer of human insulin. Importantly, this covalent dimer was capable of further oligomerization to form the structural equivalent of the classical hexamer. The covalently linked dimer neither bound to the insulin receptor, nor induced a metabolic response in vitro. However, it was extremely thermodynamically stable and did not form amyloid fibrils when subjected to mechanical stress, underlining the importance of oligomerization for insulin stability.

  10. Micellisation and immunoreactivities of dimeric beta-caseins.

    Science.gov (United States)

    Yousefi, Reza; Gaudin, Jean-Charles; Chobert, Jean-Marc; Pourpak, Zahra; Moin, Mostafa; Moosavi-Movahedi, Ali Akbar; Haertle, Thomas

    2009-12-01

    Bovine beta-casein (beta-CN) is a highly amphiphilic micellising phospho-protein showing chaperone-like activity in vitro. Recently, existence of multiple sequential epitopes on beta-CN polypeptide chain in both hydrophilic-polar (psi) and hydrophobic-apolar domains (phi) has been evidenced. In order to clarify specific contribution of polar and apolar domains in micellisation process and in shaping immunoreactivity of beta-CN, its dimeric/bi-amphiphilic "quasi palindromic" forms covalently connected by a disulfide bond linking either N-terminal (C4 beta-CND) or C-terminal domain (C208 beta-CND) were produced and studied. Depending on the C- or N-terminal position of inserted cysteine, each dimeric beta-CN contains one polar/apolar region at the centre and two external hydrophobic/hydrophilic ends. Consequently, such casein dimers have radically different polarities/hydrophobicities on their outside surfaces. Dynamic light scattering (DLS) measurements indicate that these dimeric casein molecules form micelles of different sizes depending on arrangement of polar fragments of the beta-CN mutants in their constrained dimers. Non-aggregated dimers have different hydrodynamic diameters that could be explained by their different geometries. Measurements of fluorescence showed more hydrophobic environment of Trp residues of C208 beta-CND, while in similar experimental conditions Trp residues of C4 beta-CND and native beta-CN were more exposed to the polar medium. Both fluorescence and DLS studies showed greater propensity for micellisation of the dimeric beta-CNs, suggesting that the factors inducing the formation of micelles are stronger in the bi-amphiphilic dimers. 1-anilino-naphthalene-8-sulfonate (ANS) binding studies showed different binding of ANS by these dimers as well as different exposition of ANS binding (hydrophobic) regions in the micellar states. The differences in fluorescence resonance energy transfer (FRET) profiles of C4 beta-CND and C208 beta-CND can

  11. Molecular Interplay between the Dimer Interface and the Substrate-Binding Site of Human Peptidylarginine Deiminase 4

    Science.gov (United States)

    Lee, Chien-Yun; Lin, Chu-Cheng; Liu, Yi-Liang; Liu, Guang-Yaw; Liu, Jyung-Hurng; Hung, Hui-Chih

    2017-01-01

    Our previous studies suggest that the fully active form of Peptidylarginine deiminase 4 (PAD4) should be a dimer and not a monomer. This paper provides a plausible mechanism for the control of PAD4 catalysis by molecular interplay between its dimer-interface loop (I-loop) and its substrate-binding loop (S-loop). Mutagenesis studies revealed that two hydrophobic residues, W347 and V469, are critical for substrate binding at the active site; mutating these two residues led to a severe reduction in the catalytic activity. We also identified several hydrophobic amino acid residues (L6, L279 and V283) at the dimer interface. Ultracentrifugation analysis revealed that interruption of the hydrophobicity of this region decreases dimer formation and, consequently, enzyme activity. Molecular dynamic simulations and mutagenesis studies suggested that the dimer interface and the substrate-binding site of PAD4, which consist of the I-loop and the S-loop, respectively, are responsible for substrate binding and dimer stabilization. We identified five residues with crucial roles in PAD4 catalysis and dimerization: Y435 and R441 in the I-loop, D465 and V469 in the S-loop, and W548, which stabilizes the I-loop via van der Waals interactions with C434 and Y435. The molecular interplay between the S-loop and the I-loop is crucial for PAD4 catalysis. PMID:28209966

  12. Construction of a ferritin dimer by breaking its symmetry

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, B; Uenuma, M; Uraoka, Y; Yamashita, I [Graduate School of Materials Science, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0192 (Japan); CREST, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan)

    2010-11-05

    Ferritin has a mono-dispersed structure and biomineralization properties that allow it to form various kinds of nanoparticles and play an important role in modern nanotechnology. Independent nanoparticles synthesized in ferritin are valuable, but moreover a pair of nanoparticles can bring new properties different from those of the independent nanoparticles. In this study, by breaking ferritin's symmetry, we successfully produced ferritin dimers which provide real protein frameworks for nanoparticle dimer formation. Identical nickel hydro-oxide nanoparticle dimers were produced by simply biomineralizing ferritin dimers. The method presented here can produce multi-functional ferritin dimers with different kinds of nanoparticles.

  13. Kinetics of DNA tile dimerization.

    Science.gov (United States)

    Jiang, Shuoxing; Yan, Hao; Liu, Yan

    2014-06-24

    Investigating how individual molecular components interact with one another within DNA nanoarchitectures, both in terms of their spatial and temporal interactions, is fundamentally important for a better understanding of their physical behaviors. This will provide researchers with valuable insight for designing more complex higher-order structures that can be assembled more efficiently. In this report, we examined several spatial factors that affect the kinetics of bivalent, double-helical (DH) tile dimerization, including the orientation and number of sticky ends (SEs), the flexibility of the double helical domains, and the size of the tiles. The rate constants we obtained confirm our hypothesis that increased nucleation opportunities and well-aligned SEs accelerate tile-tile dimerization. Increased flexibility in the tiles causes slower dimerization rates, an effect that can be reversed by introducing restrictions to the tile flexibility. The higher dimerization rates of more rigid tiles results from the opposing effects of higher activation energies and higher pre-exponential factors from the Arrhenius equation, where the pre-exponential factor dominates. We believe that the results presented here will assist in improved implementation of DNA tile based algorithmic self-assembly, DNA based molecular robotics, and other specific nucleic acid systems, and will provide guidance to design and assembly processes to improve overall yield and efficiency.

  14. Effect of the Principle of Activating Blood Circulation to Break Stasis on GMP-140 and D2 Dimer in Patients with Acute Cerebral Infarction

    Institute of Scientific and Technical Information of China (English)

    WANG Ning

    2005-01-01

    Objective:To explore the clinical efficacy of the principle of activating blood circulation to break stasis (ABCBS) and its influence on platelet membranous protein particle (GMP-140) and D2 dimer (Ddimer) before and after treatment. Methods: Eighty-eight patients with blood stasis syndrome (BSS) of acute cerebral infarction (ACI) were randomly divided into two groups, both of which were treated with conventional treatment, i.e. with western medicine (WM), with Salvia injection added through intravenously dripping.One of the two groups was used as the control and the other group as the treated group who had ABCBS herbs orally taken in addition. The duration of treatment course for both groups was 3 weeks. Results: There were changes in both groups over clinical symptoms, nerve function deficit scoring and GMP-140, D-dimer, but the treated group showed significantly better than that of the control group, (P<0.05). Conclusion: ABCBS principle could serve as an important auxiliary treating method for BSS of ACI, as it can effectively alter the blood of ACI patients which was viscous, condense, coagulant and aggregating.

  15. Immunoglobulin Free Light Chain Dimers in Human Diseases

    Directory of Open Access Journals (Sweden)

    Batia Kaplan

    2011-01-01

    Full Text Available Immunoglobulin free light chain (FLC kappa (κ and lambda (λ isotypes exist mainly in monomeric and dimeric forms. Under pathological conditions, the level of FLCs as well as the structure of monomeric and dimeric FLCs and their dimerization properties might be significantly altered. The abnormally high fractions of dimeric FLCs were demonstrated in the serum of patients with multiple myeloma (MM and primary systemic amyloidosis (AL, as well as in the serum of anephric patients. The presence of tetra- and trimolecular complexes formed due to dimer-dimer and dimer-monomer interactions was detected in the myeloma serum. Analysis of the amyloidogenic light chains demonstrated mutations within the dimer interface, thus raising the possibility that these mutations are responsible for amyloidogenicity. Increased κ monomer and dimer levels, as well as a high κ/λ monomer ratio, were typically found in the cerebrospinal fluid from patients with multiple sclerosis (MS. In many MS cases, the elevation of κ FLCs was accompanied by an abnormally high proportion of λ dimers. This review focuses on the disease-related changes of the structure and level of dimeric FLCs, and raises the questions regarding their formation, function, and role in the pathogenesis and diagnosis of human diseases.

  16. Large D-Dimer Fluctuation in Normal Pregnancy

    DEFF Research Database (Denmark)

    Hedengran, Katrine K; Andersen, Malene R; Stender, Steen

    2016-01-01

    pregnancies were recruited. D-dimer was repeatedly measured during pregnancy, at active labor, and on the first and second postpartum days. Percentiles for each gestational week were calculated. Each individual D-dimer was normalized by transformation into percentiles for the relevant gestational age......Introduction. D-dimer levels increase throughout pregnancy, hampering the usefulness of the conventional threshold for dismissing thromboembolism. This study investigates the biological fluctuation of D-dimer in normal pregnancy. Methods. A total of 801 healthy women with expected normal...... normal pregnancy, repeated D-dimer measurements are of no clinical use in the evaluation of thromboembolic events during pregnancy....

  17. ERα dimerization: a key factor for the weak estrogenic activity of an ERα modulator unable to compete with estradiol in binding assays.

    Science.gov (United States)

    Leclercq, Guy; Laïos, Ioanna; Elie-Caille, Céline; Leiber, Denis; Laurent, Guy; Lesniewska, Eric; Tanfin, Zahra; Jacquot, Yves

    2017-04-01

    Estrothiazine (ESTZ) is a weak estrogen sharing structural similarities with coumestrol. ESTZ failed to compete with [(3)H]17β-estradiol ([(3)H]17β-E2) for binding to the estrogen receptor α (ERα), questioning its ability to interact with the receptor. However, detection by atomic force spectroscopy (AFS) of an ESTZ-induced ERα dimerization has eliminated any remaining doubts. The effect of the compound on the proliferation of ERα-positive and negative breast cancer cells confirmed the requirement of the receptor. The efficiency of ESTZ in MCF-7 cells was weak without any potency to modify the proliferation profile of estradiol and coumestrol. Growth enhancement was associated with a proteasomal degradation of ERα without substantial recruitment of LxxLL coactivators. This may be related to an unusual delay between the acquisition by the receptor of an ERE-binding capacity and the subsequent estrogen-dependent transcription. A complementary ability to enhance TPA-induced AP-1 transcription was observed, even at concentrations insufficient to activate the ERα, suggesting a partly independent mechanism. ESTZ also rapidly and transiently activated ERK1/2 likely through membrane estrogenic pathways provoking a reorganization of the actin network. Finally, the systematic absence of biological responses with an ESTZ derivative unable to induce ERα dimerization stresses the importance of this step in the action of the compound, as reported for conventional estrogens. In view of the existence of many other ERα modulators (endocrine disruptors such as, for example, pesticides, environmental contaminants or phytoestrogens) with extremely weak or similar apparent lack of binding ability, our work may appear as pilot investigation for assessing their mechanism of action.

  18. Dimerization capacities of FGF2 purified with or without heparin-affinity chromatography.

    Directory of Open Access Journals (Sweden)

    Natalia Platonova

    Full Text Available Fibroblast growth factor-2 (FGF2 is a pleiotropic growth factor exhibiting a variety of biological activities. In this article, we studied the capacity of FGF2 purified with or without heparin affinity chromatography to self-associate. Analyzing the NMR HSQC spectra for different FGF2 concentrations, heparin-affinity purified FGF2 showed perturbations that indicate dimerization and are a higher-order oligomerization state. HSQC perturbation observed with different FGF2 concentrations revealed a heparin-binding site and two dimer interfaces. Thus, with increasing protein concentrations, FGF2 monomers make contacts with each other and form dimers or higher order oligomers. On the contrary, FGF2 purified with ion-exchange chromatography did not show similar perturbation indicating that self-association of FGF2 is eliminated if purification is done without heparin-affinity chromatography. The HSQC spectra of heparin-affinity purified FGF2 can be reproduced to some extent by adding heparin tetra-saccharide to ion exchange chromatography purified FGF2. Heparin-affinity purified FGF2 bound to acceptor and donor beads in a tagged form using His-tagged or GST-tagged proteins, also dimerized in the AlphaScreen™ assay. This assay was further validated using different experimental conditions and competitors. The assay constitutes an interesting tool to study dimerization of other FGF forms as well.

  19. Pore-forming activity of clostridial binary toxins.

    Science.gov (United States)

    Knapp, O; Benz, R; Popoff, M R

    2016-03-01

    Clostridial binary toxins (Clostridium perfringens Iota toxin, Clostridium difficile transferase, Clostridium spiroforme toxin, Clostridium botulinum C2 toxin) as Bacillus binary toxins, including Bacillus anthracis toxins consist of two independent proteins, one being the binding component which mediates the internalization into cell of the intracellularly active component. Clostridial binary toxins induce actin cytoskeleton disorganization through mono-ADP-ribosylation of globular actin and are responsible for enteric diseases. Clostridial and Bacillus binary toxins share structurally and functionally related binding components which recognize specific cell receptors, oligomerize, form pores in endocytic vesicle membrane, and mediate the transport of the enzymatic component into the cytosol. Binding components retain the global structure of pore-forming toxins (PFTs) from the cholesterol-dependent cytotoxin family such as perfringolysin. However, their pore-forming activity notably that of clostridial binding components is more related to that of heptameric PFT family including aerolysin and C. perfringens epsilon toxin. This review focuses upon pore-forming activity of clostridial binary toxins compared to other related PFTs. This article is part of a Special Issue entitled: Pore-Forming Toxins edited by Mauro Dalla Serra and Franco Gambale.

  20. Exploratory Topology Modelling of Form-Active Hybrid Structures

    DEFF Research Database (Denmark)

    Holden Deleuran, Anders; Pauly, Mark; Tamke, Martin;

    2016-01-01

    The development of novel form-active hybrid structures (FAHS) is impeded by a lack of modelling tools that allow for exploratory topology modelling of shaped assemblies. We present a flexible and real-time computational design modelling pipeline developed for the exploratory modelling of FAHS tha...

  1. Adsorption of dimeric surfactants in lamellar silicates

    Energy Technology Data Exchange (ETDEWEB)

    Balcerzak, Mateusz; Pietralik, Zuzanna [Department of Macromolecular Physics, Faculty of Physics, A. Mickiewicz University, Umultowska 85, 61-614 Poznań (Poland); Domka, Ludwik [Department of Metalorganic Chemistry, Faculty of Chemistry, A. Mickiewicz University, Grunwaldzka 6, 60-780 Poznań (Poland); Skrzypczak, Andrzej [Institute of Chemical Technology, Poznań University of Technology, Berdychowo 4, 60-965 Poznań (Poland); Kozak, Maciej, E-mail: mkozak@amu.edu.pl [Department of Macromolecular Physics, Faculty of Physics, A. Mickiewicz University, Umultowska 85, 61-614 Poznań (Poland)

    2015-12-01

    Highlights: • The intercalation of dimeric surfactants changed the morphology of MMT samples. • XRD indicated structures formed by surfactant molecules in interlayer space. • The four-step thermal decomposition of dimeric surfactant, confirms intercalation. - Abstract: The adsorption of different types of cationic surfactants in lamellar silicates changes their surface character from hydrophilic to hydrophobic. This study was undertaken to obtain lamellar silicates modified by a series of novel dimeric (gemini) surfactants of different length alkyl chains and to characterise these organophilised materials. Synthetic sodium montmorillonite SOMASIF® ME 100 (M) and enriched bentonite of natural origin (Nanoclay – hydrophilic bentonite®) were organophilised with dimeric (gemini) surfactants (1,1′-(1,4-butanediyl)bis(alkoxymethyl)imidazolium dichlorides). As a result of surfactant molecule adsorption in interlamellar space, the d-spacing (d{sub 001}) increased from 0.97 nm (for the anhydrous structure) to 2.04 nm. A Fourier transform infrared spectroscopy (FTIR) analysis of the modified systems reveals bands assigned to the stretching vibrations of the CH{sub 2} and CH{sub 3} groups and the scissoring vibrations of the NH group from the structure of the dimeric surfactants. Thermogravimetric (TG) and derivative thermogravimetric (DTG) studies imply a four-stage process of surfactant decomposition. Scanning electron microscopy (SEM) images provide information on the influence of dimeric surfactant intercalation into the silicate structures. Particles of the modified systems show a tendency toward the formation of irregularly shaped agglomerates.

  2. Structural and dynamic requirements for optimal activity of the essential bacterial enzyme dihydrodipicolinate synthase.

    Directory of Open Access Journals (Sweden)

    C F Reboul

    Full Text Available Dihydrodipicolinate synthase (DHDPS is an essential enzyme involved in the lysine biosynthesis pathway. DHDPS from E. coli is a homotetramer consisting of a 'dimer of dimers', with the catalytic residues found at the tight-dimer interface. Crystallographic and biophysical evidence suggest that the dimers associate to stabilise the active site configuration, and mutation of a central dimer-dimer interface residue destabilises the tetramer, thus increasing the flexibility and reducing catalytic efficiency and substrate specificity. This has led to the hypothesis that the tetramer evolved to optimise the dynamics within the tight-dimer. In order to gain insights into DHDPS flexibility and its relationship to quaternary structure and function, we performed comparative Molecular Dynamics simulation studies of native tetrameric and dimeric forms of DHDPS from E. coli and also the native dimeric form from methicillin-resistant Staphylococcus aureus (MRSA. These reveal a striking contrast between the dynamics of tetrameric and dimeric forms. Whereas the E. coli DHDPS tetramer is relatively rigid, both the E. coli and MRSA DHDPS dimers display high flexibility, resulting in monomer reorientation within the dimer and increased flexibility at the tight-dimer interface. The mutant E. coli DHDPS dimer exhibits disorder within its active site with deformation of critical catalytic residues and removal of key hydrogen bonds that render it inactive, whereas the similarly flexible MRSA DHDPS dimer maintains its catalytic geometry and is thus fully functional. Our data support the hypothesis that in both bacterial species optimal activity is achieved by fine tuning protein dynamics in different ways: E. coli DHDPS buttresses together two dimers, whereas MRSA dampens the motion using an extended tight-dimer interface.

  3. Strong mode coupling in InP quantum dot-based GaInP microdisk cavity dimers

    Science.gov (United States)

    Witzany, M.; Liu, T.-L.; Shim, J.-B.; Hargart, F.; Koroknay, E.; Schulz, W.-M.; Jetter, M.; Hu, E.; Wiersig, J.; Michler, P.

    2013-01-01

    We report on strong mode coupling in closely spaced GaInP microdisk dimer structures including InP quantum dots as the active medium. Using electron beam lithography and a combination of dry- and wet-etch processes, dimers with inter-disk separations down to d < 100 nm have been fabricated. Applying a photo-thermal heating scheme, we overcome the spectral mode detuning due to the size mismatch between the two disks forming the dimer. We observe signatures of mode coupling in the corresponding photoluminescence spectra with coupling energies of up to 0.66 MeV. With the aid of a numerical analysis, we specify the geometrical and physical factors of the microdisk dimer precisely, and reproduce its spectrum with good agreement.

  4. Cytochrome P450 as dimerization catalyst in diketopiperazine alkaloid biosynthesis.

    Science.gov (United States)

    Saruwatari, Takayoshi; Yagishita, Fumitoshi; Mino, Takashi; Noguchi, Hiroshi; Hotta, Kinya; Watanabe, Kenji

    2014-03-21

    As dimeric natural products frequently exhibit useful biological activities, identifying and understanding their mechanisms of dimerization is of great interest. One such compound is (−)-ditryptophenaline, isolated from Aspergillus flavus, which inhibits substance P receptor for potential analgesic and anti-inflammatory activity. Through targeted gene knockout in A. flavus and heterologous yeast gene expression, we determined for the first time the gene cluster and pathway for the biosynthesis of a dimeric diketopiperazine alkaloid. We also determined that a single cytochrome P450, DtpC, is responsible not only for pyrroloindole ring formation but also for concurrent dimerization of N-methylphenylalanyltryptophanyl diketopiperazine monomers into a homodimeric product. Furthermore, DtpC exhibits relaxed substrate specificity, allowing the formation of two new dimeric compounds from a non-native monomeric precursor, brevianamide F. A radical-mediated mechanism of dimerization is proposed.

  5. Novel disulfide bond-mediated dimerization of the CARD domain was revealed by the crystal structure of CARMA1 CARD.

    Directory of Open Access Journals (Sweden)

    Tae-ho Jang

    Full Text Available CARMA1, BCL10 and MALT1 form a large molecular complex known as the CARMA1 signalosome during lymphocyte activation. Lymphocyte activation via the CARMA1 signalosome is critical to immune response and linked to many immune diseases. Despite the important role of the CARMA1 signalosome during lymphocyte activation and proliferation, limited structural information is available. Here, we report the dimeric structure of CARMA1 CARD at a resolution of 3.2 Å. Interestingly, although CARMA1 CARD has a canonical six helical-bundles structural fold similar to other CARDs, CARMA1 CARD shows the first homo-dimeric structure of CARD formed by a disulfide bond and reveals a possible biologically important homo-dimerization mechanism.

  6. New structural forms of a mycobacterial adenylyl cyclase Rv1625c

    Directory of Open Access Journals (Sweden)

    Deivanayaga Barathy

    2014-09-01

    Full Text Available Rv1625c is one of 16 adenylyl cyclases encoded in the genome of Mycobacterium tuberculosis. In solution Rv1625c exists predominantly as a monomer, with a small amount of dimer. It has been shown previously that the monomer is active and the dimeric fraction is inactive. Both fractions of wild-type Rv1625c crystallized as head-to-head inactive domain-swapped dimers as opposed to the head-to-tail dimer seen in other functional adenylyl cyclases. About half of the molecule is involved in extensive domain swapping. The strain created by a serine residue located on a hinge loop and the crystallization condition might have led to this unusual domain swapping. The inactivity of the dimeric form of Rv1625c could be explained by the absence of the required catalytic site in the swapped dimer. A single mutant of the enzyme was also generated by changing a phenylalanine predicted to occur at the functional dimer interface to an arginine. This single mutant exists as a dimer in solution but crystallized as a monomer. Analysis of the structure showed that a salt bridge formed between a glutamate residue in the N-terminal segment and the mutated arginine residue hinders dimer formation by pulling the N-terminal region towards the dimer interface. Both structures reported here show a change in the dimerization-arm region which is involved in formation of the functional dimer. It is concluded that the dimerization arm along with other structural elements such as the N-terminal region and certain loops are vital for determining the oligomeric nature of the enzyme, which in turn dictates its activity.

  7. Structure of the dimerization domain of DiGeorge critical region 8

    Energy Technology Data Exchange (ETDEWEB)

    Senturia, R.; Faller, M.; Yin, S.; Loo, J.A.; Cascio, D.; Sawaya, M.R.; Hwang, D.; Clubb, R.T.; Guo, F. (UCLA)

    2010-09-27

    Maturation of microRNAs (miRNAs, {approx}22nt) from long primary transcripts [primary miRNAs (pri-miRNAs)] is regulated during development and is altered in diseases such as cancer. The first processing step is a cleavage mediated by the Microprocessor complex containing the Drosha nuclease and the RNA-binding protein DiGeorge critical region 8 (DGCR8). We previously reported that dimeric DGCR8 binds heme and that the heme-bound DGCR8 is more active than the heme-free form. Here, we identified a conserved dimerization domain in DGCR8. Our crystal structure of this domain (residues 298-352) at 1.7 {angstrom} resolution demonstrates a previously unknown use of a WW motif as a platform for extensive dimerization interactions. The dimerization domain of DGCR8 is embedded in an independently folded heme-binding domain and directly contributes to association with heme. Heme-binding-deficient DGCR8 mutants have reduced pri-miRNA processing activity in vitro. Our study provides structural and biochemical bases for understanding how dimerization and heme binding of DGCR8 may contribute to regulation of miRNA biogenesis.

  8. Disordered clusters of Bak dimers rupture mitochondria during apoptosis

    Science.gov (United States)

    Uren, Rachel T; O’Hely, Martin; Iyer, Sweta; Bartolo, Ray; Shi, Melissa X; Brouwer, Jason M; Alsop, Amber E; Dewson, Grant; Kluck, Ruth M

    2017-01-01

    During apoptosis, Bak and Bax undergo major conformational change and form symmetric dimers that coalesce to perforate the mitochondrial outer membrane via an unknown mechanism. We have employed cysteine labelling and linkage analysis to the full length of Bak in mitochondria. This comprehensive survey showed that in each Bak dimer the N-termini are fully solvent-exposed and mobile, the core is highly structured, and the C-termini are flexible but restrained by their contact with the membrane. Dimer-dimer interactions were more labile than the BH3:groove interaction within dimers, suggesting there is no extensive protein interface between dimers. In addition, linkage in the mobile Bak N-terminus (V61C) specifically quantified association between dimers, allowing mathematical simulations of dimer arrangement. Together, our data show that Bak dimers form disordered clusters to generate lipidic pores. These findings provide a molecular explanation for the observed structural heterogeneity of the apoptotic pore. DOI: http://dx.doi.org/10.7554/eLife.19944.001 PMID:28182867

  9. Alkane dimers interaction

    DEFF Research Database (Denmark)

    Ferrighi, Lara; Madsen, Georg Kent Hellerup; Hammer, Bjørk

    2010-01-01

    The interaction energies of a series of n-alkane dimers, from methane to decane, have been investigated with Density Functional Theory (DFT), using the MGGA-M06-L density functional. The results are compared both to the available wavefunction-based values as well as to dispersion corrected DFT...

  10. Computational studies of the structural properties of the monomer and dimer of Aβ(1-28)

    Science.gov (United States)

    Dong, Xiao; Chen, Wei; Mousseau, Normand; Derreumaux, Philippe

    2007-03-01

    Neurodegenerative diseases are linked with the self-assembly of normally soluble proteins into amyloid fibrils. In this work, in silico characterization of the structures of the monomer and dimer of Aβ(1-28) are studied with the coarse-grained OPEP model using the activation-relaxation technique (ART nouveau). We find a dominant anti-parallel β-sheet structure present for both the monomer and dimer. While the monomer does not adopt a stable conformation, it fluctuates around a well-defined structure: starting from the end point, the monomer wraps a first time around, producing a β-hairpin and returns on the other side of the N-terminal, forming a three-strand β-sheet. The dimer assembles in a similar fashion, but with the two strands interlocking. The thermodynamics of the molecular assemblies and various folding path-ways are further studied using molecular dynamics.

  11. Dimeric (isoquinoline)(N-salicylidene-D,L-glutamato)copper(II) ethanol solvate.

    Science.gov (United States)

    Langer, Vratislav; Gyepesová, Dalma; Kohútová, Mária; Valent, Aladár

    2009-05-01

    The title racemic complex, bis[mu-N-(2-oxidobenzylidene)-D,L-glutamato(2-)]bis[(isoquinoline)copper(II)] ethanol disolvate, [Cu(2)(C(12)H(11)NO(5))(2)(C(9)H(7)N)(2)].2C(2)H(6)O, adopts a square-pyramidal Cu(II) coordination mode with a tridentate N-salicylideneglutamato Schiff base dianion and an isoquinoline ligand bound in the basal plane. The apex of the pyramid is occupied by a phenolic O atom from the adjacent chelate molecule at an apical distance of 2.487 (3) A, building a dimer located on the crystallographic inversion center. The Cu...Cu spacing within the dimers is 3.3264 (12) A. The ethanol solvent molecules are hydrogen bonded to the dimeric complex molecules, forming infinite chains in the a direction. The biological activity of the title complex has been studied.

  12. Cloning, large-scale production, and purification of active dimeric rat vascular endothelial growth factor (rrVEGF-164).

    NARCIS (Netherlands)

    Geutjes, P.J.; Nillesen, S.T.M.; Lammers, G.; Daamen, W.F.; Kuppevelt, A.H.M.S.M. van

    2010-01-01

    Large-scale production of recombinant rat vascular endothelial growth factor (rrVEGF-164) is desirable for angiogenic studies. In this study, biologically active recombinant rat vascular endothelial growth factor (rrVEGF-164) was cloned and expressed in the yeast Pichia pastoris, and large-scale pro

  13. FATTY ACIDS MODULATE TOLL-LIKE RECEPTOR 4 ACTIVATION THROUGH REGULATION OF RECEPTOR DIMERIZATION AND RECRUITMENT INTO LIPID RAFTS

    Science.gov (United States)

    The saturated fatty acids acylated on Lipid A of lipopolysaccharide (LPS) or bacterial lipoproteins play critical roles in ligand recognition and receptor activation for Toll-like Receptor 4 (TLR4) and TLR2. The results from our previous studies (J Biol Chem 2003, 2004) demonstrated that saturated ...

  14. Synthesis of novel 15-membered macrolide dimers

    Institute of Scientific and Technical Information of China (English)

    Shu Tao Ma; Rui Xin Ma; Rui Qing Xian; Bo Jiao

    2009-01-01

    A series of novel dimers of 15-memhered macrolides was synthesized and evaluated. The directs exhibited excellent activity against erythromycin-susceptible S. pneumonia, but did not show any improved activity against erythromycin-resistant S. pneumoniae encoded by erm gene.

  15. Structural and Dynamic Requirements for Optimal Activity of the Essential Bacterial Enzyme Dihydrodipicolinate Synthase

    Science.gov (United States)

    Reboul, C. F.; Porebski, B. T.; Griffin, M. D. W.; Dobson, R. C. J.; Perugini, M. A.; Gerrard, J. A.; Buckle, A. M.

    2012-01-01

    Dihydrodipicolinate synthase (DHDPS) is an essential enzyme involved in the lysine biosynthesis pathway. DHDPS from E. coli is a homotetramer consisting of a ‘dimer of dimers’, with the catalytic residues found at the tight-dimer interface. Crystallographic and biophysical evidence suggest that the dimers associate to stabilise the active site configuration, and mutation of a central dimer-dimer interface residue destabilises the tetramer, thus increasing the flexibility and reducing catalytic efficiency and substrate specificity. This has led to the hypothesis that the tetramer evolved to optimise the dynamics within the tight-dimer. In order to gain insights into DHDPS flexibility and its relationship to quaternary structure and function, we performed comparative Molecular Dynamics simulation studies of native tetrameric and dimeric forms of DHDPS from E. coli and also the native dimeric form from methicillin-resistant Staphylococcus aureus (MRSA). These reveal a striking contrast between the dynamics of tetrameric and dimeric forms. Whereas the E. coli DHDPS tetramer is relatively rigid, both the E. coli and MRSA DHDPS dimers display high flexibility, resulting in monomer reorientation within the dimer and increased flexibility at the tight-dimer interface. The mutant E. coli DHDPS dimer exhibits disorder within its active site with deformation of critical catalytic residues and removal of key hydrogen bonds that render it inactive, whereas the similarly flexible MRSA DHDPS dimer maintains its catalytic geometry and is thus fully functional. Our data support the hypothesis that in both bacterial species optimal activity is achieved by fine tuning protein dynamics in different ways: E. coli DHDPS buttresses together two dimers, whereas MRSA dampens the motion using an extended tight-dimer interface. PMID:22685390

  16. 76 FR 25364 - Agency Information Collection Activities: Form I-864, Form I-864A, Form I-864EZ, and From I-864W...

    Science.gov (United States)

    2011-05-04

    ... SECURITY U.S. Citizenship and Immigration Services Agency Information Collection Activities: Form I-864, Form I- 864A, Form I-864EZ, and From I-864W; Extension of an Existing Information Collection; Comment Request. ACTION: 60-Day Notice of Information Collection Under Review; Form I- 864, Affidavit of...

  17. A Novel Dimer of α-Tocopherol

    Directory of Open Access Journals (Sweden)

    Anjan Patel

    2008-01-01

    Full Text Available Decomposition of the complex 4, formed between the α-tocopherol ortho-quinone methide (2 and NMMO, by fast heating from −78∘C to 70∘C in inert solvents produces a novel α-tocopherol dimer with 6H,12H-dibenzo[b,f][1,5]dioxocine structure (5 which—in contrast to the well-known spiro-dimer of α-tocopherol (3—is symmetrical. This is the first example of a direct reaction of the highly transient zwitterionic, aromatic precursor 2a in the formation of the ortho-quinone methide 2.

  18. Pathogenic Cysteine Removal Mutations in FGFR Extracellular Domains Stabilize Receptor Dimers and Perturb the TM Dimer Structure.

    Science.gov (United States)

    Sarabipour, Sarvenaz; Hristova, Kalina

    2016-10-09

    Missense mutations that introduce or remove cysteine residues in receptor tyrosine kinases are believed to cause pathologies by stabilizing the active receptor tyrosine kinase dimers. However, the magnitude of this stabilizing effect has not been measured for full-length receptors. Here, we characterize the dimer stabilities of three full-length fibroblast growth factor receptor (FGFR) mutants harboring pathogenic cysteine substitutions: the C178S FGFR1 mutant, the C342R FGFR2 mutant, and the C228R FGFR3 mutant. We find that the three mutations stabilize the FGFR dimers. We further see that the mutations alter the configuration of the FGFR transmembrane dimers. Thus, both aberrant dimerization and perturbed dimer structure likely contribute to the pathological phenotypes arising due to these mutations.

  19. Pathologically activated neuroprotection via uncompetitive blockade of N-methyl-D-aspartate receptors with fast off-rate by novel multifunctional dimer bis(propyl)-cognitin.

    Science.gov (United States)

    Luo, Jialie; Li, Wenming; Zhao, Yuming; Fu, Hongjun; Ma, Dik-Lung; Tang, Jing; Li, Chaoying; Peoples, Robert W; Li, Fushun; Wang, Qinwen; Huang, Pingbo; Xia, Jun; Pang, Yuanping; Han, Yifan

    2010-06-25

    Uncompetitive N-methyl-d-aspartate (NMDA) receptor antagonists with fast off-rate (UFO) may represent promising drug candidates for various neurodegenerative disorders. In this study, we report that bis(propyl)-cognitin, a novel dimeric acetylcholinesterase inhibitor and gamma-aminobutyric acid subtype A receptor antagonist, is such an antagonist of NMDA receptors. In cultured rat hippocampal neurons, we demonstrated that bis(propyl)-cognitin voltage-dependently, selectively, and moderately inhibited NMDA-activated currents. The inhibitory effects of bis(propyl)-cognitin increased with the rise in NMDA and glycine concentrations. Kinetics analysis showed that the inhibition was of fast onset and offset with an off-rate time constant of 1.9 s. Molecular docking simulations showed moderate hydrophobic interaction between bis(propyl)-cognitin and the MK-801 binding region in the ion channel pore of the NMDA receptor. Bis(propyl)-cognitin was further found to compete with [(3)H]MK-801 with a K(i) value of 0.27 mum, and the mutation of NR1(N616R) significantly reduced its inhibitory potency. Under glutamate-mediated pathological conditions, bis(propyl)-cognitin, in contrast to bis(heptyl)-cognitin, prevented excitotoxicity with increasing effectiveness against escalating levels of glutamate and much more effectively protected against middle cerebral artery occlusion-induced brain damage than did memantine. More interestingly, under NMDA receptor-mediated physiological conditions, bis(propyl)-cognitin enhanced long-term potentiation in hippocampal slices, whereas MK-801 reduced and memantine did not alter this process. These results suggest that bis(propyl)-cognitin is a UFO antagonist of NMDA receptors with moderate affinity, which may provide a pathologically activated therapy for various neurodegenerative disorders associated with NMDA receptor dysregulation.

  20. A Large Conformational Change of a Bridged β-Cyclodextrin Dimer in Aqueous Solution

    Institute of Scientific and Technical Information of China (English)

    Xiao Qi ZHENG; Yong Hui WANG; Qing Xiang GUO; Li YANG; You Cheng LIU

    2003-01-01

    A novel bridged β-CD dimer in which two β-cyclodextrins were linked by a naphthalene at positions 2 and 7 has been synthesized. 1H and 13CNMR measurements showed that a large change in the conformation of the dimer occurred in aqueous solution. The dimer interacted with methyl and ethyl orange to form stable inclusion complexes via "induced fit" mechanism.

  1. Synthesis of β,β'-Porphyrin Dimer Linked by Vinylene

    Institute of Scientific and Technical Information of China (English)

    Jiang, Xuliang; Li, Panli; Wang, Yucheng; Shen, Qi; Tao, Jingchao; Shi, Weimin

    2012-01-01

    Synthesis of a novel β,β'-tetraalkylporphyrin dimer linked by vinylene was discribed, in which the dimer was readily prepared from a porphyrin-derived Wittig reagent and a mono-formylated porphyrin via Wittig reaction. No π-conjugation between the two porphyrin rings was obserbed, and the dimer was in trans form.

  2. Detection of alpha-helical coiled-coil dimer formation by spin-labeled synthetic peptides: a model parallel coiled-coil peptide and the antiparallel coiled coil formed by a replica of the ProP C-terminus.

    Science.gov (United States)

    Hillar, Alexander; Tripet, Brian; Zoetewey, David; Wood, Janet M; Hodges, Robert S; Boggs, Joan M

    2003-12-30

    Electron paramagnetic resonance spectroscopy was used to determine relative peptide orientation within homodimeric, alpha-helical coiled-coil structures. Introduction of cysteine (Cys) residues into peptides/proteins for spin labeling allows detection of their oligomerization from exchange broadening or dipolar interactions between residues within 25 A of each other. Two synthetic peptides containing Cys substitutions were used: a 35-residue model peptide and the 30-residue ProP peptide. The model peptide is known to form a stable, parallel homodimeric coiled coil, which is partially destabilized by Cys substitutions at heptad a and d positions (peptides C30a and C33d). The ProP peptide, a 30-residue synthetic peptide, corresponds to residues 468-497 of osmoregulatory transporter ProP from Escherichia coli. It forms a relatively unstable, homodimeric coiled coil that is predicted to be antiparallel in orientation. Cys was introduced in heptad g positions of the ProP peptide, near the N-terminus (K473C, creating peptide C473g) or closer to the center of the sequence (E480C, creating peptide C480g). In contrast to the destabilizing effect of Cys substitution at the core heptad a or d positions of model peptides C30a and C33d, circular dichroism spectroscopy showed that Cys substitutions at the heptad g positions of the ProP peptide had little or no effect on coiled-coil stability. Thermal denaturation analysis showed that spin labeling increased the stability of the coiled coil for all peptides. Strong exchange broadening was detected for both C30a and C33d, in agreement with a parallel structure. EPR spectra of C480g had a large hyperfine splitting of about 90 G, indicative of strong dipole-dipole interactions and a distance between spin-labeled residues of less than 9 A. Spin-spin interactions were much weaker for C473g. These results supported the hypothesis that the ProP peptide primarily formed an antiparallel coiled coil, since formation of a parallel dimer

  3. Differential antifungal activity of isomeric forms of nystatin.

    Science.gov (United States)

    Ostrosky-Zeichner, L; Bazemore, S; Paetznick, V L; Rodriguez, J R; Chen, E; Wallace, T; Cossum, P; Rex, J H

    2001-10-01

    When nystatin is placed in RPMI and other biological fluids, there is loss of pure nystatin, with the development of two distinguishable chromatographic peaks, 1 and 2. Peak 1 appears identical to commercially prepared nystatin. By nuclear magnetic resonance (NMR) and mass spectral analysis, peak 2 appears to be an isomer of peak 1. The isomers are quantitatively and fully interconvertible. Formation of peak 2 is accelerated at a pH of >7.0 and ultimately reaches a near 55:45 (peak 1/peak 2 ratio) mixture. We sought to determine the relative activities of peaks 1 and 2 against Candida spp. Peak 2 consistently showed higher MICs when it was the predominant form during the experiment. Time-kill analyses showed that peak 2 required > or =8 x the concentration of peak 1 to produce a modest and delayed killing effect, which was never of the same magnitude as that produced by peak 1. In both types of assays, the activity of peak 2 corresponded with intra-assay formation of peak 1. Both MIC measurements and time-kill analysis suggest that peak 2 has considerably less activity, if any at all, against Candida spp. Peak 2 may serve as a reservoir for peak 1.

  4. Fluorescence Microspectroscopy for Testing the Dimerization Hypothesis of BACE1 Protein in Cultured HEK293 Cells

    Science.gov (United States)

    Gardeen, Spencer; Johnson, Joseph L.; Heikal, Ahmed A.

    2016-06-01

    Alzheimer's Disease (AD) is a neurodegenerative disorder that results from the formation of beta-amyloid plaques in the brain that trigger the known symptoms of memory loss in AD patients. The beta-amyloid plaques are formed by the proteolytic cleavage of the amyloid precursor protein (APP) by the proteases BACE1 and gamma-secretase. These enzyme-facilitated cleavages lead to the production of beta-amyloid fragments that aggregate to form plaques, which ultimately lead to neuronal cell death. Recent detergent protein extraction studies suggest that BACE1 protein forms a dimer that has significantly higher catalytic activity than its monomeric counterpart. In this contribution, we examine the dimerization hypothesis of BACE1 in cultured HEK293 cells using complementary fluorescence spectroscopy and microscopy methods. Cells were transfected with a BACE1-EGFP fusion protein construct and imaged using confocal, and differential interference contrast to monitor the localization and distribution of intracellular BACE1. Complementary fluorescence lifetime and anisotropy measurements enabled us to examine the conformational and environmental changes of BACE1 as a function of substrate binding. Using fluorescence correlation spectroscopy, we also quantified the diffusion coefficient of BACE1-EGFP on the plasma membrane as a means to test the dimerization hypothesis as a fucntion of substrate-analog inhibitition. Our results represent an important first towards examining the substrate-mediated dimerization hypothesis of BACE1 in live cells.

  5. FAK dimerization controls its kinase-dependent functions at focal adhesions

    KAUST Repository

    Brami-Cherrier, Karen

    2014-01-30

    Focal adhesion kinase (FAK) controls adhesion-dependent cell motility, survival, and proliferation. FAK has kinase-dependent and kinase-independent functions, both of which play major roles in embryogenesis and tumor invasiveness. The precise mechanisms of FAK activation are not known. Using x-ray crystallography, small angle x-ray scattering, and biochemical and functional analyses, we show that the key step for activation of FAK\\'s kinase-dependent functions-autophosphorylation of tyrosine-397-requires site-specific dimerization of FAK. The dimers form via the association of the N-terminal FERM domain of FAK and are stabilized by an interaction between FERM and the C-terminal FAT domain. FAT binds to a basic motif on FERM that regulates co-activation and nuclear localization. FAK dimerization requires local enrichment, which occurs specifically at focal adhesions. Paxillin plays a dual role, by recruiting FAK to focal adhesions and by reinforcing the FAT:FERM interaction. Our results provide a structural and mechanistic framework to explain how FAK combines multiple stimuli into a site-specific function. The dimer interfaces we describe are promising targets for blocking FAK activation. © 2014 The Authors.

  6. Experimental and computational X-ray emission spectroscopy as a direct probe of protonation states in oxo-bridged Mn(IV) dimers relevant to redox-active metalloproteins.

    Science.gov (United States)

    Lassalle-Kaiser, Benedikt; Boron, Thaddeus T; Krewald, Vera; Kern, Jan; Beckwith, Martha A; Delgado-Jaime, Mario U; Schroeder, Henning; Alonso-Mori, Roberto; Nordlund, Dennis; Weng, Tsu-Chien; Sokaras, Dimosthenis; Neese, Frank; Bergmann, Uwe; Yachandra, Vittal K; DeBeer, Serena; Pecoraro, Vincent L; Yano, Junko

    2013-11-18

    The protonation state of oxo bridges in nature is of profound importance for a variety of enzymes, including the Mn4CaO5 cluster of photosystem II and the Mn2O2 cluster in Mn catalase. A set of dinuclear bis-μ-oxo-bridged Mn(IV) complexes in different protonation states was studied by Kβ emission spectroscopy to form the foundation for unraveling the protonation states in the native complex. The valence-to-core regions (valence-to-core XES) of the spectra show significant changes in intensity and peak position upon protonation. DFT calculations were performed to simulate the valence-to-core XES spectra and to assign the spectral features to specific transitions. The Kβ(2,5) peaks arise primarily from the ligand 2p to Mn 1s transitions, with a characteristic low energy shoulder appearing upon oxo-bridge protonation. The satellite Kβ" peak provides a more direct signature of the protonation state change, since the transitions originating from the 2s orbitals of protonated and unprotonated μ-oxo bridges dominate this spectral region. The energies of the Kβ" features differ by ~3 eV and thus are well resolved in the experimental spectra. Additionally, our work explores the chemical resolution limits of the method, namely, whether a mixed (μ-O)(μ-OH2) motif can be distinguished from a symmetric (μ-OH)2 one. The results reported here highlight the sensitivity of Kβ valence-to-core XES to single protonation state changes of bridging ligands, and form the basis for further studies of oxo-bridged polymetallic complexes and metalloenzyme active sites. In a complementary paper, the results from X-ray absorption spectroscopy of the same Mn(IV) dimer series are discussed.

  7. Radiometric microbiologic assay for the biologically active forms of niacin

    Energy Technology Data Exchange (ETDEWEB)

    Kertcher, J.A.; Guilarte, T.R.; Chen, M.F.; Rider, A.A.; McIntyre, P.A.

    1979-05-01

    A radiometric microbiologic assay has been developed for the determination of niacin in biologic fluids. Lactobacillus plantarum produced /sup 14/CO/sub 2/ from L-(U-/sup 14/C) malic acid in quantities proportional to the amount of niacin present. The assay is specific for the biologically active forms of niacin in humans. Thirty normal hemolysates were analyzed and the values ranged from 13.0 to 17.8 ..mu..g niacin/ml RBC (mean = 15.27 +- 1.33 s.d.). Good recovery and reproducibility studies were obtained with this assay. On thirty blood samples, correlation was excellent between the radiometric and the conventional turbidimetric assays.

  8. The Circumnuclear Star-forming Activities along the Hubble Sequence

    CERN Document Server

    Shi, L; Peng, Z; Shi, Lei; Gu, Qiusheng; Peng, Zhixin

    2005-01-01

    In order to study the circumnuclear star-forming activity along the Hubble sequence, we cross-correlate the Sloan Digital Sky Survey Data Release 2 (SDSS DR2) with the Third Reference Catalog of Bright Galaxies (RC3) to derive a large sample of 1015 galaxies with both morphological and spectral information. Among these, 385 sources are classified as star-forming galaxies and the SDSS fibre covered the circumnuclear regions (0.2 $-$ 2.0 kpc). By using the spectral synthesis method to remove the contribution from the underlying old stellar population, we measure the emission lines fluxes accurately which are then used to estimate the star formation rates(SFRs). Our main findings are that: (1) Early-type spirals show much larger H$\\alpha$ luminosities and hence higher SFRs, they also suffer more extinctions than late-type ones. The equivalent widths (EWs) of H$\\alpha$ emission lines show the similar trend, however, the very late types (Sdm $\\sim$ Irr) do have large fractions of high EWs. (2) We confirm that D$_n...

  9. Monounsaturated fatty acid ether oligomers formed during heating of virgin olive oil show agglutination activity against human red blood cells.

    Science.gov (United States)

    Patrikios, Ioannis S; Mavromoustakos, Thomas M

    2014-01-29

    The present work focuses on the characterization of molecules formed when virgin olive oil is heated at 130 °C for 24 h open in air, which are found to be strong agglutinins. The hemagglutinating activity of the newly formed molecule isolated from the heated virgin olive oil sample was estimated against human red blood cells (RBCs). Dimers and polymers (high molecular weight molecules) were identified through thin layer chromatography (TLC) of the oil mixture. (1)H and (13)C nuclear magnetic resonance (NMR) and gas chromatography-mass spectroscopy (GC-MS) were the methods used for structural characterization. Among others, oligomerization of at least two monounsaturated fatty acids (FA) by an ether linkage between the hydrocarbon chains is involved. Light microscopy was used to characterize and visualize the agglutination process. Agglutination without fusion or lysis was observed. It was concluded that the heating of virgin olive oil open in air, among other effects, produces oligomerization as well as polymerization of unsaturated FA, possibly of monohydroxy, monounsaturated FA that is associated with strong hemagglutinating activity against human RBCs. The nutritional value and the effects on human health of such oligomers are not discussed in the literature and remain to be investigated.

  10. An alternative RNA polymerase I structure reveals a dimer hinge.

    Science.gov (United States)

    Kostrewa, Dirk; Kuhn, Claus-D; Engel, Christoph; Cramer, Patrick

    2015-09-01

    RNA polymerase I (Pol I) is the central, 14-subunit enzyme that synthesizes the ribosomal RNA (rRNA) precursor in eukaryotic cells. The recent crystal structure of Pol I at 2.8 Å resolution revealed two novel elements: the `expander' in the active-centre cleft and the `connector' that mediates Pol I dimerization [Engel et al. (2013), Nature (London), 502, 650-655]. Here, a Pol I structure in an alternative crystal form that was solved by molecular replacement using the original atomic Pol I structure is reported. The resulting alternative structure lacks the expander but still shows an expanded active-centre cleft. The neighbouring Pol I monomers form a homodimer with a relative orientation distinct from that observed previously, establishing the connector as a hinge between Pol I monomers.

  11. A putatively novel form of spontaneous coordination in neural activity.

    Science.gov (United States)

    Hermer-Vazquez, Raymond; Hermer-Vazquez, Linda; Srinivasan, Sridhar

    2009-04-06

    We simultaneously recorded local field potentials from three sites along the olfactory-entorhinal axis in rats lightly anesthetized with isoflurane, as part of another experiment. While analyzing the initial data from that experiment with spectrograms, we discovered a potentially novel form of correlated neural activity, with near-simultaneous occurrence across the three widely separated brain sites. After validating their existence further, we named these events Synchronous Frequency Bursts (SFBs). Here we report our initial investigations into their properties and their potential functional significance. In Experiment 1, we found that SFBs have highly regular properties, consisting of brief (approximately 250 ms), high amplitude bursts of LFP energy spanning frequency ranges from the delta band (1-4 Hz) to at least the low gamma band (30-50 Hz). SFBs occurred almost simultaneously across recording sites, usually with onsets sites. While the SFBs had fairly typical, exponentially decaying power spectral density plots, their coherence structure was unusual, with high peaks in several narrow frequency ranges and little coherence in other bands. In Experiment 2, we found that SFBs occurred far more often under light anesthesia than deeper anesthetic states, and were especially prevalent as the animals regained consciousness. Finally, in Experiment 3 we showed that SFBs occur simultaneously at a significant rate across brain sites from putatively different functional subsystems--olfactory versus motor pathways. We suggest that SFBs do not carry information per se, but rather, play a role in coordinating activity in different frequency bands, potentially brain-wide, as animals progress from sleep or anesthesia toward full consciousness.

  12. The acrylonitrile dimer ion

    Science.gov (United States)

    Ervasti, Henri K.; Jobst, Karl J.; Burgers, Peter C.; Ruttink, Paul J. Ae; Terlouw, Johan K.

    2007-04-01

    Large energy barriers prohibit the rearrangement of solitary acrylonitrile ions, CH2CHCN+, into their more stable hydrogen-shift isomers CH2CCNH+ or CHCH-CNH+. This prompted us to examine if these isomerizations occur by self-catalysis in acrylonitrile dimer ions. Such ions, generated by chemical ionization experiments of acrylonitrile with an excess of carbon dioxide, undergo five dissociations in the [mu]s time frame, as witnessed by peaks at m/z 53, 54, 79, 80 and 105 in their metastable ion mass spectrum. Collision experiments on these product ions, deuterium labeling, and a detailed computational analysis using the CBS-QB3 model chemistry lead to the following conclusions: (i) the m/z 54 ions are ions CH2CHCNH+ generated by self-protonation in ion-dipole stabilized hydrogen-bridged dimer ions [CH2CHCN...H-C(CN)CH2]+ and [CH2CHCN...H-C(H)C(H)CN]+; the proton shifts in these ions are associated with a small reverse barrier; (ii) dissociation of the H-bridged ions into CH2CCNH+ or CHCH-CNH+ by self-catalysis is energetically feasible but kinetically improbable: experiment shows that the m/z 53 ions are CH2CHCN+ ions, generated by back dissociation; (iii) the peaks at m/z 79, 80 and 105 correspond with the losses of HCN, C2H2 and H, respectively. The calculations indicate that these ions are generated from dimer ions that have adopted the (much more stable) covalently bound "head-to-tail" structure [CH2CHCN-C(H2)C(H)CN]+; experiments indicate that the m/z 79 (C5H5N) and m/z 105 (C6H6N2) ions have linear structures but the m/z 80 (C4H4N2) ions consist of ionized pyrimidine in admixture with its stable pyrimidine-2-ylidene isomer. Acrylonitrile is a confirmed species in interstellar space and our study provides experimental and computational evidence that its dimer radical cation yields the ionized prebiotic pyrimidine molecule.

  13. Metallothionein dimers studied by nano-spray mass spectrometry.

    Science.gov (United States)

    Hathout, Yetrib; Reynolds, Kristy J; Szilagyi, Zoltan; Fenselau, Catherine

    2002-01-15

    Both transient and stable dimers of metallothionein have been characterized, based on earlier studies using NMR, circular dichroism and size-exclusion chromatography. Here additional characterization is provided by nanospray mass spectrometry. Rapid redistribution of metal ions between monomeric Cd7- and Zn7-metallothionein 2a is monitored by nanospray. An experiment in which theses two forms of the monomeric protein are separated by a dialysis membrane, which will pass metal ions but not proteins, confirms that a transient dimer must form for metal ions to be redistributed. On the other hand, size-exclusion chromatography of reconstituted Zn7- or Cd7-metallothionein revealed the presence of monomeric and dimeric species. These dimers do not equilibrate readily to form monomers and they are shown to be covalent.

  14. Crystal structure of TNF-α-inducing protein from Helicobacter pylori in active form reveals the intrinsic molecular flexibility for unique DNA-binding.

    Directory of Open Access Journals (Sweden)

    Mingming Gao

    Full Text Available Tipα (TNF-α-inducing protein from Helicobacter pylori is a carcinogenic effector. Studies on this protein revealed that a homodimer linked by a pair of intermolecular disulfide bridges (Cys25-Cys25 and Cys27-Cys27 was absolutely necessary for its biological functions. The activities of Tipα would be abolished when both disulfide bridges were disrupted. The crystal structures of Tipα reported to date, however, were based on inactive, monomeric mutants with their N-terminal, including residues Cys25 and Cys27, truncated. Here we report the crystal structure of H. pylori Tipα protein, TipαN(25, at 2.2Å resolution, in which Cys25 and Cys27 form a pair of inter-chain disulfide bridges linking an active dimer. The disulfide bridges exhibit structural flexibility in the present structure. A series of structure-based mutagenesis, biochemical assays and molecular dynamic simulations on DNA-Tipα interactions reveal that Tipα utilizes the dimeric interface as the DNA-binding site and that residues His60, Arg77 and Arg81 located at the interface are crucial for DNA binding. Tipα could bind to one ssDNA, two ssDNA or one dsDNA in experiments, respectively, in the native or mutant states. The unique DNA-binding activities of Tipα indicate that the intrinsic flexible nature of disulfide bridges could endow certain elasticity to the Tipα dimer for its unique bioactivities. The results shed light on the possible structural mechanism for the functional performances of Tipα.

  15. A variant form of the human deleted in malignant brain tumor 1 (DMBT1 gene shows increased expression in inflammatory bowel diseases and interacts with dimeric trefoil factor 3 (TFF3.

    Directory of Open Access Journals (Sweden)

    Jens Madsen

    Full Text Available The protein deleted in malignant brain tumors (DMBT1 and the trefoil factor (TFF proteins have all been proposed to have roles in epithelial cell growth and cell differentiation and shown to be up regulated in inflammatory bowel diseases. A panel of monoclonal antibodies was raised against human DMBT1(gp340. Analysis of lung washings and colon tissue extracts by Western blotting in the unreduced state, two antibodies (Hyb213-1 and Hyb213-6 reacted with a double band of 290 kDa in lung lavage. Hyb213-6, in addition, reacted against a double band of 270 kDa in colon extract while Hyb213-1 showed no reaction. Hyb213-6 showed strong cytoplasmic staining in epithelial cells of both the small and large intestine whereas no staining was seen with Hyb213-1. The number of DMBT1(gp340 positive epithelial cells, stained with Hyb213-6, was significantly up regulated in inflammatory colon tissue sections from patients with ulcerative colitis (p<0.0001 and Crohn's disease (p = 0.006 compared to normal colon tissue. Immunohistochemical analysis of trefoil factor TFF1, 2 and 3 showed that TFF1 and 3 localized to goblet cells in both normal colon tissue and in tissue from patients with ulcerative colitis or Crohn's disease. No staining for TFF2 was seen in goblet cells in normal colon tissue whereas the majority of tissue sections in ulcerative colitis and Crohn's disease showed sparse and scattered TFF2 positive goblet cells. DMBT1 and TFF proteins did therefore not co-localize in the same cells but localized in adjacent cells in the colon. The interaction between DMBT1(gp340 and trefoil TFFs proteins was investigated using an ELISA assay. DMBT1(gp340 bound to solid-phase bound recombinant dimeric TFF3 in a calcium dependent manner (p<0.0001 but did not bind to recombinant forms of monomeric TFF3, TFF2 or glycosylated TFF2. This implies a role for DMBT1 and TFF3 together in inflammatory bowel disease.

  16. Synthesis of Macrocyclic Hexaoxazole (6OTD Dimers, Containing Guanidine and Amine Functionalized Side Chains, and an Evaluation of Their Telomeric G4 Stabilizing Properties

    Directory of Open Access Journals (Sweden)

    Keisuke Iida

    2010-01-01

    Full Text Available Structure-activity relationship studies were carried out on macrocyclic hexaoxazole (6OTD dimers, whose core structure stabilizes telomeric G-quadruplexes (G4. Two new 6OTD dimers having side chain amine and guanidine functional groups were synthesized and evaluated for their stabilizing ability against a telomeric G4 DNA sequence. The results show that the 6OTD dimers interact with the DNA to form 1:1 complexes and stabilize the antiparallel G4 structure of DNA in the presence of potassium cation. The guanidine functionalized dimer displays a potent stabilizing ability of the G4 structure, as determined by using a FRET melting assay (ΔTm=14 °C.

  17. Reengineering Antibiotics to Combat Bacterial Resistance: Click Chemistry [1,2,3]-Triazole Vancomycin Dimers with Potent Activity against MRSA and VRE.

    Science.gov (United States)

    Silverman, Steven M; Moses, John E; Sharpless, K Barry

    2017-01-01

    Vancomycin has long been considered a drug of last resort. Its efficiency in treating multiple drug-resistant bacterial infections, particularly methicillin-resistant Staphylococcus aureus (MRSA), has had a profound effect on the treatment of life-threatening infections. However, the emergence of resistance to vancomycin is a cause for significant worldwide concern, prompting the urgent development of new effective treatments for antibiotic resistant bacterial infections. Harnessing the benefits of multivalency and cooperativity against vancomycin-resistant strains, we report a Click Chemistry approach towards reengineered vancomycin derivatives and the synthesis of a number of dimers with increased potency against MRSA and vancomycin resistant Enterococci (VRE; VanB). These semi-synthetic dimeric ligands were linked together with great efficiency using the powerful CuAAC reaction, demonstrating high levels of selectivity and purity.

  18. Solution structure of the dimerization domain of the eukaryotic stalk P1/P2 complex reveals the structural organization of eukaryotic stalk complex.

    Science.gov (United States)

    Lee, Ka-Ming; Yu, Conny Wing-Heng; Chiu, Teddy Yu-Hin; Sze, Kong-Hung; Shaw, Pang-Chui; Wong, Kam-Bo

    2012-04-01

    The lateral ribosomal stalk is responsible for the kingdom-specific binding of translation factors and activation of GTP hydrolysis during protein synthesis. The eukaryotic stalk is composed of three acidic ribosomal proteins P0, P1 and P2. P0 binds two copies of P1/P2 hetero-dimers to form a pentameric P-complex. The structure of the eukaryotic stalk is currently not known. To provide a better understanding on the structural organization of eukaryotic stalk, we have determined the solution structure of the N-terminal dimerization domain (NTD) of P1/P2 hetero-dimer. Helix-1, -2 and -4 from each of the NTD-P1 and NTD-P2 form the dimeric interface that buries 2200 A(2) of solvent accessible surface area. In contrast to the symmetric P2 homo-dimer, P1/P2 hetero-dimer is asymmetric. Three conserved hydrophobic residues on the surface of NTD-P1 are replaced by charged residues in NTD-P2. Moreover, NTD-P1 has an extra turn in helix-1, which forms extensive intermolecular interactions with helix-1 and -4 of NTD-P2. Truncation of this extra turn of P1 abolished the formation of P1/P2 hetero-dimer. Systematic truncation studies suggest that P0 contains two spine-helices that each binds one copy of P1/P2 hetero-dimer. Modeling studies suggest that a large hydrophobic cavity, which can accommodate the loop between the spine-helices of P0, can be found on NTD-P1 but not on NTD-P2 when the helix-4 adopts an 'open' conformation. Based on the asymmetric properties of NTD-P1/NTD-P2, a structural model of the eukaryotic P-complex with P2/P1:P1/P2 topology is proposed.

  19. Reduction of Streptolysin O (SLO Pore-Forming Activity Enhances Inflammasome Activation

    Directory of Open Access Journals (Sweden)

    Peter A. Keyel

    2013-06-01

    Full Text Available Pore-forming toxins are utilized by bacterial and mammalian cells to exert pathogenic effects and induce cell lysis. In addition to rapid plasma membrane repair, macrophages respond to pore-forming toxins through activation of the NLRP3 inflammasome, leading to IL-1β secretion and pyroptosis. The structural determinants of pore-forming toxins required for NLRP3 activation remain unknown. Here, we demonstrate using streptolysin O (SLO that pore-formation controls IL-1β secretion and direct toxicity. An SLO mutant incapable of pore-formation did not promote direct killing, pyroptosis or IL-1β production. This indicated that pore formation is necessary for inflammasome activation. However, a partially active mutant (SLO N402C that was less toxic to macrophages than wild-type SLO, even at concentrations that directly lysed an equivalent number of red blood cells, enhanced IL-1β production but did not alter pyroptosis. This suggests that direct lysis may attenuate immune responses by preventing macrophages from successfully repairing their plasma membrane and elaborating more robust cytokine production. We suggest that mutagenesis of pore-forming toxins represents a strategy to enhance adjuvant activity.

  20. Multicritical tensor models and hard dimers on spherical random lattices

    CERN Document Server

    Bonzom, Valentin

    2012-01-01

    Random tensor models which display multicritical behaviors in a remarkably simple fashion are presented. They come with entropy exponents \\gamma = (m-1)/m, similarly to multicritical random branched polymers. Moreover, they are interpreted as models of hard dimers on a set of random lattices for the sphere in dimension three and higher. Dimers with their exclusion rules are generated by the different interactions between tensors, whose coupling constants are dimer activities. As an illustration, we describe one multicritical point, which is interpreted as a transition between the dilute phase and a crystallized phase, though with negative activities.

  1. Subsurface dimerization in III-V semiconductor (001) surfaces

    DEFF Research Database (Denmark)

    Kumpf, C.; Marks, L.D.; Ellis, D.

    2001-01-01

    We present the atomic structure of the c(8 X 2) reconstructions of InSb-, InAs-, and GaAs-(001) surfaces as determined by surface x-ray diffraction using direct methods. Contrary to common belief, group III dimers are not prominent on the surface, instead subsurface dimerization of group m atoms ...... takes place in the second bilayer, accompanied by a major rearrangement of the surface atoms above the dimers to form linear arrays. By varying the occupancies of four surface sites the (001)-c(8 X 2) reconstructions of III-V semiconductors can be described in a unified model....

  2. Developmental regulation of synthesis and dimerization of the amyloidogenic protease inhibitor cystatin C in the hematopoietic system.

    Science.gov (United States)

    Xu, Yuekang; Lindemann, Petra; Vega-Ramos, Javier; Zhang, Jian-Guo; Villadangos, Jose A

    2014-04-04

    The cysteine protease inhibitor cystatin C is thought to be secreted by most cells and eliminated in the kidneys, so its concentration in plasma is diagnostic of kidney function. Low extracellular cystatin C is linked to pathologic protease activity in cancer, arthritis, atherosclerosis, aortic aneurism, and emphysema. Cystatin C forms non-inhibitory dimers and aggregates by a mechanism known as domain swapping, a property that reportedly protects against Alzheimer disease but can also cause amyloid angiopathy. Despite these clinical associations, little is known about the regulation of cystatin C production, dimerization, and secretion. We show that hematopoietic cells are major contributors to extracellular cystatin C levels in healthy mice. Among these cells, macrophages and dendritic cells (DC) are the predominant producers of cystatin C. Both cell types synthesize monomeric and dimeric cystatin C in vivo, but only secrete monomer. Dimerization occurs co-translationally in the endoplasmic reticulum and is regulated by the levels of reactive oxygen species (ROS) derived from mitochondria. Drugs or stimuli that reduce the intracellular concentration of ROS inhibit cystatin C dimerization. The extracellular concentration of inhibitory cystatin C is thus partly dependent on the abundance of macrophages and DC, and the ROS levels. These results have implications for the diagnostic use of serum cystatin C as a marker of kidney function during inflammatory processes that induce changes in DC or macrophage abundance. They also suggest an important role for macrophages, DC, and ROS in diseases associated with the protease inhibitory activity or amyloidogenic properties of cystatin C.

  3. Dimeric c-di-GMP is required for post-translational regulation of alginate production in Pseudomonas aeruginosa.

    Science.gov (United States)

    Whitney, John C; Whitfield, Gregory B; Marmont, Lindsey S; Yip, Patrick; Neculai, A Mirela; Lobsanov, Yuri D; Robinson, Howard; Ohman, Dennis E; Howell, P Lynne

    2015-05-15

    Pseudomonas aeruginosa is an opportunistic human pathogen that secretes the exopolysaccharide alginate during infection of the respiratory tract of individuals afflicted with cystic fibrosis and chronic obstructive pulmonary disease. Among the proteins required for alginate production, Alg44 has been identified as an inner membrane protein whose bis-(3',5')-cyclic dimeric guanosine monophosphate (c-di-GMP) binding activity post-translationally regulates alginate secretion. In this study, we report the 1.8 Å crystal structure of the cytoplasmic region of Alg44 in complex with dimeric self-intercalated c-di-GMP and characterize its dinucleotide-binding site using mutational analysis. The structure shows that the c-di-GMP binding region of Alg44 adopts a PilZ domain fold with a dimerization mode not previously observed for this family of proteins. Calorimetric binding analysis of residues in the c-di-GMP binding site demonstrate that mutation of Arg-17 and Arg-95 alters the binding stoichiometry between c-di-GMP and Alg44 from 2:1 to 1:1. Introduction of these mutant alleles on the P. aeruginosa chromosome show that the residues required for binding of dimeric c-di-GMP in vitro are also required for efficient alginate production in vivo. These results suggest that the dimeric form of c-di-GMP represents the biologically active signaling molecule needed for the secretion of an important virulence factor produced by P. aeruginosa.

  4. Active star-forming galaxies in the X ray foreground

    Science.gov (United States)

    Griffiths, R. E.; Padovani, P.

    1989-01-01

    Star forming galaxies were discovered as a component of the X-ray background (XRB) in the Einstein deep surveys. Such star forming galaxies may be largely powered by superluminous Population 1 massive X-ray binaries (MXRB), formed in the wake of star formation in regions of low metallicity. The star forming galaxies with moderate numbers of MXRB may evolve into the infrared starburst galaxies found at low redshifts using IRAS (Infrared Astronomy Satellite), and may also be related to those galaxies identified with sub-mJy radio sources. A conservative contribution to the XRB of at least approximately 15 percent, without evolution is estimated. It is shown that moderate evolution leads to a contribution at least equalling that of quasars. Above 3 keV, star forming galaxies may dominate the XRB.

  5. Dynamics of a bouncing dimer

    CERN Document Server

    Dorbolo, S; Tsimring, L S; Kudrolli, A

    2005-01-01

    We investigate the dynamics of a dimer bouncing on a vertically oscillated plate. The dimer, composed of two spheres rigidly connected by a light rod, exhibits several modes depending on initial and driving conditions. The first excited mode has a novel horizontal drift in which one end of the dimer stays on the plate during most of the cycle, while the other end bounces in phase with the plate. The speed and direction of the drift depend on the aspect ratio of the dimer. We employ event-driven simulations based on a detailed treatment of frictional interactions between the dimer and the plate in order to elucidate the nature of the transport mechanism in the drift mode.

  6. A peroxiredoxin cDNA from Taiwanofungus camphorata: role of Cys31 in dimerization.

    Science.gov (United States)

    Huang, Chih-Yu; Chen, Yu-Ting; Wen, Lisa; Sheu, Dey-Chyi; Lin, Chi-Tsai

    2014-01-01

    Peroxiredoxins (Prxs) play important roles in antioxidant defense and redox signaling pathways. A Prx isozyme cDNA (TcPrx2, 745 bp, EF552425) was cloned from Taiwanofungus camphorata and its recombinant protein was overexpressed. The purified protein was shown to exist predominantly as a dimer by sodium dodecyl sulfate-polyacrylamide gel electrolysis in the absence of a reducing agent. The protein in its dimeric form showed no detectable Prx activity. However, the protein showed increased Prx activity with increasing dithiothreitol concentration which correlates with dissociation of the dimer into monomer. The TcPrx2 contains two Cys residues. The Cys(60) located in the conserved active site is the putative active peroxidatic Cys. The role of Cys(31) was investigated by site-directed mutagenesis. The C31S mutant (C(31) → S(31)) exists predominantly as a monomer with noticeable Prx activity. The Prx activity of the mutant was higher than that of the corresponding wild-type protein by nearly twofold at 12 μg/mL. The substrate preference of the mutant was H2O2 > cumene peroxide > t-butyl peroxide. The Michaelis constant (K M) value for H2O2 of the mutant was 0.11 mM. The mutant enzyme was active under a broad pH range from 6 to 10. The results suggest a role of Cys(31) in dimerization of the TcPrx2, a role which, at least in part, may be involved in determining the activity of Prx. The C(31) residue does not function as a resolving Cys and therefore the TcPrx2 must follow the reaction mechanism of 1-Cys Prx. This TcPrx2 represents a new isoform of Prx family.

  7. A common model for cytokine receptor activation: combined scissor-like rotation and self-rotation of receptor dimer induced by class I cytokine.

    Directory of Open Access Journals (Sweden)

    Xiaodong Pang

    Full Text Available The precise mechanism by which the binding of a class I cytokine to the extracellular domain of its corresponding receptor transmits a signal through the cell membrane remains unclear. Receptor activation involves a cytokine-receptor complex with a 1∶2 stoichiometry. Previously we used our transient-complex theory to calculate the rate constant of the initial cytokine-receptor binding to form a 1∶1 complex. Here we computed the binding pathway leading to the 1∶2 activation complex. Three cytokine systems (growth hormone, erythropoietin, and prolactin were studied, and the focus was on the binding of the extracellular domain of the second receptor molecule after forming the 1∶1 complex. According to the transient-complex theory, translational and rotation diffusion of the binding entities bring them together to form a transient complex, which has near-native relative separation and orientation but not the short-range specific native interactions. Subsequently conformational rearrangement leads to the formation of the native complex. We found that the changes in relative orientations between the two receptor molecules from the transient complex to the 1∶2 native complex are similar for the three cytokine-receptor systems. We thus propose a common model for receptor activation by class I cytokines, involving combined scissor-like rotation and self-rotation of the two receptor molecules. Both types of rotations seem essential: the scissor-like rotation separates the intracellular domains of the two receptor molecules to make room for the associated Janus kinase molecules, while the self-rotation allows them to orient properly for transphosphorylation. This activation model explains a host of experimental observations. The transient-complex based approach presented here may provide a strategy for designing antagonists and prove useful for elucidating activation mechanisms of other receptors.

  8. How to use D-dimer in acute cardiovascular care

    DEFF Research Database (Denmark)

    Giannitsis, Evangelos; Mair, Johannes; Christersson, Christina

    2015-01-01

    D-dimer testing is important to aid in the exclusion of venous thromboembolic events (VTEs), including deep venous thrombosis and pulmonary embolism, and it may be used to evaluate suspected aortic dissection. D-dimer is produced upon activation of the coagulation system with the generation and s...... testing. For the exclusion of pulmonary embolism/deep vein thrombosis, age-adjusted cut-offs are recommend. Clinicians must be aware of the validated use of their hospital's D-dimer assay to avoid inappropriate use of this biomarker in routine care....

  9. Dimeric Surfactants: Promising Ingredients of Cosmetics and Toiletries

    Directory of Open Access Journals (Sweden)

    Naveen Kumar

    2013-11-01

    Full Text Available Surfactants are an essential ingredient for cosmetic, toiletries and personal care products for enhancing their performance. Dimeric surfactants demonstrate superiority compared to conventional surfactants in all areas of application. Dimeric surfactants are extremely promising for utilization in various cosmetic formulations viz. shampoo, lotions, creams, conditioners etc. These surfactants possess extremely unique surface properties viz. lower surface tension, unique micellization, low critical micelle concentration (CMC and antimicrobial activity, higher solubilization etc. Dimerics enhance the performances of cosmetics in an extraordinary manner and provide eco-friendly preparations for human epidermis.

  10. Rapid dimerization of quercetin through an oxidative mechanism in the presence of serum albumin decreases its ability to induce cytotoxicity in MDA-MB-231 cells

    Energy Technology Data Exchange (ETDEWEB)

    Pham, Anh; Bortolazzo, Anthony [Department of Biological Sciences, San Jose State University, San Jose, CA 95192-0100 (United States); White, J. Brandon, E-mail: Brandon.White@sjsu.edu [Department of Biological Sciences, San Jose State University, San Jose, CA 95192-0100 (United States)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Quercetin cannot be detected intracellularly despite killing MDA-MB-231 cells. Black-Right-Pointing-Pointer Quercetin forms a heterodimer through oxidation in media with serum. Black-Right-Pointing-Pointer The quercetin heterodimer does not kill MDA-MB-231 cells. Black-Right-Pointing-Pointer Ascorbic acid stabilizes quercetin increasing cell death in quercetin treated cells. Black-Right-Pointing-Pointer Quercetin, and not a modified form, is responsible for apoptosis and cell death. -- Abstract: Quercetin is a member of the flavonoid family and has been previously shown to have a variety of anti-cancer activities. We and others have reported anti-proliferation, cell cycle arrest, and induction of apoptosis of cancer cells after treatment with quercetin. Quercetin has also been shown to undergo oxidation. However, it is unclear if quercetin or one of its oxidized forms is responsible for cell death. Here we report that quercetin rapidly oxidized in cell culture media to form a dimer. The quercetin dimer is identical to a dimer that is naturally produced by onions. The quercetin dimer and quercetin-3-O-glucopyranoside are unable to cross the cell membrane and do not kill MDA-MB-231 cells. Finally, supplementing the media with ascorbic acid increases quercetin's ability to induce cell death probably by reduction oxidative dimerization. Our results suggest that an unmodified quercetin is the compound that elicits cell death.

  11. Transparent form-active system with structural glass

    NARCIS (Netherlands)

    Nikolaou, M.S.N.; Veer, F.A.; Eigenraam, P.

    2015-01-01

    Free-form transparent wide-span spatial structures which have being constructed so far, are based on the concept of three sets of components, the structural components, usually steel elements to ensure both compressive and tensional capacity; the glass cladding elements for expressing transparency;

  12. Universal dimer-dimer scattering in lattice effective field theory

    CERN Document Server

    Elhatisari, Serdar; Lee, Dean; Meißner, Ulf-G; Rupak, Gautam

    2016-01-01

    We consider two-component fermions with short-range interactions and large scattering length. This system has universal properties that are realized in many different fields including atomic, nuclear and particle physics. In the limit of large fermion-fermion scattering length $a_\\mathrm{ff}$ and zero range interaction, all properties of the system scale proportionally with the only length scale $a_\\mathrm{ff}$. We consider the case where there are bound dimers and calculate the scattering phase shifts for the two-dimer system near threshold using lattice effective field theory. From the scattering phase shifts, we extract the universal dimer-dimer scattering length $a_\\mathrm{dd}/a_\\mathrm{ff}=0.645(89)$ and effective range $r_\\mathrm{dd}/a_\\mathrm{ff}=-0.413(79)$.

  13. A purified truncated form of yeast Gal4 expressed in Escherichia coli and used to functionalize poly(lactic acid) nanoparticle surface is transcriptionally active in cellulo.

    Science.gov (United States)

    Legaz, Sophie; Exposito, Jean-Yves; Borel, Agnès; Candusso, Marie-Pierre; Megy, Simon; Montserret, Roland; Lahaye, Vincent; Terzian, Christophe; Verrier, Bernard

    2015-09-01

    Gal4/UAS system is a powerful tool for the analysis of numerous biological processes. Gal4 is a large yeast transcription factor that activates genes including UAS sequences in their promoter. Here, we have synthesized a minimal form of Gal4 DNA sequence coding for the binding and dimerization regions, but also part of the transcriptional activation domain. This truncated Gal4 protein was expressed as inclusion bodies in Escherichia coli. A structured and active form of this recombinant protein was purified and used to cover poly(lactic acid) (PLA) nanoparticles. In cellulo, these Gal4-vehicles were able to activate the expression of a Green Fluorescent Protein (GFP) gene under the control of UAS sequences, demonstrating that the decorated Gal4 variant can be delivery into cells where it still retains its transcription factor capacities. Thus, we have produced in E. coli and purified a short active form of Gal4 that retains its functions at the surface of PLA-nanoparticles in cellular assay. These decorated Gal4-nanoparticles will be useful to decipher their tissue distribution and their potential after ingestion or injection in UAS-GFP recombinant animal models.

  14. Dimerization mediates thermo-adaptation, substrate affinity and transglycosylation in a highly thermostable maltogenic amylase of Geobacillus thermoleovorans.

    Directory of Open Access Journals (Sweden)

    Deepika Mehta

    Full Text Available BACKGROUND: Maltogenic amylases belong to a subclass of cyclodextrin-hydrolyzing enzymes and hydrolyze cyclodextrins more efficiently than starch unlike typical α-amylases. Several bacterial malto-genic amylases with temperature optima of 40-60°C have been previously characterized. The thermo-adaption, substrate preferences and transglycosylation aspects of extremely thermostable bacterial maltogenic amylases have not yet been reported. METHODOLOGY/PRINCIPAL FINDINGS: The recombinant monomeric and dimeric forms of maltogenic α-amylase (Gt-Mamy of the extremely thermophilic bacterium Geobacillus thermoleovorans are of 72.5 and 145 kDa, which are active optimally at 80°C. Extreme thermostability of this enzyme has been explained by analyzing far-UV CD spectra. Dimerization increases T1/2 of Gt-Mamy from 8.2 h to 12.63 h at 90°C and mediates its enthalpy-driven conformational thermostabilization. Furthermore, dime-rization regulates preferential substrate binding of the enzyme. The substrate preference switching of Gt-Mamy upon dimerization has been confirmed from the substrate-binding affinities of the enzyme for various high and low molecular weight substrates. There is an alteration in Km and substrate hydrolysis efficiency (Vmax/Km of the enzyme (for cyclodex-trins/starch upon dimerization. N-terminal truncation indicated the role of N-terminal 128 amino acids in the thermostabilization and modulation of substrate-binding affinity. This has been confirmed by molecular docking of β-cyclodextrin to Gt-Mamy that indicated the requirement of homodimer formation by the interaction of a few N-terminal residues of chain A with the catalytic residues of (α/β8 barrel of chain B and vice-versa for stable cyclodextrin binding. Site directed mutagenesis provided evidence for the role of N-terminal D109 at the dimeric interface in substrate affinity modulation and thermostabilization. The dimeric Gt-Mamy transglycosylates hydrolytic products of G4/G

  15. Determining equilibrium constants for dimerization reactions from molecular dynamics simulations.

    Science.gov (United States)

    De Jong, Djurre H; Schäfer, Lars V; De Vries, Alex H; Marrink, Siewert J; Berendsen, Herman J C; Grubmüller, Helmut

    2011-07-15

    With today's available computer power, free energy calculations from equilibrium molecular dynamics simulations "via counting" become feasible for an increasing number of reactions. An example is the dimerization reaction of transmembrane alpha-helices. If an extended simulation of the two helices covers sufficiently many dimerization and dissociation events, their binding free energy is readily derived from the fraction of time during which the two helices are observed in dimeric form. Exactly how the correct value for the free energy is to be calculated, however, is unclear, and indeed several different and contradictory approaches have been used. In particular, results obtained via Boltzmann statistics differ from those determined via the law of mass action. Here, we develop a theory that resolves this discrepancy. We show that for simulation systems containing two molecules, the dimerization free energy is given by a formula of the form ΔG ∝ ln(P(1) /P(0) ). Our theory is also applicable to high concentrations that typically have to be used in molecular dynamics simulations to keep the simulation system small, where the textbook dilute approximations fail. It also covers simulations with an arbitrary number of monomers and dimers and provides rigorous error estimates. Comparison with test simulations of a simple Lennard Jones system with various particle numbers as well as with reference free energy values obtained from radial distribution functions show full agreement for both binding free energies and dimerization statistics.

  16. Formation of the productive ATP-Mg2+-bound dimer of GlcV, an ABC-ATPase from Sulfolobus solfataricus

    NARCIS (Netherlands)

    Verdon, G; Albers, SV; van Oosterwijk, N; Dijkstra, BW; Driessen, AJM; Thunnissen, AMWH; Dijkstra, Bauke W.

    2003-01-01

    The ABC-ATPase GlcV from Sulfolobus solfataricus energizes an ABC transporter mediating glucose uptake. In ABC transporters, two ABC-ATPases are believed to form a head-to-tail dimer, with both monomers contributing conserved residues to each of the two productive active sites. In contrast, isolated

  17. Dimerization of a Viral SET Protein Endows its Function

    Energy Technology Data Exchange (ETDEWEB)

    H Wei; M Zhou

    2011-12-31

    Histone modifications are regarded as the most indispensible phenomena in epigenetics. Of these modifications, lysine methylation is of the greatest complexity and importance as site- and state-specific lysine methylation exerts a plethora of effects on chromatin structure and gene transcription. Notably, paramecium bursaria chlorella viruses encode a conserved SET domain methyltransferase, termed vSET, that functions to suppress host transcription by methylating histone H3 at lysine 27 (H3K27), a mark for eukaryotic gene silencing. Unlike mammalian lysine methyltransferases (KMTs), vSET functions only as a dimer, but the underlying mechanism has remained elusive. In this study, we demonstrate that dimeric vSET operates with negative cooperativity between the two active sites and engages in H3K27 methylation one site at a time. New atomic structures of vSET in the free form and a ternary complex with S-adenosyl homocysteine and a histone H3 peptide and biochemical analyses reveal the molecular origin for the negative cooperativity and explain the substrate specificity of H3K27 methyltransferases. Our study suggests a 'walking' mechanism, by which vSET acts all by itself to globally methylate host H3K27, which is accomplished by the mammalian EZH2 KMT only in the context of the Polycomb repressive complex.

  18. How Lipid Membranes Affect Pore Forming Toxin Activity.

    Science.gov (United States)

    Rojko, Nejc; Anderluh, Gregor

    2015-12-15

    Pore forming toxins (PFTs) evolved to permeate the plasma membrane of target cells. This is achieved in a multistep mechanism that usually involves binding of soluble protein monomer to the lipid membrane, oligomerization at the plane of the membrane, and insertion of part of the polypeptide chain across the lipid membrane to form a conductive channel. Introduced pores allow uncontrolled transport of solutes across the membrane, inflicting damage to the target cell. PFTs are usually studied from the perspective of structure-function relationships, often neglecting the important role of the bulk membrane properties on the PFT mechanism of action. In this Account, we discuss how membrane lateral heterogeneity, thickness, and fluidity influence the pore forming process of PFTs. In general, lipid molecules are more accessible for binding in fluid membranes due to steric reasons. When PFT specifically binds ordered domains, it usually recognizes a specific lipid distribution pattern, like sphingomyelin (SM) clusters or SM/cholesterol complexes, and not individual lipid species. Lipid domains were also suggested to act as an additional concentration platform facilitating PFT oligomerization, but this is yet to be shown. The last stage in PFT action is the insertion of the transmembrane segment across the membranes to build the transmembrane pore walls. Conformational changes are a spontaneous process, and sufficient free energy has to be available for efficient membrane penetration. Therefore, fluid bilayers are permeabilized more readily in comparison to highly ordered and thicker liquid ordered lipid phase (Lo). Energetically more costly insertion into the Lo phase can be driven by the hydrophobic mismatch between the thinner liquid disordered phase (Ld) and large protein complexes, which are unable to tilt like single transmembrane segments. In the case of proteolipid pores, membrane properties can directly modulate pore size, stability, and even selectivity. Finally

  19. Multiple forms of endopeptidase activity from jojoba seeds.

    Science.gov (United States)

    Wolf, M J; Storey, R D

    1990-01-01

    The cotyledons of 27 day post-germination jojoba seedlings (Simmondsia chinensis) contained five distinct endopeptidase activities separable by DEAE Bio-Gel and CM-cellulose ion exchange chromatography. The endopeptidases were purified 108- to 266-fold and their individuality was confirmed by activity-specific assays in native acrylamide gels along with differences in their Mr and catalytic properties. The five endopeptidases, which showed activity on model substrates and protein, were named EP Ia, EP Ib, EP II, EP III and EP IV. EP Ia was a serine proteinase with a pH optimum of ca 8 and Mr of 58,000. EP Ib, II and III were discrete cysteine proteinases showing pH optima of ca 6.8, 6.0 and 5.4 and Mr of 41,000, 47,000 and 35,000 respectively. EP IV was an aspartic acid proteinase with a ca pH optimum of 3.5 and Mr of 33,000.

  20. Conformational Heterogeneity of Bax Helix 9 Dimer for Apoptotic Pore Formation

    Science.gov (United States)

    Liao, Chenyi; Zhang, Zhi; Kale, Justin; Andrews, David W.; Lin, Jialing; Li, Jianing

    2016-07-01

    Helix α9 of Bax protein can dimerize in the mitochondrial outer membrane (MOM) and lead to apoptotic pores. However, it remains unclear how different conformations of the dimer contribute to the pore formation on the molecular level. Thus we have investigated various conformational states of the α9 dimer in a MOM model — using computer simulations supplemented with site-specific mutagenesis and crosslinking of the α9 helices. Our data not only confirmed the critical membrane environment for the α9 stability and dimerization, but also revealed the distinct lipid-binding preference of the dimer in different conformational states. In our proposed pathway, a crucial iso-parallel dimer that mediates the conformational transition was discovered computationally and validated experimentally. The corroborating evidence from simulations and experiments suggests that, helix α9 assists Bax activation via the dimer heterogeneity and interactions with specific MOM lipids, which eventually facilitate proteolipidic pore formation in apoptosis regulation.

  1. Rotational spectra of propargyl alcohol dimer: A dimer bound with three different types of hydrogen bonds

    Energy Technology Data Exchange (ETDEWEB)

    Mani, Devendra; Arunan, E., E-mail: arunan@ipc.iisc.ernet.in [Department of Inorganic and Physical Chemistry, Indian Institute of Science, Bangalore 560012 (India)

    2014-10-28

    Pure rotational spectra of the propargyl alcohol dimer and its three deuterium isotopologues have been observed in the 4 to 13 GHz range using a pulsed-nozzle Fourier transform microwave spectrometer. For the parent dimer, a total of 51 transitions could be observed and fitted within experimental uncertainty. For two mono-substituted and one bi-substituted deuterium isotopologues, a total of 14, 17, and 19 transitions were observed, respectively. The observed rotational constants for the parent dimer [A = 2321.8335(4) MHz, B = 1150.4774(2) MHz, and C = 1124.8898(2) MHz] are close to those of the most stable structure predicted by ab initio calculations. Spectra of the three deuterated isotopologues and Kraitchman analysis positively confirm this structure. Geometrical parameters and “Atoms in Molecules” analysis on the observed structure reveal that the two propargyl alcohol units in the dimer are bound by three different types of hydrogen bonds: O–H⋯O, O–H⋯π, and C–H⋯π. To the best of our knowledge, propargyl alcohol seems to be the smallest molecule forming a homodimer with three different points of contact.

  2. A High-Resolution Crystal Structure of a Psychrohalophilic α–Carbonic Anhydrase from Photobacterium profundum Reveals a Unique Dimer Interface

    Energy Technology Data Exchange (ETDEWEB)

    Somalinga, Vijayakumar; Buhrman, Greg; Arun, Ashikha; Rose, Robert B.; Grunden, Amy M. (NCSU)

    2016-12-09

    Bacterial α–carbonic anhydrases (α-CA) are zinc containing metalloenzymes that catalyze the rapid interconversion of CO2 to bicarbonate and a proton. We report the first crystal structure of a pyschrohalophilic α–CA from a deep-sea bacterium, Photobacterium profundum. Size exclusion chromatography of the purified P. profundum α–CA (PprCA) reveals that the protein is a heterogeneous mix of monomers and dimers. Furthermore, an “in-gel” carbonic anhydrase activity assay, also known as protonography, revealed two distinct bands corresponding to monomeric and dimeric forms of PprCA that are catalytically active. The crystal structure of PprCA was determined in its native form and reveals a highly conserved “knot-topology” that is characteristic of α–CA’s. Similar to other bacterial α–CA’s, PprCA also crystallized as a dimer. Furthermore, dimer interface analysis revealed the presence of a chloride ion (Cl-) in the interface which is unique to PprCA and has not been observed in any other α–CA’s characterized so far. Molecular dynamics simulation and chloride ion occupancy analysis shows 100% occupancy for the Cl- ion in the dimer interface. Zinc coordinating triple histidine residues, substrate binding hydrophobic patch residues, and the hydrophilic proton wire residues are highly conserved in PprCA and are identical to other well-studied α–CA’s.

  3. Influence of point mutations on the stability, dimerization and oligomerization of human cystatin C and its L68Q variant

    Directory of Open Access Journals (Sweden)

    Aneta eSzymanska

    2012-07-01

    Full Text Available Human cystatin C (hCC is a small but very intriguing protein. Produced by all nucleated cells is found in almost all tissues and body fluids where, at physiological conditions, plays a role of a very potent inhibitor of cysteine proteases. Biologically active hCC is a monomeric protein but during cellular trafficking it forms dimers, transiently loosing its inhibitory activity. In vitro, dimerization of cystatin C was observed for the mature protein during crystallization trials, revealing that the mechanism of this process is based on the three dimensional swapping of the protein domains. In our work we have focused on the impact of two proposed hot spots in cystatin C structure on its conformational stability. Encouraged by promising results of the theoretical calculations, we designed and produced several hCC hinge region point mutation variants that display a variety of conformational stability and propensity for dimerization and aggregation. A similar approach, i.e. rational mutagenesis, has been also applied to study the amyloidogenic L68Q variant to determine the contribution of hydrophobic interactions and steric effect on the stability of monomeric cystatin C. In this overview we would like to summarize the results of our studies. The impact of a particular mutation on the properties of the studied proteins will be presented in the context of their thermal and mechanical stability, in vitro dimerization tendency as well as the outcome of crystallization. Better understanding of the mechanism and, especially, factors affecting conformational stability of cystatin C and access to stable monomeric and dimeric versions of the protein opens new perspectives in explaining the role of dimers and the domain swapping process in hCC oligomerization, as well as designing potential inhibitors of this process.

  4. Tectonic Activity on Pluto After the Charon-Forming Impact

    CERN Document Server

    Barr, Amy C

    2014-01-01

    The Pluto-Charon system, likely formed from an impact, has reached the endpoint of its tidal evolution. During its evolution into the dual-synchronous state, the equilibrium tidal figures of Pluto and Charon would have also evolved as angular momentum was transferred from Pluto's spin to Charon's orbit. The rate of tidal evolution is controlled by Pluto's interior physical and thermal state. We examine three interior models for Pluto: an undifferentiated rock/ice mixture, differentiated with ice above rock, and differentiated with an ocean. For the undifferentiated case without an ocean, the Pluto-Charon binary does not evolve to its current state unless its internal temperature $T_i>200$ K, which would likely lead to strong tidal heating, melting, and differentiation. Without an ocean, Pluto's interior temperature must be higher than 240 K for Charon to evolve on a time scale less than the age of the solar system. Further tidal heating would likely create an ocean. If New Horizons finds evidence of ancient t...

  5. Heat of Hydration of Low Activity Cementitious Waste Forms

    Energy Technology Data Exchange (ETDEWEB)

    Nasol, D. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-07-23

    During the curing of secondary waste grout, the hydraulic materials in the dry mix react exothermally with the water in the secondary low-activity waste (LAW). The heat released, called the heat of hydration, can be measured using a TAM Air Isothermal Calorimeter. By holding temperature constant in the instrument, the heat of hydration during the curing process can be determined. This will provide information that can be used in the design of a waste solidification facility. At the Savannah River National Laboratory (SRNL), the heat of hydration and other physical properties are being collected on grout prepared using three simulants of liquid secondary waste generated at the Hanford Site. From this study it was found that both the simulant and dry mix each had an effect on the heat of hydration. It was also concluded that the higher the cement content in the dry materials mix, the greater the heat of hydration during the curing of grout.

  6. Surface active complexes formed between keratin polypeptides and ionic surfactants.

    Science.gov (United States)

    Pan, Fang; Lu, Zhiming; Tucker, Ian; Hosking, Sarah; Petkov, Jordan; Lu, Jian R

    2016-12-15

    Keratins are a group of important proteins in skin and hair and as biomaterials they can provide desirable properties such as strength, biocompatibility, and moisture regaining and retaining. The aim of this work is to develop water-soluble keratin polypeptides from sheep wool and then explore how their surface adsorption behaves with and without surfactants. Successful preparation of keratin samples was demonstrated by identification of the key components from gel electrophoresis and the reproducible production of gram scale samples with and without SDS (sodium dodecylsulphate) during wool fibre dissolution. SDS micelles could reduce the formation of disulphide bonds between keratins during extraction, reducing inter-molecular crosslinking and improving keratin polypeptide solubility. However, Zeta potential measurements of the two polypeptide batches demonstrated almost identical pH dependent surface charge distributions with isoelectric points around pH 3.5, showing complete removal of SDS during purification by dialysis. In spite of different solubility from the two batches of keratin samples prepared, very similar adsorption and aggregation behavior was revealed from surface tension measurements and dynamic light scattering. Mixing of keratin polypeptides with SDS and C12TAB (dodecyltrimethylammonium bromide) led to the formation of keratin-surfactant complexes that were substantially more effective at reducing surface tension than the polypeptides alone, showing great promise in the delivery of keratin polypeptides via the surface active complexes. Neutron reflection measurements revealed the coexistence of surfactant and keratin polypeptides at the interface, thus providing the structural support to the observed surface tension changes associated with the formation of the surface active complexes.

  7. Wound Healing Activity of Topical Application Forms Based on Ayurveda

    Directory of Open Access Journals (Sweden)

    Hema Sharma Datta

    2011-01-01

    Full Text Available The traditional Indian medicine—Ayurveda, describes various herbs, fats, oils and minerals with anti-aging as well as wound healing properties. With aging, numerous changes occur in skin, including decrease in tissue cell regeneration, decrease in collagen content, loss of skin elasticity and mechanical strength. We prepared five topical anti-aging formulations using cow ghee, flax seed oil, Phyllanthus emblica fruits, Shorea robusta resin, Yashada bhasma as study materials. For preliminary efficacy evaluation of the anti-aging activity we chose excision and incision wound healing animal models and studied the parameters including wound contraction, collagen content and skin breaking strength which in turn is indicative of the tissue cell regeneration capacity, collagenation capacity and mechanical strength of skin. The group treated with the formulations containing Yashada bhasma along with Shorea robusta resin and flax seed oil showed significantly better wound contraction (P < .01, higher collagen content (P < .05 and better skin breaking strength (P < .01 as compared to control group; thus proposing them to be effective prospective anti-aging formulations.

  8. Thermodynamic properties for the sodium dimer

    Science.gov (United States)

    Song, Xiao-Qin; Wang, Chao-Wen; Jia, Chun-Sheng

    2017-04-01

    We present a closed-form expression of the classical vibrational partition function for the improved Rosen-Morse potential energy model. We give explicit expressions for the vibrational mean energy, vibrational specific heat, vibrational free energy, and vibrational entropy for diatomic molecule systems. The properties of these thermodynamic functions for the Na2 dimer are discussed in detail. We find that the improved Rosen-Morse potential model is superior to the harmonic oscillator in calculating the heat capacity for the Na2 molecules.

  9. Role of a Putative gp41 Dimerization Domain in Human Immunodeficiency Virus Type 1 Membrane Fusion

    Energy Technology Data Exchange (ETDEWEB)

    Liu, J.; Deng, Y; Li, Q; Dey, A; Moore, J; Lu, M

    2010-01-01

    The entry of human immunodeficiency virus type 1 (HIV-1) into a target cell entails a series of conformational changes in the gp41 transmembrane glycoprotein that mediates the fusion of the viral and target cell membranes. A trimer-of-hairpins structure formed by the association of two heptad repeat (HR) regions of the gp41 ectodomain has been implicated in a late step of the fusion pathway. Earlier native and intermediate states of the protein are postulated to mediate the antiviral activity of the fusion inhibitor enfuvirtide and of broadly neutralizing monoclonal antibodies (NAbs), but the details of these structures remain unknown. Here, we report the identification and crystal structure of a dimerization domain in the C-terminal ectodomain of gp41 (residues 630 to 683, or C54). Two C54 monomers associate to form an asymmetric, antiparallel coiled coil with two distinct C-terminal {alpha}-helical overhangs. This dimer structure is conferred largely by interactions within a central core that corresponds to the sequence of enfuvirtide. The mutagenic alteration of the dimer interface severely impairs the infectivity of Env-pseudotyped viruses. Moreover, the C54 structure binds tightly to both the 2F5 and 4E10 NAbs and likely represents a potential intermediate conformation of gp41. These results should enhance our understanding of the molecular basis of the gp41 fusogenic structural transitions and thereby guide rational, structure-based efforts to design new fusion inhibitors and vaccine candidates intended to induce broadly neutralizing antibodies.

  10. 氧化偶联反应和酸催化反应制备活性买麻藤醇二聚体衍生物%Preparation of Active Gnetol Dimers by Oxidative Coupling Reaction and Acid-Catalyzed Dimerization

    Institute of Scientific and Technical Information of China (English)

    姚春所; 林茂; 杨庆云

    2013-01-01

    Oxidative coupling reaction with FeCl3·6H2O as oxidant and acid-catalyzed dimerization of natural gnetol in methanol afforded two new gnetol dimers and one new phenyl naphthalene derivative: 4-[1-(2,6-dihydroxyphenyl)-2-(3,5-dihydroxyphenyl)ethyl]-2-[(lE)-2-(3,5-dihydroxyphenyl)ethenyl]-1,3-benzenediol (1),2-[1-(2,6-dihydroxyphenyl)-2-(3,5-di-hydroxyphenyl)ethyl]-5-f(1E)-2-(2,6-dihydroxyphenyl)ethenyl]-1,3-benzenediol (2) and 4-(6,8-dimethoxyl-2-naphthalenyl)-1,3-benzenediol (3). Their structures were elucidated on the basis of spectral analysis,and their possible formation mechanisms were discussed. 1 and 2 were new linear stilbene dimers synthesized for the first time. Pharmacological tests showed 1,2 and 3 to exhibit potent anti-oxidation activity with IC50 values of 6.29×10-9,4.19× 10-6,and 2.96×10-5 mol·L-1,respectively,and 2 was shown to have potent anti-inflammatory activity.%以天然得到的买麻藤醇为原料,以FeCl3·6H2O为氧化剂进行氧化偶联反应和酸催化二聚反应,获得了2个新的买麻藤醇二聚体及一个新的苯基萘衍生物:4-[1-(2,6-二羟基苯基)-2-(3,5-二羟基苯基)乙基]-2-[(1E)-2-(3,5-二 羟基苯基)乙烯基]-1,3-苯二醇(1),2-[1-(2,6-二羟基苯基)-2-(3,5-二羟基苯基)乙基]-5-[(1E)2-(2,6-二羟基苯基)乙烯基]-1,3-苯二醇(2)和4-(6,8-二甲氧基-2-萘基)-1,3-苯二醇(3).应用波谱分析的方法确定了它们的结构,并分别讨论了它们可能的形成机理.其中,化合物1和2首次为人工合成的二苯乙烯链状二聚体.活性测试结果表明,化合物1,2和3显示有较强的抗氧化活性,其IC50值分别为6.29×10-9,4.19×10-6和2.96×10-5 mol·L-1;化合物2还显示有较强的抗炎活性.

  11. Near-ultraviolet tyrosyl circular dichroism of pig insulin monomers, dimers, and hexamers. Dipole-dipole coupling calculations in the monopole approximation.

    Science.gov (United States)

    Strickland, E H; Mercola, D

    1976-08-24

    The tyrosyl circular dichroism (CD) has been calculated using the conformation of pig insulin observed in rhombohedral crystals containing 2 zinc atoms per hexamer. These calculations predict that the tyrosyl CD at 275 nm will be enhanced disproportionally as monomers interact to form dimers and as dimers interact to form hexamers. This enhanced tyrosyl CD (delta epsilon per 5800 molecular weight) results from new coupling interactions generated in the regions of contact between monomers and between dimers. These calculations illustrate that a large CD enhancement may accompany aggregation even in the absence of a conformation change in eith monomer. The tyrosyl CD intensities calculated for monomers, dimers, and hexamers of 2-zinc pig insulin are compatible with the experimentally observed CD spectra which are enhanced about fourfold in the hexamer compared with the monomer. Zinc ions and other metals do not contribute directly to the tyrosyl CD but only influence the optical properties by promoting the hexameric state. The relation of the integrity of the molecule to dimer formation and the biological activity of the molecules are discussed. The largest calculated contributions to tryosyl CD arise from interactions with far-ultraviolet transitions of neighboring aromatic groups. In the hexamer, about half of the tyrosyl CD intensity is calculated to arise from Tyr-A14.

  12. Dimeric interactions and complex formation using direct coevolutionary couplings.

    Science.gov (United States)

    dos Santos, Ricardo N; Morcos, Faruck; Jana, Biman; Andricopulo, Adriano D; Onuchic, José N

    2015-09-04

    We develop a procedure to characterize the association of protein structures into homodimers using coevolutionary couplings extracted from Direct Coupling Analysis (DCA) in combination with Structure Based Models (SBM). Identification of dimerization contacts using DCA is more challenging than intradomain contacts since direct couplings are mixed with monomeric contacts. Therefore a systematic way to extract dimerization signals has been elusive. We provide evidence that the prediction of homodimeric complexes is possible with high accuracy for all the cases we studied which have rich sequence information. For the most accurate conformations of the structurally diverse dimeric complexes studied the mean and interfacial RMSDs are 1.95Å and 1.44Å, respectively. This methodology is also able to identify distinct dimerization conformations as for the case of the family of response regulators, which dimerize upon activation. The identification of dimeric complexes can provide interesting molecular insights in the construction of large oligomeric complexes and be useful in the study of aggregation related diseases like Alzheimer's or Parkinson's.

  13. Adventures in Holographic Dimer Models

    Energy Technology Data Exchange (ETDEWEB)

    Kachru, Shamit; /Stanford U., Phys. Dept. /SLAC; Karch, Andreas; /Washington U., Seattle; Yaida, Sho; /Stanford U., Phys. Dept.

    2011-08-12

    We abstract the essential features of holographic dimer models, and develop several new applications of these models. Firstly, semi-holographically coupling free band fermions to holographic dimers, we uncover novel phase transitions between conventional Fermi liquids and non-Fermi liquids, accompanied by a change in the structure of the Fermi surface. Secondly, we make dimer vibrations propagate through the whole crystal by way of double trace deformations, obtaining nontrivial band structure. In a simple toy model, the topology of the band structure experiences an interesting reorganization as we vary the strength of the double trace deformations. Finally, we develop tools that would allow one to build, in a bottom-up fashion, a holographic avatar of the Hubbard model.

  14. Monomer-dimer equilibrium in glutathione transferases: a critical re-examination.

    Science.gov (United States)

    Fabrini, Raffaele; De Luca, Anastasia; Stella, Lorenzo; Mei, Giampiero; Orioni, Barbara; Ciccone, Sarah; Federici, Giorgio; Lo Bello, Mario; Ricci, Giorgio

    2009-11-03

    Glutathione transferases (GSTs) are dimeric enzymes involved in cell detoxification versus many endogenous toxic compounds and xenobiotics. In addition, single monomers of GSTs appear to be involved in particular protein-protein interactions as in the case of the pi class GST that regulates the apoptotic process by means of a GST-c-Jun N-terminal kinase complex. Thus, the dimer-monomer transition of GSTs may have important physiological relevance, but many studies reached contrasting conclusions both about the modality and extension of this event and about the catalytic competence of a single subunit. This paper re-examines the monomer-dimer question in light of novel experiments and old observations. Recent papers claimed the existence of a predominant monomeric and active species among pi, alpha, and mu class GSTs at 20-40 nM dilution levels, reporting dissociation constants (K(d)) for dimeric GST of 5.1, 0.34, and 0.16 microM, respectively. However, we demonstrate here that only traces of monomers could be found at these concentrations since all these enzymes display K(d) values of <1 nM, values thousands of times lower than those reported previously. Time-resolved and steady-state fluorescence anisotropy experiments, two-photon fluorescence correlation spectroscopy, kinetic studies, and docking simulations have been used to reach such conclusions. Our results also indicate that there is no clear evidence of the existence of a fully active monomer. Conversely, many data strongly support the idea that the monomeric form is scarcely active or fully inactive.

  15. Urinary levels of thymine dimer as a biomarker of exposure to ultraviolet radiation in humans during outdoor activities in the summer.

    Science.gov (United States)

    Liljendahl, Tove Sandberg; Blomqvist, Anna; Andersson, Eva M; Barregard, Lars; Segerbäck, Dan

    2013-05-01

    The incidence of skin cancer is rising rapidly in many countries, presumably due to increased leisure time exposure to solar ultraviolet radiation (UVR). UVR causes DNA lesions, such as the thymine dimer (T=T), which have been causatively linked to the development of skin cancer. T=T is clearly detectable in urine and may, thereby, be a potentially valuable biomarker of UVR exposure. The objective of this study was to evaluate the relationship between UVR exposure and urinary levels of T=T in a field study involving outdoor workers. Daily ambient and personal exposure of 52 beach lifeguards and agricultural workers to UVR were determined (employing 656 personal polysulphone dosimeters). In 22 of these subjects, daily urinary T=T levels (120 samples) were measured, the area of skin exposed calculated and associations assessed utilizing mixed statistical models. The average daily UVR dose was approximately 600 J/m(2) (7.7 standard erythemal doses), i.e. about 20% of ambient UVR. T=T levels were correlated to UVR dose, increasing by about 6 fmol/µmol creatinine for each 100 J/m(2) increase in dose (average of the three preceding days). This is the first demonstration of a relationship between occupational UVR exposure and urinary levels of a biomarker of DNA damage. On a population level, urinary levels of T=T can be used as a biomarker for UVR exposure in the field.

  16. New Insights into Molecular Organization of Human Neuraminidase-1: Transmembrane Topology and Dimerization Ability

    Science.gov (United States)

    Maurice, Pascal; Baud, Stéphanie; Bocharova, Olga V.; Bocharov, Eduard V.; Kuznetsov, Andrey S.; Kawecki, Charlotte; Bocquet, Olivier; Romier, Beatrice; Gorisse, Laetitia; Ghirardi, Maxime; Duca, Laurent; Blaise, Sébastien; Martiny, Laurent; Dauchez, Manuel; Efremov, Roman G.; Debelle, Laurent

    2016-01-01

    Neuraminidase 1 (NEU1) is a lysosomal sialidase catalyzing the removal of terminal sialic acids from sialyloconjugates. A plasma membrane-bound NEU1 modulating a plethora of receptors by desialylation, has been consistently documented from the last ten years. Despite a growing interest of the scientific community to NEU1, its membrane organization is not understood and current structural and biochemical data cannot account for such membrane localization. By combining molecular biology and biochemical analyses with structural biophysics and computational approaches, we identified here two regions in human NEU1 - segments 139–159 (TM1) and 316–333 (TM2) - as potential transmembrane (TM) domains. In membrane mimicking environments, the corresponding peptides form stable α-helices and TM2 is suited for self-association. This was confirmed with full-size NEU1 by co-immunoprecipitations from membrane preparations and split-ubiquitin yeast two hybrids. The TM2 region was shown to be critical for dimerization since introduction of point mutations within TM2 leads to disruption of NEU1 dimerization and decrease of sialidase activity in membrane. In conclusion, these results bring new insights in the molecular organization of membrane-bound NEU1 and demonstrate, for the first time, the presence of two potential TM domains that may anchor NEU1 in the membrane, control its dimerization and sialidase activity. PMID:27917893

  17. New Insights into Molecular Organization of Human Neuraminidase-1: Transmembrane Topology and Dimerization Ability

    Science.gov (United States)

    Maurice, Pascal; Baud, Stéphanie; Bocharova, Olga V.; Bocharov, Eduard V.; Kuznetsov, Andrey S.; Kawecki, Charlotte; Bocquet, Olivier; Romier, Beatrice; Gorisse, Laetitia; Ghirardi, Maxime; Duca, Laurent; Blaise, Sébastien; Martiny, Laurent; Dauchez, Manuel; Efremov, Roman G.; Debelle, Laurent

    2016-12-01

    Neuraminidase 1 (NEU1) is a lysosomal sialidase catalyzing the removal of terminal sialic acids from sialyloconjugates. A plasma membrane-bound NEU1 modulating a plethora of receptors by desialylation, has been consistently documented from the last ten years. Despite a growing interest of the scientific community to NEU1, its membrane organization is not understood and current structural and biochemical data cannot account for such membrane localization. By combining molecular biology and biochemical analyses with structural biophysics and computational approaches, we identified here two regions in human NEU1 - segments 139-159 (TM1) and 316-333 (TM2) - as potential transmembrane (TM) domains. In membrane mimicking environments, the corresponding peptides form stable α-helices and TM2 is suited for self-association. This was confirmed with full-size NEU1 by co-immunoprecipitations from membrane preparations and split-ubiquitin yeast two hybrids. The TM2 region was shown to be critical for dimerization since introduction of point mutations within TM2 leads to disruption of NEU1 dimerization and decrease of sialidase activity in membrane. In conclusion, these results bring new insights in the molecular organization of membrane-bound NEU1 and demonstrate, for the first time, the presence of two potential TM domains that may anchor NEU1 in the membrane, control its dimerization and sialidase activity.

  18. Dimer and String Formation during Low Temperature Silicon Deposition on Si(100)

    DEFF Research Database (Denmark)

    Smith, A. P.; Jonsson, Hannes

    1996-01-01

    We present theoretical results based on density functional theory and kinetic Monte Carlo simulations of silicon deposition and address observations made in recently reported low temperature scanning tunneling microscopy studies. A mechanism is presented which explains dimer formation on top...... of the substrate's dimer rows at 160 K and up to room temperature, while between-row dimers and longer strings of adatoms (''diluted dimer rows'') form at higher temperature. A crossover occurs at around room temperature between two different mechanisms for adatom diffusion in our model....

  19. Radiation-induced tetramer-to-dimer transition of Escherichia coli lactose repressor

    Energy Technology Data Exchange (ETDEWEB)

    Goffinont, S. [Centre de Biophysique Moleculaire, CNRS, rue C. Sadron, 45071 Orleans (France); Davidkova, M. [Department of Radiation Dosimetry, Nuclear Physics Institute AS CR, Na Truhlarce 39/64, 18086, Prague 8 (Czech Republic); Spotheim-Maurizot, M., E-mail: spotheim@cnrs-orleans.fr [Centre de Biophysique Moleculaire, CNRS, rue C. Sadron, 45071 Orleans (France)

    2009-08-21

    The wild type lactose repressor of Escherichia coli is a tetrameric protein formed by two identical dimers. They are associated via a C-terminal 4-helix bundle (called tetramerization domain) whose stability is ensured by the interaction of leucine zipper motifs. Upon in vitro {gamma}-irradiation the repressor losses its ability to bind the operator DNA sequence due to damage of its DNA-binding domains. Using an engineered dimeric repressor for comparison, we show here that irradiation induces also the change of repressor oligomerisation state from tetramer to dimer. The splitting of the tetramer into dimers can result from the oxidation of the leucine residues of the tetramerization domain.

  20. Vortexlike topological defects in nematic colloids: chiral colloidal dimers and 2D crystals.

    Science.gov (United States)

    Tkalec, U; Ravnik, M; Zumer, S; Musevic, I

    2009-09-18

    We show that chiral ordering of the underlying complex fluid strongly influences defect formation and colloidal interactions. Nonsingular defect loops with a topological charge -2 are observed, with a cross section identical to hyperbolic vortices in magnetic systems. These loops are binding spontaneously formed pairs of colloidal particles and dimers, which are chiral objects. Chiral dimer-dimer interaction weakly depends on the chirality of dimers and leads to the assembly of 2D nematic colloidal crystals of pure or "mixed" chirality, intercalated with a lattice of nonsingular vortexlike defects.

  1. 75 FR 22438 - Proposed Information Collection (Health Resource Center Medical Center Payment Form) Activity...

    Science.gov (United States)

    2010-04-28

    ... AFFAIRS Proposed Information Collection (Health Resource Center Medical Center Payment Form) Activity... information technology. Title: Health Resource Center Medical Center Payment Form, VA Form 10-0505. OMB... proposed collection of certain information by the agency. Under the Paperwork Reduction Act (PRA) of...

  2. Reversible Dimerization of Acid-Denatured ACBP Controlled by Helix A4

    DEFF Research Database (Denmark)

    Fieber, Wolfgang; Kragelund, Birthe Brandt; Meldal, Morten Peter;

    2005-01-01

    of dimers and revealed a cooperative stabilization of helix A4 in this process. This emphasizes its special role in the structure formation in the denatured state of ACBP. No dimers are formed in the presence of guanidine hydrochloride, which underlines the fundamental difference between the nature...

  3. Dimeric Labdane Diterpenes: Synthesis and Antiproliferative Effects

    Directory of Open Access Journals (Sweden)

    Guillermo Schmeda-Hirschmann

    2013-05-01

    Full Text Available Several diterpenes with the labdane skeleton show biological activity, including antiproliferative effects. Most of the research work on bioactive labdanes has been carried out on naturally occurring diterpenes and semisynthetic derivatives, but much less is known on the effects of diterpene dimers. The aim of the present work was to synthesize dimeric diterpenes from the labdane imbricatolic acid using esters, ethers and the triazole ring as linkers. Some 18 new derivatives were prepared and the compounds were evaluated for antiproliferative activity on human normal fibroblasts (MRC-5 and the following human tumor cell lines: AGS, SK-MES-1, J82 and HL-60. The diethers 8–10, differing in the number of CH2 units in the linker, presented better antiproliferative activity with a maximum effect for the derivative 9. The best antiproliferative effect against HL-60 cells was found for compounds 3 and 17, with IC50 values of 22.3 and 23.2 μM, lower than that found for the reference compound etoposide (2.23 μM. The compounds 9, 17 and 11 were the most active derivatives towards AGS cells with IC50 values of 17.8, 23.4 and 26.1 μM. A free carboxylic acid function seems relevant for the effect as several of the compounds showed less antiproliferative effect after methylation.

  4. Evidence that levels of the dimeric cellular transcription factor CP2 play little role in the activation of the HIV-1 long terminal repeat in vivo or following superinfection with herpes simplex virus type 1.

    Science.gov (United States)

    Zhong, F; Swendeman, S L; Popik, W; Pitha, P M; Sheffery, M

    1994-08-19

    The dimeric transcription factor CP2 binds a sequence element found near the transcription start site of the human immunodeficiency virus (HIV-1) long terminal repeat. Several groups have suggested that cellular factors binding this element might play a role in modulating HIV-1 promoter activity in vivo. For example, induction of latent HIV-1 gene expression in response to superinfection by herpes simplex virus type 1 (HSV-1) or cytomegalovirus is thought to be mediated, in part, by factors binding the CP2 site. In this report we began to examine directly the relationship between CP2 and expression of the HIV-1 promoter. First, we tested what effect HSV-1 infection of T cells had on the cellular levels of CP2. The results showed that HSV-1 infection led to a significant reduction in the level of CP2 DNA binding activity and protein within 20 h. Next, we tested the effect of overexpressing either the wild-type factor or a dominant negative variant of CP2 on HIV-1 promoter activity in vivo. The results showed that CP2 had little effect or slightly repressed HIV-1 promoter activity in vivo. In addition, these expression constructs had little effect on the induction of HIV-1 promoter activity elicited by HSV-1 infection.

  5. Structural studies on MRG701 chromodomain reveal a novel dimerization interface of MRG proteins in green plants

    Directory of Open Access Journals (Sweden)

    Yanchao Liu

    2016-09-01

    Full Text Available Abstract MRG proteins are conserved during evolution in fungi, flies, mammals and plants, and they can exhibit diversified functions. The animal MRGs were found to form various complexes to activate gene expression. Plant MRG1/2 and MRG702 were reported to be involved in the regulation of flowering time via binding to H3K36me3-marked flowering genes. Herein, we determined the crystal structure of MRG701 chromodomain (MRG701CD. MRG701CD forms a novel dimerization fold both in crystal and in solution. Moreover, we found that the dimerization of MRG chromodomains is conserved in green plants. Our findings may provide new insights into the mechanism of MRGs in regulation of gene expression in green plants.

  6. NMR Structural Studies on Alamethicin Dimers

    Institute of Scientific and Technical Information of China (English)

    李星

    2003-01-01

    15N labeled alamethicin dimer was synthesized. The structure and dynamics of alamethicin dimers were studied with nuclear magnetic resonance (NMR) spectroscopy. The data from 15N-labeled alamethicin dimer suggest little differences in conformation between the dimer and monomer in the Aib1-Pro14 region. Significant difference in the conformation of the C-terminus are manifest in the NH chemical shifts in the Val15-Pho20 region.

  7. Increased disulphide dimer formation of latent associated peptide fusions of TGF-β by addition of L-cystine.

    Science.gov (United States)

    Mullen, Lisa M; Adams, Gill; Chernajovsky, Yuti

    2012-10-31

    The development of novel protein therapeutics relies on the ability to express appreciable amounts of correctly folded recombinant proteins. Latent IFN-β is engineered using the latency-associated peptide (LAP) of transforming growth factor β1 (TGF-β1) to maintain IFN-β in a biologically inactive form until such time as it is released at sites of inflammation by matrix metalloproteinase activity (see Adams et al., 2003). CHO cells cultured in suspension were used for expression of latent IFN-β to allow medium scale transient transfection. However, the recombinant protein expressed in this system consisted of a mixture of properly linked disulphide dimers and monomers. The ratio of dimer:monomer produced could be significantly altered towards increased dimer production by the addition of L-cystine to the CHO culture medium. The total yield of latent IFN-β was increased by co-transfection of plasmid coding for the simian virus (SV) 40 large T antigen to the plasmid with the SV40 origin of replication expressing latent IFN-β DNA. These results provide valuable new insights for developing protocols to produce substantial quantities of latent cytokine dimers in CHO cells in suspension.

  8. Mechanism of ubiquitylation by dimeric RING ligase RNF4

    Science.gov (United States)

    Plechanovová, Anna; Jaffray, Ellis G.; McMahon, Stephen A.; Johnson, Kenneth A.; Navrátilová, Iva; Naismith, James H.; Hay, Ronald T.

    2012-01-01

    Mammalian RNF4 is a dimeric RING ubiquitin E3 ligase that ubiquitylates poly-SUMOylated proteins. We found that RNF4 bound ubiquitin-charged UbcH5a tightly but free UbcH5a weakly. To provide insight into the mechanism of RING-mediated ubiquitylation we docked the UbcH5~ubiquitin thioester onto the RNF4 RING structure. This revealed that with E2 bound to one monomer of RNF4, the thioester-linked ubiquitin could reach across the dimer to engage the other monomer. In this model the “Ile44 hydrophobic patch” of ubiquitin is predicted to engage a conserved tyrosine located at the dimer interface of the RING and mutation of these residues blocked ubiquitylation activity. Thus, dimeric RING ligases are not simply inert scaffolds that bring substrate and E2-loaded ubiquitin into close proximity. Instead, they facilitate ubiquitin transfer by preferentially binding the E2~ubiquitin thioester across the dimer and activating the thioester bond for catalysis. PMID:21857666

  9. Energy landscapes of the monomer and dimer of the Alzheimer's peptide A β (1 -28 )

    Science.gov (United States)

    Dong, Xiao; Chen, Wei; Mousseau, Normand; Derreumaux, Philippe

    2008-03-01

    The cytoxicity of Alzheimer's disease has been linked to the self-assembly of the 40 /42 amino acid of the amyloid-β (A β ) peptide into oligomers. To understand the assembly process, it is important to characterize the very first steps of aggregation at an atomic level of detail. Here, we focus on the N-terminal fragment 1-28, known to form fibrils in vitro. Circular dichroism and NMR experiments indicate that the monomer of A β (1 -28 ) is α -helical in a membranelike environment and random coil in aqueous solution. Using the activation-relaxation technique coupled with the OPEP coarse grained force field, we determine the structures of the monomer and of the dimer of A β (1 -28 ) . In agreement with experiments, we find that the monomer is predominantly random coil in character, but displays a non-negligible β -strand probability in the N-terminal region. Dimerization impacts the structure of each chain and leads to an ensemble of intertwined conformations with little β -strand content in the region Leu17-Ala21. All these structural characteristics are inconsistent with the amyloid fibril structure and indicate that the dimer has to undergo significant rearrangement en route to fibril formation.

  10. Energy landscapes of the monomer and dimer of the Alzheimer's peptide Abeta(1-28).

    Science.gov (United States)

    Dong, Xiao; Chen, Wei; Mousseau, Normand; Derreumaux, Philippe

    2008-03-28

    The cytotoxicity of Alzheimer's disease has been linked to the self-assembly of the 4042 amino acid of the amyloid-beta (Abeta) peptide into oligomers. To understand the assembly process, it is important to characterize the very first steps of aggregation at an atomic level of detail. Here, we focus on the N-terminal fragment 1-28, known to form fibrils in vitro. Circular dichroism and NMR experiments indicate that the monomer of Abeta(1-28) is alpha-helical in a membranelike environment and random coil in aqueous solution. Using the activation-relaxation technique coupled with the OPEP coarse grained force field, we determine the structures of the monomer and of the dimer of Abeta(1-28). In agreement with experiments, we find that the monomer is predominantly random coil in character, but displays a non-negligible beta-strand probability in the N-terminal region. Dimerization impacts the structure of each chain and leads to an ensemble of intertwined conformations with little beta-strand content in the region Leu17-Ala21. All these structural characteristics are inconsistent with the amyloid fibril structure and indicate that the dimer has to undergo significant rearrangement en route to fibril formation.

  11. IR/UV and UV/UV double-resonance study of guaiacol and eugenol dimers

    Science.gov (United States)

    Longarte, Asier; Redondo, Carolina; Fernández, José A.; Castaño, Fernando

    2005-04-01

    Guaiacol (2-methoxyphenol) and eugenol (4-allyl-2-methoxyphenol) molecules are biologically active phenol derivatives with an intramolecular -OH⋯OCH3 hydrogen bond (H bond). Pulsed supersonic expansions of mixtures of either of the two molecules with He yield weakly bound homodimers as well as other higher-order complexes. A number of complementary and powerful laser spectroscopic techniques, including UV-UV and IR-UV double resonances, have been employed to interrogate the species formed in the expansion in order to get information on their structures and spectroscopic properties. The interpretation of the spectra of eugenol dimer is complex and required a previous investigation on a similar but simpler molecule both to gain insight into the possible structures and support the conclusions. Guaiacol (2-methoxyphenol) has been used for that purpose. The combination of the broad laser study combined with ab initio calculations at the Becke 3 Lee-Yang-Parr/6-31+G(d) level has provided the isomer structures, the potential-energy wells, and shed light on the inter- and intramolecular interactions involved. Guaiacol homodimer has been shown to have a single isomer whereas eugenol dimer has at least two. The comparison between the computed geometries of the dimers, their respective energies, and the vibrational normal modes permits the identification of the spectra.

  12. Stability improvement of the Nieuwland catalyst in the dimerization of acetylene to monovinylacetylene

    Institute of Scientific and Technical Information of China (English)

    Jianguo Liu; Yizan Zuo; Minghan Han; Zhanwen Wang; Dezheng Wang

    2012-01-01

    In the process of dimerization of acetylene to produce monovinylacetylene (MVA),the loss of active component CuCl in the Nieuwland catalyst due to the formation of a dark red precipitate was investigated.The formula of the precipitate was CuCl·2C2H2·1/5NH3,and it was presumed to be formed by the combination of NH3,C2H2 and [Cu]-acetylene π-complex,which was an intermediate in the dimerization reaction.The addition of hydrochloric acid into the catalyst can reduce the formation of precipitate,whereas excessive H+ is unfavorable to the dimerization reaction of acetylene.To balance between high acetylene conversion and low loss rate of CuCl,the optimum mass percentage of HCl in the added hydrochloric acid was determined.The result showed the optimum mass percentage of HCl decreased from 5.0% to 3.2% when the space velocity of acetylene was from 140 h-1 to 360 h-1.The result in this work also indicated the pH of the Nieuwland catalyst should be kept in the range of 5.80-5.97 during the reaction process,which was good for both catalyst life and acetylene conversion.

  13. Intrinsic Kinetic Modeling of Thermal Dimerization of C5 Fraction

    Institute of Scientific and Technical Information of China (English)

    Guo Liang; Wang Tiefeng; Li Dongfeng; Wang Jinfu

    2016-01-01

    This work aims to investigate the intrinsic kinetics of thermal dimerization of C5 fraction in the reactive distilla-tion process. Experiments are conducted in an 1000-mL stainless steel autoclave under some selected design conditions. By means of the weighted least squares method, the intrinsic kinetics of thermal dimerization of C5 fraction is established, and the corresponding pre-exponential factor as well as the activation energy are determined. For example, the pre-exponential factor A is equal to 4.39×105 and the activation energy Ea is equal to 6.58×104 J/mol for the cyclopentadiene dimerization re-action. The comparison between the experimental and calculated results shows that the kinetics model derived in this work is accurate and reliable, which can be used in the design of reactive distillation columns.

  14. The development and characterization of an ELISA specifically detecting the active form of cathepsin K

    DEFF Research Database (Denmark)

    Sun, S; Karsdal, M A; Bay-Jensen, A C;

    2013-01-01

    OBJECTIVE: Cathepsin K plays essential roles in bone resorption and is intensely investigated as a therapeutic target for the treatment of osteoporosis. Hence an assessment of the active form of cathepsin K may provide important biological information in metabolic bone diseases, such as osteoporo......OBJECTIVE: Cathepsin K plays essential roles in bone resorption and is intensely investigated as a therapeutic target for the treatment of osteoporosis. Hence an assessment of the active form of cathepsin K may provide important biological information in metabolic bone diseases......, such as osteoporosis or ankylosing spondylitis. METHODS: Presently there are no robust assays for the assessment of active cathepsin K in serum, and therefore an ELISA specifically detecting the N-terminal of the active form of cathepsin K was developed. RESULTS: The assay was technically robust, with a lowest limit...... form. Quantification of the levels of active cathepsin K in supernatants of purified human osteoclasts compared to corresponding macrophages showed a 30-fold induction (p...

  15. Ultrafast red light activation of Synechocystis phytochrome Cph1 triggers major structural change to form the Pfr signalling-competent state.

    Science.gov (United States)

    Heyes, Derren J; Khara, Basile; Sakuma, Michiyo; Hardman, Samantha J O; O'Cualain, Ronan; Rigby, Stephen E J; Scrutton, Nigel S

    2012-01-01

    Phytochromes are dimeric photoreceptors that regulate a range of responses in plants and microorganisms through interconversion of red light-absorbing (Pr) and far-red light-absorbing (Pfr) states. Photoconversion between these states is initiated by light-driven isomerization of a bilin cofactor, which triggers protein structural change. The extent of this change, and how light-driven structural changes in the N-terminal photosensory region are transmitted to the C-terminal regulatory domain to initiate the signalling cascade, is unknown. We have used pulsed electron-electron double resonance (PELDOR) spectroscopy to identify multiple structural transitions in a phytochrome from Synechocystis sp. PCC6803 (Cph1) by measuring distances between nitroxide labels introduced into the protein. We show that monomers in the Cph1 dimer are aligned in a parallel 'head-to-head' arrangement and that photoconversion between the Pr and Pfr forms involves conformational change in both the N- and C-terminal domains of the protein. Cryo-trapping and kinetic measurements were used to probe the extent and temporal properties of protein motions for individual steps during photoconversion of Cph1. Formation of the primary photoproduct Lumi-R is not affected by changes in solvent viscosity and dielectric constant. Lumi-R formation occurs at cryogenic temperatures, consistent with their being no major structural reorganization of Cph1 during primary photoproduct formation. All remaining steps in the formation of the Pfr state are affected by solvent viscosity and dielectric constant and occur only at elevated temperatures, implying involvement of a series of long-range solvent-coupled conformational changes in Cph1. We show that signalling is achieved through ultrafast photoisomerization where localized structural change in the GAF domain is transmitted and amplified to cause larger-scale and slower conformational change in the PHY and histidine kinase domains. This hierarchy of

  16. Molecular scaffolds with high propensity to form multi-target activity cliffs.

    Science.gov (United States)

    Hu, Ye; Bajorath, Jürgen

    2010-04-26

    In target-dependent activity landscapes of compound series, cliffs are formed by pairs of molecules that are structurally analogous but display significant differences in potency. The detection and analysis of such activity cliffs is a major task in structure-activity relationship analysis and compound optimization. In analogy to activity cliffs, selectivity cliffs can be defined that are formed by structural analogs having significantly different potencies against two targets. The formation of activity cliffs by analogs is generally a consequence of different R-group patterns; e.g., a specific substitution of a given scaffold might increase and another substitution decrease potency. Therefore, activity (or selectivity) cliffs are typically analyzed for a given scaffold representing an analog series, and it has thus far not been explored whether certain scaffolds might display a general tendency to yield compounds forming activity cliffs against different targets. We have exhaustively analyzed scaffolds and associated compound activity data in the ChemblDB and BindingDB databases in order to compare the availability of target-selective scaffolds in these databases and determine whether multi-target activity and multi-target selectivity cliff scaffolds exist. Perhaps unexpectedly, we have identified 143 scaffolds that are represented by multiple compounds and form activity or selectivity cliffs against different targets. These scaffolds have varying chemical complexities and are in part promiscuous binders (i.e., compounds containing these scaffolds bind to distantly related or unrelated targets). However, analogs derived from these scaffolds form steep activity cliffs against different targets. A catalog of scaffolds with high propensity to form activity or selectivity cliffs against multiple targets is provided to help identify potentially promiscuous candidate scaffolds during compound optimization efforts.

  17. Genetic predictors of fibrin D-dimer levels in healthy adults

    NARCIS (Netherlands)

    N.L. Smith (Nicholas); J.E. Huffman (Jennifer E.); D.P. Strachan (David); J. Huang (Jian); A. Dehghan (Abbas); S. Trompet (Stella); L.M. Lopez (Lorna M.); S.Y. Shin (So Youn); J. Baumert (Jens); V. Vitart (Veronique); J.C. Bis (Joshua); S.H. Wild (Sarah); A. Rumley (Ann); Q. Yang (Qiong Fang); A.G. Uitterlinden (André); D.J. Stott (David. J.); G. Davies (Gareth); A.M. Carter (Angela M.); B. Thorand (Barbara); O. Polašek (Ozren); B. McKnight (Barbara); H. Campbell (Harry); A.R. Rudnicka (Alicja); M.H. Chen (Min-hsin); B.M. Buckley (Brendan M.); S.E. Harris (Sarah); A. Peters (Annette); D. Pulanic (Drazen); T. Lumley (Thomas); A.J.M. de Craen (Anton J.M.); D.C. Liewald (David C.); C. Gieger (Christian); I. Ford (Ian); A.J. Gow (Alan J.); M. Luciano (Michelle); D.J. Porteous (David J.); X. Guo (Xiuqing); N. Sattar (Naveed); A. Tenesa (Albert); M. Cushman (Mary Ann); P.E. Slagboom (Eline); P.M. Visscher (Peter M.); T.D. Spector (Tim); T. Illig (Thomas); I. Rudan (Igor); E.G. Bovill (Edwin G.); A.F. Wright (Alan); W.L. McArdle (Wendy); G.H. Tofler (Geoffrey); A. Hofman (Albert); R.G.J. Westendorp (Rudi); J.M. Starr (John); P.J. Grant (Peter J.); M. Karakas (Mahir); N.D. Hastie (Nicholas D.); B.M. Psaty (Bruce); J.F. Wilson (James); G.D.O. Lowe (Gordon); C.J. O'Donnell (Christopher); J.C.M. Witteman (Jacqueline); J.W. Jukema (Jan Wouter); I.J. Deary (Ian); N. Soranzo (Nicole); W. Koenig (Wolfgang); C. Hayward (Caroline)

    2011-01-01

    textabstractBACKGROUND: Fibrin fragment D-dimer, one of several peptides produced when crosslinked fibrin is degraded by plasmin, is the most widely used clinical marker of activated blood coagulation. To identity genetic loci influencing D-dimer levels, we performed the first large-scale, genome-wi

  18. 77 FR 71432 - Agency Information Collection Activities: Application for Travel Document, Form Number I-131...

    Science.gov (United States)

    2012-11-30

    ... of the Form/Collection: Application for Travel Document. (3) Agency form number, if any, and the... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF HOMELAND SECURITY U.S. Citizenship and Immigration Services Agency Information Collection Activities:...

  19. Circulating intact and cleaved forms of the urokinase-type plasminogen activator receptor

    DEFF Research Database (Denmark)

    Sørensen, Tine Thurison; Christensen, Ib J; Lund, Ida K;

    2015-01-01

    BACKGROUND: High levels of circulating forms of the urokinase-type plasminogen activator receptor (uPAR) are significantly associated to poor prognosis in cancer patients. Our aim was to determine biological variations and reference intervals of the uPAR forms in blood, and in addition, to test t...

  20. Fiber optic D dimer biosensor

    Science.gov (United States)

    Glass, Robert S.; Grant, Sheila A.

    1999-01-01

    A fiber optic sensor for D dimer (a fibrinolytic product) can be used in vivo (e.g., in catheter-based procedures) for the diagnosis and treatment of stroke-related conditions in humans. Stroke is the third leading cause of death in the United States. It has been estimated that strokes and stroke-related disorders cost Americans between $15-30 billion annually. Relatively recently, new medical procedures have been developed for the treatment of stroke. These endovascular procedures rely upon the use of microcatheters. These procedures could be facilitated with this sensor for D dimer integrated with a microcatheter for the diagnosis of clot type, and as an indicator of the effectiveness, or end-point of thrombolytic therapy.

  1. Recognition of HIV TAR RNA by triazole linked neomycin dimers.

    Science.gov (United States)

    Kumar, Sunil; Arya, Dev P

    2011-08-15

    A series of neomycin dimers have been synthesized using 'click chemistry' with varying linker functionality and length to target the TAR RNA region of HIV virus. TAR (trans activation response) RNA region, a 59 base pair stem loop structure located at 5'-end of all nascent HIV-1 transcripts interacts with a key regulatory protein, Tat, and necessitates the replication of HIV-1 virus. Neomycin, an aminosugar, has been shown to exhibit more than one binding site with HIV TAR RNA. Multiple TAR binding sites of neomycin prompted us to design and synthesize a small library of neomycin dimers using click chemistry. The binding between neomycin dimers and HIV TAR RNA was characterized using spectroscopic techniques including FID (Fluorescent Intercalator Displacement) titration and UV-thermal denaturation. UV thermal denaturation studies demonstrate that neomycin dimer binding increase the melting temperature (T(m)) of the HIV TAR RNA up to 10°C. Ethidium bromide displacement titrations revealed nanomolar IC(50) between neomycin dimers and HIV TAR RNA, whereas with neomycin, a much higher IC(50) in the micromolar range is observed.

  2. Palladium dimers adsorbed on graphene: A DFT study

    Energy Technology Data Exchange (ETDEWEB)

    Kaur, Gagandeep, E-mail: gaganj1981@yahoo.com [Department of Physics and Centre of Advanced Studies in Physics, Panjab University, Chandigarh-160014 (India); Chandigarh Engineering College, Landran, Mohali, Punjab (India); Gupta, Shuchi, E-mail: sgupta@pu.ac.in [University Institute of Engineering and Technology, Panjab University, Chandigarh -160014 (India); Dharamvir, Keya, E-mail: keya@pu.ac.in [Department of Physics and Centre of Advanced Studies in Physics, Panjab University, Chandigarh-160014 (India)

    2015-05-15

    The 2D structure of graphene shows a great promise for enhanced catalytic activity when adsorbed with palladium. We performed a systematic density functional theory (DFT) study of the adsorption of palladium dimer (Pd{sub 2}) on graphene using SIESTA package, in the generalized gradient approximation (GGA). The adsorption energy, geometry, and charge transfer of Pd{sub 2}-graphene system are calculated. Both horizontal and vertical orientations of Pd{sub 2} on graphene are studied. Our calculations revealed that the minimum energy configuration for Pd dimer is parallel to the graphene sheet with its two atoms occupying centre of adjacent hexagonal rings of graphene sheet. Magnetic moment is induced for Pd dimer adsorbed on graphene in vertical orientation while horizontal orientation of Pd dimer on graphene do not exhibit magnetism. Insignificant energy differences among adsorption sites means that dimer mobility on the graphene sheet is high. There is imperceptible distortion of graphene sheet perpendicular to its plane. However, some lateral displacements are seen.

  3. Self-deactivation of water vapor - Role of the dimer

    Science.gov (United States)

    Zuckerwar, A. J.

    1984-01-01

    A phenomenological multiple-relaxation theory of the deactivation rate constant for the nu-2 (1 - 0) bending mode of water vapor is presented which incorporates the role not only of the excited monomer but also of the bound molecular complex, in particular the dimer. The deactivation takes place by means of three parallel processes: (1) collisional deexcitation of the excited monomer, (2) a two-step reaction involving association and spontaneous redissociation of an H2O collision complex, and (3) spontaneous dissociation of the stably bound H2O dimer. Oxygen, but not nitrogen or argon, serves as an effective chaperon for the formation of the activated complex. This observation explains the impurity dependence of the self-deactivation rate constant of water vapor. Analysis of an ultrasonic absorption peak based on the third process yields values for the standard entropy and enthalpy of dissociation of the stably bound H2O dimer.

  4. REVISITING THE PUTATIVE TCR Cα DIMERIZATION MODEL THROUGH STRUCTURAL ANALYSIS

    Directory of Open Access Journals (Sweden)

    Jia-huai eWang

    2013-01-01

    Full Text Available Despite major advances in T cell receptor (TCR biology and structure, how peptide-MHC complex (pMHC ligands trigger αβ TCR activation remains unresolved. Two views exist. One model postulates that monomeric TCR-pMHC ligation events are sufficient while a second proposes that TCR-TCR dimerization in cis via Cα domain interaction plus pMHC binding is critical. We scrutinized 22 known TCR/pMHC complex crystal structures, and did not find any predicted molecular Cα-Cα contacts in these crystals that would allow for physiological TCR dimerization. Moreover, the presence of conserved glycan adducts on the outer face of the Cα domain preclude the hypothesized TCR dimerization through the Cα domain. Observed functional consequences of Cα mutations are likely indirect, with TCR microclusters at the immunological synapse driven by TCR transmembrane/cytoplasmic interactions via signaling molecules, scaffold proteins and/or cytoskeletal elements.

  5. Synthesis, structural characterization and antimicrobial activities of diorganotin(IV) complexes with azo-imino carboxylic acid ligand: Crystal structure and topological study of a doubly phenoxide-bridged dimeric dimethyltin(IV) complex appended with free carboxylic acid groups

    Science.gov (United States)

    Roy, Manojit; Roy, Subhadip; Devi, N. Manglembi; Singh, Ch. Brajakishor; Singh, Keisham Surjit

    2016-09-01

    Diorganotin(IV) complexes appended with free carboxylic acids were synthesized by reacting diorganotin(IV) dichlorides [R2SnCl2; R = Me (1), Bu (2) and Ph (3)] with an azo-imino carboxylic acid ligand i.e. 2-{4-hydroxy-3-[(2-hydroxyphenylimino)methyl]phenylazo}benzoic acid in presence of triethylamine. The complexes were characterized by elemental analysis, IR and multinuclear NMR (1H, 13C and 119Sn) spectroscopy. The structure of 1 in solid state has been determined by X-ray crystallography. Crystal structure of 1 reveals that the compound crystallizes in monoclinic space group P21/c and is a dimeric dimethyltin(IV) complex appended with free carboxylic acid groups. In the structure of 1, the Sn(IV) atoms are hexacoordinated and have a distorted octahedral coordination geometry in which two phenoxy oxygen atoms and the azomethine nitrogen atom of the ligand coordinate to each tin atom. One of the phenoxy oxygen atom bridges the two tin centers resulting in a planar Sn2O2 core. Topological analysis is used for the description of molecular packing in 1. Tin NMR spectroscopy study indicates that the complexes have five coordinate geometry around tin atom in solution state. Since the complexes have free carboxylic acids, these compounds could be further used as potential metallo-ligands for the synthesis of other complexes. The synthesized diorganotin(IV) complexes were also screened for their antimicrobial activities and compound 2 showed effective antimicrobial activities.

  6. Domain-Swapped Dimers of Intracellular Lipid-Binding Proteins: Evidence for Ordered Folding Intermediates.

    Science.gov (United States)

    Assar, Zahra; Nossoni, Zahra; Wang, Wenjing; Santos, Elizabeth M; Kramer, Kevin; McCornack, Colin; Vasileiou, Chrysoula; Borhan, Babak; Geiger, James H

    2016-09-06

    Human Cellular Retinol Binding Protein II (hCRBPII), a member of the intracellular lipid-binding protein family, is a monomeric protein responsible for the intracellular transport of retinol and retinal. Herein we report that hCRBPII forms an extensive domain-swapped dimer during bacterial expression. The domain-swapped region encompasses almost half of the protein. The dimer represents a novel structural architecture with the mouths of the two binding cavities facing each other, producing a new binding cavity that spans the length of the protein complex. Although wild-type hCRBPII forms the dimer, the propensity for dimerization can be substantially increased via mutation at Tyr60. The monomeric form of the wild-type protein represents the thermodynamically more stable species, making the domain-swapped dimer a kinetically trapped entity. Hypothetically, the wild-type protein has evolved to minimize dimerization of the folding intermediate through a critical hydrogen bond (Tyr60-Glu72) that disfavors the dimeric form.

  7. Cellulase occurs in multiple active forms in ripe avocado fruit mesocarp.

    Science.gov (United States)

    Kanellis, A K; Kalaitzis, P

    1992-02-01

    The existence of multiple forms of avocado (Persea americana Mill. cv Hass) cellulase in crude protein extracts of ripe avocado fruit is reported. Cellulase was separated into at least 11 multiple forms by native isoelectric focusing in the pH range between 4 and 7 and visualized by both activity staining using Congo red and immunostaining. The enzyme components were acidic proteins with isoelectric points in the range of pH 5.10 to 6.80, the predominant forms having isoelectric points of 5.60, 5.80, 5.95, and 6.20. All 11 forms were immunologically related with molecular masses of 54 kilodaltons.

  8. Effects of Three Forms of Reading-Based Output Activity on L2 Vocabulary Learning

    Science.gov (United States)

    Rassaei, Ehsan

    2017-01-01

    The current study investigated the effects of three forms of output activity on EFL learners' recognition and recall of second language (L2) vocabulary. To this end, three groups of learners of English as a foreign language (EFL) were instructed to employ the following three output activities after reading two narrative texts: (1) summarizing the…

  9. Morphology and optical properties of aluminum oxide formed into oxalic electrolyte with addition surface active agents

    Science.gov (United States)

    Kazarkin, B.; Stsiapanau, A.; Zhilinski, V.; Chernik, A.; Bezborodov, V.; Kozak, G.; Danilovich, S.; Smirnov, A.

    2016-08-01

    The article discusses the results of investigations of porous films of alumina, formed into oxalic electrolyte with addition surface active agents, in particular, ordering structure, roughness of a surface, the optical transparency of the electrolyte concentration and surface active agents. Also discusses the features of the formation of porous films of temperature and IR radiation.

  10. 40 CFR 761.205 - Notification of PCB waste activity (EPA Form 7710-53).

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Notification of PCB waste activity..., DISTRIBUTION IN COMMERCE, AND USE PROHIBITIONS PCB Waste Disposal Records and Reports § 761.205 Notification of PCB waste activity (EPA Form 7710-53). (a)(1) All commercial storers, transporters, and disposers...

  11. Comparing Two Forms of Concept Map Critique Activities to Facilitate Knowledge Integration Processes in Evolution Education

    Science.gov (United States)

    Schwendimann, Beat A.; Linn, Marcia C.

    2016-01-01

    Concept map activities often lack a subsequent revision step that facilitates knowledge integration. This study compares two collaborative critique activities using a Knowledge Integration Map (KIM), a form of concept map. Four classes of high school biology students (n?=?81) using an online inquiry-based learning unit on evolution were assigned…

  12. Synthesis of steroidal dimers: Selective amine catalysed steroidal dimerization

    Indian Academy of Sciences (India)

    Shamsuzzaman; Mohd Gulfam Aalam; Tabassum Siddiqui

    2011-07-01

    Some new dimeric steroids namely cholest-5-en-3-spiro-[6',5'-oxa]-5'-cholest-3'-one (2), cholest-5-en-7-spiro-[4',5'-oxa]-5'-cholest-7'-one (4a) and 3-substitutedcholest-5-en-7-spiro-[4',5'-oxa]-3'-substituted-5'-cholestan-7'-ones (4b, c) are synthesized starting from cholest-5-en-3-one (1), cholest-5-en-7-one (3a) and 3-substituted-cholest-5-en-7-ones (3b, c) respectively by using DMAP and xylene. All the synthesized compounds were characterized by using IR, MS and 1H, 13C NMR spectral and elemental analysis.

  13. Microwave Measurements of Maleimide and its Doubly Hydrogen Bonded Dimer with Formic ACID*

    Science.gov (United States)

    Pejlovas, Aaron M.; Kang, Lu; Kukolich, Stephen G.

    2016-06-01

    The microwave spectra were measured for the maleimide monomer and the maleimide-formic acid doubly hydrogen bonded dimer using a pulsed-beam Fourier transform microwave spectrometer. Many previously studied doubly hydrogen bonded dimers are formed between oxygen containing species, so it is important to also characterize and study other dimers containing nitrogen, as hydrogen bonding interactions with nitrogen are found in biological systems such as in DNA. The transition state of the dimer does not exhibit C_2_V symmetry, so the tunneling motion was not expected to be observed based on the symmetry, but it would be very important to also observe the tunneling process for an asymmetric dimer. Single-line b-type transitions were observed, so the tunneling motion was not observed in our microwave spectra. The hydrogen bond lengths were determined using a nonlinear least squares fitting program. *Supported by the NSF CHE-1057796

  14. Application of Bruggeman and Maxwell Garnett homogenization formalisms to random composite materials containing dimers

    CERN Document Server

    Mackay, Tom G

    2015-01-01

    The homogenization of a composite material comprising three isotropic dielectric materials was investigated. The component materials were randomly distributed as spherical particles, with the particles of two of the component materials being coupled to form dimers. The Bruggeman and Maxwell Garnett formalisms were developed to estimate the permittivity dyadic of the homogenized composite material (HCM), under the quasi-electrostatic approximation. Both randomly oriented and identically oriented dimers were accommodated; in the former case the HCM is isotropic, whereas in the latter case the HCM is uniaxial. Representative numerical results for composite materials containing dielectric--dielectric dimers demonstrate close agreement between the estimates delivered by the Bruggeman and Maxwell Garnett formalisms. For composite materials containing metal--dielectric dimers with moderate degrees of dissipation, the estimates of the two formalisms are in broad agreement, provided that the dimer volume fractions are...

  15. Solubilization of the O2(-)-forming activity responsible for the respiratory burst in human neutrophils.

    Science.gov (United States)

    Gabig, T G; Kipnes, R S; Babior, B M

    1978-10-10

    On exposure to suitable activating agents, neutrophils sharply alter their oxygen metabolism, showing large increases in oxygen uptake, O2 and H2O2 production, and glucose consumption via the hexose monophosphate shunt. These metabolic alterations, which together are designated the "respiratory burst," are due to the activation of a system which catalyzes the reaction: 2O2 + NADPH leads to 2O2(-) + NADP. This O2(-)-forming system is found in a particulate fraction isolated from neutrophils which had been activated with opsonized zymosan. When these particles were treated with detergent under suitable conditions, the O2(-)-forming activity was released in a form which passed through a membrane filter capable of retaining species of Mr greater than 3000,000. Soluble O2(-)-forming activity was obtained from normal activated neutrophils, but not from normal resting neutrophils or from activated neutrophils obtained from patients with chronic granulomatous disease, an inherited condition in which the respiratory burst is defective. O2(-)production by the soluble system required a reduced pyridine nucleotide as electron donor, and showed a quadratic dependence on the concentration of the solubilized preparation.

  16. Self-assembly of Asymmetric Dimer Particles in Supported Copolymer Bilayer

    Institute of Scientific and Technical Information of China (English)

    Xiao-chun Qin; Chun-lai Ren

    2011-01-01

    Using self-consistent field and density functional theories, we investigate the self-assembly behavior of asymmetric dimer particles in a supported AB block copolymer bilayer. Asymmetric dimer particles are amphiphilic molecules composed by two different spheres. One prefers to A block of copolymers and the other likes B block when they are introduced into the copolymer bilayer. The two layer structure of the dimer particles is formed within the bilayer.Due to the presence of the substrate surface, the symmetry of the two leaflets of the bilayer is broken, which may lead to two different layer structures of dimer particles within each leaflet of the bilayer. With the increasing concentration of the asymmetric dimer particles,in-plane structure of the dimer particles undergoes sparse square, hexagonal, dense square, and cylindrical structures. In a further condensed packing, a bending cylindrical structure comes into being. Here we verify that the entropic effect of copolymers, the enthalpy of the system and the steric repulsion of the dimer particles are three important factors determing the self-assembly of dimer particles within the supported copolymer bilayer.

  17. Synchronized oscillations of dimers in biphasic charged fd-virus suspensions

    Science.gov (United States)

    Kang, K.; Piao, S. H.; Choi, H. J.

    2016-08-01

    Micron-sized colloidal spheres that are dispersed in an isotropic-nematic biphasic host suspension of charged rods (fd-virus particles) are shown to spontaneously form dimers, which exhibit a synchronized oscillatory motion. Dimer formation is not observed in the monophase of isotropic and nematic suspensions. The synchronized oscillations of dimers are connected to the inhomogeneous state of the host suspension of charged rods (fd viruses) where nematic domains are in coexistence with isotropic regions. The synchronization of oscillations occurs in bulk states, in the absence of an external field. With a low field strength of an applied electric field, the synchronization is rather reduced, but it recovers again when the field is turned off. In this Rapid Communication, we report this observation as an example of the strange attractor, occurring in the mixture of PS (polystyrene) dimers in an isotropic-nematic coexistence biphasic fd-virus network. Furthermore, we highlight that the synchronization of PS-dimer oscillations is the result of a global bifurcation diagram, driven by a delicate balance between the short-attractive "twisted" interaction of PS dimers and long-ranged electrostatic repulsive interactions of charged fd rods. The interest is then in the local enhancement of "twist-nematic" elasticity in reorientation of the dimer oscillations. An analysis of image-time correlations is provided with the data movies and Fourier transforms of averaged orientations for the synchronized oscillations of dimers in the biphasic I -N coexistence concentration of charged fd-virus suspensions.

  18. Synthesis of cadmium complexes of 4'-chloro-terpyridine: From discrete dimer to 1D chain polymer, crystal structure and antibacterial activity

    Indian Academy of Sciences (India)

    Lotfali Saghatforoush; Laura Valencia Matarranz; Firoozeh Chalabian; Shahriare Ghammamy; Fatemeh Katouzian

    2012-05-01

    Two new Cd(II) complexes with the ligand 4'-chloro-2,2':6',2"-terpyridine (Cltpy), [Cd(Cltpy)(N3)(CH3COO)], 1, and [Cd(Cltpy)(NCS)(CH3COO)], 2, have been synthesized and characterized by CHN elemental analyses, 1HNMR-, 13C NMR-, IR spectroscopy and structurally analysed by X-ray singlecrystal diffraction. The single crystal X-ray analyses show that the coordination number in these complexes is seven with three terpyridine (Cltpy) N-donor atoms, two acetate oxygens and two anionic bridged ligands. The crystal structure of 2 comprises a one-dimensional polymeric network bridged by NCS− anions. The antibacterial activities of Cltpy and its Cd(II) complexes are tested against different bacteria. Both complexes have shown good activity against all the tested bacteria. Against Klebsiella pneumonia and Staphylococcus aureus, antibacterial activity of complexes is higher than Cltpy ligand. The higher activity of complexes may be explained on the basis of chelation theory.

  19. Dimeric Arrangement of the Parathyroid Hormone Receptor and a Structural Mechanism for Ligand-induced Dissociation

    Energy Technology Data Exchange (ETDEWEB)

    Pioszak, Augen A.; Harikumar, Kaleeckal G.; Parker, Naomi R.; Miller, Laurence J.; Xu, H. Eric (Van Andel); (Mayo)

    2010-06-25

    The parathyroid hormone receptor (PTH1R) is a class B G protein-coupled receptor that is activated by parathyroid hormone (PTH) and PTH-related protein (PTHrP). Little is known about the oligomeric state of the receptor and its regulation by hormone. The crystal structure of the ligand-free PTH1R extracellular domain (ECD) reveals an unexpected dimer in which the C-terminal segment of both ECD protomers forms an {alpha}-helix that mimics PTH/PTHrP by occupying the peptide binding groove of the opposing protomer. ECD-mediated oligomerization of intact PTH1R was confirmed in living cells by bioluminescence and fluorescence resonance energy transfer experiments. As predicted by the structure, PTH binding disrupted receptor oligomerization. A receptor rendered monomeric by mutations in the ECD retained wild-type PTH binding and cAMP signaling ability. Our results are consistent with the hypothesis that PTH1R forms constitutive dimers that are dissociated by ligand binding and that monomeric PTH1R is capable of activating G protein.

  20. Dimerization of Human Growth Hormone by Zinc

    Science.gov (United States)

    Cunningham, Brian C.; Mulkerrin, Michael G.; Wells, James A.

    1991-08-01

    Size-exclusion chromatography and sedimentation equilibrium studies demonstrated that zinc ion (Zn2+) induced the dimerization of human growth hormone (hGH). Scatchard analysis of 65Zn2+ binding to hGH showed that two Zn2+ ions associate per dimer of hGH in a cooperative fashion. Cobalt (II) can substitute for Zn2+ in the hormone dimer and gives a visible spectrum characteristic of cobalt coordinated in a tetrahedral fashion by oxygen- and nitrogen-containing ligands. Replacement of potential Zn2+ ligands (His18, His21, and Glu174) in hGH with alanine weakened both Zn2+ binding and hGH dimer formation. The Zn2+-hGH dimer was more stable than monomeric hGH to denaturation in guanidine-HCl. Formation of a Zn2+-hGH dimeric complex may be important for storage of hGH in secretory granules.

  1. Formation and occurrence of dimer esters of pinene oxidation products in atmospheric aerosols

    Directory of Open Access Journals (Sweden)

    K. Kristensen

    2012-08-01

    Full Text Available Formation of carboxylic acids and dimer esters from α-pinene oxidation were investigated in a smog chamber and in ambient aerosol samples collected during the Biosphere Effects on Aerosols and Photochemistry Experiment (BEARPEX. Chamber experiments of α-pinene ozonolysis in dry air and at low NOx concentrations demonstrated formation of two dimer esters, pinyl-diaterpenyl (MW 358 and pinonyl-pinyl dimer ester (MW 368, under both low and high temperature conditions. Concentration levels of the pinyl-diaterpenyl dimer ester were lower than the assumed first-generation oxidation products cis-pinic and terpenylic acids, but similar to the second-generation oxidation products 3-methyl-1,2,3-butane tricarboxylic acid (MBTCA and diaterpenylic acid acetate (DTAA. Dimer esters were observed within the first 30 min, indicating rapid production simultaneous to their structural precursors. However, the sampling time resolution precluded conclusive evidence regarding formation from gas- or particle-phase processes. CCN activities of the particles formed in the smog chamber displayed a modest variation during the course of experiments with κ values in the range 0.06–0.09 (derived at a supersaturation of 0.19%.

    The pinyl-diaterpenyl dimer ester was also observed in ambient aerosol samples collected above a ponderosa pine forest in the Sierra Nevada Mountains of California during two seasonally distinct field campaigns in September 2007 and July 2009. The pinonyl-pinyl ester was observed for the first time in ambient air during the 2009 campaign, and although present at much lower concentrations, it was correlated with the abundance of the pinyl-diaterpenyl ester suggesting similarities in their formation. The maximum concentration of the pinyl-diaterpenyl ester was almost 10 times higher during the warmer 2009 campaign relative to 2007, while the concentration of cis-pinic acid was approximately the same during both

  2. Synthesis, Dimeric Crystal Structure, and Biological Activities of N-(4-Methyl-6-oxo-1,6-dihydro-pyrimidin-2-yl)-N-(2-trifluoromethyl-phenyl)-guanidine

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The title compound, N-(4-methyl-6-oxo-1,6-dihydro-pyrimidin-2-yl)-N′-(2-trifluoromethyl-phenyl)-guanidine, was synthesized and its structure was confirmed by using IR, MS, 1H NMR, and elemental analysis. The single crystal structure of the title compound was determined by X-ray diffraction. The preliminary biological test showed that the synthesized compound has a weak herbicidal activity.

  3. On Dimer Models and Closed String Theories

    OpenAIRE

    Sarkar, Tapobrata

    2007-01-01

    We study some aspects of the recently discovered connection between dimer models and D-brane gauge theories. We argue that dimer models are also naturally related to closed string theories on non compact orbifolds of $\\BC^2$ and $\\BC^3$, via their twisted sector R charges, and show that perfect matchings in dimer models correspond to twisted sector states in the closed string theory. We also use this formalism to study the combinatorics of some unstable orbifolds of $\\BC^2$.

  4. Dimer models and Calabi-Yau algebras

    CERN Document Server

    Broomhead, Nathan

    2008-01-01

    In this thesis we study dimer models, as introduced in string theory, which give a way of writing down a class of non-commutative `superpotential' algebras. Some examples are 3-dimensional Calabi-Yau algebras, as defined by Ginzburg, and some are not. We consider two types of `consistency' condition on dimer models, and show that a `geometrically consistent' model is `algebraically consistent'. Finally we prove that the algebras obtained from algebraically consistent dimer models are 3-dimensional Calabi-Yau algebras.

  5. Cargo binding activates myosin VIIA motor function in cells.

    Science.gov (United States)

    Sakai, Tsuyoshi; Umeki, Nobuhisa; Ikebe, Reiko; Ikebe, Mitsuo

    2011-04-26

    Myosin VIIA, thought to be involved in human auditory function, is a gene responsible for human Usher syndrome type 1B, which causes hearing and visual loss. Recent studies have suggested that it can move processively if it forms a dimer. Nevertheless, it exists as a monomer in vitro, unlike the well-known two-headed processive myosin Va. Here we studied the molecular mechanism, which is currently unknown, of activating myosin VIIA as a cargo-transporting motor. Human myosin VIIA was present throughout cytosol, but it moved to the tip of filopodia upon the formation of dimer induced by dimer-inducing reagent. The forced dimer of myosin VIIA translocated its cargo molecule, MyRip, to the tip of filopodia, whereas myosin VIIA without the forced dimer-forming module does not translocate to the filopodial tips. These results suggest that dimer formation of myosin VIIA is important for its cargo-transporting activity. On the other hand, myosin VIIA without the forced dimerization module became translocated to the filopodial tips in the presence of cargo complex, i.e., MyRip/Rab27a, and transported its cargo complex to the tip. Coexpression of MyRip promoted the association of myosin VIIA to vesicles and the dimer formation. These results suggest that association of myosin VIIA monomers with membrane via the MyRip/Rab27a complex facilitates the cargo-transporting activity of myosin VIIA, which is achieved by cluster formation on the membrane, where it possibly forms a dimer. Present findings support that MyRip, a cargo molecule, functions as an activator of myosin VIIA transporter function.

  6. Sputtering of dimers off a silicon surface

    Energy Technology Data Exchange (ETDEWEB)

    Nietiadi, Maureen L. [Physics Department, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Research Center OPTIMAS, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Rosandi, Yudi [Physics Department, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Research Center OPTIMAS, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Department of Physics, Universitas Padjadjaran, Jatinangor, Sumedang 45363 (Indonesia); Kopnarski, Michael [Physics Department, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Research Center OPTIMAS, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Institut fuer Oberflaechen- und Schichtanalytik IFOS GmbH, Trippstadter Strasse 120, D-67663 Kaiserslautern (Germany); Urbassek, Herbert M., E-mail: urbassek@rhrk.uni-kl.de [Physics Department, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany); Research Center OPTIMAS, University Kaiserslautern, Erwin-Schroedinger-Strasse, D-67663 Kaiserslautern (Germany)

    2012-10-15

    We present experimental and molecular-dynamics simulation results of the sputtering of a Si surface by 2 keV Ar ions. Results on both the monomer and dimer distributions are presented. In simulation, these distributions follow a generalized Thompson law with power exponent n=2 and n=3, respectively. The experimental data, obtained via plasma post-ionization in an SNMS (secondary neutral mass spectrometry) apparatus, show good agreement with respect to the dimer fraction, and the relative energy distributions of dimers and monomers. The consequences for the dimer sputtering mechanism are discussed.

  7. Enhanced Chiral Recognition by Cyclodextrin Dimers

    Directory of Open Access Journals (Sweden)

    Bart Jan Ravoo

    2011-07-01

    Full Text Available In this article we investigate the effect of multivalency in chiral recognition. To this end, we measured the host-guest interaction of a β-cyclodextrin dimer with divalent chiral guests. We report the synthesis of carbohydrate-based water soluble chiral guests functionalized with two borneol, menthol, or isopinocampheol units in either (+ or (– configuration. We determined the interaction of these divalent guests with a β-cyclodextrin dimer using isothermal titration calorimetry. It was found that—in spite of a highly unfavorable conformation—the cyclodextrin dimer binds to guest dimers with an increased enantioselectivity, which clearly reflects the effect of multivalency.

  8. Mode of action of antimicrobial proteins, pore-forming toxins and biologically active

    Directory of Open Access Journals (Sweden)

    O Schmidt

    2005-07-01

    Full Text Available Antimicrobial peptides and pore-forming toxins are important effectors in innate immune defencereactions. But their mode of action, comprising the insertion into cholesterol-containing membranes isnot known. Here we explore the mechanical implications of pore-formation by extracellular proteinassemblies that drive cellular uptake reactions by leverage-mediated (LM processes, whereoligomeric adhesion molecules bent membrane-receptors around ‘hinge’-like lipophorin particles. Theinteractions of antimicrobial peptides, pore-forming toxins and biologically active proteins with LMassembliesprovide a new paradigm for the configurational specificity and sterical selectivity ofbiologically active peptides.

  9. Ictal technetium-99m ethyl cysteinate dimer single-photon emission tomographic findings and propagation of epileptic seizure activity in patients with extratemporal epilepsies

    Energy Technology Data Exchange (ETDEWEB)

    Noachtar, S.; Arnold, S.; Werhahn, K.J. [Department Neurologie, Ludwig-Maximilians Universitaet, Muenchen (Germany); Yousry, T.A. [Muenchen Univ. (Germany). Abt. fuer Neuroradiologie; Bartenstein, P. [Department of Nuclear Medicine, Technical University of Munich, Munich (Germany); Tatsch, K. [Department of Nuclear Medicine, Ludwig-Maximilians University of Munich, Munich (Germany)

    1998-02-01

    We investigated the influence of the propagation of extratemporal epileptic seizure activity on the regional increase in cerebral blood flow, which is usually associated with epileptic seizure activity. Forty-two consecutive patients with extratemporal epilepsies were prospectively evaluated. All patients underwent ictal SPET studies with simultaneous electroencephalography (EEG) and video recordings of habitual seizures and imaging studies including cranial magnetic resonance imaging and positron emission tomography with 2-[{sup 18}F]-fluoro-2 deoxy-d-glucose. Propagation of epilptic seizure activity (PESA) was defined as the absence of hyperperfusion on ictal ECD SPET in the lobe of seizure onset, but its presence in another ipsilateral or contralateral lobe. Observers analysing the SPET images were not informed of the other results. PESA was observed in 8 of the 42 patients (19%) and was ipsilateral to the seizure onset in five (63%) of these eight patients. The time between clinical seizure onset and injection of the ECD tracer ranged from 14 to 61 s (mean 34 s). Seven patients (88%) with PESA had parieto-occipital epilepsy and one patient had a frontal epilepsy. PESA was statistically more frequent in patients with parieto-occipital lobe epilepsies (58%) than in the remaining extratemporal epilepsy syndromes (3%) (P<0.0002). These findings indicate that ictal SPET studies require simultaneous EEG-video recordings in patients with extratemporal epilepsies. PESA should be considered when interpreting ictal SPET studies in these patients. Patients with PESA are more likely to have parieto-occipital lobe epilepsy than seizure onset in other extratemporal regions. (orig./MG) (orig.) With 1 fig., 2 tabs., 23 refs.

  10. Angle-Resolved Plasmonic Properties of Single Gold Nanorod Dimers

    Institute of Scientific and Technical Information of China (English)

    Jian Wu; Xuxing Lu; Qiannan Zhu; Junwei Zhao; Qishun Shen; Li Zhan; Weihai Ni

    2014-01-01

    Through wet-chemical assembly methods, gold nanorods were placed close to each other and formed a dimer with a gap distance*1 nm, and hence degenerated plasmonic dipole modes of individual nanorods coupled together to produce hybridized bonding and antibonding resonance modes. Previous studies using a condenser for illumination result in averaged signals over all excitation angles. By exciting an individual dimer obliquely at different angles, we demonstrate that these two new resonance modes are highly tunable and sensitive to the angle between the excitation polarization and the dimer orientation, which follows cos2u dependence. Moreover, for dimer structures with various structure angles, the resonance wavelengths as well as the refractive index sensitivities were found independent of the structure angle. Cal-culated angle-resolved plasmonic properties are in good agreement with the measurements. The assembled nanostructures investigated here are important for fundamental researches as well as potential applications when they are used as building blocks in plasmon-based optical and optoelectronic devices.

  11. Centrosymmetric dimer of quinuclidine betaine and squaric acid complex

    Science.gov (United States)

    Dega-Szafran, Z.; Katrusiak, A.; Szafran, M.

    2012-12-01

    The complex of squaric acid (3,4-dihydroxy-3-cyclobuten-1,2-dion, H2SQ) with quinuclidine betaine (1-carboxymethyl-1-azabicyclo[2.2.2]octane inner salt, QNB), 1, has been characterized by single-crystal X-ray analysis, FTIR and NMR spectroscopies and by DFT calculations. In the crystal of 1, monoclinic space group P21/n, one proton from H2SQ is transferred to QNB. QNBH+ and HSQ- are linked together by a Osbnd H⋯O hydrogen bond of 2.553(2) Å. Two such QNBH+·HSQ- complexes form a centrosymmetric dimer bridged by two Osbnd H⋯O bonds of 2.536(2) Å. The FTIR spectrum is consistent with the X-ray results. The structures of monomer QNBH+·HSQ- (1a) and dimer [QNB·H2SQ]2 (2) have been optimized at the B3LYP/6-311++G(d,p) level of theory. Isolated dimer 2 optimized back to a molecular aggregate of H2SQ and QNB. The calculated frequencies for the optimized structure of dimer 2 have been used to explain the frequencies of the experimental FTIR spectrum. The interpretation of 1H and 13C NMR spectra has been based on the calculated GIAO/B3LYP/6-311++G(d,p) magnetic isotropic shielding constants for monomer 1a.

  12. Morphological and physiological retinal degeneration induced by intravenous delivery of vitamin A dimers in rabbits

    Directory of Open Access Journals (Sweden)

    Jackie Penn

    2015-02-01

    Full Text Available The eye uses vitamin A as a cofactor to sense light and, during this process, some vitamin A molecules dimerize, forming vitamin A dimers. A striking chemical signature of retinas undergoing degeneration in major eye diseases such as age-related macular degeneration (AMD and Stargardt disease is the accumulation of these dimers in the retinal pigment epithelium (RPE and Bruch’s membrane (BM. However, it is not known whether dimers of vitamin A are secondary symptoms or primary insults that drive degeneration. Here, we present a chromatography-free method to prepare gram quantities of the vitamin A dimer, A2E, and show that intravenous administration of A2E to the rabbit results in retinal degeneration. A2E-damaged photoreceptors and RPE cells triggered inflammation, induced remolding of the choroidal vasculature and triggered a decline in the retina’s response to light. Data suggest that vitamin A dimers are not bystanders, but can be primary drivers of retinal degeneration. Thus, preventing dimer formation could be a preemptive strategy to address serious forms of blindness.

  13. Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E.

    Science.gov (United States)

    Siddiqui, Shoib; Schwarz, Flavio; Springer, Stevan; Khedri, Zahra; Yu, Hai; Deng, Lingquan; Verhagen, Andrea; Naito-Matsui, Yuko; Jiang, Weiping; Kim, Daniel; Zhou, Jie; Ding, Beibei; Chen, Xi; Varki, Nissi; Varki, Ajit

    2017-01-20

    CD33-related Siglecs are a family of proteins widely expressed on innate immune cells. Binding of sialylated glycans or other ligands triggers signals that inhibit or activate inflammation. Immunomodulation by Siglecs has been extensively studied, but relationships between structure and functions are poorly explored. Here we present new data relating to the structure and function of Siglec-E, the major CD33-related Siglec expressed on mouse neutrophils, monocytes, macrophages, and dendritic cells. We generated nine new rat monoclonal antibodies specific to mouse Siglec-E, with no cross-reactivity to Siglec-F. Although all antibodies detected Siglec-E on transfected human HEK-293T cells, only two reacted with mouse bone marrow neutrophils by flow cytometry and on spleen sections by immunohistochemistry. Moreover, whereas all antibodies recognized Siglec-E-Fc on immunoblots, binding was dependent on intact disulfide bonds and N-glycans, and only two antibodies recognized native Siglec-E within spleen lysates. Thus, we further investigated the impact of Siglec-E homodimerization. Homology-based structural modeling predicted a cysteine residue (Cys-298) in position to form a disulfide bridge between two Siglec-E polypeptides. Mutagenesis of Cys-298 confirmed its role in dimerization. In keeping with the high level of 9-O-acetylation found in mice, sialoglycan array studies indicate that this modification has complex effects on recognition by Siglec-E, in relationship to the underlying structures. However, we found no differences in phosphorylation or SHP-1 recruitment between dimeric and monomeric Siglec-E expressed on HEK293A cells. Phylogenomic analyses predicted that only some human and mouse Siglecs form disulfide-linked dimers. Notably, Siglec-9, the functionally equivalent human paralog of Siglec-E, occurs as a monomer.

  14. Mechanism of Processive Movement of Monomeric and Dimeric Kinesin Molecules

    Directory of Open Access Journals (Sweden)

    Ping Xie

    2010-01-01

    Full Text Available Kinesin molecules are motor proteins capable of moving along microtubule by hydrolyzing ATP. They generally have several forms of construct. This review focuses on two of the most studied forms: monomers such as KIF1A (kinesin-3 family and dimers such as conventional kinesin (kinesin-1 family, both of which can move processively towards the microtubule plus end. There now exist numerous models that try to explain how the kinesin molecules convert the chemical energy of ATP hydrolysis into the mechanical energy to “power” their proceesive movement along microtubule. Here, we attempt to present a comprehensive review of these models. We further propose a new hybrid model for the dimeric kinesin by combining the existing models and provide a framework for future studies in this subject.

  15. Functional Specificity of the Visual Word Form Area: General Activation for Words and Symbols but Specific Network Activation for Words

    Science.gov (United States)

    Reinke, Karen; Fernandes, Myra; Schwindt, Graeme; O'Craven, Kathleen; Grady, Cheryl L.

    2008-01-01

    The functional specificity of the brain region known as the Visual Word Form Area (VWFA) was examined using fMRI. We explored whether this area serves a general role in processing symbolic stimuli, rather than being selective for the processing of words. Brain activity was measured during a visual 1-back task to English words, meaningful symbols…

  16. Interleukin-1 beta converting enzyme requires oligomerization for activity of processed forms in vivo.

    Science.gov (United States)

    Gu, Y; Wu, J; Faucheu, C; Lalanne, J L; Diu, A; Livingston, D J; Su, M S

    1995-05-01

    Interleukin-1 beta converting enzyme (ICE) is composed of 10' (p10) and 20 kDa (p20) subunits, which are derived from a common 45 kDa precursor. Recent crystallographic studies have shown that ICE exists as a tetramer (p20/p10)2 in the crystal lattice. We provide evidence that the p10 and p20 subunits of ICE associate as oligomers in transfected COS cells. Using intragenic complementation, we show that the activity of a p10/p10 interface mutant defective in autoprocessing can be restored by co-expression with active site ICE mutants. Different active site mutants can also complement each other by oligomerization to form active ICE. These studies indicate that ICE precursor polypeptides may associate in different quaternary structures and that oligomerization is required for autoprocessing. Furthermore, integenic complementation of active site mutants of ICE and an ICE homolog restores autoprocessing activity, suggesting that hetero-oligomerization occurs between ICE homologs.

  17. Project Activities as a Form of English Language Teaching Based on the Interdisciplinary Approach to Form Intercultural Communicative Competence

    Science.gov (United States)

    Redchenko, Nadezhda N.

    2016-01-01

    The authors of this article suggest a thesis about the purpose of teaching a foreign language--it is student's communicative activities, i.e. learning a foreign language in practice. The teacher's task is to encourage activities of every student and to create situations to develop their creative activities in a learning process. New information…

  18. Formation and diffusion of water dimers on rutile TiO2(110).

    Science.gov (United States)

    Matthiesen, J; Hansen, J O; Wendt, S; Lira, E; Schaub, R; Laegsgaard, E; Besenbacher, F; Hammer, B

    2009-06-05

    From an interplay of time-lapsed high resolution scanning tunneling microscopy and density functional theory calculations we reveal the formation and diffusion of water dimers on hydrated rutile TiO2(110)-(1x1) surfaces, i.e., surfaces containing OH_{br} groups. At temperatures between approximately 150 and approximately 210 K water monomers diffusing along the Ti troughs were found to form stable water dimers that diffuse faster than the water monomers. An H bond mediated rollover mechanism operating for the water dimers explains the experimental findings.

  19. 75 FR 39622 - Proposed Information Collection (Health Resource Center Medical Center Payment Form) Activity...

    Science.gov (United States)

    2010-07-09

    ... AFFAIRS Proposed Information Collection (Health Resource Center Medical Center Payment Form) Activity... collection of information abstracted below to the Office of Management and Budget (OMB) for review and comment. The PRA submission describes the nature of the information collection and its expected cost...

  20. Soluble urokinase-type plasminogen activator receptor forms in plasma as markers of atherosclerotic plaque vulnerability

    DEFF Research Database (Denmark)

    Olson, Fredrik J; Thurison, Tine; Ryndel, Mikael;

    2009-01-01

    OBJECTIVES:: To test if circulating forms of the soluble urokinase-type plasminogen activator receptor (suPAR) are potential biomarkers of plaque vulnerability. DESIGN AND METHODS:: Plasma concentrations of suPAR(I-III), suPAR(II-III) and uPAR(I) were measured by time-resolved fluorescence immuno...

  1. 76 FR 21912 - Agency Information Collection Activities: Form I-765, Extension of an Existing Information...

    Science.gov (United States)

    2011-04-19

    ... the methodology and assumptions used; (3) Enhance the quality, utility, and clarity of the information... copy of the information collection instrument, please visit: http://www.regulations.gov/search/index... SECURITY U.S. Citizenship and Immigration Services Agency Information Collection Activities: Form...

  2. 76 FR 37059 - Agency Information Collection Activities: Proposed Collection; Comment Request-Form FNS-339...

    Science.gov (United States)

    2011-06-24

    ..., Infants and Children (WIC); the WIC Farmers' Market Nutrition Program (FMNP); and/or the Senior Farmers... Food and Nutrition Service Agency Information Collection Activities: Proposed Collection; Comment Request--Form FNS-339, Federal-State Supplemental Nutrition Program(s) Agreement AGENCY: Food...

  3. 75 FR 5066 - Commission Information Collection Activities (FERC Form 60,1

    Science.gov (United States)

    2010-02-01

    ... information (including balance sheet, assets, liabilities, billing and charges for associated and non... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF ENERGY Federal Energy Regulatory Commission Commission Information Collection Activities (FERC Form 60,\\1\\ FERC-61,...

  4. Future of the Learning Activities in Teenage School: Content, Methods, and Forms

    Directory of Open Access Journals (Sweden)

    Vorontsov A.B.

    2015-11-01

    Full Text Available the early 1990s their scientific research results have been formed in the educational system and began to be used in general primary school. However, when the widespread use of developmental education in elementary school, further studies on the age possibilities of adolescents and the content of their education have not been completed. Targeted research was organized again under the leadership of B.D. Elkonin only in 2000. Designing of teenage school in the framework of the principles and ideology of this system started at the same time at the Psychological Institute of the Russian Academy of Education and many other educational institutions. The article presents the hypothetical ideas about the content, forms and methods of organization of educational process in the second stage of schooling. Particular attention is paid to the fate of the educational activity in teenage school, as well as methods and forms of organization of other activities in the adolescent school.

  5. Antimicrobial peptide protegrin-3 adopt an antiparallel dimer in the presence of DPC micelles: a high-resolution NMR study

    Energy Technology Data Exchange (ETDEWEB)

    Usachev, K. S., E-mail: k.usachev@kpfu.ru; Efimov, S. V.; Kolosova, O. A.; Klochkova, E. A.; Aganov, A. V.; Klochkov, V. V. [Kazan Federal University, NMR Laboratory, Institute of Physics (Russian Federation)

    2015-05-15

    A tendency to dimerize in the presence of lipids was found for the protegrin. The dimer formation by the protegrin-1 (PG-1) is the first step for further oligomeric membrane pore formation. Generally there are two distinct model of PG-1 dimerization in either a parallel or antiparallel β-sheet. But despite the wealth of data available today, protegrin dimer structure and pore formation is still not completely understood. In order to investigate a more detailed dimerization process of PG-1 and if it will be the same for another type of protegrins, in this work we used a high-resolution NMR spectroscopy for structure determination of protegrin-3 (RGGGL-CYCRR-RFCVC-VGR) in the presence of perdeuterated DPC micelles and demonstrate that PG-3 forms an antiparallel NCCN dimer with a possible association of these dimers. This structural study complements previously published solution, solid state and computational studies of PG-1 in various environments and validate the potential of mean force simulations of PG-1 dimers and association of dimers to form octameric or decameric β-barrels.

  6. Forms of Physical Activity of Biała Podlaska Preschool Children

    Directory of Open Access Journals (Sweden)

    Gradus Paulina

    2014-09-01

    Full Text Available Introduction. Physical activity is fundamental to children's all-round development in the first six years of their life. It is particularly significant in preschool age, when the need for movement is predominant. Psychomotor development, conditioned by children's innate abilities and their own activity, depends upon the influence of external surroundings as well as parental upbringing and preschool education. The aim of the study was to obtain data regarding physical activity (both structured and unstructured of children during their stay in a kindergarten. Material and methods. The study was conducted in five state kindergartens situated in four main housing estates of Bia³a Podlaska. It included 11 groups of older preschool children. It was a diagnostic study. A diagnostic poll method employed in the study included interviews with teachers, lesson observations (during one day and document analyses (programmes and class registers. Results. During their stay in a kindergarten children take part in physical activities for 1.5 hours a day. During this period they participate in obligatory forms of physical activity such as morning gymnastics, movement plays, outdoor activities and gymnastic exercises. Furthermore, optional movement activities are organised in kindergartens (different forms in different kindergartens. Such extra classes enable children to be physically active for over two more hours per week. Conclusions. In terms of caring about health and motor development of children, not all elements of preschool education programmes are fully realised. Training sessions, workshops and lectures showing diverse forms of physical activity for children ought to be organised for preschool teachers.

  7. THE ENTERPRISE - THE LEGAL FORM FOR CARRYING ON AN ACTIVITY HAVING A PROFESSIONAL NATURE

    Directory of Open Access Journals (Sweden)

    Stanciu D. C RPENARU

    2011-08-01

    Full Text Available The new Romanian Civil Code has institutionalised a new conception regarding the regulating system for civil and commercial legal relations. Thus, new concepts emerge to fit the new conception, concepts regarding to the persons, the professionals, and the carrying on of an organised and systematised activity that qualifies such activity as having a professional nature. As one can find the new civil code has changed the conception regarding the enterprise, as it resulted from the actual commercial code. The operation of an enterprise will represent the legal form of carrying on an activity having a professional nature.

  8. Structural studies of a cold-adapted dimeric β-D-galactosidase from Paracoccus sp. 32d.

    Science.gov (United States)

    Rutkiewicz-Krotewicz, Maria; Pietrzyk-Brzezinska, Agnieszka J; Sekula, Bartosz; Cieśliński, Hubert; Wierzbicka-Woś, Anna; Kur, Józef; Bujacz, Anna

    2016-09-01

    The crystal structure of a novel dimeric β-D-galactosidase from Paracoccus sp. 32d (ParβDG) was solved in space group P212121 at a resolution of 2.4 Å by molecular replacement with multiple models using the BALBES software. This enzyme belongs to glycoside hydrolase family 2 (GH2), similar to the tetrameric and hexameric β-D-galactosidases from Escherichia coli and Arthrobacter sp. C2-2, respectively. It is the second known structure of a cold-active GH2 β-galactosidase, and the first in the form of a functional dimer, which is also present in the asymmetric unit. Cold-adapted β-D-galactosidases have been the focus of extensive research owing to their utility in a variety of industrial technologies. One of their most appealing applications is in the hydrolysis of lactose, which not only results in the production of lactose-free dairy, but also eliminates the `sandy effect' and increases the sweetness of the product, thus enhancing its quality. The determined crystal structure represents the five-domain architecture of the enzyme, with its active site located in close vicinity to the dimer interface. To identify the amino-acid residues involved in the catalytic reaction and to obtain a better understanding of the mechanism of action of this atypical β-D-galactosidase, the crystal structure in complex with galactose (ParβDG-Gal) was also determined. The catalytic site of the enzyme is created by amino-acid residues from the central domain 3 and from domain 4 of an adjacent monomer. The crystal structure of this dimeric β-D-galactosidase reveals significant differences in comparison to other β-galactosidases. The largest difference is in the fifth domain, named Bgal_windup domain 5 in ParβDG, which contributes to stabilization of the functional dimer. The location of this domain 5, which is unique in size and structure, may be one of the factors responsible for the creation of a functional dimer and cold-adaptation of this enzyme.

  9. Activities of aluminum forms of zeolites and zeolite-containing aluminosilicates in phenol arylalkylation by styrene

    Energy Technology Data Exchange (ETDEWEB)

    Kurashev, M.V.; Kolesnichenko, N.V.; Romanovskii, B.V.; Lokshin, B.V.

    1979-03-01

    The activities of aluminum forms of zeolites and zeolite-containing aluminosilicates in phenol arylalkylation by styrene were studied in a flow microreactor at 250/sup 0/C, 1 atm, 4:1 phenol-styrene, and 2.6 sec contact time. The aluminum-exchanged forms of a NaY zeolite and a zeolite-containing amorphous aluminosilicate (the Soviet commercial AShNTs-3 grade) were prepared by the K. M. Wang technique and some of the samples were pretreated by phenol at 250/sup 0/C and 2 atm for two hours. The AShNTs-3 and Al/AShNTs-3 catalysts showed maximum steady-state activities in terms of over-all styrene conversion (67.6 and 46.7Vertical Bar3<, respectively) and the combined yield of 2- and 4-(..cap alpha..-methylbenzyl)phenols (64.1 and 32.2Vertical Bar3<, respectively). The AlNaY zeolite rapidly and irreversibly deactivated, and pure amorphous aluminosilicate (AAS) showed relatively low activity. The pretreatment with phenol improved the activities of the aluiminum forms and increased the ortho-to-para isomer ratio of the alkylate from 2.74 to 4.33 for Al/AShNTs-3 and from 2.37 to 4.00 for AAS, probably because of the formation of active surface aluminum phenolate structures, but deteriorated the catalytic properties of AShNTs-3.

  10. Binary Toxin Subunits of Lysinibacillus sphaericus Are Monomeric and Form Heterodimers after In Vitro Activation.

    Directory of Open Access Journals (Sweden)

    Wahyu Surya

    Full Text Available The binary toxin from Lysinibacillus sphaericus has been successfully used for controlling mosquito-transmitted diseases. An activation step shortens both subunits BinA and BinB before their interaction with membranes and internalization in midgut cells, but the precise role of this activation step is unknown. Herein, we show conclusively using three orthogonal biophysical techniques that protoxin subunits form only monomers in aqueous solution. However, in vitro activated toxins readily form heterodimers. This oligomeric state did not change after incubation of these heterodimers with detergent. These results are consistent with the evidence that maximal toxicity in mosquito larvae is achieved when the two subunits, BinA and BinB, are in a 1:1 molar ratio, and directly link proteolytic activation to heterodimerization. Formation of a heterodimer must thus be necessary for subsequent steps, e.g., interaction with membranes, or with a suitable receptor in susceptible mosquito species. Lastly, despite existing similarities between BinB C-terminal domain with domains 3 and 4 of pore-forming aerolysin, no aerolysin-like SDS-resistant heptameric oligomers were observed when the activated Bin subunits were incubated in the presence of detergents or lipidic membranes.

  11. Biosynthesis of intestinal microvillar proteins. Dimerization of aminopeptidase N and lactase-phlorizin hydrolase

    DEFF Research Database (Denmark)

    Danielsen, E M

    1990-01-01

    The pig intestinal brush border enzymes aminopeptidase N (EC 3.4.11.2) and lactase-phlorizin hydrolase (EC 3.2.1.23-62) are present in the microvillar membrane as homodimers. Dimethyl adipimidate was used to cross-link the two [35S]methionine-labeled brush border enzymes from cultured mucosal...... explants. For aminopeptidase N, dimerization did not begin until 5-10 min after synthesis, and maximal dimerization by cross-linking of the transient form of the enzyme required 1 h, whereas the mature form of aminopeptidase N cross-linked with unchanged efficiency from 45 min to 3 h of labeling. Formation...... of dimers of this enzyme therefore occurs prior to the Golgi-associated processing, and the slow rate of dimerization may be the rate-limiting step in the transport from the endoplasmic reticulum to the Golgi complex. For lactase-phlorizin hydrolase, the posttranslational processing includes a proteolytic...

  12. Mahler Measure, Eisenstein Series and Dimers

    NARCIS (Netherlands)

    Stienstra, J.

    2007-01-01

    This note reveals a mysterious link between the partition function of certain dimer models on 2-dimensional tori and the L-function of their spectral curves. It also relates the partition function in certain families of dimer models to Eisenstein series. http://www.arxiv.org/abs/math.NT/0502197

  13. Dimeric assembly of enterocyte brush border enzymes

    DEFF Research Database (Denmark)

    Danielsen, E M

    1994-01-01

    temperature (20 degrees C) reduced the rate of, but did not prevent, dimerization. Maltase-glucoamylase (EC 3.2.1.20) only appeared as a dimer when extracted and analyzed under low salt conditions, suggesting a weak association between the two subunits. This finding is consistent with the electronmicroscopic...

  14. Influence of small forms of active rest on intellection of students of agrarian higher institutes.

    Directory of Open Access Journals (Sweden)

    Prysjazhnuk S.I.

    2012-12-01

    Full Text Available Influence of facilities of active rest is considered on the increase of mental capacity of students. In research took part 105 students with the low level of physical preparedness. Progress of students is select the integral index of mental capacity. It is set that the most effective factors of diminishing of action of processes of fatigue and increase of mental capacity of students is the use of small forms of active rest during an educational day. It is suggested to utillize athletic pauses (minutes, micro pauses during the leadthrough of employments on theoretical disciplines. It is set that small forms of active rest in combination with a leadthrough independent health-improvement-trainings employments on physical self-perfection instrumental in the substantial improvement of the state of physical and mental capacity of student young people.

  15. Potassium Hexacyanoferrate (III-Catalyzed Dimerization of Hydroxystilbene: Biomimetic Synthesis of Indane Stilbene Dimers

    Directory of Open Access Journals (Sweden)

    Jing-Shan Xie

    2015-12-01

    Full Text Available Using potassium hexacyanoferrate (III–sodium acetate as oxidant, the oxidative coupling reaction of isorhapontigenin and resveratrol in aqueous acetone resulted in the isolation of three new indane dimers 4, 6, and 7, together with six known stilbene dimers. Indane dimer 5 was obtained for the first time by direct transformation from isorhapontigenin. The structures and relative configurations of the dimers were elucidated using spectral analysis, and their possible formation mechanisms were discussed. The results indicate that this reaction could be used as a convenient method for the semi-synthesis of indane dimers because of the mild conditions and simple reaction products.

  16. Intramolecular hydrophobic interactions are critical mediators of STAT5 dimerization

    Science.gov (United States)

    Fahrenkamp, Dirk; Li, Jinyu; Ernst, Sabrina; Schmitz-van de Leur, Hildegard; Chatain, Nicolas; Küster, Andrea; Koschmieder, Steffen; Lüscher, Bernhard; Rossetti, Giulia; Müller-Newen, Gerhard

    2016-10-01

    STAT5 is an essential transcription factor in hematopoiesis, which is activated through tyrosine phosphorylation in response to cytokine stimulation. Constitutive activation of STAT5 is a hallmark of myeloid and lymphoblastic leukemia. Using homology modeling and molecular dynamics simulations, a model of the STAT5 phosphotyrosine-SH2 domain interface was generated providing first structural information on the activated STAT5 dimer including a sequence, for which no structural information is available for any of the STAT proteins. We identified a novel intramolecular interaction mediated through F706, adjacent to the phosphotyrosine motif, and a unique hydrophobic interface on the surface of the SH2 domain. Analysis of corresponding STAT5 mutants revealed that this interaction is dispensable for Epo receptor-mediated phosphorylation of STAT5 but essential for dimer formation and subsequent nuclear accumulation. Moreover, the herein presented model clarifies molecular mechanisms of recently discovered leukemic STAT5 mutants and will help to guide future drug development.

  17. Inhibition of HIV-1 Reverse Transcriptase Dimerization by Small Molecules.

    Science.gov (United States)

    Tintori, Cristina; Corona, Angela; Esposito, Francesca; Brai, Annalaura; Grandi, Nicole; Ceresola, Elisa Rita; Clementi, Massimo; Canducci, Filippo; Tramontano, Enzo; Botta, Maurizio

    2016-04-15

    Because HIV-1 reverse transcriptase is an enzyme whose catalytic activity depends on its heterodimeric structure, this system could be a target for inhibitors that perturb the interactions between the protein subunits, p51 and p66. We previously demonstrated that the small molecule MAS0 reduced the association of the two RT subunits and simultaneously inhibited both the polymerase and ribonuclease H activities. In this study, some analogues of MAS0 were rationally selected by docking studies and evaluated in vitro for their ability to disrupt dimeric assembly. Two inhibitors were identified with improved activity compared to MAS0. This study lays the basis for the rational design of more potent inhibitors of RT dimerization.

  18. Structure of the active form of human origin recognition complex and its ATPase motor module

    Energy Technology Data Exchange (ETDEWEB)

    Tocilj, Ante; On, Kin Fan; Yuan, Zuanning; Sun, Jingchuan; Elkayam, Elad; Li, Huilin; Stillman, Bruce; Joshua-Tor, Leemor

    2017-01-23

    Binding of the Origin Recognition Complex (ORC) to origins of replication marks the first step in the initiation of replication of the genome in all eukaryotic cells. Here, we report the structure of the active form of human ORC determined by X-ray crystallography and cryo-electron microscopy. The complex is composed of an ORC1/4/5 motor module lobe in an organization reminiscent of the DNA polymerase clamp loader complexes. A second lobe contains the ORC2/3 subunits. The complex is organized as a double-layered shallow corkscrew, with the AAA+ and AAA+-like domains forming one layer, and the winged-helix domains (WHDs) forming a top layer. CDC6 fits easily between ORC1 and ORC2, completing the ring and the DNA-binding channel, forming an additional ATP hydrolysis site. Analysis of the ATPase activity of the complex provides a basis for understanding ORC activity as well as molecular defects observed in Meier-Gorlin Syndrome mutations.

  19. Structure of the active form of human origin recognition complex and its ATPase motor module

    Science.gov (United States)

    Tocilj, Ante; On, Kin Fan; Yuan, Zuanning; Sun, Jingchuan; Elkayam, Elad; Li, Huilin; Stillman, Bruce; Joshua-Tor, Leemor

    2017-01-01

    Binding of the Origin Recognition Complex (ORC) to origins of replication marks the first step in the initiation of replication of the genome in all eukaryotic cells. Here, we report the structure of the active form of human ORC determined by X-ray crystallography and cryo-electron microscopy. The complex is composed of an ORC1/4/5 motor module lobe in an organization reminiscent of the DNA polymerase clamp loader complexes. A second lobe contains the ORC2/3 subunits. The complex is organized as a double-layered shallow corkscrew, with the AAA+ and AAA+-like domains forming one layer, and the winged-helix domains (WHDs) forming a top layer. CDC6 fits easily between ORC1 and ORC2, completing the ring and the DNA-binding channel, forming an additional ATP hydrolysis site. Analysis of the ATPase activity of the complex provides a basis for understanding ORC activity as well as molecular defects observed in Meier-Gorlin Syndrome mutations. DOI: http://dx.doi.org/10.7554/eLife.20818.001 PMID:28112645

  20. Universal four-Boson states in ultracold molecular gases: resonant effects in dimer-dimer collisions.

    Science.gov (United States)

    D'Incao, J P; von Stecher, J; Greene, Chris H

    2009-07-17

    We study the manifestations of universal four-body physics in ultracold dimer-dimer collisions. We show that resonant features associated with three-body Efimov physics and dimer-dimer scattering lengths are universally related. The emergence of universal four-boson states allows for the tunability of the dimer-dimer interaction, thus enabling the future study of ultracold molecular gases with both attractive and repulsive interactions. Moreover, our study of the interconversion between dimers and Efimov trimers shows that B2+B2-->B3+B rearrangement reactions can provide an efficient trimer formation mechanism. Our analysis of the temperature dependence of this reaction provides an interpretation of the available experimental data and sheds light on the possible experimental realization of rearrangement processes in ultracold gases.

  1. Determinants of Practising Selected Forms of Physical Activity in a Group of Administrative and Office Workers

    Directory of Open Access Journals (Sweden)

    Kowalczyk Anna

    2016-03-01

    Full Text Available Introduction. In recent years, a decline in the level of physical activity has been observed all over the world. The number of professions where work is performed in a sitting position has increased. This has had many consequences for our health, the society, and the economy. The aim of this work was to determine which forms of physical activity are the most popular in administrative and office workers, depending on the motives which encourage them to be active. Material and methods. In 2014, a diagnostic survey was carried out among 937 persons in administrative and office positions using a questionnaire form designed by the authors. The study involved persons aged 18 to 65 years, and most of the respondents were female (n = 669. A qualitative analysis of the data was carried out using logistic regression, and the findings were considered statistically significant at p < 0.05. Results. Changing the shape of one’s body was found to be the main determinant of using the gym among the respondents. Persons who jogged regularly, on the other hand, did so in order to increase physical fitness, and those who practised Nordic walking were motivated by the need to care for their health. As far as swimming is concerned, persons who had friends that engaged in this form of activity undertook it almost ten times more often than those who did not have such support from their family and friends (OR = 9.58. Respondents who desired to meet new people were over five times more likely to choose team games as an active form of spending their leisure time (OR = 5.21 than other respondents. Finally, those who engaged in physical activity in order to strengthen family bonds preferred playing and playing games with children in the open air. Conclusions. The predominant forms of physical activity which were regularly performed by the respondents were walking, cycling, and doing gymnastic exercise at home. The respondents were mainly motivated to pursue these

  2. Effects of pH and temperature on dimerization rate of glycine: Evaluation of favorable environmental conditions for chemical evolution of life

    Science.gov (United States)

    Sakata, Kasumi; Kitadai, Norio; Yokoyama, Tadashi

    2010-12-01

    To evaluate favorable environmental conditions for the chemical evolution of life, we studied the effects of pH and temperature on the dimerization rate of glycine (Gly: NH 2-CH 2-COOH), one of the simplest amino acids. Gly dimerizes to form glycylglycine (GlyGly), and GlyGly further reacts to form diketopiperazine (DKP). Gly solutions with pH ranging from 3.1 to 10.9 were heated for 1-14 days at 140 °C, and changes in concentrations of Gly, GlyGly, and DKP were evaluated. At pH 9.8, the experiments were conducted at 120, 140, 160, and 180 °C. The dimerization rate of Gly was nearly constant at pH 3-7 and increased with increasing pH from 7 to 9.8 and then decreased with further increases in pH. To elucidate the reason for this pH dependency, we evaluated the role of the three dissociation states of Gly (cationic state: Gly +, zwitterionic state: Gly ±, and anionic state: Gly -). For pH >6, the dominant forms are Gly ± and Gly -, and the molar fraction of Gly ± decreases and that of Gly - increases with increasing pH. The dimerization rate was determined for each dissociation state. The reaction between Gly ± and Gly - was found to be the fastest; the rate constant of the reaction between Gly ± and Gly - was 10 times the size of that between Gly - and Gly - and 98 times that between Gly ± and Gly ±. The dimerization rate became greatest at pH 9.8 because the molar fractions of Gly ± and Gly - are approximately equal at this pH. The dimerization rate increased with temperature, and an activation energy of 88 kJ mol -1 was obtained. Based on these results and previous reports on the stability of amino acids under hydrothermal conditions, we determined that Gly dimerizes most efficiently under alkaline pH (˜9.8) at about 150 °C.

  3. Actively Star Forming Elliptical Galaxies at Low Redshifts in the Sloan Digital Sky Survey

    CERN Document Server

    Fukugita, M; Turner, E L; Helmboldt, J; Nichol, R C; Fukugita, Masataka; Nakamura, Osamu; Turner, Edwin L.; Helmboldt, Joe

    2004-01-01

    We report discovery of actively star forming elliptical galaxies in a morphologically classified sample of bright galaxies at a low redshift obtained from the Sloan Digital Sky Survey. The emission lines of these galaxies do not show the characteristics of active galactic nuclei, and thus their strong H$\\alpha$ emission is ascribed to star formation with a rate nearly as high as that is seen in typical late spiral galaxies. This is taken as evidence against the traditional view that all elliptical galaxies formed early and now evolve only passively. The frequency of such star forming elliptical galaxies is a few tenths of a percent in the sample, but increases to 3% if we include active S0 galaxies. We may identify these galaxies as probable progenitors of so-called E+A galaxies that show the strong Balmer absorption feature of A stars superimposed on an old star population. The approximate match of the abundance of active elliptical plus S0 galaxies with that of E+A galaxies indicates that the duration of su...

  4. Dimerization of ethylene in the presence of homogeneous catalysts containing alkyl-substituted nickelocene

    Energy Technology Data Exchange (ETDEWEB)

    Dzhabiyeva, Z.M.; Belov, G.P.; Mar`in, V.P. [Institute of Chemical Physics, Chernogolovka (Russian Federation)

    1993-12-31

    A study has been made of the effect of the temperature and the substituent in the cyclopentadienyl ligand of the compounds (C{sub 5}H{sub 4}R){sub 2}Ni, where R = H, C{sub 2}H{sub 5}, n-C{sub 3}H{sub 7}, iso-C{sub 3}H{sub 7} and tert-C{sub 4}H{sub 9}, and (C{sub 5}H{sub 4}R)Ni-C{sub 3}H{sub 5}, where R = H or iso-C{sub 3}H{sub 7}-, on the kinetics and selectivity of ethylene dimerization reactions. It has been established that greatest activity is exhibited by (C{sub 5}H{sub 4}- iso-C{sub 3}H{sub 7}){sub 2} Ni. The influence of the Al/Ni molar ratio on the kinetics and selectivity of dimerization in the presence of the catalytic system (C{sub 5}H{sub 4}-iso-C{sub 3}H{sub 7}){sub 2}Ni-C{sub 2}H{sub 5}AlCl{sub 2}, has been studied. The nature of the active centres formed when mono- and bicomponent catalysts containing nickelocene are used is discussed. 6 refs., 4 figs., 2 tab.

  5. Structural and mechanistic insights into cooperative assembly of dimeric Notch transcription complexes

    Energy Technology Data Exchange (ETDEWEB)

    Arnett, Kelly L.; Hass, Matthew; McArthur, Debbie G.; Ilagan, Ma Xenia G.; Aster, Jon C.; Kopan, Raphael; Blacklow, Stephen C. (WU); (BWH); (DFCI)

    2010-11-12

    Ligand-induced proteolysis of Notch produces an intracellular effector domain that transduces essential signals by regulating the transcription of target genes. This function relies on the formation of transcriptional activation complexes that include intracellular Notch, a Mastermind co-activator and the transcription factor CSL bound to cognate DNA. These complexes form higher-order assemblies on paired, head-to-head CSL recognition sites. Here we report the X-ray structure of a dimeric human Notch1 transcription complex loaded on the paired site from the human HES1 promoter. The small interface between the Notch ankyrin domains could accommodate DNA bending and untwisting to allow a range of spacer lengths between the two sites. Cooperative dimerization occurred on the human and mouse Hes5 promoters at a sequence that diverged from the CSL-binding consensus at one of the sites. These studies reveal how promoter organizational features control cooperativity and, thus, the responsiveness of different promoters to Notch signaling.

  6. Participation of the extracellular domain in (pro)renin receptor dimerization

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki-Nakagawa, Chiharu [The United Graduate School of Agricultural Sciences, Gifu University, Gifu (Japan); Nishimura, Misa; Tsukamoto, Tomoko; Aoyama, Sho [Department of Applied Life Science, Faculty of Applied Biological Sciences, Gifu University, Gifu (Japan); Ebihara, Akio; Suzuki, Fumiaki [The United Graduate School of Agricultural Sciences, Gifu University, Gifu (Japan); Department of Applied Life Science, Faculty of Applied Biological Sciences, Gifu University, Gifu (Japan); Nakagawa, Tsutomu, E-mail: nakagawa@gifu-u.ac.jp [The United Graduate School of Agricultural Sciences, Gifu University, Gifu (Japan); Department of Applied Life Science, Faculty of Applied Biological Sciences, Gifu University, Gifu (Japan)

    2014-02-21

    Highlights: • The (pro)renin receptor [(P)RR] is a regulator of the renin–angiotensin system. • The region responsible for (P)RR dimerization was investigated. • (P)RR extracellular domain constructs were retained intracellularly. • The extracellular domain of (P)RR is responsible for its dimerization. • Novel insight into the regulatory mechanism of soluble (P)RR secretion is provided. - Abstract: The (pro)renin receptor [(P)RR] induces the catalytic activation of prorenin, as well as the activation of the mitogen-activated protein kinase (MAPK) signaling pathway; as such, it plays an important regulatory role in the renin–angiotensin system. (P)RR is known to form a homodimer, but the region participating in its dimerization is unknown. Using glutathione S-transferase (GST) as a carrier protein and a GST pull-down assay, we investigated the interaction of several (P)RR constructs with full-length (FL) (P)RR in mammalian cells. GST fusion proteins with FL (P)RR (GST-FL), the C-terminal M8-9 fragment (GST-M8-9), the extracellular domain (ECD) of (P)RR (GST-ECD), and the (P)RR ECD with a deletion of 32 amino acids encoded by exon 4 (GST-ECDd4) were retained intracellularly, whereas GST alone was efficiently secreted into the culture medium when transiently expressed in COS-7 cells. Immunofluorescence microscopy showed prominent localization of GST-ECD to the endoplasmic reticulum. The GST pull-down analysis revealed that GST-FL, GST-ECD, and GST-ECDd4 bound FLAG-tagged FL (P)RR, whereas GST-M8-9 showed little or no binding when transiently co-expressed in HEK293T cells. Furthermore, pull-down analysis using His-tag affinity resin showed co-precipitation of soluble (P)RR with FL (P)RR from a stable CHO cell line expressing FL h(P)RR with a C-terminal decahistidine tag. These results indicate that the (P)RR ECD participates in dimerization.

  7. The E2 Domains of APP and APLP1 Share a Conserved Mode of Dimerization

    Energy Technology Data Exchange (ETDEWEB)

    S Lee; Y Xue; J Hulbert; Y Wang; X Liu; B Demeler; Y Ha

    2011-12-31

    Amyloid precursor protein (APP) is genetically linked to Alzheimer's disease. APP is a type I membrane protein, and its oligomeric structure is potentially important because this property may play a role in its function or affect the processing of the precursor by the secretases to generate amyloid {beta}-peptide. Several independent studies have shown that APP can form dimers in the cell, but how it dimerizes remains controversial. At least three regions of the precursor, including a centrally located and conserved domain called E2, have been proposed to contribute to dimerization. Here we report two new crystal structures of E2, one from APP and the other from APLP1, a mammalian APP homologue. Comparison with an earlier APP structure, which was determined in a different space group, shows that the E2 domains share a conserved and antiparallel mode of dimerization. Biophysical measurements in solution show that heparin binding induces E2 dimerization. The 2.1 {angstrom} resolution electron density map also reveals phosphate ions that are bound to the protein surface. Mutational analysis shows that protein residues interacting with the phosphate ions are also involved in heparin binding. The locations of two of these residues, Arg-369 and His-433, at the dimeric interface suggest a mechanism for heparin-induced protein dimerization.

  8. The REC domain mediated dimerization is critical for FleQ from Pseudomonas aeruginosa to function as a c-di-GMP receptor and flagella gene regulator.

    Science.gov (United States)

    Su, Tiantian; Liu, Shiheng; Wang, Kang; Chi, Kaikai; Zhu, Deyu; Wei, Tiandi; Huang, Yan; Guo, Liming; Hu, Wei; Xu, Sujuan; Lin, Zong; Gu, Lichuan

    2015-10-01

    FleQ is an AAA+ ATPase enhancer-binding protein that regulates both flagella and biofilm formation in the opportunistic pathogen Pseudomonas aeruginosa. FleQ belongs to the NtrC subfamily of response regulators, but lacks the corresponding aspartic acid for phosphorylation in the REC domain (FleQ(R), also named FleQ domain). Here, we show that the atypical REC domain of FleQ is essential for the function of FleQ. Crystal structure of FleQ(R) at 2.3Å reveals that the structure of FleQ(R) is significantly different from the REC domain of NtrC1 which regulates gene expression in a phosphorylation dependent manner. FleQ(R) forms a novel active dimer (transverse dimer), and mediates the dimerization of full-length FleQ in an unusual manner. Point mutations that affect the dimerization of FleQ lead to loss of function of the protein. Moreover, a c-di-GMP binding site deviating from the previous reported one is identified through structure analysis and point mutations.

  9. Biosynthesis of intestinal microvillar proteins. Dimerization of aminopeptidase N and lactase-phlorizin hydrolase

    Energy Technology Data Exchange (ETDEWEB)

    Danielsen, E.M. (Univ. of Cophenhagen (Denmark))

    1990-01-09

    The pig intestinal brush border enzymes aminopeptidase and lactase-phlorizin hydrolase are present in the microvilla membrane as homodimers. Dimethyl adipimidate was used to cross-link the two ({sup 35}S)methionine-labeled brush border enzymes from cultured mucosal explants. For aminopeptidase N, dimerization did not begin until 5-10 min after synthesis, and maximal dimerization by cross-linking of the transient form of the enzyme required 1 h, whereas the mature form of aminopeptidase N cross-linked with unchanged efficiency from 45 min to 3 h of labeling. Formation of dimers of this enzyme therefore occurs prior to the Golgi-associated processing, and the slow rate of dimerization may be the rate-limiting step in the transport from the endoplasmic reticulum to the Golgi complex. For lactase-phlorizin hydrolase, the posttranslational processing includes a proteolytic cleavage of its high molecular weight precursor. Since only the mature form and not the precursor of this enzyme could be cross-linked, formation of tightly associated dimers only takes place after transport out of the endoplasmic reticulum. Dimerization of the two brush border enzymes therefore seems to occur in different organelles of the enterocyte.

  10. Validity of the international physical activity questionnaire short form (IPAQ-SF: A systematic review

    Directory of Open Access Journals (Sweden)

    Stewart Sunita M

    2011-10-01

    Full Text Available Abstract Background The International Physical Activity Questionnaire - Short Form (IPAQ-SF has been recommended as a cost-effective method to assess physical activity. Several studies validating the IPAQ-SF have been conducted with differing results, but no systematic review of these studies has been reported. Methods The keywords "IPAQ", "validation", and "validity" were searched in PubMed and Scopus. Studies published in English that validated the IPAQ-SF against an objective physical activity measuring device, doubly labeled water, or an objective fitness measure were included. Results Twenty-three validation studies were included in this review. There was a great deal of variability in the methods used across studies, but the results were largely similar. Correlations between the total physical activity level measured by the IPAQ-SF and objective standards ranged from 0.09 to 0.39; none reached the minimal acceptable standard in the literature (0.50 for objective activity measuring devices, 0.40 for fitness measures. Correlations between sections of the IPAQ-SF for vigorous activity or moderate activity level/walking and an objective standard showed even greater variability (-0.18 to 0.76, yet several reached the minimal acceptable standard. Only six studies provided comparisons between physical activity levels derived from the IPAQ-SF and those obtained from objective criterion. In most studies the IPAQ-SF overestimated physical activity level by 36 to 173 percent; one study underestimated by 28 percent. Conclusions The correlation between the IPAQ-SF and objective measures of activity or fitness in the large majority of studies was lower than the acceptable standard. Furthermore, the IPAQ-SF typically overestimated physical activity as measured by objective criterion by an average of 84 percent. Hence, the evidence to support the use of the IPAQ-SF as an indicator of relative or absolute physical activity is weak.

  11. Ultrafast red light activation of Synechocystis phytochrome Cph1 triggers major structural change to form the Pfr signalling-competent state.

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    Derren J Heyes

    Full Text Available Phytochromes are dimeric photoreceptors that regulate a range of responses in plants and microorganisms through interconversion of red light-absorbing (Pr and far-red light-absorbing (Pfr states. Photoconversion between these states is initiated by light-driven isomerization of a bilin cofactor, which triggers protein structural change. The extent of this change, and how light-driven structural changes in the N-terminal photosensory region are transmitted to the C-terminal regulatory domain to initiate the signalling cascade, is unknown. We have used pulsed electron-electron double resonance (PELDOR spectroscopy to identify multiple structural transitions in a phytochrome from Synechocystis sp. PCC6803 (Cph1 by measuring distances between nitroxide labels introduced into the protein. We show that monomers in the Cph1 dimer are aligned in a parallel 'head-to-head' arrangement and that photoconversion between the Pr and Pfr forms involves conformational change in both the N- and C-terminal domains of the protein. Cryo-trapping and kinetic measurements were used to probe the extent and temporal properties of protein motions for individual steps during photoconversion of Cph1. Formation of the primary photoproduct Lumi-R is not affected by changes in solvent viscosity and dielectric constant. Lumi-R formation occurs at cryogenic temperatures, consistent with their being no major structural reorganization of Cph1 during primary photoproduct formation. All remaining steps in the formation of the Pfr state are affected by solvent viscosity and dielectric constant and occur only at elevated temperatures, implying involvement of a series of long-range solvent-coupled conformational changes in Cph1. We show that signalling is achieved through ultrafast photoisomerization where localized structural change in the GAF domain is transmitted and amplified to cause larger-scale and slower conformational change in the PHY and histidine kinase domains. This

  12. Palmitoylated APP Forms Dimers, Cleaved by BACE1

    OpenAIRE

    Bhattacharyya, Raja; Fenn, Rebecca H.; Barren, Cory; Tanzi, Rudolph E.; Kovacs, Dora M.

    2016-01-01

    A major rate-limiting step for Aβ generation and deposition in Alzheimer’s disease brains is BACE1-mediated cleavage (β-cleavage) of the amyloid precursor protein (APP). We previously reported that APP undergoes palmitoylation at two cysteine residues (Cys186 and Cys187) in the E1-ectodomain. 8–10% of total APP is palmitoylated in vitro and in vivo. Palmitoylated APP (palAPP) shows greater preference for β-cleavage than total APP in detergent resistant lipid rafts. Protein palmitoylation is k...

  13. Phenylalanine binding is linked to dimerization of the regulatory domain of phenylalanine hydroxylase.

    Science.gov (United States)

    Zhang, Shengnan; Roberts, Kenneth M; Fitzpatrick, Paul F

    2014-10-28

    Analytical ultracentrifugation has been used to analyze the oligomeric structure of the isolated regulatory domain of phenylalanine hydroxylase. The protein exhibits a monomer-dimer equilibrium with a dissociation constant of ~46 μM; this value is unaffected by the removal of the 24 N-terminal residues or by phosphorylation of Ser16. In contrast, phenylalanine binding (Kd = 8 μM) stabilizes the dimer. These results suggest that dimerization of the regulatory domain of phenylalanine hydroxylase is linked to allosteric activation of the enzyme.

  14. 78 FR 3900 - Generic Drug User Fee-Active Pharmaceutical Ingredient and Finished Dosage Form Facility Fee...

    Science.gov (United States)

    2013-01-17

    ... drug active pharmaceutical ingredient (API) and finished dosage form (FDF) facilities user fees for... applications in the backlog as of October 1, 2012, on finished dosage form (FDF) and active pharmaceutical... Finished Dosage Form Facility Fee Rates for Fiscal Year 2013 AGENCY: Food and Drug Administration,...

  15. Substrate-Induced Dimerization of Engineered Monomeric Variants of Triosephosphate Isomerase from Trichomonas vaginalis.

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    Samuel Lara-Gonzalez

    Full Text Available The dimeric nature of triosephosphate isomerases (TIMs is maintained by an extensive surface area interface of more than 1600 Å2. TIMs from Trichomonas vaginalis (TvTIM are held in their dimeric state by two mechanisms: a ball and socket interaction of residue 45 of one subunit that fits into the hydrophobic pocket of the complementary subunit and by swapping of loop 3 between subunits. TvTIMs differ from other TIMs in their unfolding energetics. In TvTIMs the energy necessary to unfold a monomer is greater than the energy necessary to dissociate the dimer. Herein we found that the character of residue I45 controls the dimer-monomer equilibrium in TvTIMs. Unfolding experiments employing monomeric and dimeric mutants led us to conclude that dimeric TvTIMs unfold following a four state model denaturation process whereas monomeric TvTIMs follow a three state model. In contrast to other monomeric TIMs, monomeric variants of TvTIM1 are stable and unexpectedly one of them (I45A is only 29-fold less active than wild-type TvTIM1. The high enzymatic activity of monomeric TvTIMs contrast with the marginal catalytic activity of diverse monomeric TIMs variants. The stability of the monomeric variants of TvTIM1 and the use of cross-linking and analytical ultracentrifugation experiments permit us to understand the differences between the catalytic activities of TvTIMs and other marginally active monomeric TIMs. As TvTIMs do not unfold upon dimer dissociation, herein we found that the high enzymatic activity of monomeric TvTIM variants is explained by the formation of catalytic dimeric competent species assisted by substrate binding.

  16. Sintering activation of 316L powder using a liquid phase forming powder

    Directory of Open Access Journals (Sweden)

    Nattaya Tosangthum

    2010-03-01

    Full Text Available It was found that the addition of a liquid forming powder (up to 6 wt.% of a gas-atomized tin powder to 316L powdercould activate the sintering process. Sintering activation could be observed by an increase of the sintered density and selected mechanical properties. When optimized tin powder content was used, shorter sintering time and lower sintering temperaturecould produce sintered 316L+tin materials with excellent mechanical properties. Electron dispersive spectroscopy analyses across 316L-tin-316L grains indicated that Ni transportation during the sintering process was enhanced by the presence of liquid tin.

  17. Forming a negative impression of another person correlates with activation in medial prefrontal cortex and amygdala.

    Science.gov (United States)

    Iidaka, Tetsuya; Harada, Tokiko; Sadato, Norihiro

    2011-09-01

    Neural correlates involved in the formation of negative impression from face were investigated using event-related functional magnetic resonance imaging and a partial conditioning paradigm. Eighteen normal volunteers underwent imaging while they viewed the faces of two unfamiliar individuals: one individual's face was partially accompanied by negative emotion but the other's was not. After the volunteers learned the relationship between the faces and the emotion, they formed a more negative impression of the person's face when the emotion was presented. Subtraction analysis of the individuals' neutral faces revealed activation in the dorsal anterior cingulate cortex and superior temporal sulcus, but this activity did not correlate with the change of impression from face. On the other hand, the response in the left amygdala negatively correlated with the change of impression from face in the first run. Time modulation analysis revealed that activity in the dorsomedial prefrontal cortex associated with negative emotion was the largest in the initial part of the acquisition. These results suggest that a negative impression from face may be formed by orchestrated activity in the dorsomedial prefrontal cortex, dorsal anterior cingulate cortex and amygdala, and that the activity has a prominent role in the initial acquisition of negative emotion.

  18. Antimicrobial and Antioxidant Activity of Chitosan/Hydroxypropyl Methylcellulose Film-Forming Hydrosols Hydrolyzed by Cellulase

    Directory of Open Access Journals (Sweden)

    Anna Zimoch-Korzycka

    2016-09-01

    Full Text Available The aim of this study was to evaluate the impact of cellulase (C on the biological activity of chitosan/hydroxypropyl methylcellulose (CH/HPMC film-forming hydrosols. The hydrolytic activity of cellulase in two concentrations (0.05% and 0.1% was verified by determination of the progress of polysaccharide hydrolysis, based on viscosity measurement and reducing sugar-ends assay. The 2,2-diphenyl-1-picrylhydrazyl (DPPH free radical scavenging effect, the ferric reducing antioxidant power (FRAP, and microbial reduction of Pseudomonas fluorescens, Yersinia enterocolitica, Bacillus cereus, and Staphylococcus aureus were studied. During the first 3 h of reaction, relative reducing sugar concentration increased progressively, and viscosity decreased rapidly. With increasing amount of enzyme from 0.05% to 0.1%, the reducing sugar concentration increased, and the viscosity decreased significantly. The scavenging effect of film-forming solutions was improved from 7.6% at time 0 and without enzyme to 52.1% for 0.1% cellulase after 20 h of reaction. A significant effect of cellulase addition and reaction time on antioxidant power of the tested film-forming solutions was also reported. Film-forming hydrosols with cellulase exhibited a bacteriostatic effect on all tested bacteria, causing a total reduction.

  19. Acylphenols and dimeric acylphenols from Myristica maxima Warb.

    Science.gov (United States)

    Othman, Muhamad Aqmal; Sivasothy, Yasodha; Looi, Chung Yeng; Ablat, Abdulwali; Mohamad, Jamaludin; Litaudon, Marc; Awang, Khalijah

    2016-06-01

    Giganteone E (1), a new dimeric acylphenol was isolated as a minor constituent from the bark of Myristica maxima Warb. The structure of 1 was established on the basis of 1D and 2D NMR techniques and LCMS-IT-TOF analysis. Malabaricones A-C (2-4), giganteones A and C (5 and 6), maingayones A and B (7 and 8), maingayic acid B (9) and β-sitosteryl oleate (10) were also characterized in this plant for the first time. Compound 10 was identified for the first time in the Myristicaceae. Compounds 2 and 5 were active against human prostate cancer cell-lines, thus making this the first report on the prostate cancer inhibiting potential of acylphenols and dimeric acylphenols. Compounds 1, 4, 5, 7 and 8 exhibited potent DPPH free radical scavenging activity. This is the first report on their free radical scavenging capacity.

  20. A Natural Chimeric Pseudomonas Bacteriocin with Novel Pore-Forming Activity Parasitizes the Ferrichrome Transporter.

    Science.gov (United States)

    Ghequire, Maarten G K; Kemland, Lieselore; Anoz-Carbonell, Ernesto; Buchanan, Susan K; De Mot, René

    2017-02-21

    Modular bacteriocins represent a major group of secreted protein toxins with a narrow spectrum of activity, involved in interference competition between Gram-negative bacteria. These antibacterial proteins include a domain for binding to the target cell and a toxin module at the carboxy terminus. Self-inhibition of producers is provided by coexpression of linked immunity genes that transiently inhibit the toxin's activity through formation of bacteriocin-immunity complexes or by insertion in the inner membrane, depending on the type of toxin module. We demonstrate strain-specific inhibitory activity for PmnH, a Pseudomonas bacteriocin with an unprecedented dual-toxin architecture, hosting both a colicin M domain, potentially interfering with peptidoglycan synthesis, and a novel colicin N-type domain, a pore-forming module distinct from the colicin Ia-type domain in Pseudomonas aeruginosa pyocin S5. A downstream-linked gene product confers PmnH immunity upon susceptible strains. This protein, ImnH, has a transmembrane topology similar to that of Pseudomonas colicin M-like and pore-forming immunity proteins, although homology with either of these is essentially absent. The enhanced killing activity of PmnH under iron-limited growth conditions reflects parasitism of the ferrichrome-type transporter for entry into target cells, a strategy shown here to be used as well by monodomain colicin M-like bacteriocins from pseudomonads. The integration of a second type of toxin module in a bacteriocin gene could offer a competitive advantage against bacteria displaying immunity against only one of both toxic activities.IMPORTANCE In their continuous struggle for ecological space, bacteria face a huge load of contenders, including phylogenetically related strains that compete for the same niche. One important group of secreted antibacterial proteins assisting in eliminating these rivals are modular bacteriocins of Gram-negative bacteria, comprising a domain for docking onto the

  1. Aberrant phenotypes of transgenic mice expressing dimeric human erythropoietin

    Directory of Open Access Journals (Sweden)

    Yun Seong-Jo

    2012-01-01

    Full Text Available Abstract Background Dimeric human erythropoietin (dHuEPO peptides are reported to exhibit significantly higher biological activity than the monomeric form of recombinant EPO. The objective of this study was to produce transgenic (tg mice expressing dHuEPO and to investigate the characteristics of these mice. Methods A dHuEPO-expressing vector under the control of the goat beta-casein promoter, which produced a dimer of human EPO molecules linked by a 2-amino acid peptide linker (Asp-Ile, was constructed and injected into 1-cell fertilized embryos by microinjection. Mice were screened using genomic DNA samples obtained from tail biopsies. Blood samples were obtained by heart puncture using heparinized tubes, and hematologic parameters were assessed. Using the microarray analysis tool, we analyzed differences in gene expression in the spleens of tg and control mice. Results A high rate of spontaneous abortion or death of the offspring was observed in the recipients of dHuEPO embryos. We obtained 3 founder lines (#4, #11, and #47 of tg mice expressing the dHuEPO gene. However, only one founder line showed stable germline integration and transmission, subsequently establishing the only transgenic line (#11. We obtained 2 F1 mice and 3 F2 mice from line #11. The dHuEPO protein could not be obtained because of repeated spontaneous abortions in the tg mice. Tg mice exhibited symptoms such as short lifespan and abnormal blood composition. The red blood cell count, white blood cell count, and hematocrit levels in the tg mice were remarkably higher than those in the control mice. The spleens of the tg mice (F1 and F2 females were 11- and -21-fold larger than those of the control mice. Microarray analysis revealed 2,672 spleen-derived candidate genes; more genes were downregulated than upregulated (849/764. Reverse transcriptase-polymerase chain reaction (RT-PCR and quantitative real-time PCR (qRT-PCR were used for validating the results of the microarray

  2. A hydrodynamic analysis of APOBEC3G reveals a monomer-dimer-tetramer self-association that has implications for anti-HIV function.

    Science.gov (United States)

    Salter, Jason D; Krucinska, Jolanta; Raina, Jay; Smith, Harold C; Wedekind, Joseph E

    2009-11-17

    The innate antiviral factor APOBEC3G (A3G) possesses RNA binding activity and deaminates HIV-1 DNA. High-molecular mass forms of A3G can be isolated from a variety of cell types but exhibit limited deaminase activity relative to low-molecular mass species prepared under RNA-depleted conditions. To investigate the fundamental oligomeric state and shape of A3G, we conducted sedimentation velocity analyses of the pure enzyme under RNA-deficient conditions. The results reveal a predominant dimer in equilibrium with minor monomeric and tetrameric species. Hydrodynamic modeling of the dimer supports an extended cylindrical shape that assembles into an elongated tetramer. Overall, the results provide physical restraints for the A3G quaternary structure that have implications for modulating antiviral function.

  3. Pore-forming activity of alpha-toxin is essential for clostridium septicum-mediated myonecrosis.

    Science.gov (United States)

    Kennedy, Catherine L; Lyras, Dena; Cordner, Leanne M; Melton-Witt, Jody; Emmins, John J; Tweten, Rodney K; Rood, Julian I

    2009-03-01

    Clostridium septicum alpha-toxin is a beta-barrel pore-forming cytolysin that is functionally similar to aerolysin. Residues important in receptor binding, oligomerization, and pore formation have been identified; however, little is known about the activity of the toxin in an infection, although it is essential for disease. We have now shown that deletion of a small portion of the transmembrane domain, so that the toxin is no longer able to form pores, completely abrogates its ability to contribute to disease, as does replacement of the sole cysteine residue with leucine. However, although previous biochemical and cytotoxicity assays clearly indicated that mutations in residues important in oligomerization, binding, and prepore conversion greatly reduced activity or rendered the toxin inactive, once the mutated toxins were overexpressed by the natural host in the context of an infection it was found they were able to cause disease in a mouse model of myonecrosis. These results highlight the importance of testing the activity of virulence determinants in the normal host background and in an infectious disease context and provide unequivocal evidence that it is the ability of alpha-toxin to form a pore that confers its toxicity in vivo.

  4. Effects of pharmaceutical processing on pepsin activity during the formulation of solid dosage forms.

    Science.gov (United States)

    Kristó, Katalin; Pintye-Hódi, Klára

    2013-02-01

    The main aim of this study was to investigate the effects of pharmaceutical technological methods on pepsin activity during the formulation of solid dosage forms. The circumstances of direct compression and wet granulation were modeled. During direct compression, the heat and the compression force must be taken into consideration. The effects of these parameters were investigated in three materials (pure pepsin, and 1:1 (w/w) pepsin-tartaric acid and 1:1 (w/w) pepsin-citric acid powder mixtures). It was concluded that direct compression is appropriate for the formulation of solid dosage forms containing pepsin through application without acids or with acids at low compression force. The effects of wet granulation were investigated with a factorial design for the same three materials. The factors were time, temperature and moisture content. There was no significant effect of the factors when acids were not applied. Temperature was a significant factor when acids were applied. The negative effect was significantly higher for citric acid than for tartaric acid. It was found that wet granulation can be utilized for the processing of pepsin into solid dosage forms under well-controlled circumstances. The application of citric acid is not recommended during the formulation of solid dosage forms through wet granulation. A mathematically based optimization may be necessary for preformulation studies of the preparation of dosage forms containing sensitive enzymes.

  5. Structural and thermodynamic insight into the process of "weak" dimerization of the ErbB4 transmembrane domain by solution NMR.

    Science.gov (United States)

    Bocharov, Eduard V; Mineev, Konstantin S; Goncharuk, Marina V; Arseniev, Alexander S

    2012-09-01

    Specific helix-helix interactions between the single-span transmembrane domains of receptor tyrosine kinases are believed to be important for their lateral dimerization and signal transduction. Establishing structure-function relationships requires precise structural-dynamic information about this class of biologically significant bitopic membrane proteins. ErbB4 is a ubiquitously expressed member of the HER/ErbB family of growth factor receptor tyrosine kinases that is essential for the normal development of various adult and fetal human tissues and plays a role in the pathobiology of the organism. The dimerization of the ErbB4 transmembrane domain in membrane-mimicking lipid bicelles was investigated by solution NMR. In a bicellar DMPC/DHPC environment, the ErbB4 membrane-spanning α-helices (651-678)(2) form a right-handed parallel dimer through the N-terminal double GG4-like motif A(655)GxxGG(660) in a fashion that is believed to permit proper kinase domain activation. During helix association, the dimer subunits undergo a structural adjustment (slight bending) with the formation of a network of inter-monomeric polar contacts. The quantitative analysis of the observed monomer-dimer equilibrium provides insights into the kinetics and thermodynamics of the folding process of the helical transmembrane domain in the model environment that may be directly relevant to the process that occurs in biological membranes. The lipid bicelles occupied by a single ErbB4 transmembrane domain behave as a true ("ideal") solvent for the peptide, while multiply occupied bicelles are more similar to the ordered lipid microdomains of cellular membranes and appear to provide substantial entropic enhancement of the weak helix-helix interactions, which may be critical for membrane protein activity.

  6. Naturally occurring pentaoxygenated, hexaoxygenated and dimeric xanthones: a literature survey

    Directory of Open Access Journals (Sweden)

    V. Peres

    1997-08-01

    Full Text Available This review gives information on the chemical study of 71 pentaoxygenated, 11 hexaoxygenated and 9 dimeric and more complex xanthones naturally occurring in 7 families, 29 genus and 62 species of higher plants, and 11 described as fern and fungal metabolites. The value of these groups of substances in the connection with the pharmacological activity and the therapeutic use of some species is shown. The structural formulas of 23 isolated compounds and their distribution in the species studied are given.

  7. Quantum dimer model for the pseudogap metal.

    Science.gov (United States)

    Punk, Matthias; Allais, Andrea; Sachdev, Subir

    2015-08-04

    We propose a quantum dimer model for the metallic state of the hole-doped cuprates at low hole density, p. The Hilbert space is spanned by spinless, neutral, bosonic dimers and spin S = 1/2, charge +e fermionic dimers. The model realizes a "fractionalized Fermi liquid" with no symmetry breaking and small hole pocket Fermi surfaces enclosing a total area determined by p. Exact diagonalization, on lattices of sizes up to 8 × 8, shows anisotropic quasiparticle residue around the pocket Fermi surfaces. We discuss the relationship to experiments.

  8. Functional role of dimerization of human peptidylarginine deiminase 4 (PAD4.

    Directory of Open Access Journals (Sweden)

    Yi-Liang Liu

    Full Text Available Peptidylarginine deiminase 4 (PAD4 is a homodimeric enzyme that catalyzes Ca²⁺-dependent protein citrullination, which results in the conversion of arginine to citrulline. This paper demonstrates the functional role of dimerization in the regulation of PAD4 activity. To address this question, we created a series of dimer interface mutants of PAD4. The residues Arg8, Tyr237, Asp273, Glu281, Tyr435, Arg544 and Asp547, which are located at the dimer interface, were mutated to disturb the dimer organization of PAD4. Sedimentation velocity experiments were performed to investigate the changes in the quaternary structures and the dissociation constants (K(d between wild-type and mutant PAD4 monomers and dimers. The kinetic data indicated that disrupting the dimer interface of the enzyme decreases its enzymatic activity and calcium-binding cooperativity. The K(d values of some PAD4 mutants were much higher than that of the wild-type (WT protein (0.45 µM and were concomitant with lower k(cat values than that of WT (13.4 s⁻¹. The K(d values of the monomeric PAD4 mutants ranged from 16.8 to 45.6 µM, and the k(cat values of the monomeric mutants ranged from 3.3 to 7.3 s⁻¹. The k(cat values of these interface mutants decreased as the K(d values increased, which suggests that the dissociation of dimers to monomers considerably influences the activity of the enzyme. Although dissociation of the enzyme reduces the activity of the enzyme, monomeric PAD4 is still active but does not display cooperative calcium binding. The ionic interaction between Arg8 and Asp547 and the Tyr435-mediated hydrophobic interaction are determinants of PAD4 dimer formation.

  9. Variability of the calcium ion activity with pH in stone-forming and non-stone-forming urine.

    Science.gov (United States)

    Thode, J; Holgersen, R B; Gerstenberg, T

    1993-01-01

    In recurrent renal stone-formers (N = 20) and matched healthy adults (N = 20), the actual activity of ionized calcium (alpha Ca2+) and pH were determined in whole urine with an ion-selective electrode. No significant difference was found for the actual median activity of ionized calcium, however the actual median pH was significantly higher in stone-formers compared to healthy adults (pH = 5.57 vs. pH = 5.24; p titration with HCl/NaOH. In all urines the Ca2+ activity decreased with increasing pH in a typical bifasic manner. All curves showed a characteristic "breaking point" at a similar median pH in the stone-formers and in the healthy adults (pH = 6.81 vs. pH = 6.77) (NS). However the slope of the curves in the stone-formers and healthy adults changed from a median value of delta lg alpha Ca2+/delta pH of -0.139 and -0.173 (NS) respectively, to a highly significant difference of -1.326 and -1.053 (p < 0.0001) between the groups, indicating increased binding/precipitation of Ca2+ in stone-formers than in healthy adults supporting the theory of the lack of inhibitors in stone-formers. The strong relationship between the activity of ionized calcium and pH, combined with a higher actual pH and a higher decrease of ionized calcium with pH in stone-formers than in healthy adults, indicates hydrogen ion as a major factor in stone-formation. The close relationship between Ca2+ activity and pH indicates the need for simultaneous measurements of the pH in order to interpret data for the Ca2+ activity. In order to preserve a low urinary pH, where Ca2+ is predominantly in a free ionic state, our results suggest that treatment with acidifying salts could be a logical choice in order to prevent stone-formation.

  10. The Effects of Two Forms of Physical Activity on Eyeblink Classical Conditioning

    Science.gov (United States)

    Green, John T.; Chess, Amy C.; Burns, Montana; Schachinger, Kira M.; Thanellou, Alexandra

    2011-01-01

    Voluntary exercise, in the form of free access to a running wheel in the home cage, has been shown to improve several forms of learning and memory. Acrobatic training, in the form of learning to traverse an elevated obstacle course, has been shown to induce markers of neural plasticity in the cerebellar cortex in rodents. In three experiments, we examined the effects of these two forms of physical activity on delay eyeblink conditioning in rats. In Experiment 1, exercising rats were given 17 days of free access to a running wheel in their home cage prior to 10 days of delay eyeblink conditioning. Rats that exercised conditioned significantly better and showed a larger reflexive eyeblink unconditioned response to the periocular stimulation unconditioned stimulus than rats that did not exercise. In Experiment 2, exercising rats were given 17 days of free access to a running wheel in their home cage prior to 10 days of explicitly unpaired stimulus presentations. Rats that exercised responded the same to tone, light, and periocular stimulation as rats that did not exercise. In Experiment 3, acrobatic training rats were given 15 days of daily training on an elevated obstacle course prior to 10 days of eyeblink conditioning. Activity control rats underwent 15 days of yoked daily running in an open field. Rats that underwent acrobatic training did not differ in eyeblink conditioning from activity control rats. The ability to measure the learned response precisely, and the well-mapped neural circuitry of eyeblink conditioning offer some advantages for the study of exercise effects on learning and memory. PMID:21238502

  11. The effects of two forms of physical activity on eyeblink classical conditioning.

    Science.gov (United States)

    Green, John T; Chess, Amy C; Burns, Montana; Schachinger, Kira M; Thanellou, Alexandra

    2011-05-16

    Voluntary exercise, in the form of free access to a running wheel in the home cage, has been shown to improve several forms of learning and memory. Acrobatic training, in the form of learning to traverse an elevated obstacle course, has been shown to induce markers of neural plasticity in the cerebellar cortex in rodents. In three experiments, we examined the effects of these two forms of physical activity on delay eyeblink conditioning in rats. In Experiment 1, exercising rats were given 17 days of free access to a running wheel in their home cage prior to 10 days of delay eyeblink conditioning. Rats that exercised conditioned significantly better and showed a larger reflexive eyeblink unconditioned response to the periocular stimulation unconditioned stimulus than rats that did not exercise. In Experiment 2, exercising rats were given 17 days of free access to a running wheel in their home cage prior to 10 days of explicitly unpaired stimulus presentations. Rats that exercised responded the same to tone, light, and periocular stimulation as rats that did not exercise. In Experiment 3, acrobatic training rats were given 15 days of daily training on an elevated obstacle course prior to 10 days of eyeblink conditioning. Activity control rats underwent 15 days of yoked daily running in an open field. Rats that underwent acrobatic training did not differ in eyeblink conditioning from activity control rats. The ability to measure the learned response precisely, and the well-mapped neural circuitry of eyeblink conditioning offer some advantages for the study of exercise effects on learning and memory.

  12. Brain activation patterns resulting from learning letter forms through active self-production and passive observation in young children

    Directory of Open Access Journals (Sweden)

    Alyssa J Kersey

    2013-09-01

    Full Text Available Although previous literature suggests that writing practice facilitates neural specialization for letters, it is unclear if this facilitation is driven by the perceptual feedback from the act of writing or the actual execution of the motor act. The present study addresses this issue by measuring the change in BOLD signal in response to hand-printed letters, unlearned cursive letters, and cursive letters that 7 year-old children learned actively, by writing, and passively, by observing an experimenter write. Brain activation was assessed using fMRI while perceiving letters – in both cursive and manuscript forms. Results showed that active training led to increased recruitment of the sensori-motor network associated with letter perception as well as the insula and claustrum, but passive observation did not. This suggests that perceptual networks for newly learned cursive letters are driven by motor execution rather than by perceptual feedback.

  13. Multifunctional porous titanium oxide coating with apatite forming ability and photocatalytic activity on a titanium substrate formed by plasma electrolytic oxidation.

    Science.gov (United States)

    Akatsu, T; Yamada, Y; Hoshikawa, Y; Onoki, T; Shinoda, Y; Wakai, F

    2013-12-01

    Plasma electrolytic oxidation (PEO) was used to make a multifunctional porous titanium oxide (TiO2) coating on a titanium substrate. The key finding of this study is that a highly crystalline TiO2 coating can be made by performing the PEO in an ammonium acetate (CH3COONH4) solution; the PEO coating was formed by alternating between rapid heating by spark discharges and quenching in the solution. The high crystallinity of the TiO2 led to the surface having multiple functions, including apatite forming ability and photocatalytic activity. Hydroxyapatite formed on the PEO coating when it was soaked in simulated body fluid. The good apatite forming ability can be attributed to the high density of hydroxyl groups on the anatase and rutile phases in the coating. The degradation of methylene blue under ultraviolet radiation indicated that the coating had high photocatalytic activity.

  14. A Note on Dimer Models and D-brane Gauge Theories

    CERN Document Server

    Agarwal, Prarit; Sarkar, Tapobrata

    2008-01-01

    The connection between quiver gauge theories and dimer models has been well studied. It is known that the matter fields of the quiver gauge theories can be represented using the perfect matchings of the dimer model. We conjecture that the perfect matchings give information about the charge matrix of the quiver gauge theory. Further, we perform explicit computations on some aspects of partial resolutions of toric singularities using dimer models. We analyse these with graph theory techniques, using the perfect matchings of orbifolds of the form $\\BC^3/\\Gamma$, where the orbifolding group $\\Gamma$ may be noncyclic. Using these, we study the construction of the superpotential of gauge theories living on D-branes which probe these singularities, including the case where one or more adjoint fields are present upon partial resolution. Applying a combination of open and closed string techniques to dimer models, we also study some aspects of their symmetries.

  15. Dimers of cyclic carbonates: chirality recognition in battery solvents and energy storage.

    Science.gov (United States)

    Kollipost, Franz; Hesse, Susanne; Lee, Juhyon J; Suhm, Martin A

    2011-08-21

    Dimers of ethylene carbonate and propylene carbonate are created in supersonic jet expansions and characterized by FTIR spectroscopy. Fermi resonances are switched on and off by dimerization. There is a unique centrosymmetric dimer of ethylene carbonate in a pronounced case of complementary chirality synchronization, contributing to its energy storage capacity at melting. Two chiral propylene carbonate molecules combine in more intricate ways. If they have the same handedness, one of them is forced into an axial conformation and the binding partner stays in the more stable equatorial structure. If they have opposite handedness, centrosymmetric dimers of either axial or equatorial conformations are formed. This suggests the usefulness of chirality control in elucidating ionic transport mechanisms in battery solvents and asymmetric catalysis in such solvents.

  16. Dimerization of the transmembrane domain of amyloid precursor proteins and familial Alzheimer's disease mutants

    Directory of Open Access Journals (Sweden)

    Fraser Paul E

    2008-01-01

    Full Text Available Abstract Background Amyloid precursor protein (APP is enzymatically cleaved by γ-secretase to form two peptide products, either Aβ40 or the more neurotoxic Aβ42. The Aβ42/40 ratio is increased in many cases of familial Alzheimer's disease (FAD. The transmembrane domain (TM of APP contains the known dimerization motif GXXXA. We have investigated the dimerization of both wild type and FAD mutant APP transmembrane domains. Results Using synthetic peptides derived from the APP-TM domain, we show that this segment is capable of forming stable transmembrane dimers. A model of a dimeric APP-TM domain reveals a putative dimerization interface, and interestingly, majority of FAD mutations in APP are localized to this interface region. We find that FAD-APP mutations destabilize the APP-TM dimer and increase the population of APP peptide monomers. Conclusion The dissociation constants are correlated to both the Aβ42/Aβ40 ratio and the mean age of disease onset in AD patients. We also show that these TM-peptides reduce Aβ production and Aβ42/Aβ40 ratios when added to HEK293 cells overexpressing the Swedish FAD mutation and γ-secretase components, potentially revealing a new class of γ-secretase inhibitors.

  17. Special Form Testing of Sealed Source Encapsulation for High-Alpha-Activity Actinide Materials

    Energy Technology Data Exchange (ETDEWEB)

    Martinez, Oscar A [ORNL

    2016-01-01

    In the United States all transportation of radioactive material is regulated by the U.S. Department of Transportation (DOT). Beginning in 2008 a new type of sealed-source encapsulation package was developed and tested by Oak Ridge National Laboratory (ORNL). These packages contain high-alpha-activity actinides and are regulated and transported in accordance with the requirements for DOT Class 7 hazardous material. The DOT provides specific regulations pertaining to special form encapsulation designs. The special form designation indicates that the encapsulated radioactive contents have a very low probability of dispersion even when subjected to significant structural events. The special form designs have been shown to simplify the delivery, transport, acceptance, and receipt processes. It is intended for these sealed-source encapsulations to be shipped to various facilities making it very advantageous for them to be certified as special form. To this end, DOT Certificates of Competent Authority (CoCAs) have been sought for the design suitable for containing high-alpha-activity actinide materials. This design consists of the high-alpha-activity material encapsulated within a triangular zirconia canister, referred to as a ZipCan, tile that is then enclosed by a spherical shell. The spherical shell design, with ZipCan tile inside, was tested for compliance with the special form regulations found in 49 CFR 173.469. The spherical enclosure was subjected to 9-m impact, 1 m percussion, and 10-minute thermal tests at the Packaging Evaluation Facility located at the National Transportation Research Center in Knoxville, TN USA and operated by ORNL. Before and after each test, the test units were subjected to a helium leak check and a bubble test. The ZipCan tiles and core were also subjected to the tests required for ISO 2919:2012(E), including a Class IV impact test and heat test and subsequently subjected to helium leakage rate tests [49 CFR 173.469(a)(4)(i)]. The impact

  18. Quantitative analysis of cyclic dimer fatty acid content in the dimerization product by proton NMR spectroscopy.

    Science.gov (United States)

    Park, Kyun Joo; Kim, Minyoung; Seok, Seunghwan; Kim, Young-Wun; Kim, Do Hyun

    2015-01-01

    In this work, (1)H NMR is utilized for the quantitative analysis of a specific cyclic dimer fatty acid in a dimer acid mixture using the pseudo-standard material of mesitylene on the basis of its structural similarity. Mesitylene and cyclic dimer acid levels were determined using the signal of the proton on the cyclic ring (δ=6.8) referenced to the signal of maleic acid (δ=6.2). The content of the cyclic dimer fatty acid was successfully determined through the standard curve of mesitylene and the reported equation. Using the linearity of the mesitylene curve, the cyclic dimer fatty acid in the oil mixture was quantified. The results suggest that the proposed method can be used to quantify cyclic compounds in mixtures to optimize the dimerization process.

  19. Structure-activity relationship of a recombinant hybrid Manganese superoxide dismutase of Staphylococcus saprophyticus/S. equorum.

    Science.gov (United States)

    Retnoningrum, Debbie S; Arumsari, Sekar; Artarini, Anita; Ismaya, Wangsa T

    2017-05-01

    Recombinant hybrid Manganese superoxide dismutase from Staphyloccus saphropyticus/S. equorum (rMnSODSeq) exhibits stability at high temperatures. The enzyme occurs as a dimer that dissociates around 52°C prior to unfolding of the monomer around 64°C, demonstrating contribution of the dimeric form to stability. Here, structure - activity relationship of rMnSODSeq was evaluated on the basis of its activity and stability in the presence of inhibitors, NaCl, denaturants, detergents, reducing agents, and at different pH values. The activity was evaluated at both 37°C and 52°C, which the latter is the temperature for dissociation of the dimer. Dimer to monomer transition coincided with significant decrease in residual activity at 52°C. However, the activity assay results at 52°C and 37°C suggest spontaneous re-association of the monomer into dimer. Intriguingly, various new species with melting temperature (TM) values other than those of the dimer or monomer were observed. These species displayed medium to comparable level of residual activities to the native at 37°C. This report suggests that dimer to monomer transition may be not the only explanation for activity loss or decrease.

  20. Photoreactivation of pyrimidine dimers in DNA from thyroid cells of the teleost, Poecilia formosa

    Energy Technology Data Exchange (ETDEWEB)

    Achey, P.M.; Woodhead, A.D.; Setlow, R.B.

    1979-01-01

    We have developed and used a simple technique to estimate the quantity of pyrimidine dimers in unlabeled cellular DNA. DNA is extracted from cells, treated with an endonuclease specific for dimers, and its molecular weight estimated by its electrophoretic mobility on alkaline agarose slab gels. The technique is used to show that cells from thyroid tissue of the fish Poecilia formosa have photoreactivating activity towards dimmers in the cellular DNA.

  1. Formation of cystine slipknots in dimeric proteins.

    Directory of Open Access Journals (Sweden)

    Mateusz Sikora

    Full Text Available We consider mechanical stability of dimeric and monomeric proteins with the cystine knot motif. A structure based dynamical model is used to demonstrate that all dimeric and some monomeric proteins of this kind should have considerable resistance to stretching that is significantly larger than that of titin. The mechanisms of the large mechanostability are elucidated. In most cases, it originates from the induced formation of one or two cystine slipknots. Since there are four termini in a dimer, there are several ways of selecting two of them to pull by. We show that in the cystine knot systems, there is strong anisotropy in mechanostability and force patterns related to the selection. We show that the thermodynamic stability of the dimers is enhanced compared to the constituting monomers whereas machanostability is either lower or higher.

  2. A New Dimeric Phthalide from Angelica sinensis

    Institute of Scientific and Technical Information of China (English)

    Ling YI; Ping LI; Zhi Ming BI

    2006-01-01

    A new dimeric phthalide named Z, Z'-3.3'a, 7.7'a-diligustilide was isolated from the roots of Angelica sinensis. Its structure was determined using spectroscopic methods and X-ray crystallographic diffraction analysis.

  3. Formation of Cystine Slipknots in Dimeric Proteins

    OpenAIRE

    Mateusz Sikora; Marek Cieplak

    2013-01-01

    We consider mechanical stability of dimeric and monomeric proteins with the cystine knot motif. A structure based dynamical model is used to demonstrate that all dimeric and some monomeric proteins of this kind should have considerable resistance to stretching that is significantly larger than that of titin. The mechanisms of the large mechanostability are elucidated. In most cases, it originates from the induced formation of one or two cystine slipknots. Since there are four termini in a dim...

  4. Designing Stable Antiparallel Coiled Coil Dimers

    Institute of Scientific and Technical Information of China (English)

    曾宪纲; 周海梦

    2001-01-01

    The history of antiparallel coiled coil dimer design is briefly reviewed and the main principles governing the successful designs are explained. They include analysis of the inter-subunit electrostatic repulsion for determining partners for dimerization and of the buried polar interaction for determining the relative orientation of the partners. A theory is proposed to explain the lack of antiparallel coiled coil homodimers in nature.

  5. [Suppression of telomerase activity leukemic cells by mutant forms of Rhodospirillum rubrum L-asparaginase].

    Science.gov (United States)

    Pokrovskaya, M V; Zhdanov, D D; Eldarov, M A; Aleksandrova, S S; Veselovskiy, A V; Pokrovskiy, V S; Grishin, D V; Gladilina, Ju A; Sokolov, N N

    2017-01-01

    The active and stable mutant forms of short chain cytoplasmic L-asparaginase type I of Rhodospirillum rubrum (RrA): RrA+N17, D60K, F61L, RrA+N17, A64V, E67K, RrA+N17, E149R, V150P, RrAE149R, V150P and RrAE149R, V150P, F151T were obtained by the method of site-directed mutagenesis. It is established that variants RrA-N17, E149R, V150P, F151T and RrАE149R, V150P are capable to reduce an expression hTERT subunit of telomerase and, hence, activity of telomeres in Jurkat cells, but not in cellular lysates. During too time, L-asparaginases of Escherichia coli, Erwinia carotovora and Wolinella succinogenes, mutant forms RrА+N17, D60K, F61L and RrА+N17, A64V, E67K do not suppress of telomerase activity. The assumption of existence in structure RrA of areas (amino acids residues in the position 146-164, 1-17, 60-67) which are responsible for suppression of telomerase activity is made. The received results show that antineoplastic activity of some variants RrA is connected both with reduction of concentration of free L-asparagine, and with expression suppression of hTERT telomerase subunit, that opens new prospects for antineoplastic therapy.

  6. Structure of dimeric, recombinant Sulfolobus solfataricus phosphoribosyl diphosphate synthase

    DEFF Research Database (Denmark)

    Andersen, Rune W.; Lo Leggio, Leila; Hove-Jensen, Bjarne

    2015-01-01

    PRPP synthase as a search model. The two amino acid sequences share 35 % identity. The resulting asymmetric unit consists of three separated dimers. The protein was co-crystallised in the presence of AMP and ribose 5-phosphate, but in the electron density map of the active site only AMP and a sulphate......The enzyme 5-phosphoribosyl-1-α-diphosphate (PRPP) synthase (EC 2.7.6.1) catalyses the Mg2+-dependent transfer of a diphosphoryl group from ATP to the C1 hydroxyl group of ribose 5-phosphate resulting in the production of PRPP and AMP. A nucleotide sequence specifying Sulfolobus solfataricus PRPP....... A bent dimer oligomerisation was revealed, which seems to be an abundant feature among PRPP synthases for defining the adenine specificity of the substrate ATP. Molecular replacement was used to determine the S. solfataricus PRPP synthase structure with a monomer subunit of Methanocaldococcus jannaschii...

  7. Dimerization effect of sucrose octasulfate on rat FGF1

    DEFF Research Database (Denmark)

    Kulahin, Nikolaj; Kiselyov, Vladislav; Kochoyan, Artur

    2008-01-01

    Fibroblast growth factors (FGFs) constitute a family of at least 23 structurally related heparin-binding proteins that are involved in regulation of cell growth, survival, differentiation and migration. Sucrose octasulfate (SOS), a chemical analogue of heparin, has been demonstrated to activate FGF...... signalling pathways. The structure of rat FGF1 crystallized in the presence of SOS has been determined at 2.2 A resolution. SOS-mediated dimerization of FGF1 was observed, which was further supported by gel-filtration experiments. The major contributors to the sulfate-binding sites in rat FGF1 are Lys113......, Lys118, Arg122 and Lys128. An arginine at position 116 is a consensus residue in mammalian FGF molecules; however, it is a serine in rat FGF1. This difference may be important for SOS-mediated FGF1 dimerization in rat....

  8. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    Directory of Open Access Journals (Sweden)

    George Kourouniotis

    2016-07-01

    Full Text Available The binding of epidermal growth factor (EGF to EGF receptor (EGFR stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internalization is still very controversial. In this study, we constructed a chimeric EGFR by replacing its extracellular domain with leucine zipper (LZ and tagged a green fluorescent protein (GFP at its C-terminus. We showed that the chimeric LZ-EGFR-GFP was constitutively dimerized. The LZ-EGFR-GFP dimer autophosphorylated each of its five well-defined C-terminal tyrosine residues as the ligand-induced EGFR dimer does. Phosphorylated LZ-EGFR-GFP was localized to both the plasma membrane and endosomes, suggesting it is capable of endocytosis. We also showed that LZ-EGFR-GFP activated major signalling proteins including Src homology collagen-like (Shc, extracellular signal-regulated kinase (ERK and Akt. Moreover, LZ-EGFR-GFP was able to stimulate cell proliferation. These results indicate that non-ligand induced dimerization is sufficient to activate EGFR and initiate cell signalling and EGFR endocytosis. We conclude that receptor dimerization is a critical event in EGF-induced cell signalling and EGFR endocytosis.

  9. On the photophysics and photochemistry of the water dimer

    Energy Technology Data Exchange (ETDEWEB)

    Segarra-Marti, Javier; Merchan, Manuela [Instituto de Ciencia Molecular, Universitat de Valencia, P.O. Box 22085, 46071 Valencia (Spain); Roca-Sanjuan, Daniel; Lindh, Roland [Department of Chemistry - Angstroem, Theoretical Chemistry Program, Uppsala University, Box 518, 75120 Uppsala (Sweden)

    2012-12-28

    The photochemistry of the water dimer irradiated by UV light is studied by means of the complete active space perturbation theory//complete active space self-consistent field (CASPT2//CASSCF) method and accurate computational approaches like as minimum energy paths. Both electronic structure computations and ab initio molecular dynamics simulations are carried out. The results obtained show small shifts relative to a single water molecule on the vertical excitation energies of the dimer due to the hydrogen bond placed between the water donor (W{sub D}) and the water acceptor (W{sub A}). A red-shift and a blue-shift are predicted for the W{sub D} and W{sub A}, respectively, supporting previous theoretical and experimental results. The photoinduced chemistry of the water dimer is described as a process occurring between two single water molecules in which the effect of the hydrogen bond plays a minor role. Thus, the photoinduced decay routes correspond to two photodissociation processes, one for each water molecule. The proposed mechanism for the decay channels of the lowest-lying excited states of the system is established as the photochemical production of a hydrogen-bonded H{sub 2}O Horizontal-Ellipsis HO species plus a hydrogen H atom.

  10. Dynamic interplay between adhesive and lateral E-cadherin dimers

    DEFF Research Database (Denmark)

    Klingelhöfer, Jörg; Laur, Oscar Y; Troyanovsky, Regina B;

    2002-01-01

    M. The disappearance of adhesive dimers was counterbalanced by an increase in Trp156-dependent lateral dimers. Increasing the calcium concentration to a normal level rapidly restored the original balance between adhesive and lateral dimers. We also present evidence that E-cadherin dimers in vivo have a short lifetime...

  11. Dimer Models, Free Fermions and Super Quantum Mechanics

    CERN Document Server

    Dijkgraaf, R; Reffert, S

    2007-01-01

    This note relates topics in statistical mechanics, graph theory and combinatorics, lattice quantum field theory, super quantum mechanics and string theory. We give a precise relation between the dimer model on a graph embedded on a torus and the massless free Majorana fermion living on the same lattice. A loop expansion of the fermion determinant is performed, where the loops turn out to be compositions of two perfect matchings. These loop states are sorted into co-chain groups using categorification techniques similar to the ones used for categorifying knot polynomials. The Euler characteristic of the resulting co-chain complex recovers the Newton polynomial of the dimer model. We re-interpret this system as supersymmetric quantum mechanics, where configurations with vanishing net winding number form the ground states. Finally, we make use of the quiver gauge theory - dimer model correspondence to obtain an interpretation of the loops in terms of the physics of D-branes probing a toric Calabi-Yau singularity...

  12. Structure and dimerization of translation initiation factor aIF5B in solution

    Energy Technology Data Exchange (ETDEWEB)

    Rasmussen, Louise Caroe Vohlander [Department of Molecular Biology, Aarhus University, Gustav Wieds Vej 10, DK-8000 Aarhus C (Denmark); Oliveira, Cristiano Luis Pinto [Department of Chemistry, Centre for mRNP Biogenesis and Metabolism, and iNANO Interdisciplinary Nanoscience Center, Aarhus University, Langelandsgade 140, DK-8000 Aarhus C (Denmark); Byron, Olwyn [Glasgow Biomedical Research Center, University of Glasgow, Glasgow G12 8QQ, Scotland (United Kingdom); Jensen, Janni Mosgaard [Department of Molecular Biology, Aarhus University, Gustav Wieds Vej 10, DK-8000 Aarhus C (Denmark); Pedersen, Jan Skov [Department of Chemistry, Centre for mRNP Biogenesis and Metabolism, and iNANO Interdisciplinary Nanoscience Center, Aarhus University, Langelandsgade 140, DK-8000 Aarhus C (Denmark); Sperling-Petersen, Hans Uffe [Department of Molecular Biology, Aarhus University, Gustav Wieds Vej 10, DK-8000 Aarhus C (Denmark); Mortensen, Kim Kusk, E-mail: kkm@science.au.dk [Department of Molecular Biology, Aarhus University, Gustav Wieds Vej 10, DK-8000 Aarhus C (Denmark)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer aIF5B forms maximum 5.0-6.8% irreversible dimers in solution. Black-Right-Pointing-Pointer Sedimentation coefficients for monomer and dimer are 3.64 and 5.51 {+-} 0.29 S. Black-Right-Pointing-Pointer Adding only 2% glycerol prevents dimerization. Black-Right-Pointing-Pointer SAXS on aIF5B monomer gave an R{sub g} of 37.5 {+-} 0.2 A and a D{sub max} of {approx}130 A. Black-Right-Pointing-Pointer There are universal structural differences between aIF5B and Escherichia coli IF2. -- Abstract: Translation initiation factor 5B (IF5B) is required for initiation of protein synthesis. The solution structure of archaeal IF5B (aIF5B) was analysed by small-angle X-ray scattering (SAXS) and dynamic light scattering (DLS) and was indicated to be in both monomeric and dimeric form. Sedimentation equilibrium (SE) analytical ultracentrifugation (AUC) of aIF5B indicated that aIF5B forms irreversible dimers in solution but only to a maximum of 5.0-6.8% dimer. Sedimentation velocity (SV) AUC at higher speed also indicated the presence of two species, and the sedimentation coefficients s{sub 20,w}{sup 0} were determined to be 3.64 and 5.51 {+-} 0.29 S for monomer and dimer, respectively. The atomic resolution (crystallographic) structure of aIF5B (Roll-Mecak et al. ) was used to model monomer and dimer, and theoretical sedimentation coefficients for these models were computed (3.89 and 5.63 S, respectively) in good agreement with the sedimentation coefficients obtained from SV analysis. Thus, the structure of aIF5B in solution must be very similar to the atomic resolution structure of aIF5B. SAXS data were acquired in the same buffer with the addition of 2% glycerol to inhibit dimerization, and the resultant monomeric aIF5B in solution did indeed adopt a structure very similar to the one reported earlier for the protein in crystalline form. The p(r) function indicated an elongated conformation supported by a radius of gyration of 37.5 {+-} 0.2 A

  13. Theoretical study of the phototoxicity of naproxen and the active form of nabumetone.

    Science.gov (United States)

    Musa, Klefah A K; Eriksson, Leif A

    2008-10-30

    Density functional theory using the hybrid functional B3LYP has been employed in order to study the mechanisms of photoinduced decomposition of the closely related nonsteroidal anti-inflammatory drugs naproxen (NP) and 6-methoxy-2-naphthylacetic acid (MNAA; the active form of nabumetone). The photochemical properties and computed energies of various species obtained in this study show that both drugs dominate in their deprotonated forms at physiological pH. The deprotonated acids are unable to decarboxylate from their excited singlets; instead, they decarboxylate from their first excited triplet states with high efficiency, overcoming energy barriers less than 3 and 1 kcal/mol for MNAA and NP, respectively. The ultraviolet and visible spectra of the neutral, deprotonated, and decarboxylated moieties of MNAA and NP are more-or-less similar but with higher probabilites (oscillator strength) for the latter. This fact, as well as the higher reactivity of NP, is explained in terms of the electron-donating effect of the additional methyl group present in NP. Singlet oxygen, superoxide radical anion, and corresponding peroxyl radical species are expected to be formed in different steps throughout the proposed photodegradation pathways of both drugs, which give rise to their effects on biomolecules, for example, lipid peroxidation.

  14. Trypanocidal activity of the essential oils in their conventional and nanoemulsion forms: in vitro tests.

    Science.gov (United States)

    Baldissera, Matheus D; Da Silva, Aleksandro S; Oliveira, Camila B; Zimmermann, Carine E P; Vaucher, Rodrigo A; Santos, Roberto C V; Rech, Virginia C; Tonin, Alexandre A; Giongo, Janice L; Mattos, Cristiane B; Koester, Letícia; Santurio, Janio M; Monteiro, Silvia G

    2013-07-01

    The aim of this study was to investigate the susceptibility in vitro of Trypanosoma evansi to the essential oils of andiroba (Carapa guaianensis) and aroeira (Schinus molle), in their conventional and nanostructured forms. For that, pure oils at concentrations of 0.5%, 1.0% and 2.0% were used. A negative control (untreated) and a positive control (diminazene aceturate 0.5%) were used as comparative parameters. Later, the same tests were performed, using nanoemulsions oils at concentrations of 0.5% and 1.0%. The tests were carried out in triplicates and the numbers of parasites were quantified on 1, 3 and 6 h from onset of the study. A dose-dependent reduction in the number of parasites to the forms of two oils tested was observed after 1 h. The concentration of parasites was significantly reduced at low concentrations after 3 h, as well as at 6 h no alive parasites were observed for the essential oils tested. Ours findings indicate, for the first time, that oils of andiroba and aroeira (in their conventional and nanoemulsion forms) have high activity against T. evansi in vitro, leading to the suggestion that these oils may be applied as an alternative treatment for this disease.

  15. Overlapping patterns of neural activity for different forms of novelty in fMRI

    Directory of Open Access Journals (Sweden)

    Colin Shaun Hawco

    2014-09-01

    Full Text Available When stimuli are presented multiple times, the neural response to repeated stimuli is reduced relative to novel stimuli (repetition suppression. Responses to different types of novelty were examined. Stimulus novelty was examined by contrasting first vs. second presentation of triads of objects during memory encoding. Semantic novelty was contrasted by comparing unrelated (semantically novel triads of objects to triads in which all three objects were related (e.g. all objects were tools. In recognition, associative novelty was examined by contrasting rearranged triads (previously seen objects in a new association with intact triads. Activity was observed in posterior regions (occipital and fusiform, with the largest extent of activity for stimulus novelty and smallest for associational novelty. Frontal activity was also observed in stimulus and semantic novelty. Additional analysis indicated that the hemodynamic response in voxels identified in the stimulus and semantic novelty contrasts was modulated by reaction time on a trial-by-trial basis. That is, the duration of the hemodynamic response was driven by reaction time. This was not the case for associative novelty. The high level of overlap across different forms of novelty suggests a similar mechanism for reduced neural activity, which may be related to reduced visual processing time. This is consistent with a facilitation model of repetition suppression, which posits a reduced peak and duration of neuronal firing for repeated stimuli.

  16. Dynamin Forms a Src Kinase–sensitive Complex with Cbl and Regulates Podosomes and Osteoclast Activity

    Science.gov (United States)

    Bruzzaniti, Angela; Neff, Lynn; Sanjay, Archana; Horne, William C.; De Camilli, Pietro; Baron, Roland

    2005-01-01

    Podosomes are highly dynamic actin-containing adhesion structures found in osteoclasts, macrophages, and Rous sarcoma virus (RSV)-transformed fibroblasts. After integrin engagement, Pyk2 recruits Src and the adaptor protein Cbl, forming a molecular signaling complex that is critical for cell migration, and deletion of any molecule in this complex disrupts podosome ring formation and/or decreases osteoclast migration. Dynamin, a GTPase essential for endocytosis, is also involved in actin cytoskeleton remodeling and is localized to podosomes where it has a role in actin turnover. We found that dynamin colocalizes with Cbl in the actin-rich podosome belt of osteoclasts and that dynamin forms a complex with Cbl in osteoclasts and when overexpressed in 293VnR or SYF cells. The association of dynamin with Cbl in osteoclasts was decreased by Src tyrosine kinase activity and we found that destabilization of the dynamin-Cbl complex involves the recruitment of Src through the proline-rich domain of Cbl. Overexpression of dynamin increased osteoclast bone resorbing activity and migration, whereas overexpression of dynK44A decreased osteoclast resorption and migration. These studies suggest that dynamin, Cbl, and Src coordinately participate in signaling complexes that are important in the assembly and remodeling of the actin cytoskeleton, leading to changes in osteoclast adhesion, migration, and resorption. PMID:15872089

  17. Dynamin forms a Src kinase-sensitive complex with Cbl and regulates podosomes and osteoclast activity.

    Science.gov (United States)

    Bruzzaniti, Angela; Neff, Lynn; Sanjay, Archana; Horne, William C; De Camilli, Pietro; Baron, Roland

    2005-07-01

    Podosomes are highly dynamic actin-containing adhesion structures found in osteoclasts, macrophages, and Rous sarcoma virus (RSV)-transformed fibroblasts. After integrin engagement, Pyk2 recruits Src and the adaptor protein Cbl, forming a molecular signaling complex that is critical for cell migration, and deletion of any molecule in this complex disrupts podosome ring formation and/or decreases osteoclast migration. Dynamin, a GTPase essential for endocytosis, is also involved in actin cytoskeleton remodeling and is localized to podosomes where it has a role in actin turnover. We found that dynamin colocalizes with Cbl in the actin-rich podosome belt of osteoclasts and that dynamin forms a complex with Cbl in osteoclasts and when overexpressed in 293VnR or SYF cells. The association of dynamin with Cbl in osteoclasts was decreased by Src tyrosine kinase activity and we found that destabilization of the dynamin-Cbl complex involves the recruitment of Src through the proline-rich domain of Cbl. Overexpression of dynamin increased osteoclast bone resorbing activity and migration, whereas overexpression of dynK44A decreased osteoclast resorption and migration. These studies suggest that dynamin, Cbl, and Src coordinately participate in signaling complexes that are important in the assembly and remodeling of the actin cytoskeleton, leading to changes in osteoclast adhesion, migration, and resorption.

  18. Volatile sulphur compounds-forming abilities of lactic acid bacteria: C-S lyase activities.

    Science.gov (United States)

    Bustos, Irene; Martínez-Bartolomé, Miguel A; Achemchem, Fouad; Peláez, Carmen; Requena, Teresa; Martínez-Cuesta, M Carmen

    2011-08-01

    Volatile sulphur compounds (VSCs) are of prime importance in the overall aroma of cheese and make a significant contribution to their typical flavours. Thus, the control of VSCs formation offers considerable potential for industrial applications. Here, lactic acid bacteria (LAB) from different ecological origins were screened for their abilities to produce VSCs from L-methionine. From the data presented, VSC-forming abilities were shown to be strain-specific and were correlated with the C-S lyase enzymatic activities determined using different approaches. High VSCs formation were detected for those strains that were also shown to possess high thiol-producing abilities (determined either by agar plate or spectrophotometry assays). Moreover, differences in C-S lyase activities were shown to correspond with the enzymatic potential of the strains as determined by in situ gel visualization. Therefore, the assessment of the C-S lyase enzymatic potential, by means of either of these techniques, could be used as a valuable approach for the selection of LAB strains with high VSC-producing abilities thus, representing an effective way to enhance cheese sulphur aroma compounds synthesis. In this regard, this study highlights the flavour forming potential of the Streptococcus thermophilus STY-31, that therefore could be used as a starter culture in cheese manufacture. Furthermore, although C-S lyases are involved in both biosynthetic and catabolic pathways, an association between methionine and cysteine auxotrophy of the selected strains and their VSCs-producing abilities could not be found.

  19. Piezo proteins are pore-forming subunits of mechanically activated channels.

    Science.gov (United States)

    Coste, Bertrand; Xiao, Bailong; Santos, Jose S; Syeda, Ruhma; Grandl, Jörg; Spencer, Kathryn S; Kim, Sung Eun; Schmidt, Manuela; Mathur, Jayanti; Dubin, Adrienne E; Montal, Mauricio; Patapoutian, Ardem

    2012-02-19

    Mechanotransduction has an important role in physiology. Biological processes including sensing touch and sound waves require as-yet-unidentified cation channels that detect pressure. Mouse Piezo1 (MmPiezo1) and MmPiezo2 (also called Fam38a and Fam38b, respectively) induce mechanically activated cationic currents in cells; however, it is unknown whether Piezo proteins are pore-forming ion channels or modulate ion channels. Here we show that Drosophila melanogaster Piezo (DmPiezo, also called CG8486) also induces mechanically activated currents in cells, but through channels with remarkably distinct pore properties including sensitivity to the pore blocker ruthenium red and single channel conductances. MmPiezo1 assembles as a ∼1.2-million-dalton homo-oligomer, with no evidence of other proteins in this complex. Purified MmPiezo1 reconstituted into asymmetric lipid bilayers and liposomes forms ruthenium-red-sensitive ion channels. These data demonstrate that Piezo proteins are an evolutionarily conserved ion channel family involved in mechanotransduction.

  20. SOME INNOVATIVE FORMS OF ORGANIZATION OF INDEPENDENT ACTIVITY OF STUDENTS IN CONSTRUCTIVE LEARNING

    Directory of Open Access Journals (Sweden)

    Shatalova N. P.

    2015-11-01

    Full Text Available Conducted in our country reforms require active participation in their realization of all citizens. The development of the economic components of the state, will allow not only to strengthen the defense potential of the country, but also to raise the quality of life of the population. To solve this problem the country needs competent specialists, real, educated professionals, masters of their craft. The most important task of each University is to prepare such graduates. Training at the University is done not only through training and education, but also in the process of independent activity of students. Independent work is the activity performed by the students of the University without direct contact with the teacher or teacher-driven mediated through specific training materials; it is an integral, compulsory element of the learning process, providing primarily individual work of students in accordance with the setting of teacher or textbook curricula. In the modern didactics, independent work of students is, on the one hand, a kind of educational work carried out without direct intervention, but under the guidance of a teacher, and as a means of engaging students in independent cognitive activity, the formation methods of the organization of such activities. The effect from independent work of students can be obtained only when it is organized and implemented in the educational process as a holistic system that runs through all the stages of learning of students in high school. In the article, on the basis of experience, analysis, research and publications of the Russian scientists, we considered the problems arising at the organization of independent activity of students as recommendations are offered innovative forms and methods of education based on the theory of constructive learning

  1. Beyerane derivatives and a sesquiterpene dimer from Japanese Cypress (Chamaecyparis obtusa).

    Science.gov (United States)

    Gao, Hui Yuan; Wu, Li Jun; Muto, Norio; Fuchino, Hiroyuki; Nakane, Takahisa; Shirota, Osamu; Sano, Toshikazu; Kuroyanagi, Masanori

    2008-07-01

    In the course of studies on biological active constituents from woody plants, we previously reported the isolation of many lignan derivatives as neurite outgrowth-promoting compounds from an ethyl acetate soluble fraction of Japanese Cypress (Chamaecyparis obtusa). Further chemical investigation on the residual parts of the ethyl acetate soluble fraction of a methanol extract of Japanese Cypress resulted in the isolation of four new beyerene type derivatives and a novel sesquiterpene dimer formed between cryptomeridiol and hinokiic acid. Their structures were elucidated as 18-O-(Z)-p-coumaroylbeyer-15-ene-18-ol (1), 18-O-(E)-p-coumaroylbeyer-15-ene-18-ol (2), 18-O-(E)-p-coumaroylbeyer-15-ene-11beta,18-diol (3), 18-O-(Z)-p-coumaroylbeyer-15-ene-11beta,18-diol (4) and ent-cryptomeridiol-4-yl-hinokiiate (5) by (1)H-NMR, (13)C-NMR, 2D-NMR, and HR-MS spectral analysis.

  2. Investigation of the hydrated 7-hydroxy-4-methylcoumarin dimer by combined IR/UV spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Stamm, A.; Schwing, K.; Gerhards, M., E-mail: gerhards@chemie.uni-kl.de [TU Kaiserslautern, Fachbereich Chemie and Research Center Optimas, Erwin-Schroedinger-Straße 52, D-67663 Kaiserslautern (Germany)

    2014-11-21

    The first molecular beam investigations on a coumarin dimer and clusters of a coumarin dimer with water both in the neutral (S{sub 0}) and cationic (D{sub 0}) electronic ground state are performed. The structure and structural changes due to ionization of the isolated 7-hydroxy-4-methylcoumarin dimer (7H4MC){sub 2} as well as its mono- and dihydrate (7H4MC){sub 2}(H{sub 2}O){sub 1-2} are analyzed by applying combined IR/UV spectroscopy compared with density functional theory calculations. In case of the neutral dimer of 7H4MC a doubly hydrogen-bonded structure is formed. This doubly hydrogen-bonded arrangement opens to a singly hydrogen-bonded structure in the ion presenting a rearrangement reaction within an isolated dimer. By attaching one or two water molecules to the neutral 7H4MC dimer water is inserted into the hydrogen bonds. In contrast to the non-hydrated species this general binding motif with water in a bridging function does not change via ionization but especially for the dihydrate the spatial arrangement of the two 7H4MC units changes strengthening the interaction between the aromatic chromophores. The presented analyses illustrate the strong dependence of binding motifs as a function of successive hydration and charge including a rearrangement reaction.

  3. Investigation of the hydrated 7-hydroxy-4-methylcoumarin dimer by combined IR/UV spectroscopy.

    Science.gov (United States)

    Stamm, A; Schwing, K; Gerhards, M

    2014-11-21

    The first molecular beam investigations on a coumarin dimer and clusters of a coumarin dimer with water both in the neutral (S0) and cationic (D0) electronic ground state are performed. The structure and structural changes due to ionization of the isolated 7-hydroxy-4-methylcoumarin dimer (7H4MC)2 as well as its mono- and dihydrate (7H4MC)2(H2O)1-2 are analyzed by applying combined IR/UV spectroscopy compared with density functional theory calculations. In case of the neutral dimer of 7H4MC a doubly hydrogen-bonded structure is formed. This doubly hydrogen-bonded arrangement opens to a singly hydrogen-bonded structure in the ion presenting a rearrangement reaction within an isolated dimer. By attaching one or two water molecules to the neutral 7H4MC dimer water is inserted into the hydrogen bonds. In contrast to the non-hydrated species this general binding motif with water in a bridging function does not change via ionization but especially for the dihydrate the spatial arrangement of the two 7H4MC units changes strengthening the interaction between the aromatic chromophores. The presented analyses illustrate the strong dependence of binding motifs as a function of successive hydration and charge including a rearrangement reaction.

  4. Role of antibodies in developing drugs that target G-protein-coupled receptor dimers.

    Science.gov (United States)

    Hipser, Chris; Bushlin, Ittai; Gupta, Achla; Gomes, Ivone; Devi, Lakshmi A

    2010-01-01

    G-protein-coupled receptors are important molecular targets in drug discovery. These receptors play a pivotal role in physiological signaling pathways and are targeted by nearly 50% of currently available drugs. Mounting evidence suggests that G-protein-coupled receptors form dimers, and various studies have shown that dimerization is necessary for receptor maturation, signaling, and trafficking. However, the physiological implications of dimerization in vivo have not been well explored because detection of GPCR dimers in endogenous systems has been a challenging task. One exciting new approach to this challenge is the generation of antibodies against specific G-protein-coupled receptor dimers. Such antibodies could be used as tools for characterization of heteromer-specific function; as reagents for their purification, tissue localization, and regulation in vivo; and as probes for mapping their functional domains. In addition, such antibodies could serve as alternative ligands for G-protein-coupled receptor heteromers. Thus, heteromer-specific antibodies represent novel tools for the exploration and manipulation of G-protein-coupled receptor-dimer pharmacology.

  5. Dislocations and vacancies in two-dimensional mixed crystals of spheres and dimers

    KAUST Repository

    Gerbode, Sharon J.

    2010-10-15

    In colloidal crystals of spheres, dislocation motion is unrestricted. On the other hand, recent studies of relaxation in crystals of colloidal dimer particles have demonstrated that the dislocation dynamics in such crystals are reminiscent of glassy systems. The observed glassy dynamics arise as a result of dislocation cages formed by certain dimer orientations. In the current study, we use experiments and simulations to investigate the transition that arises when a pure sphere crystal is doped with an increasing concentration of dimers. Specifically, we focus on both dislocation caging and vacancy motion. Interestingly, we find that any nonzero fraction of dimers introduces finite dislocation cages, suggesting that glassy dynamics are present for any mixed crystal. However, we have also identified a vacancy-mediated uncaging mechanism for releasing dislocations from their cages. This mechanism is dependent on vacancy diffusion, which slows by orders of magnitude as the dimer concentration is increased. We propose that in mixed crystals with low dimer concentrations vacancy diffusion is fast enough to uncage dislocations and delay the onset of glassy dislocation dynamics. © 2010 The American Physical Society.

  6. Molecular Insight into Affinities of Gallated and Nongallated Proanthocyanidins Dimers to Lipid Bilayers

    Science.gov (United States)

    Zhu, Wei; Xiong, Le; Peng, Jinming; Deng, Xiangyi; Gao, Jun; Li, Chun-Mei

    2016-11-01

    Experimental studies have proved the beneficial effects of proanthocyanidins (Pas) relating to interaction with the cell membrane. But the detailed mechanisms and structure-function relationship was unclear. In present study, molecular dynamics (MD) simulations were used to study the interactions of four PA dimers with a lipid bilayer composed of 1:1 mixed 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) and 1-palmitoyl-2-oleoyl phosphatidylethanolamine (POPE). The results showed that the gallated PA dimers had much higher affinities to the bilayer with lower binding free energies compared with nongallated PA dimers. The gallated PA dimers penetrated deeper into the bilayer and formed more hydrogen bonds (H-bonds) with bilayer oxygen atoms, especially the deeper oxygen atoms of the lipids simultaneously, thus inducing stronger lateral expansion of the membrane and lipid tails disorder. The present results provided molecular insights into the interactions between PA dimers and bio-membranes and agreed with our experimental results well. These molecular interactions helped to elucidate the structure-function relationship of the PA dimers and provided a foundation for a better understanding of the underlying mechanisms of the bioactivities of PA oligomers.

  7. Concentration of D-dimers in healthy cats and sick cats with and without disseminated intravascular coagulation (DIC).

    Science.gov (United States)

    Tholen, Inger; Weingart, Christiane; Kohn, Barbara

    2009-10-01

    The objective of this prospective study was to measure concentrations of D-dimers in 48 cats with various diseases and in 20 healthy cats to evaluate the sensitivity and specificity for D-dimers to diagnose disseminated intravascular coagulation (DIC). The cats were classified as having DIC if an underlying disease and at least three of the following criteria were present: thrombocytopenia, prolonged activated partial thromboplastin time, prothrombin time or thrombin time, schistocytes and/or a reduced antithrombin activity. D-dimer concentrations were measured using a semi-quantitative latex agglutination (LA) test (Accuclot D-Dimer, Sigma Diagnostics). The D-dimer test was positive for 8/12 cats with DIC and for 16/36 sick cats without DIC. D-dimers were negative for all healthy control cats. The comparison of the sick cats with DIC and those without DIC revealed a specificity and sensitivity of the D-dimer test of 56% and 67%; a comparison of the results for healthy cats and cats with DIC revealed a specificity and sensitivity of 100% and 67%, respectively. The D-dimer LA test is only of limited value for the diagnosis of DIC in cats.

  8. Necrosis is an active and controlled form of programmed cell death.

    Science.gov (United States)

    Proskuryakov, S Ya; Gabai, V L; Konoplyannikov, A G

    2002-04-01

    In all studies on programmed cell death (PCD) and apoptosis as its most showy form, this process was considered to be a paradigmatic antithesis to necrotic cell death. On one hand, a concept on necrosis as a cellular cataclysm, an uncontrolled and passive phenomenon, had been provoked by an enormous bulk of experimental data on its inducibility by superphysiological exposures. On the other hand, much attention was attracted to a rapidly expanding (from nematodes) field of genetic studies on PCD. However, the findings accumulated which suggested a likeness rather than the opposition of the necrotic and apoptotic forms of elimination of "unwanted" cells. 1. Very diverse pathophysiological exposures (stimuli, stresses), such as heat, ionizing radiation, pathogens, cytokines cause both forms of cell death in the same cell population. 2. Anti-apoptotic mechanisms (e.g., Bcl-2) can protect cells from both necrotic and apoptotic destruction. 3. Biochemical interventions (e.g., with inhibitors of poly-(ADP-riboso)-polymerase) into the signal and executive mechanisms of PCD can change the choice of the cell death form. 4. During both necrosis and epigenetic programs of apoptotic cell death that need no macromolecular synthesis (e.g., the CD95-dependent death), the nucleus plays a passive role. Therefore, necrosis, similarly to apoptosis, is suggested to be a form of the programmed cell death. However, for the whole body the physiological consequences of apoptosis and necrosis are quite different. In the case of apoptosis, all constituents of the nucleus and cytoplasm are isolated by an undamaged membrane and then by phagocytes together with the membrane-bound "eat me" markers (phosphatidylserine, etc.). In other words, the elimination of the cell which has realized its apoptotic program remains virtually unnoticed by the body. In the case of necrosis, the cytoplasmic content released into the intercellular space provokes an inflammatory response, i.e., an activation of

  9. Biofilm-forming activity of bacteria isolated from toilet bowl biofilms and the bactericidal activity of disinfectants against the isolates.

    Science.gov (United States)

    Mori, Miho; Gomi, Mitsuhiro; Matsumune, Norihiko; Niizeki, Kazuma; Sakagami, Yoshikazu

    2013-01-01

    To evaluate the sanitary conditions of toilets, the bacterial counts of the toilet bowl biofilms in 5 Kansai area and 11 Kansai and Kanto area homes in Japan were measured in winter and summer seasons, respectively. Isolates (128 strains) were identified by analyzing 16S ribosomal RNA sequences. The number of colonies and bacterial species from biofilms sampled in winter tended to be higher and lower, respectively, than those in summer. Moreover, the composition of bacterial communities in summer and winter samples differed considerably. In summer samples, biofilms in Kansai and Kanto areas were dominated by Blastomonas sp. and Mycobacterium sp., respectively. Methylobacterium sp. was detected in all toilet bowl biofilms except for one sample. Methylobacterium sp. constituted the major presence in biofilms along with Brevundimonas sp., Sphingomonas sp., and/or Pseudomonas sp. The composition ratio of the sum of their genera was 88.0 from 42.9% of the total bacterial flora. The biofilm formation abilities of 128 isolates were investigated, and results suggested that Methylobacterium sp. and Sphingomonas sp. were involved in biofilm formation in toilet bowls. The biofilm formation of a mixed bacteria system that included bacteria with the highest biofilm-forming ability in a winter sample was greater than mixture without such bacteria. This result suggests that isolates possessing a high biofilm-forming activity are involved in the biofilm formation in the actual toilet bowl. A bactericidal test against 25 strains indicated that the bactericidal activities of didecyldimethylammonium chloride (DDAC) tended to be higher than those of polyhexamethylene biguanide (PHMB) and N-benzyl-N,N-dimethyldodecylammonium chloride (ADBAC). In particular, DDAC showed high bactericidal activity against approximately 90% of tested strains under the 5 h treatment.

  10. The Atypical Response Regulator Protein ChxR Has Structural Characteristics and Dimer Interface Interactions That Are Unique within the OmpR/PhoB Subfamily

    Energy Technology Data Exchange (ETDEWEB)

    Hickey, John M.; Lovell, Scott; Battaile, Kevin P.; Hu, Lei; Middaugh, C. Russell; Hefty, P. Scott (Kansas); (HWMRI)

    2013-05-29

    Typically as a result of phosphorylation, OmpR/PhoB response regulators form homodimers through a receiver domain as an integral step in transcriptional activation. Phosphorylation stabilizes the ionic and hydrophobic interactions between monomers. Recent studies have shown that some response regulators retain functional activity in the absence of phosphorylation and are termed atypical response regulators. The two currently available receiver domain structures of atypical response regulators are very similar to their phospho-accepting homologs, and their propensity to form homodimers is generally retained. An atypical response regulator, ChxR, from Chlamydia trachomatis, was previously reported to form homodimers; however, the residues critical to this interaction have not been elucidated. We hypothesize that the intra- and intermolecular interactions involved in forming a transcriptionally competent ChxR are distinct from the canonical phosphorylation (activation) paradigm in the OmpR/PhoB response regulator subfamily. To test this hypothesis, structural and functional studies were performed on the receiver domain of ChxR. Two crystal structures of the receiver domain were solved with the recently developed method using triiodo compound I3C. These structures revealed many characteristics unique to OmpR/PhoB subfamily members: typical or atypical. Included was the absence of two {alpha}-helices present in all other OmpR/PhoB response regulators. Functional studies on various dimer interface residues demonstrated that ChxR forms relatively stable homodimers through hydrophobic interactions, and disruption of these can be accomplished with the introduction of a charged residue within the dimer interface. A gel shift study with monomeric ChxR supports that dimerization through the receiver domain is critical for interaction with DNA.

  11. The Structure of the Periplasmic Sensor Domain of the Histidine Kinase CusS Shows Unusual Metal Ion Coordination at the Dimeric Interface.

    Science.gov (United States)

    Affandi, Trisiani; Issaian, Aaron V; McEvoy, Megan M

    2016-09-20

    In bacteria, two-component systems act as signaling systems to respond to environmental stimuli. Two-component systems generally consist of a sensor histidine kinase and a response regulator, which work together through histidyl-aspartyl phosphorelay to result in gene regulation. One of the two-component systems in Escherichia coli, CusS-CusR, is known to induce expression of cusCFBA genes at increased periplasmic Cu(I) and Ag(I) concentrations to help maintain metal ion homeostasis. CusS is a membrane-associated histidine kinase with a periplasmic sensor domain connected to the cytoplasmic ATP binding and catalytic domains through two transmembrane helices. The mechanism of how CusS senses increasing metal ion concentrations and activates CusR is not yet known. Here, we present the crystal structure of the Ag(I)-bound periplasmic sensor domain of CusS at a resolution of 2.15 Å. The structure reveals that CusS forms a homodimer with four Ag(I) binding sites per dimeric complex. Two symmetric metal binding sites are found at the dimeric interface, which are each formed by two histidines and one phenylalanine with an unusual cation-π interaction. The other metal ion binding sites are in a nonconserved region within each monomer. Functional analyses of CusS variants with mutations in the metal sites suggest that the metal ion binding site at the dimer interface is more important for function. The structural and functional data provide support for a model in which metal-induced dimerization results in increases in kinase activity in the cytoplasmic domains of CusS.

  12. The Structure of the Periplasmic Sensor Domain of the Histidine Kinase CusS Shows Unusual Metal Ion Coordination at the Dimeric Interface

    Science.gov (United States)

    Affandi, Trisiani; Issaian, Aaron V.; McEvoy, Megan M.

    2016-01-01

    In bacteria, two-component systems act as signaling systems to respond to environmental stimuli. Two-component systems generally consist of a sensor histidine kinase and a response regulator, which work together through histidyl-aspartyl phospho-relay to result in gene regulation. One of the two-component systems in Escherichia coli, CusS-CusR, is known to induce expression of cusCFBA genes under increased periplasmic Cu(I) and Ag(I) concentrations to help maintain metal ion homeostasis. CusS is a membrane-associated histidine kinase with a periplasmic sensor domain connected to the cytoplasmic ATP-binding and catalytic domains through two transmembrane helices. The mechanism of how CusS senses increasing metal ion concentrations and activates CusR is not yet known. Here, we present the crystal structure of the Ag(I)-bound periplasmic sensor domain of CusS at a resolution of 2.15 Å. The structure reveals that CusS forms a homodimer with four Ag(I) binding sites per dimeric complex. Two symmetric metal binding sites are found at the dimeric interface, which are each formed by two histidines and one phenylalanine with an unusual cation-π interaction. The other metal ion binding sites are in a non-conserved region within each monomer. Functional analyses of CusS variants with mutations in the metal sites suggest that the metal ion binding site at the dimer interface is more important for function. The structural and functional data provide support for a model in which metal-induced dimerization results in increases in kinase activity in the cytoplasmic domains of CusS. PMID:27583660

  13. 环戊二烯基钌配合物催化的高选择性苯乙炔二聚反应%HIGHLY SELECTIVE CATALYTIC DIMERIZATION OF PHENYLACETYLENE BY CYCLOPENTADIENYL RUTHENIUM COMPLEXES

    Institute of Scientific and Technical Information of China (English)

    金军挺; 黄吉玲; 陶晓春; 钱延龙

    1999-01-01

    @@ Transition metal vinylidene complexes (M=C=CHR) have attracted a great deal of attention in recent years as a new type of organometallic intermediates that may have unusual reactivity[1]. Their reactivity has been explored and their application to organic synthesis is developed[2]. Recent reports on the ruthenium-vinylidene complexes[3]suggest that the reaction of ruthenium-vinylidene complexes with a base generates the coordinatively unsaturated ruthenium acetylide species, which are involved in a number of catalytic and stoichiometric reactions of alkynes. For example,the coordinatively unsaturated ruthenium acetylide species C5Me5Ru(PPh3)-C≡CPh,formed from the reaction of the vinylidene complex C5Me5Ru(PPh3) (Cl)=C=CHPh with a base was reactive toward a variety of small molecules and active in catalytic dimerization of terminal alkynes[4]. The dimerization of terminal alkyne is an effective method of forming enynes, but its synthetic application in organic synthesis has been limited dueto low selectivity for dimeric products[5]. In this communication, we report that three ruthenium complexes were used as catalysts for the highly selective dimerization of phenylacetylene.

  14. Formation of H-type liquid crystal dimer at air-water interface

    Energy Technology Data Exchange (ETDEWEB)

    Karthik, C., E-mail: karthik.c@pilani.bits-pilani.ac.in; Gupta, Adbhut, E-mail: karthik.c@pilani.bits-pilani.ac.in; Joshi, Aditya, E-mail: karthik.c@pilani.bits-pilani.ac.in; Manjuladevi, V., E-mail: karthik.c@pilani.bits-pilani.ac.in; Gupta, Raj Kumar, E-mail: karthik.c@pilani.bits-pilani.ac.in [Department of Physics, Birla Institute of Technology and Science, Pilani, Rajasthan -333031 (India); Varia, Mahesh C.; Kumar, Sandeep [Raman Research Institute, Sadashivanagar, Bangalore - 560080 (India)

    2014-04-24

    We have formed the Langmuir monolayer of H-shaped Azo linked liquid crystal dimer molecule at the air-water interface. Isocycles of the molecule showed hysteresis suggesting the ir-reversible nature of the monolayer formed. The thin film deposited on the silicon wafer was characterized using Atomic Force Microscopy (AFM) and Field Emission Scanning Electron Microscopy (FESEM). The images showed uniform domains of the dimer molecule. We propose that these molecules tend to take book shelf configuration in the liquid phase.

  15. N-terminal processing of membrane-targeted MnSOD and formation of multiple active superoxide dismutase dimers in the nitrogen-fixing cyanobacterium Anabaena sp. strain PCC7120.

    Science.gov (United States)

    Raghavan, Prashanth S; Rajaram, Hema; Apte, Shree K

    2013-10-01

    Anabaena sp. strain PCC7120 expresses a 30 kDa manganese-dependent superoxide dismutase (MnSOD) comprising a hydrophobic region (signal peptide + linker peptide) attached to a catalytic unit. Bioinformatics predicted cleavage of the signal peptide at (25)CQPQ by signal peptidase and of the linker peptide by an Arg-C-like protease at the Arg52/Arg59 residue. The three predicted forms of MnSOD were immunodetected in Anabaena, with the 30 kDa MnSOD found exclusively in the membrane and the shorter 27 and 24 kDa forms found both in the membrane and soluble fractions. The corresponding sodA gene was truncated for (a) the first eight residues, or, (b) the signal peptide, or (c) the entire hydrophobic region, or (d) the Arg52/Arg59 residues were modified to serine. Overexpression of these MnSOD variants in recombinant Anabaena strains revealed that (a) the 30 kDa membrane-targeted MnSOD was cleaved by membrane-localized signal peptidase either during or after its transport through the membrane to release the 27 kDa form, either in the cytosol or in the periplasmic/thylakoid lumen, (b) the 27 kDa form was further cleaved to the 24 kDa form by Arg-C-like protease, both in the cytosol and in the periplasmic/thylakoid lumen, (c) deletion of signal peptide localized the MnSOD forms in the cytosol, and (d) alteration of the signal/linker peptide cleavage sites interfered with MnSOD localization and processing. Homo/heterodimerization of the 24 and 27 kDa forms of MnSOD and the cytosolic iron-dependent SOD results in multiple SOD activities, from a single MnSOD gene (sodA), in different cellular compartments of Anabaena.

  16. 76 FR 9806 - Agency Information Collection Activities: Passenger List/Crew List (CBP Form I-418)

    Science.gov (United States)

    2011-02-22

    ... List (CBP Form I-418) AGENCY: U.S. Customs and Border Protection, Department of Homeland Security... review and approval in accordance with the Paperwork Reduction Act: Passenger List/Crew List (CBP Form I... forms of information. Title: Passenger List/Crew List. OMB Number: 1651-0103. Form Number: CBP Form...

  17. 77 FR 2561 - Agency Information Collection Activities: Passenger List/Crew List (CBP Form I-418)

    Science.gov (United States)

    2012-01-18

    ... List (CBP Form I-418) AGENCY: U.S. Customs and Border Protection (CBP), Department of Homeland Security... concerning the Passenger List/Crew List (CBP Form I-418). This request for comment is being made pursuant to...: Passenger List/Crew List. OMB Number: 1651-0103. Form Number: CBP Form I-418. Abstract: CBP Form I-418...

  18. 78 FR 26648 - Agency Information Collection Activities: Passenger List/Crew List (CBP Form I-418)

    Science.gov (United States)

    2013-05-07

    ... List (CBP Form I-418) AGENCY: U.S. Customs and Border Protection (CBP), Department of Homeland Security... concerning the Passenger List/Crew List (CBP Form I-418). This request for comment is being made pursuant to...: Passenger List/Crew List. OMB Number: 1651-0103. Form Number: CBP Form I-418. Abstract: CBP Form I-418...

  19. Molybdenum(VI) coordination chemistry of the N,N-disubstituted bis(hydroxylamido)-1,3,5-triazine ligand, H2bihyat. Water-assisted activation of the Mo(VI)═O bond and reversible dimerization of cis-[Mo(VI)O2(bihyat)] to [Mo(VI)2O4(bihyat)2(H2O)2].

    Science.gov (United States)

    Stylianou, Marios; Nikolakis, Vladimiros A; Chilas, George I; Jakusch, Tamas; Vaimakis, Tiverios; Kiss, Tamas; Sigalas, Michael P; Keramidas, Anastasios D; Kabanos, Themistoklis A

    2012-12-17

    Reaction of the N,N-disubstituted bis(hydroxylamino) ligand 2,6-bis[hydroxy(methyl)amino]-4-morpholino-1,3,5-triazine (H(2)bihyat) with cis-[Mo(VI)O(2)(acac)(2)] in tetrahydrofuran resulted in isolation of the mononuclear compound cis-[Mo(VI)O(2)(bihyat)] (1). The treatment of Na(2)Mo(VI)O(4)·2H(2)O with the ligand H(2)bihyat in aqueous solution gave the dinuclear compounds cis-[Mo(VI)(2)O(4)(bihyat)(2)(H(2)O)(2)] (2) and trans-[Mo(VI)(2)O(4)(bihyat)(2)(H(2)O)(2)] (3) at pH values of 3.5 and 5.5, respectively. The structures for the three molybdenum(VI) compounds were determined by X-ray crystallography. Compound 1 has a square-pyramidal arrangement around molybdenum, while in the two dinuclear compounds, each molybdenum atom is in a distorted pentagonal-bipyramidal environment of two bridging and one terminal oxido groups, a tridentate (O,N,O) bihyat(2-) ligand that forms two five-membered chelate rings, and a water molecule trans to the terminal oxido group. The dinuclear compounds constitute rare examples containing the {Mo(2)(VI)O(2)(μ(2)-O(2))}(4+) moiety. The potentiometry revealed that the Mo(VI)bihyat(2-) species exhibit high hydrolytic stability in aqueous solution at a narrow range of pH values, 3-5. A subtle change in the coordination environment of the five-coordinate compound 1 with ligation of a weakly bound water molecule trans to the oxido ligand (1w) renders the equatorial oxido group in 1w more nucleophilic than that in 1, and this oxido group attacks a molybdenum atom and thus the dinuclear compounds 2 and 3 are formed. This process might be considered as the first step of the oxido group nucleophilic attack on organic substrates, resulting in oxidation of the substrate, in the active site of molybdenum enzymes such as xanthine oxidase. Theoretical calculations in the gas phase were performed to examine the influence of water on the dimerization process (1 → 2/3). In addition, the molecular structures, cis/trans geometrical isomerism for the

  20. Cysteine 904 is required for maximal insulin degrading enzyme activity and polyanion activation.

    Directory of Open Access Journals (Sweden)

    Eun Suk Song

    Full Text Available Cysteine residues in insulin degrading enzyme have been reported as non-critical for its activity. We found that converting the twelve cysteine residues in rat insulin degrading enzyme (IDE to serines resulted in a cysteine-free form of the enzyme with reduced activity and decreased activation by polyanions. Mutation of each cysteine residue individually revealed cysteine 904 as the key residue required for maximal activity and polyanion activation, although other cysteines affect polyanion binding to a lesser extent. Based on the structure of IDE, Asn 575 was identified as a potential hydrogen bond partner for Cys904 and mutation of this residue also reduced activity and decreased polyanion activation. The oligomerization state of IDE did not correlate with its activity, with the dimer being the predominant form in all the samples examined. These data suggest that there are several conformational states of the dimer that affect activity and polyanion activation.

  1. Experiment study of forming and activation of conductive film of the surface conduction electron emitter display

    Institute of Scientific and Technical Information of China (English)

    LEI Xin; XU Wei-jun; LIU Chun-liang; LIANG Zhi-hu

    2007-01-01

    The forming and activation of the conductive films are studied experimentally. The power supply,a peak-to-peak 30 V triangle profile voltage,is applied to three kinds of conductive films that contain 0.25%,0.5%,and 1% of palladium respectively. In the experiments we contrasted the values of related parameter in different conditions,observed the lumi nous spots on the anode panel,dealt with and analyzed the related data,and compared the positions and the amount of the luminous spots. We have gotten the conclusion that there is a threshold value Uth. The emission current Ie will increase rapidly when the device voltage Uf is greater than Uth. And the emission current Ie could be controlled by the device voltage Uf.The positions of the luminous spots on the anode panel are related with the device voltage Uf.

  2. Microstructure evolution and surface cleaning of Cu nanoparticles during micro-forming fields activated sintering technology

    Directory of Open Access Journals (Sweden)

    Wu Mingxia

    2015-01-01

    Full Text Available For the purpose of extensive utilization of powder metallurgy to micro/nano- fabrication of materials, the micro gear was prepared by a novel method, named as micro- forming fields activated sintering technology (Micro-FAST. Surface-cleaning of particles, especially during the initial stage of sintering, is a crucial issue for the densification mechanism. However, up to date, the mechanism of surface-cleaning is too complicated to be known. In this paper, the process of surface-cleaning of Micro-FAST was studied, employing the high resolution transmission electron microscopy (HRTEM for observation of microstructure of micro-particles. According to the evolution of the microstructure, surface-cleaning is mainly ascribed to the effect of electro-thermal focusing. The process of surface-cleaning is achieved through rearrangement of grains, formation of vacancy, migration of vacancy and enhancement of electro-thermal focusing.

  3. Identification of Triclosan-O-Sulfate and other transformation products of Triclosan formed by activated sludge.

    Science.gov (United States)

    Chen, Xijuan; Casas, Mònica Escolà; Nielsen, Jeppe Lund; Wimmer, Reinhard; Bester, Kai

    2015-02-01

    Aerobic degradation experiments of Triclosan were performed in activated sludge to identify possible transformation products for this compound. During 7 days, the formation of biotransformation products such as 2,4-Dichlorophenol, 4-Chlorocatechol, 5-Hydroxy-Triclosan and other Monohydroxy-Triclosan derivatives as well as Dihydroxy-Triclosan-derivatives were observed. The structure of 5-Hydroxy-Triclosan was elucidated by NMR data for the first time in sludge degradation experiments. Additionally the production of a hitherto unknown transformation product in sludge, i.e., Triclosan-O-Sulfate was detected. During the incubations, the concentrations of this transformation product changed from zero to 330 μg L(-1). Based on the analysis of the biodegradation products, three types of reactions were identified: 1) chemical scission of ether bond to form phenols and catechols, 2) addition of OH moieties to the aromatic ring, and 3) adding of methyl or sulfate groups to the original hydroxyl group.

  4. The role of biofilms in the sedimentology of actively forming gypsum deposits at Guerrero Negro, Mexico.

    Science.gov (United States)

    Vogel, Marilyn B; Des Marais, David J; Turk, Kendra A; Parenteau, Mary N; Jahnke, Linda L; Kubo, Michael D Y

    2009-11-01

    Actively forming gypsum deposits at the Guerrero Negro sabkha and saltern system provided habitats for stratified, pigmented microbial communities that exhibited significant morphological and phylogenetic diversity. These deposits ranged from meter-thick gypsum crusts forming in saltern seawater concentration ponds to columnar microbial mats with internally crystallized gypsum granules developing in natural anchialine pools. Gypsum-depositing environments were categorized as forming precipitation surfaces, biofilm-supported surfaces, and clastic surfaces. Each surface type was described in terms of depositional environment, microbial diversity, mineralogy, and sedimentary fabrics. Precipitation surfaces developed in high-salinity subaqueous environments where rates of precipitation outpaced the accumulation of clastic, organic, and/or biofilm layers. These surfaces hosted endolithic biofilms comprised predominantly of oxygenic and anoxygenic phototrophs, sulfate-reducing bacteria, and bacteria from the phylum Bacteroidetes. Biofilm-supported deposits developed in lower-salinity subaqueous environments where light and low water-column turbulence supported dense benthic microbial communities comprised mainly of oxygenic phototrophs. In these settings, gypsum granules precipitated in the extracellular polymeric substance (EPS) matrix as individual granules exhibiting distinctive morphologies. Clastic surfaces developed in sabkha mudflats that included gypsum, carbonate, and siliclastic particles with thin gypsum/biofilm components. Clastic surfaces were influenced by subsurface brine sheets and capillary evaporation and precipitated subsedimentary gypsum discs in deeper regions. Biofilms appeared to influence both chemical and physical sedimentary processes in the various subaqueous and subaerially exposed environments studied. Biofilm interaction with chemical sedimentary processes included dissolution and granularization of precipitation surfaces, formation of

  5. Multipole analysis of unidirectional light scattering from plasmonic dimers

    Science.gov (United States)

    Poutrina, E.; Urbas, A.

    2014-11-01

    We analyze unidirectional scattering produced by sub-wavelength plasmonic dimers formed by two silver strips separated by a thin dielectric spacer and embedded in a uniform dielectric medium. Achieving the Kerker condition, which requires matching the strengths of the electric and magnetic-type contributions of the same multipolar order, is possible with such structures for both forward and backward unidirectional scattering by matching the geometric shape-leveraged resonant magnetic dipolar response with the off-resonant electric dipolar contribution. However, unidirectionality is strongly affected by coupling between the two elements in the dimer structure, leading to the manifestation of the electric quadrupole response in the far field. We develop an approach allowing for an easy inverse scattering retrieval of various multipole contributions to the far-field pattern produced by this type of geometry. The retrieval shows unambiguously that the electric quadrupole response contributes up to 30% of the scattered far-field intensity, in addition to strong manifestation of both electric and magnetic dipolar modes. A modified condition for unidirectionality can be developed based on the principle that suppression of radiation in either the forward or backward direction can be achieved whenever the combined strength of multipolar modes of a certain parity, radiating along the propagation direction, matches that of an opposite parity, and noting that parities of electric and magnetic modes interchange with increasing multipole order. With this condition satisfied, unidirectionality of 26 dB/17 dB for forward/backward scattering, respectively, can be achieved with dimer geometries. We also perform a detailed quantitative analysis of scattering cross sections of dimer structures compared to those of Si and gold spheres, accounting for the actual material losses. We show that dimer structures allow for improving backscattering unidirectionality by 10 dB compared to what

  6. Cooperation of Doxycycline with Phytochemicals and Micronutrients Against Active and Persistent Forms of Borrelia sp.

    Science.gov (United States)

    Goc, Anna; Niedzwiecki, Alexandra; Rath, Matthias

    2016-01-01

    Phytochemicals and micronutrients represent a growing theme in antimicrobial defense; however, little is known about their anti-borreliae effects of reciprocal cooperation with antibiotics. A better understanding of this aspect could advance our knowledge and help improve the efficacy of current approaches towards Borrelia sp. In this study, phytochemicals and micronutrients such as baicalein, luteolin, 10-HAD, iodine, rosmarinic acid, and monolaurin, as well as, vitamins D3 and C were tested in a combinations with doxycycline for their in vitro effectiveness against vegetative (spirochetes) and latent (rounded bodies, biofilm) forms of Borrelia burgdorferi and Borrelia garinii. Anti-borreliae effects were evaluated according to checkerboard assays and supported by statistical analysis. The results showed that combination of doxycycline with flavones such as baicalein and luteolin exhibited additive effects against all morphological forms of studied Borrelia sp. Doxycycline combined with iodine demonstrated additive effects against spirochetes and biofilm, whereas with fatty acids such as monolaurin and 10-HAD it produced FICIs of indifference. Additive anti-spirochetal effects were also observed when doxycycline was used with rosmarinic acid and both vitamins D3 and C. Antagonism was not observed in any of the cases. This data revealed the intrinsic anti-borreliae activity of doxycycline with tested phytochemicals and micronutrients indicating that their addition may enhance efficacy of this antibiotic in combating Borrelia sp. Especially the addition of flavones balcalein and luteolin to a doxycycline regimen could be explored further in defining more effective treatments against these bacteria.

  7. Microwave-assisted extraction of active pharmaceutical ingredient from solid dosage forms.

    Science.gov (United States)

    Hoang, T H; Sharma, R; Susanto, D; Di Maso, M; Kwong, E

    2007-07-13

    The microwave assisted extraction (MAE) technique has been evaluated for the extraction of active pharmaceutical ingredients (API) from various solid dosage forms. Using immediate release tablets of Compound A as a model, optimization of the extraction method with regards to extraction solvent composition, extraction time and temperature was briefly discussed. Complete recovery of Compound A was achieved when samples were extracted using acetonitrile as the extraction solvent under microwave heating at a constant cell temperature of 50 degrees C for 5 min. The optimized MAE method was applied for content uniformity (single tablet extraction) and potency (multiple tablets extraction) assays of release and stability samples of two products of Compound A (5 and 25mg dose strength) stored at various conditions. To further demonstrate the applicability of MAE, the instrumental extraction conditions (50 degrees C for 5 min) were adopted for the extraction of montelukast sodium (Singulair) from various solid dosage forms using methanol-water (75:25, v/v) as the extraction solvent. The MAE procedure demonstrated an extraction efficiency of 97.4-101.9% label claim with the greatest RSD at 1.4%. The results compare favorably with 97.6-102.3% label claim with the greatest RSD at 2.9% obtained with validated mechanical extraction procedures. The system is affordable, user-friendly and simple to operate and troubleshoot. Rapid extraction process (7 min/run) along with high throughput capacity (up to 23 samples simultaneously) would lead to reduced cycle time and thus increased productivity.

  8. Molecular Hydrogen and [Fe II] in Active Galactic Nuclei III: LINERS and Star Forming Galaxies

    CERN Document Server

    Riffel, R; Aleman, I; Brotherton, M S; Pastoriza, M G; Bonatto, C J; Dors, O L

    2013-01-01

    We study the kinematics and excitation mechanisms of H2 and [Fe II] lines in a sample of 67 emission-line galaxies with Infrared Telescope Facility SpeX near-infrared (NIR, 0.8-2.4 micrometers) spectroscopy together with new photoionisation models. H2 emission lines are systematically narrower than narrow-line region (NLR) lines, suggesting that the two are, very likely, kinematically disconnected. The new models and emission-line ratios show that the thermal excitation plays an important role not only in active galactic nuclei but also in star forming galaxies. The importance of the thermal excitation in star forming galaxies may be associated with the presence of supernova remnants close to the region emitting H2 lines. This hypothesis is further supported by the similarity between the vibrational and rotational temperatures of H2. We confirm that the diagram involving the line ratios H2 2.121/Br_gamma and [Fe II] 1.257/Pa_beta is an efficient tool for separating emission-line objects according to their dom...

  9. Dimer formation in radiation-irradiated aqueous solution of lysozyme studied by light-scattering-intensity measurement

    Energy Technology Data Exchange (ETDEWEB)

    Hashimoto, S.; Masuda, T.; Kondo, M. (Institute of Physical and Chemical Research, Wako, Saitama (Japan); Tokyo Metropolitan Univ. (Japan). Faculty of Science); Seki, H.; Imamura, M. (Institute of Physical and Chemical Research, Wako, Saitama (Japan))

    1981-07-01

    The reaction of lysozyme with OH radical, Br/sub 2/ anion radical, and e/sup -/sub(aq), produced in an aqueous solution by pulsed electrons and ..gamma..-rays, were investigated. Irradiated enzymes showed an increase in the light scattering intensity (LSI) which is proportional to the absorbed dose. Results obtained from SDS gel electrophoresis confirm dimerization of lysozyme, which is considered to be responsible for the increase in LSI. It was found that the rate constant of the dimerization of protein radicals produced in the reaction with OH radical is 2k = (1.0+-0.3) x 10/sup 6/M/sup -1/s/sup -1/ and the yield of the dimerization is 0.6 in G. The enzymatic activity of the dimer is shown to be reduced to about 30 per cent of that of the intact enzyme. It is concluded that the radiation-induced inactivation of lysozyme is largely due to dimerization.

  10. Dimer formation in radiation-irradiated aqueous solution of lysozyme studied by light-scattering-intensity measurement.

    Science.gov (United States)

    Hashimoto, S; Seki, H; Masuda, T; Imamura, M; Kondo, M

    1981-07-01

    The reaction of lysozyme with OH., Br.-2 and e-aq, produced in an aqueous solution by pulsed electrons and gamma-rays, were investigated. Irradiated enzymes showed an increase in the light scattering intensity (LSI) which is proportional to the absorbed dose. Results obtained from SDS gel electrophoresis confirm dimerization of lysozyme, which is considered to be responsible for the increase in LSI. It was found that the rate constant of the dimerization of protein radicals produced in the reaction with OH. is 2K=(1.0 +/- 0.3) X 10(6)M-1 s-1 and the yield of the dimerization is 0.6 in G. The enzymatic activity of the dimer is shown to be reduced to about 30 per cent of that of the intact enzyme. It is concluded that the radiation-induced inactivation of lysozyme is largely due to dimerization.

  11. Activation of human natural killer cells by the soluble form of cellular prion protein

    Energy Technology Data Exchange (ETDEWEB)

    Seong, Yeon-Jae [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Hafis Clinic, Seoul (Korea, Republic of); Sung, Pil Soo; Jang, Young-Soon; Choi, Young Joon [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Park, Bum-Chan [Aging Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Park, Su-Hyung [Laboratory of Translational Immunology and Vaccinology, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of); Park, Young Woo [Aging Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon (Korea, Republic of); Shin, Eui-Cheol, E-mail: ecshin@kaist.ac.kr [Laboratory of Immunology and Infectious Diseases, Graduate School of Medical Science and Engineering, KAIST, Daejeon (Korea, Republic of)

    2015-08-21

    Cellular prion protein (PrP{sup C}) is widely expressed in various cell types, including cells of the immune system. However, the specific roles of PrP{sup C} in the immune system have not been clearly elucidated. In the present study, we investigated the effects of a soluble form of recombinant PrP{sup C} protein on human natural killer (NK) cells. Recombinant soluble PrP{sup C} protein was generated by fusion of human PrP{sup C} with the Fc portion of human IgG{sub 1} (PrP{sup C}-Fc). PrP{sup C}-Fc binds to the surface of human NK cells, particularly to CD56{sup dim} NK cells. PrP{sup C}-Fc induced the production of cytokines and chemokines and the degranulation of granzyme B from NK cells. In addition, PrP{sup C}-Fc facilitated the IL-15-induced proliferation of NK cells. PrP{sup C}-Fc induced phosphorylation of ERK-1/2 and JNK in NK cells, and inhibitors of the ERK or the JNK pathways abrogated PrP{sup C}-Fc-induced cytokine production in NK cells. In conclusion, the soluble form of recombinant PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways. - Highlights: • Recombinant soluble PrP{sup C} (PrP{sup C}-Fc) was generated by fusion of human PrP{sup C} with IgG1 Fc portion. • PrP{sup C}-Fc protein induces the production of cytokines and degranulation from human NK cells. • PrP{sup C}-Fc protein enhances the IL-15-induced proliferation of human NK cells. • PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways.

  12. Surface characterization and orientation interaction between diamond- like carbon layer structure and dimeric liquid crystals

    Science.gov (United States)

    Naradikian, H.; Petrov, M.; Katranchev, B.; Milenov, T.; Tinchev, S.

    2017-01-01

    Diamond-like carbon (DLC) and amorphous carbon films are very promising type of semiconductor materials. Depending on the hybridization sp2/sp3 ratio, the material’s band gap varies between 0.8 and 3 eV. Moreover carbon films possess different interesting for practice properties: comparable to the Silicon, Diamond like structure has 22-time better thermal conductivity etc. Here we present one type of implementation of such type nanostructure. That is one attempt for orientation of dimeric LC by using of pre-deposited DLC layer with different ratio of sp2/sp3 hybridized carbon content. It could be expected a pronounced π1-π2interaction between s and p orbital levels on the surface and the dimeric ring of LC. We present comparison of surface anchoring strengths of both orientation inter-surfaces DLC/dimeric LC and single wall carbon nanotubes (SWCNT)/dimeric LC. The mechanism of interaction of dimeric LC and activated surfaces with DLC or SWCNT will be discussed. In both cases we have π-π interaction, which in combination with hydrogen bonding, typical for the dimeric LCs, influence the LC alignment. The Raman spectroscopy data evidenced the presence of charge transfer between contacting hexagonal rings of DLC and the C = O groups of the LC molecules.

  13. Quaternary organization of a phytochrome dimer as revealed by cryoelectron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Li, H.; Li, H.; Zhang, J.; Vierstra, R. D.

    2010-06-15

    Phytochromes are a collection of dimeric photoreceptors that direct a diverse array of responses in plants and microorganisms through photoconversion between a red light-absorbing ground state Pr, and a far-red light-absorbing photoactivated state Pfr. Photoconversion from Pr to Pfr is initiated by a light-driven rotation within the covalently attached bilin, which then triggers a series of protein conformational changes in the binding pocket. These movements ultimately affect an appended output module, which often has reversible protein kinase activity. Propagation of the light signal from the bilin to the output module likely depends on the dimerization interface but its architecture and response to phototransformation remain unclear. Here, we used single particle cryoelectron microscopy to determine the quaternary arrangement of the phytochrome dimer as Pr, using the bacteriophytochrome (BphP) from Deinococcus radiodurans. Contrary to the long-standing view that the two monomers are held together solely via their C-terminal region, we provide unambiguous evidence that the N-terminal bilin-binding region of BphP also provides a dimerization interface with the C-terminal kinase domain appearing as a more flexible appendage. The BphP monomers dimerize in parallel with the polypeptides intimately twisting around each other in a right-handed fashion. Based on this electron microscopic picture, we propose that the light-driven conformational changes transmitted from the chromophore to the output module along the spine of this extensive dimer interface is the central feature underpinning phytochrome signaling.

  14. Partition-DFT on the Water Dimer

    CERN Document Server

    Gómez, Sara; Restrepo, Albeiro; Wasserman, Adam

    2016-01-01

    As is well known, the ground-state symmetry group of the water dimer switches from its equilibrium $C_{s}$-character to $C_{2h}$-character as the distance between the two oxygen atoms of the dimer decreases below $R_{\\rm O-O}\\sim 2.5$ \\AA{}. For a range of $R_{\\rm O-O}$ between 1 and 5 \\AA{}, and for both symmetries, we apply Partition Density Functional Theory (PDFT) to find the unique monomer densities that sum to the correct dimer densities while minimizing the sum of the monomer energies. We calculate the work involved in deforming the isolated monomer densities and find that it is slightly larger for the $C_s$ geometry for all $R_{\\rm O-O}$. We discuss how the PDFT densities and the corresponding partition potentials support the orbital-interaction picture of hydrogen-bond formation.

  15. Photoionization of helium dimers; Photoionisation von Heliumdimeren

    Energy Technology Data Exchange (ETDEWEB)

    Havermeier, Tilo

    2010-06-09

    The helium dimer is one of the most weakly bound systems in the universe. This makes it an interesting quantum mechanical object for investigation. These Van der Waals Clusters can be produced in an expansion of a cryogenic gas jet through a small nozzle into vacuum. In the present experiment we examine the interaction of He dimers with synchrotron radiation at an energy range from 64 to 78 eV. We observed different pathways leading to single ionization of both He atoms of the dimer compound. This two close standing ions begin now to dissociate in cause of their coulomb potential. All charged fragments were detected in coincidence with a COLTRIMS system. Especially Interatomic Coulombic Decay (ICD) and the two step process (TS1) were clearly identified. Furthermore a distribution of the internuclear distance was obtained from the measured Kinetic Energy Release (KER). (orig.)

  16. Kosterlitz Thouless Universality in Dimer Models

    CERN Document Server

    Chandrasekharan, S; Chandrasekharan, Shailesh; Strouthos, Costas G.

    2003-01-01

    Using the monomer-dimer representation of strongly coupled U(N) lattice gauge theories with staggered fermions, we study finite temperature chiral phase transitions in (2+1) dimensions. A new cluster algorithm allows us to compute monomer-monomer and dimer-dimer correlations at zero monomer density (chiral limit) accurately on large lattices. This makes it possible to show convincingly, for the first time, that these models undergo a finite temperature phase transition which belongs to the Kosterlitz-Thouless universality class. We find that this universality class is unaffected even in the large N limit. This shows that the mean field analysis often used in this limit breaks down in the critical region.

  17. Structure of isothiocyanic acid dimers. Theoretical and FTIR matrix isolation studies

    Science.gov (United States)

    Krupa, Justyna; Wierzejewska, Maria

    2016-05-01

    A quantum mechanical study of the potential energy surface for the HNCS dimer is reported. The calculations were performed using DFT and ab initio MP2 methods. The most stable is a cyclic form with a double N-H⋯S interaction and the interaction energy in the range of 16.91-18.92 kJ mol-1. An open HNCS dimer bound by the N-H⋯N hydrogen bond is by ca 3.3-5.1 kJ mol-1 less stable. Experimental FTIR matrix isolation studies of HNCS/N2 system show that exclusively less stable open dimer is formed in solid nitrogen. Possible reasons for the observed discrepancy between theory and experiment are discussed.

  18. Widespread and Hidden Active Galactic Nuclei in Star-Forming Galaxies at Redshift > 0.3

    CERN Document Server

    Juneau, Stéphanie; Bournaud, Frédéric; Alexander, David M; Daddi, Emanuele; Mullaney, James R; Magnelli, Benjamin; Kartaltepe, Jeyhan S; Hwang, Ho Seong; Willner, S P; Coil, Alison L; Rosario, David J; Trump, Jonathan R; Weiner, Benjamin J; Willmer, Christopher N A; Cooper, Michael C; Elbaz, David; Faber, S M; Frayer, David T; Kocevski, Dale D; Laird, Elise S; Monkiewicz, Jacqueline A; Nandra, Kirpal; Newman, Jeffrey; Salim, Samir; Symeonidis, Myrto

    2012-01-01

    We characterize the incidence of active galactic nuclei (AGNs) is 0.3 < z < 1 star-forming galaxies by applying multi-wavelength AGN diagnostics (X-ray, optical, mid-infrared, radio) to a sample of galaxies selected at 70-micron from the Far-Infrared Deep Extragalactic Legacy survey (FIDEL). Given the depth of FIDEL, we detect "normal" galaxies on the specific star formation rate (sSFR) sequence as well as starbursting systems with elevated sSFR. We find an overall high occurrence of AGN of 37+/-3%, more than twice as high as in previous studies of galaxies with comparable infrared luminosities and redshifts but in good agreement with the AGN fraction of nearby (0.05 < z < 0.1) galaxies of similar infrared luminosities. The more complete census of AGNs comes from using the recently developed Mass-Excitation (MEx) diagnostic diagram. This optical diagnostic is also sensitive to X-ray weak AGNs and X-ray absorbed AGNs, and reveals that absorbed active nuclei reside almost exclusively in infrared-lum...

  19. WIDESPREAD AND HIDDEN ACTIVE GALACTIC NUCLEI IN STAR-FORMING GALAXIES AT REDSHIFT >0.3

    Energy Technology Data Exchange (ETDEWEB)

    Juneau, Stephanie; Bournaud, Frederic; Daddi, Emanuele; Elbaz, David [CEA-Saclay, DSM/IRFU/SAp, F-91191 Gif-sur-Yvette (France); Dickinson, Mark; Kartaltepe, Jeyhan S. [National Optical Astronomy Observatory, 950 North Cherry Avenue, Tucson, AZ 85719 (United States); Alexander, David M.; Mullaney, James R. [Department of Physics, Durham University, Durham DH1 3LE (United Kingdom); Magnelli, Benjamin [Max-Planck-Instituet fuer extraterrestrische Physik, Postfach 1312, D-85741 Garching bei Muenchen (Germany); Hwang, Ho Seong; Willner, S. P. [Harvard-Smithsonian Center for Astrophysics, 60 Garden Street, Cambridge, MA 02138 (United States); Coil, Alison L. [Department of Physics, Center for Astrophysics and Space Sciences, University of California, 9500 Gilman Dr., La Jolla, San Diego, CA 92093 (United States); Rosario, David J. [Max-Planck-Instituet fuer extraterrestrische Physik, Giessenbachstrasse, D-85748 Garching bei Muenchen (Germany); Trump, Jonathan R.; Faber, S. M.; Kocevski, Dale D. [University of California Observatories/Lick Observatory, University of California, Santa Cruz, CA 95064 (United States); Weiner, Benjamin J.; Willmer, Christopher N. A. [Steward Observatory, University of Arizona, Tucson, AZ 85721 (United States); Cooper, Michael C. [Center for Galaxy Evolution, Department of Physics and Astronomy, University of California-Irvine, 4129 Frederick Reines Hall, Irvine, CA 92697 (United States); Frayer, David T., E-mail: stephanie.juneau@cea.fr [National Radio Astronomy Observatory, P.O. Box 2, Green Bank, WV 24944 (United States); and others

    2013-02-20

    We characterize the incidence of active galactic nuclei (AGNs) in 0.3 < z < 1 star-forming galaxies by applying multi-wavelength AGN diagnostics (X-ray, optical, mid-infrared, radio) to a sample of galaxies selected at 70 {mu}m from the Far-Infrared Deep Extragalactic Legacy survey (FIDEL). Given the depth of FIDEL, we detect 'normal' galaxies on the specific star formation rate (sSFR) sequence as well as starbursting systems with elevated sSFR. We find an overall high occurrence of AGN of 37% {+-} 3%, more than twice as high as in previous studies of galaxies with comparable infrared luminosities and redshifts but in good agreement with the AGN fraction of nearby (0.05 < z < 0.1) galaxies of similar infrared luminosities. The more complete census of AGNs comes from using the recently developed Mass-Excitation (MEx) diagnostic diagram. This optical diagnostic is also sensitive to X-ray weak AGNs and X-ray absorbed AGNs, and reveals that absorbed active nuclei reside almost exclusively in infrared-luminous hosts. The fraction of galaxies hosting an AGN appears to be independent of sSFR and remains elevated both on the sSFR sequence and above. In contrast, the fraction of AGNs that are X-ray absorbed increases substantially with increasing sSFR, possibly due to an increased gas fraction and/or gas density in the host galaxies.

  20. Alarin but not its alternative-splicing form, GALP (Galanin-like peptide) has antimicrobial activity

    Energy Technology Data Exchange (ETDEWEB)

    Wada, Akihiro, E-mail: a-wada@nagasaki-u.ac.jp [Department of Bacteriology, Institute of Tropical Medicine, Nagasaki University, Nagasaki 8528523 (Japan); Wong, Pooi-Fong [Department of Pharmacology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur (Malaysia); Hojo, Hironobu [Department of Applied Biochemistry, Institute of Glycoscience, Tokai University, Kanagawa 2591292 (Japan); Hasegawa, Makoto [Department of Bioscience, Faculty of Bioscience, Nagahama Institute of Bio-Science and Technology, Shiga 5260829 (Japan); Ichinose, Akitoyo [Electron Microscopy Shop Central Laboratory, Institute of Tropical Medicine, Nagasaki University, Nagasaki 8528523 (Japan); Llanes, Rafael [Institute Pedro Kouri, Havana (Cuba); Kubo, Yoshinao [Division of Cytokine Signaling, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 8528523 (Japan); Senba, Masachika [Department of Pathology, Institute of Tropical Medicine, Nagasaki University, Nagasaki 8528523 (Japan); Ichinose, Yoshio [Kenya Research Station, Institute of Tropical Medicine, Nagasaki University, Nagasaki 8528523 (Japan)

    2013-05-03

    Highlights: • Alarin inhibits the growth of E. coli but not S. aureus. • Alarin’s potency is comparable to LL-37 in inhibiting the growth of E. coli. • Alarin can cause bacterial membrane blebbing. • Alalin does not induce hemolysis on erythrocytes. -- Abstract: Alarin is an alternative-splicing form of GALP (galanin-like peptide). It shares only 5 conserved amino acids at the N-terminal region with GALP which is involved in a diverse range of normal brain functions. This study seeks to investigate whether alarin has additional functions due to its differences from GALP. Here, we have shown using a radial diffusion assay that alarin but not GALP inhibited the growth of Escherichia coli (strain ML-35). The conserved N-terminal region, however, remained essential for the antimicrobial activity of alarin as truncated peptides showed reduced killing effect. Moreover, alarin inhibited the growth of E. coli in a similar potency as human cathelicidin LL-37, a well-studied antimicrobial peptide. Electron microscopy further showed that alarin induced bacterial membrane blebbing but unlike LL-37, it did not cause hemolysis of erythrocytes. In addition, alarin is only active against the gram-negative bacteria, E. coli but not the gram-positive bacteria, Staphylococcus aureus. Thus, these data suggest that alarin has potentials as an antimicrobial and should be considered for the development in human therapeutics.

  1. Solvent Effects on the Monomer/Hydrogen-Bonded Dimer Equilibrium in Carboxylic Acids: (+)-(S)-Ketopinic Acid as a Case Study.

    Science.gov (United States)

    Rodríguez Ortega, Pilar Gema; Montejo Gámez, Manuel; Márquez López, Fernando; López González, Juan Jesús

    2016-06-21

    The hydrogen-bond-assisted self-association process of a chiral semirigid carboxylic acid, namely, (+)-(S)-ketopinic acid, has been studied. The multiconformational monomer/dimer equilibrium has been evaluated by means of a concentration-dependent FTIR study that enabled the experimental equilibrium constants of the dimer formation reaction (Kdim ) to be determined in two solvents of different polarity. In CDCl3 , dimeric forms predominate, even in diluted solutions (KdimCF =5.074), whereas in CD3 CN the self-association process is hindered and monomers are always the main species, irrespective of solute concentration (KdimAN =0.194). The reliability of the dimerization constants and the derived mono- and dimeric experimental fractions have been proven by means of accurate matching between the experimental vibrational circular dichroism spectra of the species and the theoretical spectra generated by considering the simultaneous weighted contributions of the concomitant monomers and dimers.

  2. Dimerization and DNA recognition rules of mithramycin and its analogues.

    Science.gov (United States)

    Weidenbach, Stevi; Hou, Caixia; Chen, Jhong-Min; Tsodikov, Oleg V; Rohr, Jürgen

    2016-03-01

    The antineoplastic and antibiotic natural product mithramycin (MTM) is used against cancer-related hypercalcemia and, experimentally, against Ewing sarcoma and lung cancers. MTM exerts its cytotoxic effect by binding DNA as a divalent metal ion (Me(2+))-coordinated dimer and disrupting the function of transcription factors. A precise molecular mechanism of action of MTM, needed to develop MTM analogues selective against desired transcription factors, is lacking. Although it is known that MTM binds G/C-rich DNA, the exact DNA recognition rules that would allow one to map MTM binding sites remain incompletely understood. Towards this goal, we quantitatively investigated dimerization of MTM and several of its analogues, MTM SDK (for Short side chain, DiKeto), MTM SA-Trp (for Short side chain and Acid), MTM SA-Ala, and a biosynthetic precursor premithramycin B (PreMTM B), and measured the binding affinities of these molecules to DNA oligomers of different sequences and structural forms at physiological salt concentrations. We show that MTM and its analogues form stable dimers even in the absence of DNA. All molecules, except for PreMTM B, can bind DNA with the following rank order of affinities (strong to weak): MTM=MTM SDK>MTM SA-Trp>MTM SA-Ala. An X(G/C)(G/C)X motif, where X is any base, is necessary and sufficient for MTM binding to DNA, without a strong dependence on DNA conformation. These recognition rules will aid in mapping MTM sites across different promoters towards development of MTM analogues as useful anticancer agents.

  3. Study of the erythropoiesis activity of nano-encapsulated forms of erythropoietin

    Directory of Open Access Journals (Sweden)

    Zhanagul Khasenbekova

    2014-01-01

    Full Text Available Introduction: The recombinant human erythropoietin (rhEPO is used in the treatment of anemia. In order to improve its pharmacokinetic properties, nanoparticles of biodegradable polymers of natural or synthetic origin were used. The aim of this study was to investigate the effect of new nano-encapsulated forms of recombinant human erythropoietin for oral use on the erythropoiesis in the cyclophosphamide immunosuppression model. Material and methods: The CHOpE immortalized cells culture (a primary producer of rhEPO "Vector" in Russia was used. The following biodegradable polymers were chosen: 0.05% and 0.005% carbopol, 0.05% and 0.005% kollidon, and 0.05% and 0.005% pectin. Immunosuppression was obtained by a single dose of i.p. injection of cyclophosphamide (250 mg/kg in white mice (18-20 g. During the next 5 days, the nano-encapsulated erythropoietin (100 ED/mouse was administered orally to each mouse. After 5 and 10 days, the cell count of the number of blood reticulocytes and the myelogram of bone marrow were performed. The control group of mice received injections of Eprex. Results: On the 5th day of the experiment, the highest level of reticulocyte was observed in the samples of erythropoietin with kollidon (0.05% and pectin (0.005% nanoparticles. On the 10th day, the highest activity was observed in the samples of erythropoietin substance with pectin at 0.05% and 0.005% concentrations. The levels of reticulocytes in these groups reached 13.53% and 14.55%, respectively. The results of the myelogram during immunosuppression showed some activity of erythropoietin in conjunction with both concentrations of pectin when a two-fold increase in the number of erythroblasts was observed on the 5th day. High degrees of erythrokaryocytes in the state of mitosis were observed in the 0.05% pectin samples. Similar results were observed in equivalent groups of control animals on the 10th day of the experiment, which is compatible with the data on Eprex

  4. 75 FR 61508 - Agency Information Collection Activities: Canadian Border Boat Landing Permit (CBP Form I-68)

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    2010-10-05

    ... Landing Permit (CBP Form I-68) AGENCY: U.S. Customs and Border Protection (CBP), Department of Homeland... concerning the Canadian Border Boat Landing Permit (Form I- ] 68). This request for comment is being made... Form I-68. Abstract: The Canadian Border Boat Landing Permit (CBP Form I-68) allows...

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    2011-01-18

    ... Landing Permit (CBP Form I-68) AGENCY: U.S. Customs and Border Protection, Department of Homeland Security... (CBP Form I-68). This is a proposed extension of an information collection that was previously approved... Landing Permit. OMB Number: 1651-0108. Form Number: CBP Form I-68. Abstract: The Canadian Border...

  6. 78 FR 73875 - Agency Information Collection Activities: Canadian Border Boat Landing Permit (CBP Form I-68)

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    2013-02-27

    ... for Employment Authorization, Form I-765; Form I-765 Work Sheet, Form I- 765WS; Revision of a... collection, the categories of respondents, and the estimated burden (i.e. the time, effort, and resources used by the respondents to respond). For Forms I-765 and I-765WS, USCIS is especially interested in...

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    2012-12-17

    ... for Employment Authorization, Form I-765; Form I-765 Work Sheet, Form I- 765WS; Revision of a... the estimated burden (i.e. the time, effort, and resources used by the respondents to respond). For Forms I-765 and I-765WS, USCIS is especially interested in the public's experience, input, and...

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    2010-01-20

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    2012-10-30

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    2013-05-13

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  12. Measuring patient activation in the Netherlands: translation and validation of the American short form Patient Activation Measure (PAM13

    Directory of Open Access Journals (Sweden)

    Rademakers Jany

    2012-07-01

    Full Text Available Abstract Background The American short form Patient Activation Measure (PAM is a 13-item instrument which assesses patient (or consumer self-reported knowledge, skills and confidence for self-management of one’s health or chronic condition. In this study the PAM was translated into a Dutch version; psychometric properties of the Dutch version were established and the instrument was validated in a panel of chronically ill patients. Methods The translation was done according to WHO guidelines. The PAM 13-Dutch was sent to 4178 members of the Dutch National Panel of people with Chronic illness or Disability (NPCD in April 2010 (study A and again to a sub sample of this group (N = 973 in June 2010 (study B. Internal consistency, test-retest reliability and cross-validation with the SBSQ-D (a measure for Health literacy were computed. The Dutch results were compared to similar Danish and American data. Results The psychometric properties of the PAM 13-Dutch were generally good. The level of internal consistency is good (α = 0.88 and item-rest correlations are moderate to strong. The Dutch mean PAM score (61.3 is comparable to the American (61.9 and lower than the Danish (64.2. The test-retest reliability was moderate. The association with Health literacy was weak to moderate. Conclusions The PAM-13 Dutch is a reliable instrument to measure patient activation. More research is needed into the validity of the Patient Activation Measure, especially with respect to a more comprehensive measure of Health literacy.

  13. Refined conformal spectra in the dimer model

    CERN Document Server

    Rasmussen, Jorgen

    2012-01-01

    Working with Lieb's transfer matrix for the dimer model, we point out that the full set of dimer configurations may be partitioned into disjoint subsets (sectors) closed under the action of the transfer matrix. These sectors are labelled by an integer or half-integer quantum number we call the variation index. In the continuum scaling limit, each sector gives rise to a representation of the Virasoro algebra. We determine the corresponding conformal partition functions and their finitizations, and observe an intriguing link to the Ramond and Neveu-Schwarz sectors of the critical dense polymer model as described by a conformal field theory with central charge c=-2.

  14. Structure of Cyclic Aryl Thiosester Dimer Based on o—Phthaloyl Dichloride and Bis(4—mercaptophenyl)Sulfide

    Institute of Scientific and Technical Information of China (English)

    郭庆中; 王红华; 陈天禄

    2003-01-01

    A Cyclic aryl thioester dimer was prepared by the reaction of o-phthaloyl dichloride and bis(4-mercaptophenyl)sulfide in good yield under pseudo-high dilution conditions via interfacial polycondensation.The structure of the cyclic dimer was confirmed by a conmbination of MALDI-TOF-Ms,FTIR,gel permeation chromatography and NMR analyses.The X-ray diffraction study of the single crystal of cyclic thioester dimer obtained form two sotutions reveals no severe internal strain on the cyclic structure.

  15. Round-robin testing of a reference glass for low-activity waste forms

    Energy Technology Data Exchange (ETDEWEB)

    Ebert, W. L.; Wolf, S. F.

    1999-12-06

    A round robin test program was conducted with a glass that was developed for use as a standard test material for acceptance testing of low-activity waste glasses made with Hanford tank wastes. The glass is referred to as the low-activity test reference material (LRM). The program was conducted to measure the interlaboratory reproducibility of composition analysis and durability test results. Participants were allowed to select the methods used to analyze the glass composition. The durability tests closely followed the Product Consistency Test (PCT) Method A, except that tests were conducted at both 40 and 90 C and that parallel tests with a reference glass were not required. Samples of LRM glass that had been crushed, sieved, and washed to remove fines were provided to participants for tests and analyses. The reproducibility of both the composition and PCT results compare favorably with the results of interlaboratory studies conducted with other glasses. From the perspective of reproducibility of analysis results, this glass is acceptable for use as a composition standard for nonradioactive components of low-activity waste forms present at >0.1 elemental mass % and as a test standard for PCTS at 40 and 90 C. For PCT with LRM glass, the expected test results at the 95% confidence level are as follows: (1) at 40 C: pH = 9.86 {+-} 0.96; [B] = 2.30 {+-} 1.25 mg/L; [Na] = 19.7 {+-} 7.3 mg/L; [Si] = 13.7 {+-} 4.2 mg/L; and (2) at 90 C: pH = 10.92 {+-} 0.43; [B] = 26.7 {+-} 7.2 mg/L; [Na] = 160 {+-} 13 mg/L; [Si] = 82.0 {+-} 12.7 mg/L. These ranges can be used to evaluate the accuracy of PCTS conducted at other laboratories.

  16. The pretranslocation ribosome is targeted by GTP-bound EF-G in partially activated form

    Science.gov (United States)

    Hauryliuk, Vasili; Mitkevich, Vladimir A.; Eliseeva, Natalia A.; Petrushanko, Irina Yu.; Ehrenberg, Måns; Makarov, Alexander A.

    2008-01-01

    Translocation of the tRNA·mRNA complex through the bacterial ribosome is driven by the multidomain guanosine triphosphatase elongation factor G (EF-G). We have used isothermal titration calorimetry to characterize the binding of GDP and GTP to free EF-G at 4°C, 20°C, and 37°C. The binding affinity of EF-G is higher to GDP than to GTP at 4°C, but lower at 37°C. The binding enthalpy and entropy change little with temperature in the case of GDP binding but change greatly in the case of GTP binding. These observations are compatible with a large decrease in the solvent-accessible hydrophobic surface area of EF-G on GTP, but not GDP, binding. The explanation we propose is the locking of the switch 1 and switch 2 peptide loops in the G domain of EF-G to the γ-phosphate of GTP. From these data, in conjunction with previously reported structural data on guanine nucleotide-bound EF-G, we suggest that EF-G enters the pretranslocation ribosome as an “activity chimera,” with the G domain activated by the presence of GTP but the overall factor conformation in the inactive form typical of a GDP-bound multidomain guanosine triphosphatase. We propose that the active overall conformation of EF-G is attained only in complex with the ribosome in its “ratcheted state,” with hybrid tRNA binding sites. PMID:18836081

  17. Cellulose production, activated by cyclic di-GMP through BcsA and BcsZ, is a virulence factor and an essential determinant of the three-dimensional architectures of biofilms formed by Erwinia amylovora Ea1189.

    Science.gov (United States)

    Castiblanco, Luisa F; Sundin, George W

    2016-10-18

    Bacterial biofilms are multicellular aggregates encased in an extracellular matrix mainly composed of exopolysaccharides (EPSs), protein and nucleic acids, which determines the architecture of the biofilm. Erwinia amylovora Ea1189 forms a biofilm inside the xylem of its host, which results in vessel plugging and water transport impairment. The production of the EPSs amylovoran and levan is critical for the formation of a mature biofilm. In addition, cyclic dimeric GMP (c-di-GMP) has been reported to positively regulate amylovoran biosynthesis and biofilm formation in E. amylovora Ea1189. In this study, we demonstrate that cellulose is synthesized by E. amylovora Ea1189 and is a major modulator of the three-dimensional characteristics of biofilms formed by this bacterium, and also contributes to virulence during systemic host invasion. In addition, we demonstrate that the activation of cellulose biosynthesis in E. amylovora is a c-di-GMP-dependent process, through allosteric binding to the cellulose catalytic subunit BcsA. We also report that the endoglucanase BcsZ is a key player in c-di-GMP activation of cellulose biosynthesis. Our results provide evidence of the complex composition of the extracellular matrix produced by E. amylovora and the implications of cellulose biosynthesis in shaping the architecture of the biofilm and in the expression of one of the main virulence phenotypes of this pathogen.

  18. Time-resolved EPR studies with DNA photolyase: excited-state FADH0 abstracts an electron from Trp-306 to generate FADH-, the catalytically active form of the cofactor.

    Science.gov (United States)

    Kim, S T; Sancar, A; Essenmacher, C; Babcock, G T

    1993-09-01

    Photolyase repairs UV-induced cyclobutane-pyrimidine dimers in DNA by photoinduced electron transfer. The enzyme isolated from Escherichia coli contains 5,10-methenyltetrahydrofolate, which functions as the light-harvesting chromophore, and fully reduced flavin adenine dinucleotide (FAD), which functions as the redox catalyst. During enzyme preparation, the flavin is oxidized to FADH0, which is catalytically inert. Illumination of the enzyme with 300- to 600-nm light converts the flavin to the fully reduced form in a reaction that involves photooxidation of an amino acid in the apoenzyme. The results of earlier optical studies had indicated that the redox-active amino acid in this photoactivation process was tryptophan. We have now used time-resolved electron paramagnetic resonance (EPR) spectroscopy to investigate the photoactivation reaction. Excitation of the flavin-radical-containing inactive enzyme produces a spin-polarized radical that we identify by 2H and 15N labeling as originating from a tryptophan residue, confirming the inferences from the optical work. These results and Trp-->Phe replacement by site-directed mutagenesis reveal that flavin radical photoreduction is achieved by electron abstraction from Trp-306 by the excited-state FADH0. Analysis of the hyperfine couplings and spin density distribution deduced from the isotopic-labeling results shows that the product of the light-driven redox chemistry is the Trp-306 cation radical. The results strongly suggest that the active form of photolyase contains FADH- and not FADH2.

  19. Structural basis for the photoconversion of a phytochrome to the activated Pfr form.

    Science.gov (United States)

    Ulijasz, Andrew T; Cornilescu, Gabriel; Cornilescu, Claudia C; Zhang, Junrui; Rivera, Mario; Markley, John L; Vierstra, Richard D

    2010-01-14

    Phytochromes are a collection of bilin-containing photoreceptors that regulate numerous photoresponses in plants and microorganisms through their ability to photointerconvert between a red-light-absorbing, ground state (Pr) and a far-red-light-absorbing, photoactivated state (Pfr). Although the structures of several phytochromes as Pr have been determined, little is known about the structure of Pfr and how it initiates signalling. Here we describe the three-dimensional solution structure of the bilin-binding domain as Pfr, using the cyanobacterial phytochrome from Synechococcus OSB'. Contrary to predictions, light-induced rotation of the A pyrrole ring but not the D ring is the primary motion of the chromophore during photoconversion. Subsequent rearrangements within the protein then affect intradomain and interdomain contact sites within the phytochrome dimer. On the basis of our models, we propose that phytochromes act by propagating reversible light-driven conformational changes in the bilin to altered contacts between the adjacent output domains, which in most phytochromes direct differential phosphotransfer.

  20. Chiroptical properties, binding affinity, and photostability of a conjugated zinc porphyrin dimer complexed with left-handed Z-DNA and right-handed B-DNA.

    Science.gov (United States)

    Choi, Jung Kyu; Reed, Aisha; Balaz, Milan

    2014-01-14

    We have studied the UV-vis absorption and chiroptical properties, binding affinity and photostability of a conjugated positively charged butadiyne-linked Zn(ii) porphyrin dimer bound to DNA sequence poly(dG-dC)2. Right-handed B-DNA, spermine-induced Z-DNA and Co(iii)-induced Z-DNA have been explored. Resonance light scattering (RLS) spectra showed formation of porphyrin aggregates in the presence of all DNA forms with the largest aggregates formed with B-DNA. The porphyrin dimer gave rise to induced bisignate circular dichroism (CD) signals in the presence of the left-handed Z-DNA conformations. On the other hand, the dimer stayed nearly chiroptically silent when complexed with the B-form of poly(dG-dC)2. Our results indicated that the conjugated Zn(ii) porphyrin dimer can be used as a sensor for the chiroptical detection of Z-DNA in the visible (400-500 nm) and near-infrared region of the electromagnetic spectrum (700-800 nm). The helicity of DNA had little effect on the dimer binding affinities. The photostability of the porphyrin dimer complexed with any form of DNA was higher than that of the free molecule. The porphyrin dimer bound to Z-DNA exhibited slower photobleaching than the B-DNA dimer complex.

  1. Surprising Impact of Remote Groups on the Folding-Unfolding and Dimer-Chain Equilibria of Bifunctionl H-Bonding Unimers

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Rui; Cheng, Shuang; Baker, Erin Shammel; Smith, Richard D.; Zeng, Xiao Cheng; Gong, Bing

    2016-01-28

    Oligoamide 1, consisting of two H-bonding units linked by a trimethylene linker, was previously found to form a very stable, folded dimer. In this work, replacing the side chains and end groups of 1 led to derivatives that show the surprising impact of end groups on the folding and dimer-chain equilibria of the resultant molecules.

  2. Amyloid-beta(29-42) dimer formations studied by a multicanonical-multioverlap molecular dynamics simulation.

    Science.gov (United States)

    Itoh, Satoru G; Okamoto, Yuko

    2008-03-13

    Amyloid-beta peptides are known to form amyloid fibrils and are considered to play an important role in Alzheimer's disease. Amyloid-beta(29-42) is a fragment of the amyloid-beta peptide and also has a tendency to form amyloid fibrils. In order to study the mechanism of amyloidogenesis of this fragment, we applied one of the generalized-ensemble algorithms, the multicanonical-multioverlap algorithm, to amyloid-beta(29-42) dimer in aqueous solution. We obtained a detailed free-energy landscape of the dimer system. From the detailed free-energy landscape, we examined monomer and dimer formations of amyloid-beta(29-42) and deduced dimerization processes, which correspond to seeding processes in the amyloidogenesis of amyloid-beta(29-42).

  3. Residues involved in the pore-forming activity of the Clostridium perfringens iota toxin.

    Science.gov (United States)

    Knapp, Oliver; Maier, Elke; Waltenberger, Eva; Mazuet, Christelle; Benz, Roland; Popoff, Michel R

    2015-02-01

    Clostridium perfringens iota toxin is a binary toxin that is organized into enzyme (Ia) and binding (Ib) components. Ib forms channels in lipid bilayers and mediates the transport of Ia into the target cells. Here we show that Ib residues 334-359 contain a conserved pattern of alternating hydrophobic and hydrophilic residues forming two amphipathic β-strands involved in membrane insertion and channel formation. This stretch of amino acids shows remarkable structural and functional analogies with the β-pore-forming domain of C. perfringens epsilon toxin. Several mutations within the two amphipathic β-strands affected pore formation, single-channel conductance and ion selectivity (S339E-S341E, Q345H N346E) confirming their involvement in channel formation. F454 of Ib corresponds to the Φ-clamp F427 of anthrax protective antigen and F428 of C2II binary toxins. The mutation F454A resulted in a loss of cytotoxicity and strong increase in single-channel conductance (500 pS as compared with 85 pS in 1 M KCl) with a slight decrease in cation selectivity, indicating that the Φ-clamp is highly conserved and crucial for binary toxin activity. In contrast, the mutants Q367D, N430D, L443E had no or only minor effects on Ib properties, while T360I, T360A and T360W caused a dramatic effect on ion selectivity and single-channel conductance, indicating gross disturbance of the oligomer structure. This suggests that, at least in the iota toxin family, T360 has a structural role in the pore organization. Moreover, introduction of charged residues within the channel (S339E-S341E) or in the vestibule (Q367D, N430D and L443E) had virtually no effect on chloroquine or Ia binding, whereas F454A, T360I, T360A and T360W strongly decreased the chloroquine and Ia affinity to Ib. These results support that distinct residues within the vestibule interact with chloroquine and Ia or are responsible for channel structure, while the channel lining amino acids play a less important role.

  4. Plasmid control of 6-aminohexanoic acid cyclic dimer degradation enzymes of Flavobacterium sp. KI72.

    Science.gov (United States)

    Negoro, S; Shinagawa, H; Nakata, A; Kinoshita, S; Hatozaki, T; Okada, H

    1980-07-01

    Flavobacterium sp. K172, which is able to grow on 6-aminohexanoic acid cyclic dimer as the sole source of carbon and nitrogen, and plasmid control of the responsible enzymes, 6-aminohexanoic acid cyclic dimer hydrolase and 6-aminohexanoic acid linear oligomer hydrolase, were studied. The wild strain of K172 harbors three kinds of plasmid, pOAD1 (26.2 megadaltons), pOAD2 (28.8 megadaltons), and pOAD3 (37.2 megadaltons). The wild strain K172 was readily cured of its ability to grow on the cyclic dimer by mitomycin C, and the cyclic dimer hydrolase could not be detected either as catalytic activity or by antibody precipitation. No reversion of the cured strains was detected. pOAD2 was not detected in every cured strain tested but was restored in a transformant. The transformant recovered both of the enzyme activities, and the cyclic dimer hydrolase of the transformant was immunologically identical with that of the wild strain. All of the strains tested, including the wild, cured, and transformant ones, possessed identical pOAD3 irrespective of the metabolizing activity. Some of the cured strains possessed pOAD1 identical with the wild strain, but the others harbored plasmids with partially altered structures which were likely to be derived from pOAD1 by genetic rearrangements such as deletion, insertion, or substitution. These results suggested that the genes of the enzymes were borne on pOAD2.

  5. 78 FR 44957 - Agency Information Collection Activities: BioWatch Filter Holder Log, Filter Holder Log DHS Form...

    Science.gov (United States)

    2013-07-25

    ... SECURITY Agency Information Collection Activities: BioWatch Filter Holder Log, Filter Holder Log DHS Form... maintenance of required documentation. The filter holder log form is part of the documentation required by federal law enforcement for the BioWatch Program. The filter holder log is required to create a...

  6. Structure of a Longitudinal Actin Dimer Assembled by Tandem W Domains: Implications for Actin Filament Nucleation

    Energy Technology Data Exchange (ETDEWEB)

    Rebowski, Grzegorz; Namgoong, Suk; Boczkowska, Malgorzata; Leavis, Paul C.; Navaza, Jorge; Dominguez, Roberto (IBS); (BBRI); (UPENN-MED)

    2013-11-20

    Actin filament nucleators initiate polymerization in cells in a regulated manner. A common architecture among these molecules consists of tandem WASP homology 2 domains (W domains) that recruit three to four actin subunits to form a polymerization nucleus. We describe a low-resolution crystal structure of an actin dimer assembled by tandem W domains, where the first W domain is cross-linked to Cys374 of the actin subunit bound to it, whereas the last W domain is followed by the C-terminal pointed end-capping helix of thymosin {beta}4. While the arrangement of actin subunits in the dimer resembles that of a long-pitch helix of the actin filament, important differences are observed. These differences result from steric hindrance of the W domain with intersubunit contacts in the actin filament. We also determined the structure of the first W domain of Vibrio parahaemolyticus VopL cross-linked to actin Cys374 and show it to be nearly identical with non-cross-linked W-Actin structures. This result validates the use of cross-linking as a tool for the study of actin nucleation complexes, whose natural tendency to polymerize interferes with most structural methods. Combined with a biochemical analysis of nucleation, the structures may explain why nucleators based on tandem W domains with short inter-W linkers have relatively weak activity, cannot stay bound to filaments after nucleation, and are unlikely to influence filament elongation. The findings may also explain why nucleation-promoting factors of the Arp2/3 complex, which are related to tandem-W-domain nucleators, are ejected from branch junctions after nucleation. We finally show that the simple addition of the C-terminal pointed end-capping helix of thymosin {beta}4 to tandem W domains can change their activity from actin filament nucleation to monomer sequestration.

  7. Structure of a longitudinal actin dimer assembled by tandem w domains: implications for actin filament nucleation.

    Science.gov (United States)

    Rebowski, Grzegorz; Namgoong, Suk; Boczkowska, Malgorzata; Leavis, Paul C; Navaza, Jorge; Dominguez, Roberto

    2010-10-15

    Actin filament nucleators initiate polymerization in cells in a regulated manner. A common architecture among these molecules consists of tandem WASP homology 2 domains (W domains) that recruit three to four actin subunits to form a polymerization nucleus. We describe a low-resolution crystal structure of an actin dimer assembled by tandem W domains, where the first W domain is cross-linked to Cys374 of the actin subunit bound to it, whereas the last W domain is followed by the C-terminal pointed end-capping helix of thymosin β4. While the arrangement of actin subunits in the dimer resembles that of a long-pitch helix of the actin filament, important differences are observed. These differences result from steric hindrance of the W domain with intersubunit contacts in the actin filament. We also determined the structure of the first W domain of Vibrio parahaemolyticus VopL cross-linked to actin Cys374 and show it to be nearly identical with non-cross-linked W-Actin structures. This result validates the use of cross-linking as a tool for the study of actin nucleation complexes, whose natural tendency to polymerize interferes with most structural methods. Combined with a biochemical analysis of nucleation, the structures may explain why nucleators based on tandem W domains with short inter-W linkers have relatively weak activity, cannot stay bound to filaments after nucleation, and are unlikely to influence filament elongation. The findings may also explain why nucleation-promoting factors of the Arp2/3 complex, which are related to tandem-W-domain nucleators, are ejected from branch junctions after nucleation. We finally show that the simple addition of the C-terminal pointed end-capping helix of thymosin β4 to tandem W domains can change their activity from actin filament nucleation to monomer sequestration.

  8. [Sports activity as a form of increasing efficiency in rehabilitation of handicapped people].

    Science.gov (United States)

    Sobiecka, Joanna

    2002-01-01

    The aim of the research was to find out whether systematic sport training by disabled, fulfills its therapeutic aim in rehabilitation process and gives them back a place in healthy people community. Polish paralympians who took active part in Paralympic Games in years 1992-1998 were questioned (group I). 89% of the respondents of the whole community filled in the questionnaire. The data were compared to the results of the researches among participants of 1972-1988 Paralympic games (group II). The analysis of the influence of systematic training shown that 70% of athletes group I and II, except training, did not have any treatment and sport for 81% from group I and 94% from group II was the only form of rehabilitation and continuation of that process. In spite of professional practising sport, none of paralympians from 1992-1998 noticed worsening of their general health state. Thanks to training, paralympians accepted their disability, and for 62% of respondents in group I and 70% in group II, it was not a barrier to get married. Furthermore, one showed that sport was an agent helping to get better education (23% in years 1992-1998 and 16% in years 1972-1988). 65% respondents (group I) and 60% (group II) assumed that the experience which they got by sport, is helpful in occupational or school duties. The results suggest a need of showing disabled people--if there are no contraindications to physical activity--as early as possible even on hospital rehabilitation state, possibility of continuation of rehabilitation by sport, according to their own interests and physical ability. Medical personnel working in health service should be obliged to qualify patients to training groups, and giving them information on the place where training take part.

  9. 78 FR 2415 - Agency Information Collection Activities: Application for Family Unity Benefits, Form Number I...

    Science.gov (United States)

    2013-01-11

    ... for Family Unity Benefits, Form Number I-817; Revision of a Currently Approved Collection ACTION: 30... approved information. (2) Title of the Form/Collection: Application for Family Unity Benefits. (3)...

  10. A p-quinodimethane-bridged porphyrin dimer.

    Science.gov (United States)

    Zeng, Wangdong; Ishida, Masatoshi; Lee, Sangsu; Sung, Young Mo; Zeng, Zebing; Ni, Yong; Chi, Chunyan; Kim, Dongho; Wu, Jishan

    2013-12-01

    A p-quinodimethane (p-QDM)-bridged porphyrin dimer 1 has been prepared for the first time. An unexpected Michael addition reaction took place when we attempted to synthesize compound 1 by reaction of the cross-conjugated keto-linked porphyrin dimers 8a and 8b with alkynyl/aryl Grignard reagents. Alternatively, compound 1 could be successfully prepared by intramolecular Friedel-Crafts alkylation of the diol-linked porphyrin dimer 14 with concomitant oxidation in air. Compound 1 shows intense one-photon absorption (OPA, λ(max)=955 nm, ε=45400 M(-1) cm(-1)) and a large two-photon absorption (TPA) cross-section (σ((2))(max)=2080 GM at 1800 nm) in the near-infrared (NIR) region due to its extended π-conjugation and quinoidal character. It also exhibits a short singlet excited-state lifetime of 25 ps. The cyclic voltammogram of 1 displays multiple redox waves with a small electrochemical energy gap of 0.86 eV. The ground-state geometry, electronic structure, and optical properties of 1 have been further studied by density functional theory (DFT) calculations and compared with those of the keto-linked dimer 8b. This research has revealed that incorporation of a p-QDM unit into the porphyrin framework had a significant impact on its optical and electronic properties, leading to a novel NIR OPA and TPA chromophore.

  11. The Diamagnetic Susceptibility of the Tubulin Dimer

    Directory of Open Access Journals (Sweden)

    Wim Bras

    2014-01-01

    Full Text Available An approximate value of the diamagnetic anisotropy of the tubulin dimer, Δχdimer, has been determined assuming axial symmetry and that only the α-helices and β-sheets contribute to the anisotropy. Two approaches have been utilized: (a using the value for the Δχα for an α-helical peptide bond given by Pauling (1979 and (b using the previously determined anisotropy of fibrinogen as a calibration standard. The Δχdimer≈4×10-27 JT−2 obtained from these measurements are similar to within 20%. Although Cotton-Mouton measurements alone cannot be used to estimate Δχ directly, the value we measured, CMdimer=1.41±0.03×10-8 T−2cm2mg−1, is consistent with the above estimate for Δχdimer. The method utilized for the determination of the tubulin dimer diamagnetic susceptibility is applicable to other proteins and macromolecular assemblies as well.

  12. Amplitude enhancement by a gold dimer

    Science.gov (United States)

    Hong, Xin; Wang, Jingxin; Jin, Zheng

    2016-10-01

    The unique optical properties such as brightness, non-bleaching, good bio-compatibility make gold particles ideal label candidates for molecular probes. Due to the strongly enhanced field, aggregation of gold nanoparticles finds themselves plenty of applications in bio-imaging. But limited by its small cross-section associated with nanometer sized particle, it is a big challenge to employ it in a single molecular detection. The field enhancement results from the effect of plasmonic coupling between two closely attached gold nanoparticle under the right excitation condition. With the aim to apply the gold dimer probe to find the molecules in our recently established optical detection method, we compared of the amplitude enhancement by the dimer relative to a single particle. The amplitude distribution under a highly focused illumination objective was calculated, whose results suggest that at the optimized excitation condition, the local field can be enhanced 190 fold. In consequence, experimental detection was carried out. Gold dimers were linked together by the hybridization of two single chain DNAs. Dimer and single particle probes were mixed together in one detection. Overwhelming contrast between these two kinds of probes were clearly exhibited in the experimental detection image. This method can provide a way to a high specific detection in early diagnosis.

  13. A new lignan dimer from Mallotus philippensis.

    Science.gov (United States)

    Mai, Nguyen Thi; Cuong, Nguyen Xuan; Thao, Nguyen Phuong; Nam, Nguyen Hoai; Khoi, Nguyen Huu; Minh, Chau Van; Heyden, Yvan Vander; Thuan, Ngo Thi; Tuyen, Nguyen Van; Quetin-Leclercq, Joëlle; Kiem, Phan Van

    2010-03-01

    A new lignan dimer, bilariciresinol (1), was isolated from the leaves of Mallotus philippensis, along with platanoside (2), isovitexin (3), dihydromyricetin (4), bergenin (5), 4-O-galloylbergenin (6), and pachysandiol A (7). Their structures were elucidated by spectroscopic experiments including 1D and 2D NMR and FTICR-MS.

  14. Biological consequences of cyclobutane pyrimidine dimers

    NARCIS (Netherlands)

    Vink, A.A.; Roza, L.

    2001-01-01

    In the skin many molecules may absorb ultraviolet (UV) radiation upon exposure. In particular, cellular DNA strongly absorbs shorter wavelength solar UV radiation, resulting in various types of DNA damage. Among the DNA photoproducts produced the cyclobutane pyrimidine dimers (CPDs) are predominant.

  15. Geometric Reid's recipe for dimer models

    NARCIS (Netherlands)

    Bocklandt, R.; Craw, A.; Quintero Vélez, A.

    2015-01-01

    Crepant resolutions of three-dimensional toric Gorenstein singularities are derived equivalent to noncommutative algebras arising from consistent dimer models. By choosing a special stability parameter and hence a distinguished crepant resolution Y, this derived equivalence generalises the Fourier-M

  16. 75 FR 74070 - Agency Information Collection Activities: Form I-829, Extension of a Currently Approved...

    Science.gov (United States)

    2010-11-30

    ... information collection under review: Form I- 829, Petition by Entrepreneur to Remove Conditions; OMB Control... the Form/Collection: Petition by Entrepreneur to Remove Conditions. (3) Agency form number, if any... conditional resident alien entrepreneur who obtained such status through a qualifying investment, to apply...

  17. 77 FR 33758 - Agency Information Collection Activities: Form I-829, Extension of a Currently Approved...

    Science.gov (United States)

    2012-06-07

    ... Information Collection Under Review: Form I- 829, Petition by Entrepreneur to Remove Conditions. The... collection. (2) Title of the Form/Collection: Petition by Entrepreneur to Remove Conditions. (3) Agency form... a conditional resident alien entrepreneur who obtained such status through a qualifying...

  18. Mid- to far infrared properties of star-forming galaxies and active galactic nuclei

    CERN Document Server

    Magdis, G E; Helou, G; Farrah, D; Hurley, P; Alonso-Herrero, A; Bock, J; Burgarella, D; Chapman, S; Charmandaris, V; Cooray, A; Dai, Y S; Dale, D; Elbaz, D; Feltre, A; Hatziminaoglou, E; Huang, J-S; Morrison, G; Oliver, S; Page, M; Scott, D; Shi, Y

    2013-01-01

    We study the mid- to far-IR properties of a 24um-selected flux-limited sample (S24 > 5mJy) of 154 intermediate redshift (~0.15), infrared luminous galaxies, drawn from the 5MUSES survey. By combining existing mid-IR spectroscopy and new Herschel SPIRE submm photometry from the HerMES program, we derived robust total infrared luminosity (LIR) and dust mass (Md) estimates and infered the relative contribution of the AGN to the infrared energy budget of the sources. We found that the total infrared emission of galaxies with weak 6.2um PAH emission (EW0.2um more than 50% of the LIR arises from star formation. We also found that for galaxies detected in the 250-500um Herschel bands an AGN has a statistically insignificant effect on the temperature of the cold dust and the far-IR colours of the host galaxy, which are primarily shaped by star formation activity. For star-forming galaxies we reveal an anti-correlation between the LIR-to-rest-frame 8um luminosity ratio, IR8 = LIR\\L8, and the strength of PAH features. ...

  19. Sejong open cluster survey (SOS) - V. The active star forming region Sh 2-255 -- 257

    CERN Document Server

    Lim, Beomdu; Hur, Hyeonoh; Lee, Byeong-Cheol; Bessell, Michael S; Kim, Jinyoung S; Lee, Kang Hwan; Park, Byeong-Gon; Jeong, Gwanghui

    2015-01-01

    There is much observational evidence that active star formation is taking place in the HII regions Sh 2-255 -- 257. We present a photometric study of this star forming region (SFR) using imaging data obtained in passbands from the optical to the mid-infrared in order to study the star formation process. A total of 218 members were identified using various selection criteria based on their observational properties. The SFR is reddened by at least E(B-V) = 0.8 mag, and the reddening law toward the region is normal (R_V = 3.1). From the zero-age main sequence fitting method it is confirmed that the SFR is 2.1 +/- 0.3 kpc from the Sun. The median age of the identified members is estimated to be about 1.3 Myr from comparison of the Hertzsprung-Russell diagram (HRD) with stellar evolutionary models. The initial mass function (IMF) is derived from the HRD and the near-infrared (J, J-H) color-magnitude diagram. The slope of the IMF is about Gamma = -1.6 +/- 0.1, which is slightly steeper than that of the Salpeter/Kro...

  20. Heterotetrameric forms of human phenylalanine hydroxylase: co-expression of wild-type and mutant forms in a bicistronic system.

    Science.gov (United States)

    Leandro, João; Leandro, Paula; Flatmark, Torgeir

    2011-05-01

    Hybrid forms of human phenylalanine hydroxylase (hPAH) mutants have been found to present catalytic activities lower than predicted from the individual recombinant forms, indicating that interallelic complementation could be a major determinant of the metabolic phenotype of compound heterozygous phenylketonuric (PKU) patients. To provide a molecular explanation for interallelic complementation we have here developed a bicistronic expression system and a purification strategy to obtain isolated hPAH heteromeric forms. On co-expression of WT-hPAH (~50% tetramer; ~10% dimer) and the N- and C-terminally truncated form ΔN102/ΔC24-hPAH (~80% dimer) no heterodimers were recovered. Moreover, by co-expression of WT-hPAH and the N-terminally truncated form ΔN102-hPAH (~95% tetramer), heterotetramers, as a result of an assembly of two different homodimers, were isolated. The recovered (WT)/(ΔN102)-hPAH heterotetramers revealed a catalytic activity deviating significantly from that calculated by averaging the respective recombinant homotetrameric forms. The heterotetramer assembly also results in conformational changes in the WT-hPAH protomer, as detected by trypsin limited proteolysis. The finding that the presence of two homodimers with different kinetic parameters influences the properties of the resulting heterotetrameric protein indicates that the dimers exhibit interactions which are transmitted across the assembled tetramer. The bicistronic expression system developed here allowed the isolation of hybrid forms that exhibit negative interallelic complementation, and may represent a model system for studying the molecular pathogenic mechanisms of PAH gene mutations in compound heterozygous PKU patients, providing the rationale to understand the observed inconsistencies both in genotype/phenotype correlations and in the response to BH(4) supplementation.

  1. The dimer interface of the membrane type 1 matrix metalloproteinase hemopexin domain: crystal structure and biological functions.

    Science.gov (United States)

    Tochowicz, Anna; Goettig, Peter; Evans, Richard; Visse, Robert; Shitomi, Yasuyuki; Palmisano, Ralf; Ito, Noriko; Richter, Klaus; Maskos, Klaus; Franke, Daniel; Svergun, Dmitri; Nagase, Hideaki; Bode, Wolfram; Itoh, Yoshifumi

    2011-03-04

    Homodimerization is an essential step for membrane type 1 matrix metalloproteinase (MT1-MMP) to activate proMMP-2 and to degrade collagen on the cell surface. To uncover the molecular basis of the hemopexin (Hpx) domain-driven dimerization of MT1-MMP, a crystal structure of the Hpx domain was solved at 1.7 Å resolution. Two interactions were identified as potential biological dimer interfaces in the crystal structure, and mutagenesis studies revealed that the biological dimer possesses a symmetrical interaction where blades II and III of molecule A interact with blades III and II of molecule B. The mutations of amino acids involved in the interaction weakened the dimer interaction of Hpx domains in solution, and incorporation of these mutations into the full-length enzyme significantly inhibited dimer-dependent functions on the cell surface, including proMMP-2 activation, collagen degradation, and invasion into the three-dimensional collagen matrix, whereas dimer-independent functions, including gelatin film degradation and two-dimensional cell migration, were not affected. These results shed light on the structural basis of MT1-MMP dimerization that is crucial to promote cellular invasion.

  2. Dielectric relaxation and birefringence study of 7.O5O.7 dimeric liquid crystal compound

    Science.gov (United States)

    Bhattacharjee, Debanjan; Paul-Choudhury, Sandip; Alapati, Parameswara Rao; Bhattacharjee, Ayon

    2016-05-01

    Measurement of dielectric relaxation and birefringence phenomenon of dimeric liquid crystal compound with the dependence of temperature was reported in this paper. Homogeneous (HG) and homeotropic (HT) alignment of the cell are introduced to investigate the dielectric relaxation, activation energy and birefringence. Cole-Cole plots analyzed the dielectric relaxation of the dimeric compound. The observed Cole-Cole plots were semi-circular, and the relaxation mechanism obeys the non-Debye type of relaxation behaviour. Slater's perturbation equations have been used to analysis the activation energy of the compound. The birefringence of the compound has positively anisotropy and thin prism mechanism was used to study the anisotropy of the compound.

  3. Universal bosonic tetramers of dimer-atom-atom structure

    OpenAIRE

    Deltuva, A.

    2012-01-01

    Unstable four-boson states having an approximate dimer-atom-atom structure are studied using momentum-space integral equations for the four-particle transition operators. For a given Efimov trimer the universal properties of the lowest associated tetramer are determined. The impact of this tetramer on the atom-trimer and dimer-dimer collisions is analyzed. The reliability of the three-body dimer-atom-atom model is studied.

  4. The intrinsically disordered RNR inhibitor Sml1 is a dynamic dimer

    DEFF Research Database (Denmark)

    Danielsson, Jens; Liljedahl, Leena; Ba´ra´ny-Wallje, Elsa

    2008-01-01

    degrees C could be determined. The hydrodynamic radius for the monomeric form of Sml1 was determined to be 23.4 A, corresponding to a protein size between those of a globular protein and a coil. Formation of a dimer results in a hydrodynamic radius of 34.4 A. The observed chemical shifts showed...

  5. Integral equation study of soft-repulsive dimeric fluids

    Science.gov (United States)

    Munaò, Gianmarco; Saija, Franz

    2017-03-01

    We study fluid structure and water-like anomalies of a system constituted by dimeric particles interacting via a purely repulsive core-softened potential by means of integral equation theories. In our model, dimers interact through a repulsive pair potential of inverse-power form with a softened repulsion strength. By employing the Ornstein–Zernike approach and the reference interaction site model (RISM) theory, we study the behavior of water-like anomalies upon progressively increasing the elongation λ of the dimers from the monomeric case (λ =0 ) to the tangent configuration (λ =1 ). For each value of the elongation we consider two different values of the interaction potential, corresponding to one and two length scales, with the aim to provide a comprehensive description of the possible fluid scenarios of this model. Our theoretical results are systematically compared with already existing or newly generated Monte Carlo data: we find that theories and simulations agree in providing the picture of a fluid exhibiting density and structural anomalies for low values of λ and for both the two values of the interaction potential. Integral equation theories give accurate predictions for pressure and radial distribution functions, whereas the temperatures where anomalies occur are underestimated. Upon increasing the elongation, the RISM theory still predicts the existence of anomalies; the latter are no longer observed in simulations, since their development is likely precluded by the onset of crystallization. We discuss our results in terms of the reliability of integral equation theories in predicting the existence of water-like anomalies in core-softened fluids.

  6. Dimerization deficiency of enigmatic retinitis pigmentosa-linked rhodopsin mutants

    Science.gov (United States)

    Ploier, Birgit; Caro, Lydia N.; Morizumi, Takefumi; Pandey, Kalpana; Pearring, Jillian N.; Goren, Michael A.; Finnemann, Silvia C.; Graumann, Johannes; Arshavsky, Vadim Y.; Dittman, Jeremy S.; Ernst, Oliver P.; Menon, Anant K.

    2016-10-01

    Retinitis pigmentosa (RP) is a blinding disease often associated with mutations in rhodopsin, a light-sensing G protein-coupled receptor and phospholipid scramblase. Most RP-associated mutations affect rhodopsin's activity or transport to disc membranes. Intriguingly, some mutations produce apparently normal rhodopsins that nevertheless cause disease. Here we show that three such enigmatic mutations--F45L, V209M and F220C--yield fully functional visual pigments that bind the 11-cis retinal chromophore, activate the G protein transducin, traffic to the light-sensitive photoreceptor compartment and scramble phospholipids. However, tests of scramblase activity show that unlike wild-type rhodopsin that functionally reconstitutes into liposomes as dimers or multimers, F45L, V209M and F220C rhodopsins behave as monomers. This result was confirmed in pull-down experiments. Our data suggest that the photoreceptor pathology associated with expression of these enigmatic RP-associated pigments arises from their unexpected inability to dimerize via transmembrane helices 1 and 5.

  7. Universal dimer in a collisionally opaque medium: experimental observables and Efimov resonances.

    Science.gov (United States)

    Machtey, Olga; Kessler, David A; Khaykovich, Lev

    2012-03-30

    A universal dimer is subject to secondary collisions with atoms when formed in a cloud of ultracold atoms via three-body recombination. We show that in a collisionally opaque medium, the value of the scattering length that results in the maximum number of secondary collisions may not correspond to the Efimov resonance at the atom-dimer threshold and thus cannot be automatically associated with it. This result explains a number of controversies in recent experimental results on universal three-body states and supports the emerging evidence for the significant finite range corrections to the first excited Efimov energy level.

  8. Dimerization of HCN in the gas phase: A theoretical mechanistic study

    Science.gov (United States)

    Yim, Min Kyoung; Choe, Joong Chul

    2012-06-01

    Potential energy surfaces for the formation of covalently bound HCN dimers from two molecules of HCN or HNC were determined from CBS-QB3/APNO calculations. Several novel pathways, with and without the aid of protons, were found for the formation of iminoacetonitrile (HNdbnd CHCN), an intermediate in adenine synthesis from HCN by oligomerization. Covalent C-C or C-N bonds between the two monomers were formed after rearrangement of bimolecular complex intermediates. Energetic and kinetic analyses suggest that the proton-catalyzed dimerizations substantially lower reaction barriers but cannot occur efficiently under interstellar conditions.

  9. Tour as a form of cognitive activity of children of preschool age

    OpenAIRE

    Evgeniya Shanc

    2016-01-01

    In article the author considers theoretical approaches to the concept of "activity", "cognitive activity", especially the development of cognitive activity in preschool age; reveals the role of excursions in the development of cognitive activity of preschool children, justifies the need for the organization of excursion activity in the conditions of preschool educational organization.

  10. Kinetic analysis of histamine release due to covalently linked IgE dimers

    Energy Technology Data Exchange (ETDEWEB)

    Dembo, M. (Los Alamos Scientific Lab., NM); Kagey-Sobotka, A.; Lichtenstein, L.M.; Goldstein, B.

    1982-01-01

    We present a kinetic model of histamine release from human basophils due to covalently linked IgE dimers. Comparison of theory with experiment shows that the model gives a good description of histamine release by IgE dimers and allows a number of the parameters of the model to be determined. Comparison with previous models of release by conventional antigens indicates that despite their covalent structure, IgE dimers are subject to the same laws governing inactivation as are antigen produced crosslinks. In addition, the kinetic equation which relates the rate of histamine release to the number of crosslinked Fc/sub e/ receptors per cell is the same for crosslinks formed by IgE dimers as for antigen induced crosslinks. Quantitative fitting of histamine release data also yields a value for the rate constant for crosslink formation by IgE dimer on the cell surface (r/sub x/ approx. = to 5 x 10/sup -10/ cm/sup 2//sec). This rate constant is remarkably high and indicates that the reaction is diffusion controlled.

  11. Stability of functionalized C{sub 60} paramagnetic dimers and monomers

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Michael [Armament Research Development and Engineering Center, Picatinny, NJ 07806-5000 (United States); Owens, Frank J., E-mail: owensfj@gmail.com [Department of Physics, Hunter College, City University of New York, 695 Park Ave., NY 10065 (United States)

    2012-02-06

    Highlights: Black-Right-Pointing-Pointer DFT is used to calculate the bond dissociation energy of functionalized C{sub 60} dimers. Black-Right-Pointing-Pointer The results show the dimers would not be stable above room temperature. Black-Right-Pointing-Pointer The calculations indicate the observed magnetism cannot be due to C{sub 60} dimers. Black-Right-Pointing-Pointer Because of their higher stability the ferromagnetism is likely due to X-C{sub 60} monomers. - Abstract: Density functional theory is used to calculate the bond dissociation energy to cleave the C{sub 60}=C{sub 60} bond of the paramagnetic X-C{sub 60}=C{sub 60}-X and X-C{sub 60}=C{sub 60} dimers where X is F, OH, O and H. The results show that these dimers would not be stable much above room temperature and therefore cannot constitute the paramagnetic phase needed to form the observed ferromagnetism which has been shown to be stable up to 800 K. The calculated bond dissociation energies to remove an F, OH or H from a single C{sub 60} are large suggesting that they could be the source of the unpaired spin needed for the high temperature ferromagnetism.

  12. Viscosity and sedimentation behaviors of the magnetorheological suspensions with oleic acid/dimer acid as surfactants

    Science.gov (United States)

    Yang, Jianjian; Yan, Hua; Hu, Zhide; Ding, Ding

    2016-11-01

    This work deals with the role of polar interactions on the viscosity and sedimentation behaviors of magnetorheological suspensions with micro-sized magnetic particles dispersed in oil carriers. The oleic acid and dimer acid were employed to make an adjustment of the hydrophobicity of iron particles, in the interest of performing a comparative evaluation of the contributions of the surface polarity. The viscosity tests show that the adsorbed surfactant layer may impose a hindrance to the movement of iron particles in the oil medium. The polar attractions between dimer acid covered particles gave rise to a considerable increase in viscosity, indicating flocculation structure developed in the suspensions. The observed plateau-like region in the vicinity of 0.1 s-1 for MRF containing dimer acid is possibly due to the flocculation provoked by the carboxylic polar attraction, in which the structure is stable against fragmentation. Moreover, a quick recovery of the viscosity and a higher viscosity-temperature index also suggest the existence of particle-particle polar interaction in the suspensions containing dimer acid. The sedimentation measurements reveal that the steric repulsion of oleic acid plays a limited role in the stability of suspensions only if a large quantity of surfactant was used. The sedimentation results observed in the dimer acid covered particles confirm that loose and open flocculation was formed and enhanced sedimentation stability.

  13. Longiberine and O-methyllongiberine, dimeric protoberberine-benzyl tetrahydroisoquinoline alkaloids from Thalictrum longistylum(1).

    Science.gov (United States)

    Lee, S S; Wu, W N; Wilton, J H; Beal, J L; Doskotch, R W

    1999-10-01

    Two benzyltetrahydroisoquinoline-protoberberine dimers, longiberine (1) and O-methyllongiberine (2), were isolated from the roots of Thalictrum longistylum and represent a new class of dimeric alkaloids. The structure of longiberine (1) was established by spectral and chemical methods. Reductive cleavage of O-ethyllongiberine (4) with Na/liquid NH(3) yielded (+)-(S)-N-methylcoclaurine (5), which determined one-half of the dimer, and 1D and 2D NMR studies arranged the substituents on the protoberberine nucleus. Chemical conversion of thalidezine (6) to 1 via the O-acetyl N,N-didemethyl derivative 9, which was methylenated in the Mannich reaction and N-methylated by the Eschweiler-Clarke procedure, established the second asymmetric center as S and confirmed the ring size and the order of the substituents for 1. Methylation of 1 with diazomethane formed the O-methyl derivative 2, identical with the natural product.

  14. The research of neurospecific proteins and lysosomal peptidehydrolases in frontal neocortex during forming conditioned reaction of active avoiding of rats

    Directory of Open Access Journals (Sweden)

    Vyatkin O. K.

    2009-04-01

    Full Text Available Dynamics of the activity of neuronal cell adhesion molecule (NCAM and lysosomal cysteine cathepsins B, L, H was researched in frontal neocortex of rat brain during forming a conditioned reaction of active avoiding. The quantitative estimation of NCAM content in the neocortex membrane fraction was carried on by ELISA in 3, 7, 14 and 21 days after starting animals’ training. The dynamics correlation between the NCAM content increasing and cysteine cathepsins activity was obtained, especially for aminopeptidase cathepsin H during the process of memory engram forming in frontal neocortex of rat brain.

  15. Influence of motive activity on forming of proof skills of healthy way of life of students of higher educational establishments.

    Directory of Open Access Journals (Sweden)

    Trotsenko V.V.

    2010-08-01

    Full Text Available The estimation and the analysis of impellent activity of students is considered. The importance of impellent activity in forming a healthy way of life of students is shown. In research the data of questionnaire of students are used. sports priorities of students are revealed should to undertake in attention by development of the program of physical training. The wide spectrum of the reasons which interfere with exercises is revealed. These reasons should be considered during individual work with each student. It is proved, that impellent activity influences forming of proof skills of a healthy way of life of students.

  16. D-dimer assay in Egyptian patients with Gaucher disease: correlation with bone and lung involvement.

    Science.gov (United States)

    Sherif, Eman M; Tantawy, Azza A G; Adly, Amira A M; Kader, Hossam A; Ismail, Eman A R

    2011-04-01

    Gaucher disease is the most frequent lysosomal storage disorder. Bone and lung involvement are two major causes of morbidity in this disease. D-dimer is a reliable indicator of active microvascular thrombosis, even in patients without overt hypercoagulation. This study aimed to assess D-dimer levels in Gaucher disease, correlating this marker to clinical characteristics and radiological parameters to investigate its role as a potential predictor for the occurrence and severity of skeletal and pulmonary manifestations. The study population consisted of 56 Egyptian patients with Gaucher disease, 36 had type 1 Gaucher disease (64.3%) and 20 had type 3 Gaucher disease (35.7%). Thirty healthy individuals were enrolled as a control group. D-dimer levels were significantly higher in all patients with Gaucher disease compared with controls (P < 0.001). Patients with type 3 showed significantly higher D-dimer concentrations compared with type 1 (P < 0.001). Pulmonary involvement was present in a significant proportion among type 3 Gaucher patients (P < 0.05), whereas bone changes were present in a higher percentage in type 1 compared with type 3 Gaucher patients. D-dimers were significantly higher in patients with abnormal MRI findings of the long bones and in those with ground glass appearance on high-resolution computerized tomography of the chest compared with patients with normal radiology (P < 0.001). Splenectomized patients displayed significantly higher D-dimer levels compared with nonsplenectomized patients (P < 0.001). Our results suggest that D-dimer is significantly elevated in Gaucher disease, particularly type 3, and may be considered as a potential marker of risk prediction of bone and lung involvement that could be used to monitor treatment response.

  17. FtsK-dependent dimer resolution on multiple chromosomes in the pathogen Vibrio cholerae.

    Directory of Open Access Journals (Sweden)

    Marie-Eve Val

    Full Text Available Unlike most bacteria, Vibrio cholerae harbors two distinct, nonhomologous circular chromosomes (chromosome I and II. Many features of chromosome II are plasmid-like, which raised questions concerning its chromosomal nature. Plasmid replication and segregation are generally not coordinated with the bacterial cell cycle, further calling into question the mechanisms ensuring the synchronous management of chromosome I and II. Maintenance of circular replicons requires the resolution of dimers created by homologous recombination events. In Escherichia coli, chromosome dimers are resolved by the addition of a crossover at a specific site, dif, by two tyrosine recombinases, XerC and XerD. The process is coordinated with cell division through the activity of a DNA translocase, FtsK. Many E. coli plasmids also use XerCD for dimer resolution. However, the process is FtsK-independent. The two chromosomes of the V. cholerae N16961 strain carry divergent dimer resolution sites, dif1 and dif2. Here, we show that V. cholerae FtsK controls the addition of a crossover at dif1 and dif2 by a common pair of Xer recombinases. In addition, we show that specific DNA motifs dictate its orientation of translocation, the distribution of these motifs on chromosome I and chromosome II supporting the idea that FtsK translocation serves to bring together the resolution sites carried by a dimer at the time of cell division. Taken together, these results suggest that the same FtsK-dependent mechanism coordinates dimer resolution with cell division for each of the two V. cholerae chromosomes. Chromosome II dimer resolution thus stands as a bona fide chromosomal process.

  18. Active form Notch4 promotes the proliferation and differentiation of 3T3-L1 preadipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Peng-Yeh [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Tsai, Chong-Bin [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Department of Ophthalmology, Chiayi Christian Hospital, Chiayi 600, Taiwan, ROC (China); Tseng, Min-Jen, E-mail: biomjt@ccu.edu.tw [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China)

    2013-01-18

    Highlights: ► Notch4IC modulates the ERK pathway and cell cycle to promote 3T3-L1 proliferation. ► Notch4IC facilitates 3T3-L1 differentiation by up-regulating proadipogenic genes. ► Notch4IC promotes proliferation during the early stage of 3T3-L1 adipogenesis. ► Notch4IC enhances differentiation during subsequent stages of 3T3-L1 adipogenesis. -- Abstract: Adipose tissue is composed of adipocytes, which differentiate from precursor cells in a process called adipogenesis. Many signal molecules are involved in the transcriptional control of adipogenesis, including the Notch pathway. Previous adipogenic studies of Notch have focused on Notch1 and HES1; however, the role of other Notch receptors in adipogenesis remains unclear. Q-RT-PCR analyses showed that the augmentation of Notch4 expression during the differentiation of 3T3-L1 preadipocytes was comparable to that of Notch1. To elucidate the role of Notch4 in adipogenesis, the human active form Notch4 (N4IC) was transiently transfected into 3T3-L1 cells. The expression of HES1, Hey1, C/EBPδ and PPARγ was up-regulated, and the expression of Pref-1, an adipogenic inhibitor, was down-regulated. To further characterize the effect of N4IC in adipogenesis, stable cells expressing human N4IC were established. The expression of N4IC promoted proliferation and enhanced differentiation of 3T3-L1 cells compared with those of control cells. These data suggest that N4IC promoted proliferation through modulating the ERK pathway and the cell cycle during the early stage of 3T3-L1 adipogenesis and facilitated differentiation through up-regulating adipogenic genes such as C/EBPα, PPARγ, aP2, LPL and HSL during the middle and late stages of 3T3-L1 adipogenesis.

  19. Sejong Open Cluster Survey (SOS) - V. The Active Star Forming Region SH 2-255-257

    Science.gov (United States)

    Lim, Beomdu; Sung, Hwankyung; Hur, Hyeonoh; Lee, Byeong-Cheol; Bessell, Michael S.; Kim, Jinyoung S.; Lee, Kang Hwan; Park, Byeong-Gon; Jeong, Gwanghui

    2015-12-01

    There is much observational evidence that active star formation is taking place in the H II regions Sh 2-255-257. We present a photometric study of this star forming region (SFR) using imaging data obtained in passbands from the optical to the mid-infrared, in order to study the star formation process. A total of 218 members were identified using various selection criteria based on their observational properties. The SFR is reddened by at least E(B-V) = 0.8 mag, and the reddening law toward the region is normal (R_V = 3.1). From the zero-age main sequence fitting method it is confirmed that the SFR is 2.1 ± 0.3 kpc from the Sun. The median age of the identified members is estimated to be about 1.3 Myr from a comparison of the Hertzsprung-Russell diagram (HRD) with stellar evolutionary models. The initial mass function (IMF) is derived from the HRD and the near-infrared (J, J-H) color-magnitude diagram. The slope of the IMF is about Γ = -1.6 ± 0.1, which is slightly steeper than that of the Salpeter/Kroupa IMF. It implies that low-mass star formation is dominant in the SFR. The sum of the masses of all the identified members provides the lower limit of the cluster mass (169 M_{⊙}). We also analyzed the spectral energy distribution (SED) of pre-main sequence stars using the SED fitting tool of Robitaille et al., and confirm that there is a significant discrepancy between stellar mass and age obtained from two different methods based on the SED fitting tool and the HRD.

  20. DNA Recombinase Proteins, their Function and Structure in the Active Form, a Computational Study

    Science.gov (United States)

    Carra, Claudio; Cucinotta, Francis A.

    2007-01-01

    Homologous recombination is a crucial sequence of reactions in all cells for the repair of double strand DNA (dsDNA) breaks. While it was traditionally considered as a means for generating genetic diversity, it is now known to be essential for restart of collapsed replication forks that have met a lesion on the DNA template (Cox et al., 2000). The central stage of this process requires the presence of the DNA recombinase protein, RecA in bacteria, RadA in archaea, or Rad51 in eukaryotes, which leads to an ATP-mediated DNA strand-exchange process. Despite many years of intense study, some aspects of the biochemical mechanism, and structure of the active form of recombinase proteins are not well understood. Our theoretical study is an attempt to shed light on the main structural and mechanistic issues encountered on the RecA of the e-coli, the RecA of the extremely radio resistant Deinococcus Radiodurans (promoting an inverse DNA strand-exchange repair), and the homolog human Rad51. The conformational changes are analyzed for the naked enzymes, and when they are linked to ATP and ADP. The average structures are determined over 2ns time scale of Langevian dynamics using a collision frequency of 1.0 ps(sup -1). The systems are inserted in an octahedron periodic box with a 10 Angstrom buffer of water molecules explicitly described by the TIP3P model. The corresponding binding free energies are calculated in an implicit solvent using the Poisson-Boltzmann solvent accessible surface area, MM-PBSA model. The role of the ATP is not only in stabilizing the interaction RecA-DNA, but its hydrolysis is required to allow the DNA strand-exchange to proceed. Furthermore, we extended our study, using the hybrid QM/MM method, on the mechanism of this chemical process. All the calculations were performed using the commercial code Amber 9.