WorldWideScience

Sample records for behring alpha proteinase

  1. Production of alpha 1-proteinase inhibitor (human).

    Science.gov (United States)

    Hein, R H; Van Beveren, S M; Shearer, M A; Coan, M H; Brockway, W J

    1990-03-01

    A method for large scale isolation of alpha 1-proteinase inhibitor (alpha 1-PI) is described. This method employs waste Cohn Fraction IV-1 as the starting material and involves fractional precipitation with polyethylene glycol followed by ion exchange chromatography on diethylaminoethanol (DEAE)-Sepharose. The process also incorporates a ten hour, at 60 degrees C, heat-treatment step to reduce or eliminate the risk of transmission of viral disease. The final product, having a purity of approximately 60%, is freeze-dried. This preparation behaves almost identically to the alpha 1-PI in plasma and is suitable for replacement therapy in hereditary emphysema.

  2. [Behring's personal papers--Behring's lives].

    Science.gov (United States)

    Enke, Ulrike

    2014-09-01

    The article wants to show the connection between the enriched personal papers of Emil von Behring (1854-1917) in the Behring archives in Marburg (established in 1927) and the history of the first biography of the scientist, which was published by Heinz Zeiss and Richard Bieling during Nazi era in 1940. One focus is placed on Behring's widow Else von Behring (1876-1936), who was active in arranging Behring's papers in proper order and in searching a biographer of her husband's life. The paper also presents new discoveries from the Behring Works archives in Marburg which show Behring--founder of the serum therapy and first winner of the Nobel Prize for medicine in 1901--as an entrepreneur who was fighting for control and influence in the field of science and of business: maybe another narration of Behring's life.

  3. Neutrophils degrade subendothelial matrices in the presence of alpha-1-proteinase inhibitor. Cooperative use of lysosomal proteinases and oxygen metabolites.

    OpenAIRE

    Weiss, S J; Regiani, S

    1984-01-01

    Triggered neutrophils rapidly degraded labeled matrices secreted by cultured, venous endothelial cells via a process dependent on elastase but not oxygen metabolites. In the presence of high concentrations of alpha-1-proteinase inhibitor, the ability of the stimulated neutrophil to solubilize the matrix was impaired. However, at lower concentrations of alpha-1-proteinase inhibitor the neutrophil could enhance the degradative potential of its released elastase by a H2O2-dependent process. Coin...

  4. The thiol proteinases from the latex of Carica papaya L. IV. Proteolytic specificities of chymopapain and papaya proteinase omega determined by digestion of alpha-globin chains.

    Science.gov (United States)

    Jacquet, A; Kleinschmidt, T; Dubois, T; Schnek, A G; Looze, Y; Braunitzer, G

    1989-08-01

    The proteolytic specificities of chymopapain and papaya proteinase omega were investigated by using the alpha-chains of manatee and mole haemoglobin, whose primary structures are known, as substrates. The resulting peptides from each enzymatic cleavage were isolated by gel filtration on Sephadex G-25, followed by reversed-phase HPLC of the separated peaks and, in some cases, further purified by preparative thin-layer electrophoresis. The purified peptides were then identified on the basis of their amino-acid composition. The proteolytic specificities of chymopapain and papaya proteinase omega, deduced from the experimental cleavage patterns, are compared to that of papain. As in the case of papain, the specificity-determining factor is the amino-acid residue of the substrate that will be bound in subsite S2 (the next but one from the scissible bond). Aromatic residues in this position, preferred by papain, are not important for chymopapain and papaya proteinase omega. Cleavages preferentially occur when S2 is occupied by leucine, valine or threonine. For chymopapain, proline in position S2 also causes cleavage.

  5. Pulmonary penetration of alpha 1-proteinase inhibitor administered parenterally to dogs

    International Nuclear Information System (INIS)

    Smith, R.M.; Spragg, R.G.; Moser, K.M.; Cochrane, C.G.; McCarren, J.P.

    1987-01-01

    To study the penetration of alpha 1-proteinase inhibitor (A1Pl) into the lungs of healthy dogs, 83 mg/kg of active A1Pl was administered intravenously over 30 min followed by a bolus of 131 I-A1Pl. Animals were lavaged 2 to 72 h after infusion, sequential gamma camera scans were acquired, and urine was analyzed for the excretion of desmosine. After a distribution phase, infused A1Pl left the bloodstream with a half-life of 103 +/- 24 h. Analysis of plasma antiprotease activity demonstrated preservation of function of the infused A1Pl. Lavage fluid A1Pl concentration and activity were significantly increased 24 h after infusion. Gamma camera scans demonstrated that lung, liver, and spleen acquired 131 I-A1Pl similarly; radioactivities per gram of tissue of these organs were similar at autopsy. Excretion of desmosine did not decrease from a baseline of 157 +/- 59 nmol/24 h after A1Pl infusion, indicating no effect of A1Pl infusion on background elastolysis. These data suggest that intravenous administration of A1Pl can raise lung antiproteinase levels within 24 h despite the absence of preferential uptake by the lung of the infused protein

  6. Bioequivalence of a Liquid Formulation of Alpha1-Proteinase Inhibitor Compared with Prolastin®-C (Lyophilized Alpha1-PI) in Alpha1-Antitrypsin Deficiency.

    Science.gov (United States)

    Barker, Alan F; Campos, Michael A; Brantly, Mark L; Stocks, James M; Sandhaus, Robert A; Lee, Douglas; Steinmann, Kimberly; Lin, Jiang; Sorrells, Susan

    2017-12-01

    This study evaluated the bioequivalence, safety, and immunogenicity of a new liquid formulation of human plasma-derived alpha 1 -proteinase inhibitor, Liquid Alpha 1 -PI, compared with the Lyophilized Alpha 1 -PI formulation (Prolastin®-C), for augmentation therapy in patients with alpha 1 -antitrypsin deficiency (AATD). In this double-blind, randomized, 20-week crossover study, 32 subjects with AATD were randomized to receive 8 weekly infusions of 60 mg/kg of Liquid Alpha 1 -PI or Lyophilized Alpha 1 -PI. Serial blood samples were drawn for 7 days after the last dose followed by 8 weeks of the alternative treatment. The primary endpoint was bioequivalence at steady state, as measured by area under the concentration versus time curve from 0 to 7 days (AUC 0-7 days ) postdose using an antigenic content assay. Bioequivalence was defined as 90% confidence interval (CI) for the ratio of the geometric least squares (LS) mean of AUC 0-7 days for both products within the limits of 0.80 and 1.25. Safety and immunogenicity were assessed. Mean alpha 1 -PI concentration versus time curves for both formulations were superimposable. Mean AUC 0-7 days was 20 320 versus 19 838 mg × h/dl for Liquid Alpha 1 -PI and Lyophilized Alpha 1 -PI, respectively. The LS mean ratio of AUC 0-7 days (90% CI) for Liquid Alpha 1 -PI versus Lyophilized Alpha 1 -PI was 1.05 (1.03-1.08), indicating bioequivalence. Liquid Alpha 1 -PI was well tolerated and adverse events were consistent with Lyophilized Alpha 1 -PI. Immunogenicity to either product was not detected. In conclusion, Liquid Alpha 1 -PI is bioequivalent to Lyophilized Alpha 1 -PI, with a similar safety profile. The liquid formulation would eliminate the need for reconstitution and shorten preparation time for patients receiving augmentation therapy for AATD.

  7. The Anders Behring Breivik Trial: Performing Justice, Defending Democracy

    NARCIS (Netherlands)

    de Graaf, B.A.; van der Heide, E.J.; Wanmaker, S.; Weggemans, D.J.

    On 24 August 2012, the judges of the Oslo District Court passed their final verdict in the case of Anders Behring Breivik, declaring Breivik criminally sane and legally responsible for the killing of 77 people during the bombing of government buildings in Oslo and the shooting spree on the island of

  8. The Importance of Music to Anders Behring Breivik

    Directory of Open Access Journals (Sweden)

    Joe Stroud

    2013-06-01

    Full Text Available This article considers the significance of popular music to the Norwegian terrorist Anders Behring Breivik, particularly the people he names in his manifesto: Helene Bøksle, Clint Mansell and Saga. Breivik saw these musicians, operating outwith the mainstream music industry, as conforming to his ideology, therefore playing a crucial role in making him feel part of a community during his period of isolation in preparation for his attack. This music also helped to motivate Breivik and maintain his morale. This article considers why this music in particular appealed to Breivik, and what he saw in it to confirm his ideology.

  9. Clinical utility of alpha-1 proteinase inhibitor in the management of adult patients with severe alpha-1 antitrypsin deficiency: a review of the current literature

    Directory of Open Access Journals (Sweden)

    Parr DG

    2017-07-01

    Full Text Available David G Parr, Beatriz Lara Department of Respiratory Medicine, Cardio-Respiratory Division, University Hospital Coventry, Coventry, UK Abstract: Alpha-1 antitrypsin (AAT functions primarily to inhibit neutrophil elastase, and its deficiency predisposes individuals to the development of chronic obstructive pulmonary disease (COPD. The putative protective serum concentration is generally considered to be above a threshold of 11 µM/L, and therapeutic augmentation of AAT above this value is believed to retard the progression of emphysema. Several AAT preparations, all derived from human donor plasma, have been commercialized since approval by the US Food and Drug Administration (FDA in 1987. Biochemical efficacy has been demonstrated by augmentation of pulmonary antiprotease activity, but demonstration of clinical efficacy in randomized, placebo-controlled trials has been hampered by the practical difficulties of performing conventional studies in a rare disease with a relatively long natural history. Computed tomography has been applied to measure lung density as a more specific and sensitive surrogate outcome measure of emphysema than physiologic indices, such as forced expiratory volume in 1 second, and studies consistently show a therapeutic reduction in the rate of lung density decline. However, convincing evidence of benefit using traditional clinical measures remains elusive. Intravenous administration of AAT at a dose of 60 mg/kg/week is the commonest regime in use and has well-documented safety and tolerability. International and national guidelines on the management of AAT deficiency recommend intravenous augmentation therapy to supplement optimized usual COPD treatment in patients with severe deficiency and evidence of lung function impairment. Keywords: alpha-1 antitrypsin deficiency, augmentation or replacement therapy, computed tomography, emphysema, COPD

  10. Phage display of the serpin alpha-1 proteinase inhibitor randomized at consecutive residues in the reactive centre loop and biopanned with or without thrombin.

    Directory of Open Access Journals (Sweden)

    Benjamin M Scott

    Full Text Available In spite of the power of phage display technology to identify variant proteins with novel properties in large libraries, it has only been previously applied to one member of the serpin superfamily. Here we describe phage display of human alpha-1 proteinase inhibitor (API in a T7 bacteriophage system. API M358R fused to the C-terminus of T7 capsid protein 10B was directly shown to form denaturation-resistant complexes with thrombin by electrophoresis and immunoblotting following exposure of intact phages to thrombin. We therefore developed a biopanning protocol in which thrombin-reactive phages were selected using biotinylated anti-thrombin antibodies and streptavidin-coated magnetic beads. A library consisting of displayed API randomized at residues 357 and 358 (P2-P1 yielded predominantly Pro-Arg at these positions after five rounds of thrombin selection; in contrast the same degree of mock selection yielded only non-functional variants. A more diverse library of API M358R randomized at residues 352-356 (P7-P3 was also probed, yielding numerous variants fitting a loose consensus of DLTVS as judged by sequencing of the inserts of plaque-purified phages. The thrombin-selected sequences were transferred en masse into bacterial expression plasmids, and lysates from individual colonies were screening for API-thrombin complexing. The most active candidates from this sixth round of screening contained DITMA and AAFVS at P7-P3 and inhibited thrombin 2.1-fold more rapidly than API M358R with no change in reaction stoichiometry. Deep sequencing using the Ion Torrent platform confirmed that over 800 sequences were significantly enriched in the thrombin-panned versus naïve phage display library, including some detected using the combined phage display/bacterial lysate screening approach. Our results show that API joins Plasminogen Activator Inhibitor-1 (PAI-1 as a serpin amenable to phage display and suggest the utility of this approach for the selection

  11. The Anders Behring Breivik Trial: Performing Justice, Defending Democracy

    Directory of Open Access Journals (Sweden)

    Beatrice de Graaf

    2013-08-01

    Full Text Available On 24 August 2012, the judges of the Oslo District Court passed their final verdict in the case of Anders Behring Breivik, declaring Breivik criminally sane and legally responsible for the killing of 77 people during the bombing of government buildings in Oslo and the shooting spree on the island of Utøya on 22 July 2011. This Research Paper examines to what extent the Breivik trial attained the goals of criminal justice: retribution, prevention, restoring democratic order and upholding the rule of law. Furthermore, it aims to determine if the trial contributed to the need for closure in society. The Research Paper concludes that the trial did indeed have a positive impact on the coping mechanisms in Norwegian society and that most Norwegians viewed the trial as a positive counter-weight to the brutality of Breivik’s acts. Overall, the trial was viewed as an example of justice and as a trial that upheld the democratic values of Norwegian society – in stark contrast to Breivik’s values.

  12. Remembering Emil von Behring: from Tetanus Treatment to Antibody Cooperation with Phagocytes

    Directory of Open Access Journals (Sweden)

    Stefan H. E. Kaufmann

    2017-02-01

    Full Text Available A century ago, Emil von Behring passed away. He was the first to be honored by the Nobel Prize for Medicine in 1901 for the successful therapy of diphtheria and tetanus, which he had developed from the bench to the bed. He also contributed to the foundation of immunology, since his therapy was based on passive immunization with specific antisera. Being an ambitious character, he did not shy away from friction with his colleagues Paul Ehrlich and Elias Metchnikoff and his mentor, Robert Koch. Behring was not only an excellent translational researcher but also a successful entrepreneur and early proponent of public-private partnerships.

  13. Cysteine proteinases and cystatins

    Directory of Open Access Journals (Sweden)

    Adeliana S. Oliveira

    2003-01-01

    Full Text Available This review describeds the definition, localization, functions and examples of cysteine proteinases and their protein inhibitors in vertebrate, non-vertebrate animals and plants. These inhibitors are related with defense mechanisms of plant against pests. It also describes the factors involved in the specific cysteine proteinase-cystatin interaction and high degree of affinity and large specificity in this interaction which are not only represented by the compatibility between amino acid residues of the active site involved in catalysis, but also of all amino acid residues that participante in the enzyme-inhibitor interaction.Nesta revisão foram descritas definições, localizações, funções e exemplos de proteinases cisteínicas e suas proteinas inibidoras em animais vertebrados e invertebrados e plantas. Tratamos principalmente com aqueles inibidores que são relatados com o mecanismo de defesa da planta contra pestes. Em adição, comentamos sobre recentes trabalhos que contribuíram para uma melhor compreenção dos fatores envolvidos na interação específica proteinase cisteínica-cistatina. Por outro lado, chamamos atenção para o alto grau de afinidade e grande especificidade na interação que não são apenas representadas pela compatibilidade entre os residuos de aminoácidos do sítio ativo envolvidos na catalise, mas também de todos os resíduos de aminoácidos que participam da interação enzima-inibidor.

  14. Activation of proteinase 3 contributes to Non-alcoholic Fatty Liver Disease (NAFLD) and insulin resistance.

    Science.gov (United States)

    Toonen, Erik J M; Mirea, Andreea-Manuela; Tack, Cees J; Stienstra, Rinke; Ballak, Dov B; van Diepen, Janna A; Hijmans, Anneke; Chavakis, Triantafyllos; Dokter, Wim H; Pham, Christine T N; Netea, Mihai G; Dinarello, Charles A; Joosten, Leo A B

    2016-05-24

    Activation of inflammatory pathways is known to accompany development of obesity-induced non-alcoholic fatty liver disease (NAFLD), insulin resistance and type 2 diabetes. In addition to caspase-1, the neutrophil serine proteases proteinase 3, neutrophil elastase and cathepsin G are able to process the inactive pro-inflammatory mediators IL-1β and IL-18 to their bioactive forms, thereby regulating inflammatory responses. In the present study, we investigated whether proteinase 3 is involved in obesity-induced development of insulin resistance and NAFLD. We investigated the development of NAFLD and insulin resistance in mice deficient for neutrophil elastase/proteinase 3 and neutrophil elastase/cathepsin G and in wild-type mice treated with the neutrophil serine proteinase inhibitor human alpha-1 antitrypsin. Expression profiling of metabolically relevant tissues obtained from insulin resistant mice showed that expression of proteinase 3 was specifically upregulated in the liver, whereas neutrophil elastase, cathepsin G and caspase-1 were not. Neutrophil elastase/proteinase 3 deficient mice showed strongly reduced levels of lipids in the liver after fed a high fat diet. Moreover, these mice were resistant to high fat diet-induced weight gain, inflammation and insulin resistance. Injection of proteinase 3 exacerbated insulin resistance in caspase-1(-/-) mice, indicating that proteinase 3 acts independently of caspase-1. Treatment with alpha-1 antitrypsin during the last 10 days of a 16 week high fat diet reduced hepatic lipid content and decreased fasting glucose levels. We conclude that proteinase 3 is involved in NAFLD and insulin resistance and that inhibition of proteinase 3 may have therapeutic potential.

  15. Effects of herbicides on Behr's metalmark butterfly, a surrogate species for the endangered butterfly, Lange's metalmark

    International Nuclear Information System (INIS)

    Stark, John D.; Chen Xuedong; Johnson, Catherine S.

    2012-01-01

    Lange's metalmark butterfly, Apodemia mormo langei Comstock, is in danger of extinction due to loss of habitat caused by invasive exotic plants which are eliminating its food, naked stem buckwheat. Herbicides are being used to remove invasive weeds from the dunes; however, little is known about the potential effects of herbicides on butterflies. To address this concern we evaluated potential toxic effects of three herbicides on Behr's metalmark, a close relative of Lange's metalmark. First instars were exposed to recommended field rates of triclopyr, sethoxydim, and imazapyr. Life history parameters were recorded after exposure. These herbicides reduced the number of adults that emerged from pupation (24–36%). Each herbicide has a different mode of action. Therefore, we speculate that effects are due to inert ingredients or indirect effects on food plant quality. If these herbicides act the same in A. mormo langei, they may contribute to the decline of this species. - Highlights: ► We evaluated the effects of three herbicides on the butterfly, Behr's metalmark. ► These herbicides are used to control invasive weeds in butterfly habitat. ► The herbicides reduced adult butterfly emergence. - Herbicides are used to remove invasive weeds from butterfly habitat. Certain herbicides may be having a negative effect on butterflies.

  16. [On the Awarding of the First Nobel Prize for Physiology or Medicine to Emil von Behring].

    Science.gov (United States)

    Hansson, Nils; Enke, Ulrike

    2015-12-01

    In his will of 1895, the Swedish inventor Alfred Nobel laid the foundation for prizes in physics, chemistry, physiology or medicine, literature, and peace to those who had "conferred the greatest benefit on mankind" during the last year. The Nobel Prize is today widely considered as the most prestigious international symbol of scientific excellence, but it still is an exciting research question how it gained such prestige. Drawing on files from the Emil von Behring Archive in Marburg, Germany, and the Archive of the Nobel Assembly for Physiology or Medicine in Stockholm this essay aims at shedding light on why the first Nobel Prize for Physiology or Medicine in 1901 was awarded the German immunologist Emil von Behring, and how this decision was viewed at that time. This study is part of a research project that explores mechanisms leading to scientific recognition by using the example of the Nobel Prize for Physiology or Medicine. © Georg Thieme Verlag KG Stuttgart · New York.

  17. Serum proteinase inhibitors and other serum proteins in protein-energy malnutrition

    NARCIS (Netherlands)

    Schelp, F.P.; Migasena, P.; Pongpaew, P.; SCHREURS W.H.P

    1977-01-01

    1. The concentrations of serum protein albumin, prealbumin and transferrin were determined in twenty-eight cases of protein-energy malnutrition (PEM) with infection, together with the levels of serum proteinase inhibitors (PI), alpha1-antitrypsin (AT), alpha1-antichymotrypsin (Ach),

  18. Comparison of concentrations of two proteinase inhibitors, porcine pancreatic elastase inhibitory capacity, and cell profiles in sequential bronchoalveolar lavage samples.

    Science.gov (United States)

    Morrison, H M; Kramps, J A; Dijkman, J H; Stockley, R A

    1986-01-01

    Bronchoalveolar lavage is used to obtain cells and proteins from the lower respiratory tract for diagnosis and research. Uncertainity exists about which site in the lung is sampled by the lavage fluid and what effect different lavage volumes have on recovery of the constituents of lavage fluid. Dilution of alveolar lining fluid by lavage fluid is variable and results are usually expressed as protein ratios to surmount this problem. We have compared cell profiles and the concentrations of two proteinase inhibitors--the low molecular weight bronchial protease inhibitor antileucoprotease and alpha 1 proteinase inhibitor, together with alpha 1 proteinase inhibitor function and its relationship to the cell profile in sequential bronchoalveolar lavage fluid samples from patients undergoing bronchoscopy. There was no difference in total or differential cell counts or albumin or alpha 1 proteinase inhibitor concentrations between the first and second halves of the lavage. Both the concentration of antileucoprotease and the ratio of antileucoprotease to albumin were, however, lower in the second half of the lavage (2p less than 0.01 and 2p less than 0.05 respectively). There was no difference in the function of alpha 1 proteinase inhibitor (assessed by inhibition of porcine pancreatic elastase--PPE) between aliquots (0.28 mole PPE inhibited/mol alpha 1 proteinase inhibitor; range 0-1.19 for the first half and 0.37 mol PPE inhibited/mol alpha 1 proteinase inhibitor; range 0.10-0.80 for the second half). About 60-70% of alpha 1 proteinase inhibitor in each half of the lavage fluid was inactive as an inhibitor. The function of alpha 1 proteinase inhibitor did not differ between bronchitic smokers and ex-smokers. Alpha 1 proteinase inhibitor function was not related to the number of total white cells, macrophages, or neutrophils in the lavage fluid. Contamination of lavage by red blood cells was found to alter the concentration of alpha 1 proteinase inhibitor but not its

  19. What Have We Learned about Lone Wolves from Anders Behring Breivik?

    Directory of Open Access Journals (Sweden)

    Raffaello Pantucci

    2011-12-01

    Full Text Available Anders Behring Breivik’s massacre on July 22, 2011 showed the danger that a well-organized Lone Wolf could cause. The methodical and calculated way with which he prepared and justified his act awoke security services the world over as to the potential menace that this form of terrorism can pose. As they revise their strategies, this article casts a preliminary eye on the case using a particular Lone Wolf prism of analysis to try to see what lessons can be learned from the case. Drawing on Breivik’s own writing and public sources, the article analyses his biography, the ideology he used to justify his act, the degree to which he seems to have been connected to others, his effectiveness, what role the Internet played and his mental competence all to try to draw some early lessons from the case. In concluding it offers some possible lessons learned that might offer practitioners some ideas of how to counter this sort of a threat in the future.

  20. [Ulysses retrotransposon aspartate proteinase (Drosophila virilis)].

    Science.gov (United States)

    Volkov, D A; Savvateeva, L V; Dergousova, N I; Rumsh, L D

    2002-01-01

    Retrotransposones are mobile genetic elements occurring in genomes of bacteria, plants or animals. Retrotransposones were found to contain nucleotide sequences encoding proteins which are homological to retroviral aspartic proteinases. Our research has been focused on Ulysses which is mobile genetic element found in Drosophila virilis. We suggested a primary structure of Ulysses proteinase using comparative analysis of amino acid sequences of retroviral proteinases and proteinases from retrotransposones. The appropriate cDNA fragment has been cloned and expressed in E. coli. The purification of recombinant protein (12 kD) has been carried out by affinity chromatography using pepstatine-agarose. The obtained protein has proteolytic activity at optimum pH 5.5 like the majority of aspartic proteinases.

  1. The aspartic proteinase from Saccharomyces cerevisiae folds its own inhibitor into a helix

    DEFF Research Database (Denmark)

    Li, M; Phylip, L H; Lees, W E

    2000-01-01

    , upon contact with the enzyme, residues 2-32 become ordered and adopt a near-perfect alpha-helical conformation. Thus, the proteinase acts as a folding template, stabilizing the helical conformation in the inhibitor, which results in the potent and specific blockage of the proteolytic activity........2 and 1.8 A, respectively, for complexes of proteinase A with full-length IA3 and with a truncated form consisting only of residues 2-34, reveal an unprecedented mode of inhibitor-enzyme interactions. Neither form of the free inhibitor has detectable intrinsic secondary structure in solution. However...

  2. The aspartic proteinase from Saccharomyces cerevisiae folds its own inhibitor into a helix

    DEFF Research Database (Denmark)

    Li, M; Phylip, L H; Lees, W E

    2000-01-01

    .2 and 1.8 A, respectively, for complexes of proteinase A with full-length IA3 and with a truncated form consisting only of residues 2-34, reveal an unprecedented mode of inhibitor-enzyme interactions. Neither form of the free inhibitor has detectable intrinsic secondary structure in solution. However......, upon contact with the enzyme, residues 2-32 become ordered and adopt a near-perfect alpha-helical conformation. Thus, the proteinase acts as a folding template, stabilizing the helical conformation in the inhibitor, which results in the potent and specific blockage of the proteolytic activity.......Aspartic proteinase A from yeast is specifically and potently inhibited by a small protein called IA3 from Saccharomyces cerevisiae. Although this inhibitor consists of 68 residues, we show that the inhibitory activity resides within the N-terminal half of the molecule. Structures solved at 2...

  3. Effect of Recombinant alpha1-Antitrypsin Fc-Fused (AAT-Fc)Protein on the Inhibition of Inflammatory Cytokine Production and Streptozotocin-Induced Diabetes

    NARCIS (Netherlands)

    Lee, S.; Lee, Y.; Hong, K.; Hong, J.; Bae, S.; Choi, J.; Jhun, H.; Kwak, A.; Kim, E.; Jo, S.; Dinarello, C.A.; Kim, S.

    2013-01-01

    alpha1-Antitrypsin (AAT) is a member of the serine proteinase inhibitor family that impedes the enzymatic activity of serine proteinases, including human neutrophil elastase, cathepsin G and neutrophil proteinase 3. Here, we expressed recombinant AAT by fusing the intact AAT gene to the constant

  4. Serine proteinases and their inhibitors in fertilization

    Czech Academy of Sciences Publication Activity Database

    Jonáková, Věra; Jelínková-Slavíčková, Petra

    2004-01-01

    Roč. 8, 3,4 (2004), s. 108-110 ISSN 1211-8869. [Central European Conference on Human Tumor Markers /5./. Praha, 01.10.2004-03.10.2004] R&D Projects: GA ČR GA303/02/0433; GA ČR GP303/02/P069; GA ČR GP303/04/P070; GA MZd NJ7463 Institutional research plan: CEZ:AV0Z5052915 Keywords : serine proteinase * proteinase inhibitors * fertilization Subject RIV: CE - Biochemistry

  5. Autoactivation of proteinase A initiates activation of yeast vacuolar zymogens

    DEFF Research Database (Denmark)

    van den Hazel, H B; Kielland-Brandt, Morten; Winther, Jakob R.

    1992-01-01

    The Saccharomyces cerevisiae PEP4 gene encodes proteinase A, an aspartyl protease. pep4 mutants are defective in the activation of many vacuolar hydrolases, including proteinase B. We have expressed a pep4 mutation which directs the accumulation of pro-proteinase A with a defective active site. C...

  6. Proteinase genes of cheese starter cultures

    NARCIS (Netherlands)

    Kok, Jan

    The proteolytic enzymes of lactococci are of eminent importance for milk fermentations. By the combined action of proteinases and peptidases milk protein is degraded to peptides and amino acids which are required for cell growth and contribute to the organoleptic properties of the foods. The

  7. The Trolls Disappear in the Light: Swedish Experiences of Mediated Sexualised Hate Speech in the Aftermath of Behring Breivik

    Directory of Open Access Journals (Sweden)

    Maria Edstrom

    2016-06-01

    Full Text Available Feminist journalists have come to expect special resistance, and even threats, from men’s groups as part of their work as journalists. However, the biggest threats might not originate in men’s groups’ activities. A big threat currently comes from Internet trolls’ responses to individuals who engage in hate-provoked and hate-provoking attacks on women as women. This is exemplified in the case of Anders Behring Breivik, who blew up government buildings in Oslo in 2011 and murdered youth from the Labour Party at Utøya as part of his explicitly articulated xenophobic and misogynist campaign against the Islamification of Norway. His ideas are still being shared in social media responses to this tragedy across Nordic countries. This paper argues that this demonstrates that the harms to women and to society go well beyond the individual victims of an identifiable incident. Largely because of their role in condemning and rejecting the hateful ideas advanced across social media forums, troll responses to the Breivik tragedy constitute a particular threat to female and especially feminist journalists.

  8. On the Path of Election and Martyrdom: Some Psychic Mechanisms Involved in the Anders Behring Breivik's Determination as a Terrorist.

    Science.gov (United States)

    Cotti, Patricia

    2015-08-01

    On 22 July 2011, the Norwegian Anders Behring Breivik carried out two attacks in Oslo that cost the lives of 77 people, injured many others, and plunged the entire Norwegian nation into mourning. When he was arrested, Breivik presented himself as a member of the Knights Templar, whose mission is to defend the Christian Western world. He considers that he has sacrificed himself by his actions for his people and says that he has prepared himself for martyrdom. In analysing Breivik's words and writings, this article attempts to identify the thought mechanisms involved in Breivik's idea of election (megalomania) and martyrology. It highlights the importance of a mechanism of "return to the sender," whereby Breivik returns the reproaches directed at him by an agency of judgment (ego ideal or superegoic object). It emphasizes the existence of a "burning desire" and yearning (Sehnsucht) for this same persecuting superegoic object, an object that Breivik constantly wants to find again, even if in death. Taking into consideration Searles's hypothesis that the sense of being persecuted is a defence against the impossibility of mourning, and also H. Blum's hypothesis that persecutory feelings are indicative of fears of a "regressive loss of object constancy," the different psychic mechanisms and modes of functioning underlying Breivik's terrorist determination are related here to what we know about his affective development and infantile relationships.

  9. Substrate and inhibitor studies on proteinase 3

    NARCIS (Netherlands)

    Kam, C. M.; Kerrigan, J. E.; Dolman, K. M.; Goldschmeding, R.; von dem Borne, A. E.; Powers, J. C.

    1992-01-01

    Various amino acid and peptide thioesters were tested as substrates for human proteinase 3 and the best substrate is Boc-Ala-Ala-Nva-SBzl with a kcat/Km value of 1.0 x 10(6) M-1.s-1. Boc-Ala-Ala-AA-SBzl (AA = Val, Ala, or Met) are also good substrates with kcat/Km values of (1-4) x 10(5) M-1.s-1.

  10. Coffee cysteine proteinases and related inhibitors with high expression during grain maturation and germination

    OpenAIRE

    Lepelley Maud; Amor Mohamed; Martineau Nelly; Cheminade Gerald; Caillet Victoria; McCarthy James

    2012-01-01

    Abstract Background Cysteine proteinases perform multiple functions in seeds, including participation in remodelling polypeptides and recycling amino acids during maturation and germination. Currently, few details exist concerning these genes and proteins in coffee. Furthermore, there is limited information on the cysteine proteinase inhibitors which influence the activities of these proteinases. Results Two cysteine proteinase (CP) and four cysteine proteinase inhibitor (CPI) gene sequences ...

  11. The main proteinases in Dermatobia hominis second and third instars larvae are serine-proteinases.

    Science.gov (United States)

    Pires, F A; Moya-Borja, G E; Barreira, J D; Pinho, R T; Alves, C R

    2007-04-30

    We performed a combination of proteinase assay, either in solution or immobilized in sodium dodecyl sulfate-polyacrylamide gel copolymerized with gelatin, to detect and quantify proteinases of Dermatobia hominis second (L2) and third (L3) instar larvae. In the quantitative assay, we examined proteinase activity by hydrolysis of a panel of peptide bonds specific for the main proteinase classes. We verified that the pGlu-Phe-Leu p-nitroanilide substrate was hydrolyzed by crude extracts of L2 (3.0+/-0.2 nmol h(-1)mg of protein(-1)) and L3 (7.7+/-0.1 nmol h(-1)mg of protein(-1)) and that both activities were partially inhibited by trans-epoxysuccinyl-l-leucylamido-(4-guanidino)butane, 15% and 3%, respectively. Also, we demonstrated that the Nalpha-p-Tosyl-l-Arg methyl ester substrate was hydrolyzed by crude extracts of L2 (117+/-24 nmol h(-1)mg of protein(-1)) and L3 (111+/-10 nmol h(-1)mg of protein(-1)), suggesting a predominance of esterase activity in the crude larval preparation. Interestingly, the specific activity of serine-proteinases was totally inhibited by phenylmethylsulphonyl fluoride in the L3 crude extract, while only 10% of this enzyme class activity was inhibited in the L2 crude extract. The results of the qualitative assays with substrate gels suggested that L2 and L3 larvae express serine-proteinases with similar (13 and 22 kDa) and distinct (50 kDa in L2 and 30 kDa in L3) relative molecular masses. These findings contribute to the biochemical characterization of D. hominis L2 and L3 larvae.

  12. A cytotoxic serine proteinase isolated from mouse submandibular gland.

    Science.gov (United States)

    Shimamura, T; Nagumo, N; Ikigai, H; Murakami, K; Okubo, S; Toda, M; Ohnishi, R; Tomita, M

    1989-08-01

    We have isolated a novel cytotoxic factor from the submandibular glands of male BALB/c mice by Sephadex G-50 gel filtration chromatography and reverse-phase HPLC. The cytotoxic factor is a serine proteinase, which belongs to the mouse glandular kallikrein (mGK) family, with an Mr of approximately 27,000. The purified serine proteinase showed cytotoxic activity against mouse thymocytes in a dose-dependent manner, and a serine proteinase inhibitor, diisopropyl fluorophosphate, blocked its cytotoxic activity.

  13. Multiple pathways for vacuolar sorting of yeast proteinase A

    DEFF Research Database (Denmark)

    Westphal, V; Marcusson, E G; Winther, Jakob R.

    1996-01-01

    The sorting of the yeast proteases proteinase A and carboxypeptidase Y to the vacuole is a saturable, receptor-mediated process. Information sufficient for vacuolar sorting of the normally secreted protein invertase has in fusion constructs previously been found to reside in the propeptide...... of proteinase A. We found that sorting of such a hybrid protein is dependent on the vacuolar protein-sorting receptor Vps10p. This was unexpected, as strains disrupted for VPS10 sort more than 85% of the proteinase A to the vacuole. Consistent with a role for Vps10p in sorting of proteinase A, we found that 1...

  14. Autoactivation of proteinase A initiates activation of yeast vacuolar zymogens

    DEFF Research Database (Denmark)

    van den Hazel, H B; Kielland-Brandt, Morten; Winther, Jakob R.

    1992-01-01

    The Saccharomyces cerevisiae PEP4 gene encodes proteinase A, an aspartyl protease. pep4 mutants are defective in the activation of many vacuolar hydrolases, including proteinase B. We have expressed a pep4 mutation which directs the accumulation of pro-proteinase A with a defective active site. Co......-expression with PEP4 leads to normal processing, i.e. the mutant zymogen is functional as a substrate for the maturation reaction in trans. We conclude that wild-type pro-proteinase A has the ability to mediate its own activation. Elimination of the co-expressed PEP4 gene did not effectively stop the processing...

  15. New aspartic proteinase of Ulysses retrotransposon from Drosophila virilis.

    Science.gov (United States)

    Volkov, D A; Dergousova, N I; Rumsh, L D

    2004-06-01

    This work is focused on the investigation of a proteinase of Ulysses mobile genetic element from Drosophila virilis. The primary structure of this proteinase is suggested based on comparative analysis of amino acid sequences of aspartic proteinases from retroviruses and retrotransposons. The corresponding cDNA fragment has been cloned and expressed in E. coli. The protein accumulated in inclusion bodies. The recombinant protein (12 kD) was subjected to refolding and purified by affinity chromatography on pepstatin-agarose. Proteolytic activity of the protein was determined using oligopeptide substrates melittin and insulin B-chain. It was found that the maximum of the proteolytic activity is displayed at pH 5.5 as for the majority of aspartic proteinases. We observed that hydrolysis of B-chain of insulin was totally inhibited by pepstatin A in the micromolar concentration range. The molecular weight of the monomer of the Ulysses proteinase was determined by MALDI-TOF mass-spectrometry.

  16. Cleavage by trypsin and by the proteinase from Armillaria mellea at epsilon-N-formyl-lysine residues.

    Science.gov (United States)

    Barry, F P; Doonan, S; Ross, C A

    1981-01-01

    Kinetic studies were made of the hydrolysis by trypsin of alpha-N-acetylglycyl-L-lysine methyl ester and of its neutral analogue alpha-N-acetylglycyl-epsilon-N-formyl-L-lysine methyl ester. The latter substance is a moderately good substrate for trypsin, and this observation is discussed in terms of the substrate specifically of the enzyme. The actions of trypsin and of the lysine-specific proteinase from Armillaria mellea on both a native and a formylated polypeptide substrate were compared. Both enzymes were found to hydrolyse specifically bonds to epsilon-N-formyl-lysine in the formylated substrate. PMID:6796050

  17. Fractionation and purification of the thiol proteinases from papaya latex.

    Science.gov (United States)

    Dekeyser, P M; De Smedt, S; Demeester, J; Lauwers, A

    1994-06-03

    Three cysteine proteinases, i.e. chymopapain, papaya proteinase IV and proteinase III, were purified to homogeneity from papaya latex using a combination of ion-exchange chromatography and hydrophobic interaction chromatography. During the purification procedure, the thiol-groups of the active center were reversibly blocked as mixed disulfides with 2-thiopyridone. Homogeneity was proved electrophoretically by native polyacrylamide gel electrophoresis (PAGE), sodium dodecyl sulfate (SDS)-PAGE and rechromatography on a Mono S 5/5 column at pH 5.0.

  18. A zymography analysis of proteinase activity present in Leptospira.

    Science.gov (United States)

    Madathiparambil, Madanan G; Cattavarayane, Sandhanakrishnan; Manickam, Gayathri D; Singh, Kavita; Perumana, Sudhakaran R; Sehgal, Subhash C

    2011-03-01

    Leptospirosis is a major public health problem caused by spirochete Leptospira which is an extracellular pathogen. During infection and invasion, the bacteria cross the physical barriers and later it encounter with the host defence mechanism. These processes may involve proteolytic degradation of the host tissue biomatrix. In an effort to understand the production and nature of Leptospiral proteinases, investigations were carried out using zymograpic methods. The results showed that the leptospires degrades different kind of protein substances such as gelatin, casein, and albumin. Gelatin zymography reveals that different serovars contain multiple gelatinases in the molecular weight range from 240 to 32 kDa. Studies using inhibitors suggested that the Leptospiral proteinases include metalloproteinases, serine or cysteine proteinases. The temperature sensitivity suggests that some of these proteinases are stable even at high temperatures. The presence of multiple gelatinases in Leptospira serovars suggests a critical role for these enzymes in Leptospiral invasion and pathogenesis.

  19. Detection and Characterization of Bacterial Proteinases Using Zymography.

    Science.gov (United States)

    Madanan, Madathiparambil G; Mechoor, Ambili

    2017-01-01

    Proteinases play a crucial role in invasion and pathogenesis of bacteria, especially the extracellular and membrane-bound forms. Analysis of these proteinases demands the isolation by retaining the enzymatic activity. The isolation procedures maintaining the native structure of the enzyme in its soluble form are also of extreme importance. The qualitative analyses of these proteinases are carried out by electrophoresis and zymography. Enzymatic characterization based on the effect of inhibitors and activators on gelatinase activity also can be assessed using this zymography. The membrane-bound proteinases can be isolated in their native and soluble form, still retaining the activity using 6-aminocaproic acid and sodium deoxycholate; the procedure of which is explained in this chapter.

  20. Candida albicans Secreted Aspartyl Proteinases in Virulence and Pathogenesis

    OpenAIRE

    Naglik, Julian R.; Challacombe, Stephen J.; Hube, Bernhard

    2003-01-01

    Candida albicans is the most common fungal pathogen of humans and has developed an extensive repertoire of putative virulence mechanisms that allows successful colonization and infection of the host under suitable predisposing conditions. Extracellular proteolytic activity plays a central role in Candida pathogenicity and is produced by a family of 10 secreted aspartyl proteinases (Sap proteins). Although the consequences of proteinase secretion during human infections is not precisely known,...

  1. Action of plant proteinase inhibitors on enzymes of physiopathological importance

    Directory of Open Access Journals (Sweden)

    Maria Luiza V. Oliva

    2009-09-01

    Full Text Available Obtained from leguminous seeds, various plant proteins inhibit animal proteinases, including human, and can be considered for the development of compounds with biological activity. Inhibitors from the Bowman-Birk and plant Kunitz-type family have been characterized by proteinase specificity, primary structure and reactive site. Our group mostly studies the genus Bauhinia, mainly the species bauhinioides, rufa, ungulata and variegata. In some species, more than one inhibitor was characterized, exhibiting different properties. Although proteins from this group share high structural similarity, they present differences in proteinase inhibition, explored in studies using diverse biological models.Obtidas de sementes leguminosas, várias proteínas inibem proteinases de origem animal, incluindo humanas, e podem ser consideradas para o desenvolvimento de compostos com atividade biológica. Inibidores da família Bowman-Birk e da família Kunitz vegetal tem sido caracterizados em relação a especificidade para proteinase, estrutura primária e sitio reativo. O nosso grupo majoritariamente vem estudando o gênero Bauhinia, principalmente as espécies bauhinioides, rufa, ungulatae variegata. Em algumas espécies, mais de um inibidor com propriedades diferentes foi caracterizado. Embora tais proteínas apresentem alta similaridade estrutural, diferem quanto à inibição de proteinases, e foram exploradas em estudos utilizando diversos modelos biológicos.

  2. The induction of proteinases in corn and soybean by anoxia

    International Nuclear Information System (INIS)

    VanToai, T.; Hwang, Shihying

    1989-01-01

    This study characterized the anaerobic changes in proteinase activities in corn and soybean roots and to investigate the possibility that these changes might contribute to the differential anaerobiosis tolerance of the two species. After 24 h of anoxia, crude protein extracts from H60 corn and Keller soybean root tips (10cm) were assayed for proteinase activities at pH range from 4.5 to 9.5. Turnover of aberrant proteins was studied in seedlings labelled with 3 H-leucine for 12 h under: (a) puromycin (0.64 mM) in air, (b) ethanol (1%) in air, (c) nitrogen and (d) air. After the treatment, the labelled proteins remaining in roots were determined every 2 h for 6 h. In both corn and soybean, activities of alkali proteinases increased, and activities of acid proteinases declined under anoxia. Neutral proteinases increase in anoxic corn roots, but decline in anoxic soybean roots. The protein turnover rate in corn treated with puromycin, ethanol and nitrogen was much higher than in control roots. The protein turnover rate in soybean roots treated with puromycin, ethanol was similar to the rate of the control. The results indicated that: (a) anoxic corn can degrade aberrant proteins, but anoxic soybean cannot, (b) the degradation of aberrant proteins in anoxic corn is accomplished by neutral proteinases, and (c) the accumulation of aberrant proteins in soybean might contribute to the susceptibility of this species to anoxia

  3. Electrophoretic analysis of plant cysteine and serine proteinases using gelatin-containing polyacrylamide gels and class-specific proteinase inhibitors.

    Science.gov (United States)

    Michaud, D; Faye, L; Yelle, S

    1993-01-01

    Inclusion of gelatin in polyacrylamide gels provides a sensitive way of detecting multiple proteolytic activities in crude extracts from any source. The present study describes a method allowing discrimination between cysteine and serine proteinases in plant extracts, using gelatin-containing gels in combination with class-specific proteinase inhibitors. Preincubation of extracts with 4 mM phenylmethylsulfonyl fluoride, a serine proteinase inhibitor, or with 25 microM L-trans-epoxysuccinyl-L-leucylamido(4-guanidino) butane, a cysteine proteinase inhibitor, allowed the identification of enzymes from both classes in extracts of tomato fruit and papaya latex. The efficiency of the two low molecular weight inhibitors used was very high, and the irreversibility of the inhibiting effect was maintained during electrophoresis conducted in the presence of sodium dodecyl sulfate. The analytic procedure described here, with a detection threshold of less than 100 pg enzyme, is the first that allows quick and accurate discrimination of plant cysteine and serine proteinases separated in electrophoretic gels. This simple and rapid technique could be of interest for studying the evolution of class-specific proteinases in plant extracts during various developmental, physiological, and pathogenic processes. It is also potentially applicable to the majority of eucaryotic and procaryotic systems.

  4. Characterization of proteinases from the midgut of Rhipicephalus (Boophilus microplus involved in the generation of antimicrobial peptides

    Directory of Open Access Journals (Sweden)

    Craik Charles S

    2010-07-01

    Full Text Available Abstract Background Hemoglobin is a rich source of biologically active peptides, some of which are potent antimicrobials (hemocidins. A few hemocidins have been purified from the midgut contents of ticks. Nonetheless, how antimicrobials are generated in the tick midgut and their role in immunity is still poorly understood. Here we report, for the first time, the contribution of two midgut proteinases to the generation of hemocidins. Results An aspartic proteinase, designated BmAP, was isolated from the midgut of Rhipicephalus (Boophilus microplus using three chromatographic steps. Reverse transcription-quantitative polymerase chain reaction revealed that BmAP is restricted to the midgut. The other enzyme is a previously characterized midgut cathepsin L-like cysteine proteinase designated BmCL1. Substrate specificities of native BmAP and recombinant BmCL1 were mapped using a synthetic combinatorial peptide library and bovine hemoglobin. BmCL1 preferred substrates containing non-polar residues at P2 subsite and polar residues at P1, whereas BmAP hydrolysed substrates containing non-polar amino acids at P1 and P1'. Conclusions BmAP and BmCL1 generate hemocidins from hemoglobin alpha and beta chains in vitro. We postulate that hemocidins may be important for the control of tick pathogens and midgut flora.

  5. Candida albicans secreted aspartyl proteinases in virulence and pathogenesis.

    Science.gov (United States)

    Naglik, Julian R; Challacombe, Stephen J; Hube, Bernhard

    2003-09-01

    Candida albicans is the most common fungal pathogen of humans and has developed an extensive repertoire of putative virulence mechanisms that allows successful colonization and infection of the host under suitable predisposing conditions. Extracellular proteolytic activity plays a central role in Candida pathogenicity and is produced by a family of 10 secreted aspartyl proteinases (Sap proteins). Although the consequences of proteinase secretion during human infections is not precisely known, in vitro, animal, and human studies have implicated the proteinases in C. albicans virulence in one of the following seven ways: (i) correlation between Sap production in vitro and Candida virulence, (ii) degradation of human proteins and structural analysis in determining Sap substrate specificity, (iii) association of Sap production with other virulence processes of C. albicans, (iv) Sap protein production and Sap immune responses in animal and human infections, (v) SAP gene expression during Candida infections, (vi) modulation of C. albicans virulence by aspartyl proteinase inhibitors, and (vii) the use of SAP-disrupted mutants to analyze C. albicans virulence. Sap proteins fulfill a number of specialized functions during the infective process, which include the simple role of digesting molecules for nutrient acquisition, digesting or distorting host cell membranes to facilitate adhesion and tissue invasion, and digesting cells and molecules of the host immune system to avoid or resist antimicrobial attack by the host. We have critically discussed the data relevant to each of these seven criteria, with specific emphasis on how this proteinase family could contribute to Candida virulence and pathogenesis.

  6. The proteolytic activities of chymopapain, papain, and papaya proteinase III.

    Science.gov (United States)

    Zucker, S; Buttle, D J; Nicklin, M J; Barrett, A J

    1985-04-05

    The three proteinases present in papaya latex: papain (EC 3.4.22.2) chymopapain and papaya proteinase III (EC 3.4.22.6), were standardized by active-site titration, and compared in proteolytic activity against azocasein, serum albumin and cartilage proteoglycan. The activities were all of the same order, although there were differences in pH dependence. SDS-polyacrylamide gel electrophoresis of the early products of digestion of albumin and phosphorylase a showed very similar patterns for the three papaya proteinases. Kinetic parameters for hydrolysis of benzyloxycarbonyl-phenylalanyl-arginyl-7(4-methyl)coumarylamide were determined for the three enzymes. Values for kcat/Km varied only within a factor of 2, but the individual constants were much higher for papain than for chymopapain and papaya proteinase III. In contrast to the results obtained with the synthetic substrate, the kinetic parameters for the initial hydrolysis of succinyl-albumin were very similar for the three papaya proteinases. This was consistent with their similar proteolytic activities in other assays.

  7. Alpha1 and Alpha2 Integrins Mediate Invasive Activity of Mouse Mammary Carcinoma Cells through Regulation of Stromelysin-1 Expression

    Energy Technology Data Exchange (ETDEWEB)

    Lochter, Andre; Navre, Marc; Werb, Zena; Bissell, Mina J

    1998-06-29

    Tumor cell invasion relies on cell migration and extracellular matrix proteolysis. We investigated the contribution of different integrins to the invasive activity of mouse mammary carcinoma cells. Antibodies against integrin subunits {alpha}6 and {beta}1, but not against {alpha}1 and {alpha}2, inhibited cell locomotion on a reconstituted basement membrane in two-dimensional cell migration assays, whereas antibodies against {beta}1, but not against a6 or {alpha}2, interfered with cell adhesion to basement membrane constituents. Blocking antibodies against {alpha}1 integrins impaired only cell adhesion to type IV collagen. Antibodies against {alpha}1, {alpha}2, {alpha}6, and {beta}1, but not {alpha}5, integrin subunits reduced invasion of a reconstituted basement membrane. Integrins {alpha}1 and {alpha}2, which contributed only marginally to motility and adhesion, regulated proteinase production. Antibodies against {alpha}1 and {alpha}2, but not {alpha}6 and {beta}1, integrin subunits inhibited both transcription and protein expression of the matrix metalloproteinase stromelysin-1. Inhibition of tumor cell invasion by antibodies against {alpha}1 and {alpha}2 was reversed by addition of recombinant stromelysin-1. In contrast, stromelysin-1 could not rescue invasion inhibited by anti-{alpha}6 antibodies. Our data indicate that {alpha}1 and {alpha}2 integrins confer invasive behavior by regulating stromelysin-1 expression, whereas {alpha}6 integrins regulate cell motility. These results provide new insights into the specific functions of integrins during tumor cell invasion.

  8. Cytotoxic T-Lymphocyte Antigen-2 alpha participates in axial ...

    African Journals Online (AJOL)

    Cytotoxic T-lymphocyte antigen-2 alpha (CTLA-2α) has been discovered and expressed in mouse activated T-cells and mast cells. Structurally, it is homologous to the proregion of mouse cathepsin L, a lysosomal cystein proteinase. Expressed recombinant CTLA-2α is shown to exhibit selective inhibition to cathepsin L and ...

  9. Detection of a papaya cysteine proteinase inhibitor under different environmental conditions

    OpenAIRE

    2012-01-01

    M.Sc. Proteinases are involved in many cellular reactions involving protein degradation, such as degradation of storage proteins and protein degradation during senescence processes. Their action can be inhibited by proteinase inhibitors. Information is still limited about the regulation of these inhibitors in plants and their possible interaction with proteinases under stress conditions. To obtain a better understanding of the physiological role of a proteinase inhibitor in plants under st...

  10. Multiple pathways for vacuolar sorting of yeast proteinase A

    DEFF Research Database (Denmark)

    Westphal, V; Marcusson, E G; Winther, Jakob R.

    1996-01-01

    The sorting of the yeast proteases proteinase A and carboxypeptidase Y to the vacuole is a saturable, receptor-mediated process. Information sufficient for vacuolar sorting of the normally secreted protein invertase has in fusion constructs previously been found to reside in the propeptide......) Vps10p and proteinase A interact directly and specifically as shown by chemical cross-linking. Interestingly, overexpression of two telomere-linked VPS10 homologues, VTH1 and VTH2 suppressed the missorting phenotypes of a deltavps10 strain. However, disruption of the VTH1 and VTH2 genes did not affect...

  11. The interaction of α2-macroglobulin with proteinases. Binding and inhibition of mammalian collagenases and other metal proteinases

    Science.gov (United States)

    Werb, Zena; Burleigh, Mary C.; Barrett, Alan J.; Starkey, Phyllis M.

    1974-01-01

    1. Experiments were performed to determine whether the specific collagenases and other metal proteinases are bound and inhibited by α2-macroglobulin, as are endopeptidases of other classes. 2. A specific collagenase from rabbit synovial cells was inhibited by human serum. The inhibition could be attributed entirely to α2-macroglobulin; α1-trypsin inhibitor was not inhibitory. α2-Macroglobulin presaturated with trypsin or cathepsin B1 did not inhibit collagenase, and pretreatment of α2-macroglobulin with collagenase prevented subsequent reaction with trypsin. The binding of collagenase by α2-macroglobulin was not reversible in gel chromatography. 3. The collagenolytic activity of several rheumatoid synovial fluids was completely inhibited by incubation of the fluids with α2-macroglobulin. 4. The collagenase of human polymorphonuclear-leucocyte granules showed time-dependent inhibition by α2-macroglobulin. 5. The collagenolytic metal proteinase of Crotalus atrox venom was inhibited by α2-macroglobulin. 6. The collagenase of Clostridium histolyticum was bound by α2-macroglobulin, and inhibited more strongly with respect to collagen than with respect to a peptide substrate. 7. Thermolysin, the metal proteinase of Bacillus thermoproteolyticus, was bound and inhibited by α2-macroglobulin. 8. It was shown by polyacrylamidegel electrophoresis of reduced α2-macroglobulin in the presence of sodium dodecyl sulphate that synovial-cell collagenase, clostridial collagenase and thermolysin cleave the quarter subunit of α2-macroglobulin near its mid-point, as do serine proteinases. 9. The results are discussed in relation to previous work, and it is concluded that the characteristics of interaction of the metal proteinases with α2-macroglobulin are the same as those of other proteinases. ImagesPLATE 2PLATE 1 PMID:4374931

  12. The King of Norway: negative individuation, the hero myth and psychopathic narcissism in extreme violence and the life of Anders Behring Breivik.

    Science.gov (United States)

    Virtanen, Harri

    2013-11-01

    The paper discusses negative individuation and the hero myth as developmental concepts. It is suggested that in negative individuation healthy psychological development is hindered and goes astray. Aggression then becomes the central psychic system. Repressed anger is the core element in psychopathic narcissism (Diamond) and malignant narcissism (Kernberg). Both Diamond and Kernberg extend narcissistic personality structure to antisocial, psychopathic personality in an effort to better understand extreme violence. According to Freud, love (libido) and hate (the death drive) are the major motivational systems in the human psyche. In contrast to Freud, Jung sees libido as a life force in general, not simply as a sexual drive. Jung writes about evil and the shadow but does not present a comprehensive theory of the negative development of an individual's life. The concept of negative individuation connects the shadow and the death drive with psychopathology, psychiatry and psychotherapy. In this paper, I explore these concepts in the light of contemporary affect theory according to Kernberg. I also ask how ideology is tied to extreme violence and how it is possible that narcissistic personality structures can lead to such radically different outcomes as were manifested in the lives of Anders Behring Breivik and Steve Jobs. © 2013, The Society of Analytical Psychology.

  13. Comparisons of genetic diversity in captive versus wild populations of the federally endangered Quino checkerspot butterfly (Euphydryas editha quino Behr; Lepidoptera: Nymphalidae)

    Science.gov (United States)

    Miller, Mark P.; Pratt, Gordon F.; Mullins, Thomas D.; Haig, Susan M.

    2014-01-01

    Captive populations can play a significant role in threatened and endangered species management. An important consideration when developing and managing captive populations, however, is the maintenance of genetic diversity to ensure that adequate variation exists to avoid the negative consequences of inbreeding. In this investigation, we compared genetic diversity patterns within captive and wild populations of the federally endangered Quino checkerspot butterfly (Euphydryas editha quino Behr [Lepidoptera: Nymphalidae]), a taxon with a restricted distribution to chaparral and sage shrublands within Riverside and San Diego counties, California. Our analyses revealed that medium to high-frequency alleles from the wild populations were also present in the captive populations. While there was no significant difference in genetic diversity as quantified by expected heterozygosity, the captive populations showed tendencies toward significantly lower allelic richness than their wild counterparts. Given that alleles from the wild populations were occasionally not detected in captive populations, periodic incorporation of new wild specimens into the captive population would help ensure that allelic diversity is maintained to the extent possible. If performed in advance, genetic surveys of wild populations may provide the clearest insights regarding the number of individuals needed in captivity to adequately reflect wild populations.

  14. In vitro evaluation of proteinase, phospholipase and haemolysin ...

    African Journals Online (AJOL)

    Aim: The present study aimed to determine phospholipase, proteinase and haemolysin activities in Candida species isolated from various clinical samples. Material and Method: A total of 110 Candida species isolated from various clinical specimens were identified up to species level by standard mycological techniques ...

  15. Production of a heterologous proteinase A by Saccharomyces kluyveri

    DEFF Research Database (Denmark)

    Møller, Kasper; Tidemand, L.D.; Winther, J.R.

    2001-01-01

    In order to evaluate the potential of Saccharomyces kluyveri for heterologous protein production, S. kluyveri Y159 was transformed with a S. cerevisiae-based multi-copy plasmid containing the S. cerevisiae PEP4 gene, which encodes proteinase A, under the control of its native promoter. As a refer...

  16. The resistance of insects to plant proteinase inhibitors

    NARCIS (Netherlands)

    Jongsma, M.A.

    1995-01-01

    The research reported in this thesis describes the induction of proteinase inhibitor synthesis in solanaceous plants (tobacco and tomato), when lepidopteran larvae (Manduca sexta and Spodoptera exigua) are feeding on leaves. It is shown that the

  17. Human seminal proteinase and prostate-specific antigen are the ...

    Indian Academy of Sciences (India)

    ... natural and synthetic substrates showed that bovine serum albumin was more readily hydrolysed than casein. The results of this study should be useful in purifying and assaying this protein. Based on published studies and the present results, the broad proteolytic specificity of human seminal proteinase suggests a role ...

  18. Purification and characterization of cell-envelope proteinase from ...

    African Journals Online (AJOL)

    user

    2012-10-18

    Oct 18, 2012 ... phenylmethylsulfonyl fluoride;. ACE, angiotensin-I-converting enzyme. Poolman, 1998). Cell-envelope proteinase (CEP) play an important role in the lactobacillus proteolytic system. CEPs are the critical enzyme in the system (Kunji et al., 1996), since it is the only enzyme that can initiate the breakdown of.

  19. Identification, classification and expression pattern analysis of sugarcane cysteine proteinases

    Directory of Open Access Journals (Sweden)

    Gustavo Coelho Correa

    2001-12-01

    Full Text Available Cysteine proteases are peptidyl hydrolyses dependent on a cysteine residue at the active center. The physical and chemical properties of cysteine proteases have been extensively characterized, but their precise biological functions have not yet been completely understood, although it is known that they are involved in a number of events such as protein turnover, cancer, germination, programmed cell death and senescence. Protein sequences from different cysteine proteinases, classified as members of the E.C.3.4.22 sub-sub-class, were used to perform a T-BLAST-n search on the Brazilian Sugarcane Expressed Sequence Tags project (SUCEST data bank. Sequence homology was found with 76 cluster sequences that corresponded to possible cysteine proteinases. The alignments of these SUCEST clusters with the sequence of cysteine proteinases of known origins provided important information about the classification and possible function of these sugarcane enzymes. Inferences about the expression pattern of each gene were made by direct correlation with the SUCEST cDNA libraries from which each cluster was derived. Since no previous reports of sugarcane cysteine proteinases genes exists, this study represents a first step in the study of new biochemical, physiological and biotechnological aspects of sugarcane cysteine proteases.Proteinases cisteínicas são peptidil-hidrolases dependentes de um resíduo de cisteína em seu sítio ativo. As propriedades físico-químicas destas proteinases têm sido amplamente caracterizadas, entretanto suas funções biológicas ainda não foram completamente elucidadas. Elas estão envolvidas em um grande número de eventos, tais como: processamento e degradação protéica, câncer, germinação, morte celular programada e processos de senescência. Diferentes proteinases cisteínicas, classificadas pelo Comitê de Nomenclatura da União Internacional de Bioquímica e Biologia Molecular (IUBMB como pertencentes à sub

  20. Detection of Aspartic Proteinase Activities Using Gel Zymography.

    Science.gov (United States)

    Perera, Handunge Kumudu Irani

    2017-01-01

    Gel zymography is a two-stage process where the proteins from the test sample are first separated by electrophoresis followed by the detection of the activity of hydrolytic enzymes. Many zymography procedures use sodium dodecyl sulfate (SDS) polyacrylamide gels copolymerized with an appropriate substrate. The procedure described here uses native polyacrylamide gel electrophoresis (PAGE) in the absence of both SDS and substrate. In order to visualize aspartic proteinase activity, the gel is impregnated in bovine hemoglobin at pH 3.0 for 15 min after the electrophoresis procedure. Subsequently, the gel is incubated in a humid container in the absence of hemoglobin for 1 h at 37 °C. At the end, the gel is stained with amido black and destained. Clear areas against a dark background corresponding to aspartic proteinase activities can be detected.

  1. Proteinase inhibitors in aggregated forms of boar seminal plasma proteins

    Czech Academy of Sciences Publication Activity Database

    Jelínková, Petra; Maňásková, Pavla; Tichá, M.; Jonáková, Věra

    2003-01-01

    Roč. 32, - (2003), s. 99-107 ISSN 0141-8130 R&D Projects: GA ČR GA303/02/0433; GA ČR GP303/02/P069; GA MZd NJ7463 Institutional research plan: CEZ:AV0Z5052915; CEZ:MSM 113100001 Keywords : proteinase inhibitors, aggregated forms, boar seminal plasma proteins Subject RIV: CE - Biochemistry Impact factor: 1.427, year: 2003

  2. Production of a heterologous proteinase A by Saccharomyces kluyveri

    DEFF Research Database (Denmark)

    Møller, K; Tidemand, L D; Winther, Jakob R.

    2001-01-01

    In order to evaluate the potential of Saccharomyces kluyveri for heterologous protein production, S. kluyveri Y159 was transformed with a S. cerevisiae-based multi-copy plasmid containing the S. cerevisiae PEP4 gene, which encodes proteinase A, under the control of its native promoter......, compared to a yield of 0.40 g/g in S. cerevisiae. Overexpression of PEP4 led to the secretion of active proteinase A in both S. kluyveri and S. cerevisiae. The yield of active proteinase A during growth on glucose was found to be 3.6-fold higher in S. kluyveri than in the S. cerevisiae reference strain........ As a reference, S. cerevisiae CEN.PK 113-5D was transformed with the same plasmid and the two strains were characterised in batch cultivations on glucose. The glucose metabolism was found to be less fermentative in S. kluyveri than in S. cerevisiae. The yield of ethanol on glucose was 0.11 g/g in S. kluyveri...

  3. Mechanism and ion-dependence of in vitro autoactivation of yeast proteinase A

    DEFF Research Database (Denmark)

    Van Den Hazel, H; Wolff, A M; Kielland-Brandt, Morten

    1997-01-01

    Yeast proteinase A is synthesized as a zymogen which transits through the endoplasmic reticulum, the Golgi complex and the endosome to the vacuole. On arrival in the vacuole, activation takes place. It has previously been found that proteinase A can activate autocatalytically; however......, the propeptide of proteinase A shows essentially no similarity to other known aspartic proteinase propeptides. To understand why proteinase A activation occurs rapidly in the vacuole but not at all in earlier compartments, we have purified the zymogen and investigated the conditions that trigger autoactivation...... the pH- and ionic-strength-dependence and the predominance of a product-catalysed mechanism are well adapted to the situation in vivo, since slow activation in the absence of active proteinase A helps to prevent activation in prevacuolar compartments, whereas, on delivery to the vacuole, lower p...

  4. Wound and methyl jasmonate induced pigeon pea defensive proteinase inhibitor has potency to inhibit insect digestive proteinases.

    Science.gov (United States)

    Lomate, Purushottam R; Hivrale, Vandana K

    2012-08-01

    Wounding of plants by chewing insects or other damage induces the synthesis of defensive proteinase inhibitors (PI) in both wounded and distal unwounded leaves. In the present paper we report the characterization of inducible defensive PI from pigeon pea (Cajanus cajan) and its in vitro interaction with Helicoverpa armigera gut proteinases (HGP). We found that PI activity was induced in local as well as systemic leaves of pigeon pea by the wounding and methyl jasmonate (MeJA) application. Consistent induction of PI was observed in two wild cultivars of pigeon pea at various growth stages. The estimated molecular weight of inducible PI was ~16.5 kDa. Electrophoretic analysis and enzyme assays revealed that the induced PI significantly inhibited total gut proteinase as well as trypsin-like activity from the midgut of H. armigera. The induced PI was found to be inhibitor of trypsin as well as chymotrypsin. Study could be important to know the further roles of defensive PIs. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  5. [Lactic acid bacteria proteinase and quality of fermented dairy products--A review].

    Science.gov (United States)

    Zhang, Shuang; Zhang, Lanwei; Han, Xue

    2015-12-04

    Lactic acid bacteria (LAB) could synthesize cell envelope proteinase with weak activity, which primarily degrades casein. In addition to its crucial role in the rapid growth of LAB in milk, LAB proteinases are also of industrial importance due to their contribution to the formation of texture and flavor of many fermented dairy products. The proteolytic system, properties of proteinase, the degradation product of casein and its effect on the quality of fermented dairy products were reviewed in this manuscript.

  6. Redox regulation of peroxiredoxin and proteinases by ascorbate and thiols during pea root nodule senescence.

    Science.gov (United States)

    Groten, Karin; Dutilleul, Christelle; van Heerden, Philippus D R; Vanacker, Hélène; Bernard, Stéphanie; Finkemeier, Iris; Dietz, Karl-Josef; Foyer, Christine H

    2006-02-20

    Redox factors contributing to nodule senescence were studied in pea. The abundance of the nodule cytosolic peroxiredoxin but not the mitochondrial peroxiredoxin protein was modulated by ascorbate. In contrast to redox-active antioxidants such as ascorbate and cytosolic peroxiredoxin that decreased during nodule development, maximal extractable nodule proteinase activity increased progressively as the nodules aged. Cathepsin-like activities were constant throughout development but serine and cysteine proteinase activities increased during senescence. Senescence-induced cysteine proteinase activity was inhibited by cysteine, dithiotreitol, or E-64. Senescence-dependent decreases in redox-active factors, particularly ascorbate and peroxiredoxin favour decreased redox-mediated inactivation of cysteine proteinases.

  7. The role of proteinase enzymes in the process of conversion of muscle to meat

    Directory of Open Access Journals (Sweden)

    Dümen Emek

    2006-01-01

    Full Text Available Post mortem meat tenderization is a complex mechanism and unfortunately it has not been fully identified scientifically. It is known that endogenous proteinases have an important role in this mechanism. Detailed studies are being performed about the destructive effects of lysosomal proteinases and calcium dependent proteinases on the myofibrils and these are most common topics that are being investigated about meat tenderization processes by the scientists. The aim of this paper is to review the role of proteinase enzymes in the process of conversion of muscle to meat. .

  8. Ovostatin: a novel proteinase inhibitor from chicken egg white. II. Mechanism of inhibition studied with collagenase and thermolysin.

    Science.gov (United States)

    Nagase, H; Harris, E D

    1983-06-25

    The inhibition mechanism of ovostatin was studied using rabbit synovial collagenase and thermolysin. When enzymes were complexed with ovostatin, only the proteolytic activity towards high molecular weight substrates was inhibited. Activity towards low molecular weight substrates was partially modified: the catalytic activity of collagenase bound to ovostatin was inhibited by only 40% towards 2,4-dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-D-Arg and that of thermolysin bound to ovostatin was activated about 2.6-fold towards benzyloxycarbonyl-Gly-Leu-NH2 and benzyloxycarbonyl-Gly-Phe-NH2. Collagenase-ovostatin complexes failed to react with anti-(collagenase) antibody. Saturation of ovostatin with thermolysin prevented the subsequent binding of collagenase. Ovostatin-proteinase complexes ran faster than free ovostatin on 5% polyacrylamide gel electrophoresis. Complexing ovostatin with either collagenase or thermolysin resulted in the cleavage of the quarter-subunit of ovostatin (Mr = 165,000) into two fragments with Mr = 88,000 and 78,000. On the other hand, when the inhibitory capacity of ovostatin was tested with trypsin, chymotrypsin, and papain, only partial inhibition of their proteolytic activities was observed towards azocasein. Stronger inhibition was noted when Azocoll was a substrate, however. Analyses of ovostatin-enzyme complexes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the quarter-subunit of ovostatin was cleaved into several fragments by those enzymes. These results led us to propose that ovostatin inhibits metalloproteinases in preference to proteinases of other classes in a manner similar to alpha 2-macroglobulin; hydrolysis of a peptide bond by a proteinase in the susceptible region of the ovostatin polypeptide chain triggers a conformational change in the ovostatin molecule and the enzyme becomes bound to ovostatin in such a way that the proteinase is sterically hindered from access to large protein substrates and yet

  9. Alpha-1 antitrypsin reduces ovariectomy-induced bone loss in mice

    Science.gov (United States)

    Alpha-1antitrypsin (AAT) is a multifunctional protein with proteinase inhibitor and anti-inflammatory activities. Recent studies showed that AAT has therapeutic effect for diseases associated with inflammation, such as type 1 diabetes and arthritis. Proinflammatory cytokines are primary mediators of...

  10. Midgut proteinases of Sitotroga cerealella (Oliver) (Lepidoptera:Gelechiidae): Characterization and relationship to resistance in cereals

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Lan.

    1989-01-01

    Midgut proteinases are vital to the insects which digest ingested food in the midgut. Insect midgut proteinases, therefore, have been considered as possible targets for the control of insect pests. Proteinaceous proteinase inhibitors are very attractive for their potential use in developing insect resistant plant varieties via genetic engineering. Sitotroga cerealella is one of the major storage pests of cereals, and no antibiotic resistance in wheat against this insect has been identified to date. A series of diagnostic inhibitors, thiol-reducing agents and a metal-ion chelator were used in the identification of proteinases in crude extracts from S. cerealella larval midguts with both protein and ester substrates. The partial inhibition of proteolytic activity in crude midgut extract toward ({sup 3}H)-methemoglobin by pepstatin A suggested the presence of another proteinase which was sensitive to pepstatin A. The optimum pH range for the proteolytic activity, however, indicated that the major midgut proteinases were not carboxyl proteinases. Two proteinases were successfully purified by a combination of fractionation with ammonium sulfate, gel permeation and anion exchange chromatography. Characterization of the enzymes with the purified enzyme preparations confirmed that the two major proteinases were serine endoproteinases with trypsin-like and chymotrypsin-like specificities respectively. Bioassays were conducted using the artificial seeds to test naturally occurring proteinaceous proteinase inhibitors of potential value. Soybean trypsin inhibitor and the Bowman-Birk proteinase inhibitor had adverse effects on the development of the insect. A predictive model was constructed to evaluate effects of seed resistance in conjunction with other control methods on S. cerealella population dynamics.

  11. Midgut proteinases of Sitotroga cerealella (Oliver) (Lepidoptera:Gelechiidae): Characterization and relationship to resistance in cereals

    International Nuclear Information System (INIS)

    Wu, Lan.

    1989-01-01

    Midgut proteinases are vital to the insects which digest ingested food in the midgut. Insect midgut proteinases, therefore, have been considered as possible targets for the control of insect pests. Proteinaceous proteinase inhibitors are very attractive for their potential use in developing insect resistant plant varieties via genetic engineering. Sitotroga cerealella is one of the major storage pests of cereals, and no antibiotic resistance in wheat against this insect has been identified to date. A series of diagnostic inhibitors, thiol-reducing agents and a metal-ion chelator were used in the identification of proteinases in crude extracts from S. cerealella larval midguts with both protein and ester substrates. The partial inhibition of proteolytic activity in crude midgut extract toward [ 3 H]-methemoglobin by pepstatin A suggested the presence of another proteinase which was sensitive to pepstatin A. The optimum pH range for the proteolytic activity, however, indicated that the major midgut proteinases were not carboxyl proteinases. Two proteinases were successfully purified by a combination of fractionation with ammonium sulfate, gel permeation and anion exchange chromatography. Characterization of the enzymes with the purified enzyme preparations confirmed that the two major proteinases were serine endoproteinases with trypsin-like and chymotrypsin-like specificities respectively. Bioassays were conducted using the artificial seeds to test naturally occurring proteinaceous proteinase inhibitors of potential value. Soybean trypsin inhibitor and the Bowman-Birk proteinase inhibitor had adverse effects on the development of the insect. A predictive model was constructed to evaluate effects of seed resistance in conjunction with other control methods on S. cerealella population dynamics

  12. Proteinase K processing of rabbit muscle creatine kinase

    DEFF Research Database (Denmark)

    Leydier, C; Andersen, Jens S.; Couthon, F

    1997-01-01

    Proteinase K cleaves selectively both cytosolic and mitochondrial isoforms of creatine kinase leading to the appearance of two fragments, a large N-terminal one (K1) and a small C-terminal peptide (K2) which remain associated together. The loss of enzymatic activity correlates with the extent...... of monomer cleavage. N-terminal sequencing of the K2 fragments from rabbit cytosolic and pig mitochondrial creatine kinase shows that these peptides begin with A328 and A324, respectively. Electrospray ionization mass spectrometry demonstrates that K2 peptide is composed of 53 residues (A328-K380). However...

  13. Engineering of the Lactococcus lactis serine proteinase by construction of hybrid enzymes

    NARCIS (Netherlands)

    Boerrigter, Ingrid J.; Buist, Girbe; Haandrikman, Alfred J.; Nijhuis, Monique; Reuver, Marjon B. de; Siezen, Roland J.; Venema, Gerhardus; Vos, Willem M. de; Kok, Jan

    Plasmids containing wild-type and hybrid proteinase genes were constructed from DNA fragments of the prtP genes of Lactococcus lactis strains Wg2 and SK11. These plasmids were introduced into the plasmid-free strain L. lactis MG1363. The serine proteinases produced by these L. lactis strains were

  14. Lipid interaction converts prion protein to a PrPSc-like proteinase K-resistant conformation under physiological conditions.

    Science.gov (United States)

    Wang, Fei; Yang, Fan; Hu, Yunfei; Wang, Xu; Wang, Xinhe; Jin, Changwen; Ma, Jiyan

    2007-06-12

    The conversion of prion protein (PrP) to the pathogenic PrPSc conformation is central to prion disease. Previous studies revealed that PrP interacts with lipids and the interaction induces PrP conformational changes, yet it remains unclear whether in the absence of any denaturing treatment, PrP-lipid interaction is sufficient to convert PrP to the classic proteinase K-resistant conformation. Using recombinant mouse PrP, we analyzed PrP-lipid interaction under physiological conditions and followed lipid-induced PrP conformational change with proteinase K (PK) digestion. We found that the PrP-lipid interaction was initiated by electrostatic contact and followed by hydrophobic interaction. The PrP-lipid interaction converted full-length alpha-helix-rich recombinant PrP to different forms. A significant portion of PrP gained a conformation reminiscent of PrPSc, with a PrPSc-like PK-resistant core and increased beta-sheet content. The efficiency for lipid-induced PrP conversion depended on lipid headgroup structure and/or the arrangement of lipids on the surface of vesicles. When lipid vesicles were disrupted by Triton X-100, PrP aggregation was necessary to maintain the lipid-induced PrPSc-like conformation. However, the PK resistance of lipid-induced PrPSc-like conformation does not depend on amyloid fiber formation. Our results clearly revealed that the lipid interaction can overcome the energy barrier and convert full-length alpha-helix-rich PrP to a PrPSc-like conformation under physiological conditions, supporting the relevance of lipid-induced PrP conformational change to in vivo PrP conversion.

  15. NaCl-activated extracellular proteinase from Virgibacillus sp. SK37 isolated from fish sauce fermentation.

    Science.gov (United States)

    Sinsuwan, S; Rodtong, S; Yongsawatdigul, J

    2007-06-01

    Virgibacillus sp. SK37 exhibited high extracellular proteolytic activity in skim milk broth containing 10% NaCl. Optimum conditions of the crude proteinase were at pH 8.0 and 65 degrees C. The proteinase was strongly inhibited by phenylmethanesulfonyl fluoride (PMSF) and preferably hydrolyzed Suc-Ala-Ala-Pro-Phe-AMC, suggesting the serine proteinase with a subtilisin-like characteristic. Proteolytic activity increased with NaCl concentration up to 20%. Ca(2+) activated the enzyme activity but reduced enzyme stability at 65 degrees C. Several proteinases with dominant molecular mass (MW) of 81, 67, 63, 50, 38, and 18 kDa were detected on native-polyacrylamide gel electrophoresis (native-PAGE) activity staining in the absence and presence of 25% NaCl. These results demonstrated that Virgibacillus sp. SK37 produced salt-activated extracellular proteinases. Virgibacillus sp. SK37 could be a promising strain for starter culture development used in fish sauce fermentation.

  16. Diisopropyl fluorophosphate labeling of sperm-associated proteinases

    International Nuclear Information System (INIS)

    Odem, R.R.; Willand, J.L.; Polakoski, K.L.

    1990-01-01

    Proteinase inhibitors have been shown to be capable of preventing various aspects of fertilization. Diisopropyl fluorophosphate (DFP) is an irreversible inhibitor of trypsin-like enzymes that is commercially available in a radiolabeled form. The experiments described herein were designed to determine if DFP would prevent sperm function in live, motile sperm and to identify the sperm proteins bound with DFP. DFP at 5 mM concentrations had no observable effect on sperm motility, but inhibited the penetration of zona-free hamster ova by human sperm (5.5%) compared to controls (33.5%). Acid extracts of motile sperm that had been incubated with radiolabeled DFP and collected by the swim-up procedure demonstrated the presence of radiolabeled DFP, and the autoradiography of the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels of these extracts localized the uptake of radiolabeled DFP to proteins in the molecular weight region of the proacrosin-acrosin system. Acid-extracted proteinases from semen samples incubated with DFP demonstrated a concentration-dependent inhibition of both esterolytic hydrolysis of benzoyl-arginine ethyl ester on spectrophotometric analysis and proteolytic activity on gelatin SDS-PAGE zymography. DFP-labeled proteins were precipitated by highly specific antibodies to proacrosin. These results demonstrated that DFP is capable of inhibiting sperm function, and that it associates with the proacrosin-acrosin system in live motile sperm

  17. Diisopropyl fluorophosphate labeling of sperm-associated proteinases

    Energy Technology Data Exchange (ETDEWEB)

    Odem, R.R.; Willand, J.L.; Polakoski, K.L. (Washington Univ. School of Medicine, St. Louis, MO (USA))

    1990-02-01

    Proteinase inhibitors have been shown to be capable of preventing various aspects of fertilization. Diisopropyl fluorophosphate (DFP) is an irreversible inhibitor of trypsin-like enzymes that is commercially available in a radiolabeled form. The experiments described herein were designed to determine if DFP would prevent sperm function in live, motile sperm and to identify the sperm proteins bound with DFP. DFP at 5 mM concentrations had no observable effect on sperm motility, but inhibited the penetration of zona-free hamster ova by human sperm (5.5%) compared to controls (33.5%). Acid extracts of motile sperm that had been incubated with radiolabeled DFP and collected by the swim-up procedure demonstrated the presence of radiolabeled DFP, and the autoradiography of the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels of these extracts localized the uptake of radiolabeled DFP to proteins in the molecular weight region of the proacrosin-acrosin system. Acid-extracted proteinases from semen samples incubated with DFP demonstrated a concentration-dependent inhibition of both esterolytic hydrolysis of benzoyl-arginine ethyl ester on spectrophotometric analysis and proteolytic activity on gelatin SDS-PAGE zymography. DFP-labeled proteins were precipitated by highly specific antibodies to proacrosin. These results demonstrated that DFP is capable of inhibiting sperm function, and that it associates with the proacrosin-acrosin system in live motile sperm.

  18. Zymography in Multiwells for Quality Assessment of Proteinases.

    Science.gov (United States)

    Mechoor, Ambili; Madanan, Madathiparambil G

    2017-01-01

    Zymography is a well-standardized protocol for the qualitative assessment and analysis of proteinases under specified conditions. However, analysis of a large number of samples simultaneously becomes a challenge when the zymography is carried out by the usual protocol of electrophoresis. This can be overcome by assaying the matrix-degrading proteinases in substrate-impregnated gels in multiwells. Enzymes are copolymerized with 300 mL of 10% acrylamide impregnated with gelatin substrate and incubated for 16 h. The gels are then stained with Coomassie blue, destained with water, and visualized with the naked eye. The intensity; if needed can be measured with a densitometer or gel documentation system. This method has been tested for bacterial collagenases as well as some matrix-degrading metalloproteinases that were purified from rat mammary gland. It can also be used to characterize the enzymes with respect to the type and concentration of the cations required for activity and the role of other regulatory molecules that may affect the enzyme activity. The added advantage of this method is that the electrophoresis set up and electricity is not needed for the procedure.

  19. 1 October 2013 - British Minister of State for Trade and Investment Lord Green of Hurstpierpoint signing the guest book with Head of Internationals Relations R. Voss; visiting the LHC tunnel at Point 1 and the ATLAS experimental cavern with ATLAS Collaboration Members K. Behr and J. Catmore.

    CERN Multimedia

    Jean-Claude Gadmer

    2013-01-01

    1 October 2013 - British Minister of State for Trade and Investment Lord Green of Hurstpierpoint signing the guest book with Head of Internationals Relations R. Voss; visiting the LHC tunnel at Point 1 and the ATLAS experimental cavern with ATLAS Collaboration Members K. Behr and J. Catmore.

  20. The M358R variant of α{sub 1}-proteinase inhibitor inhibits coagulation factor VIIa

    Energy Technology Data Exchange (ETDEWEB)

    Sheffield, William P., E-mail: sheffiel@mcmaster.ca [Canadian Blood Services, Centre for Innovation, Hamilton, Ontario (Canada); Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario (Canada); Bhakta, Varsha [Canadian Blood Services, Centre for Innovation, Hamilton, Ontario (Canada)

    2016-02-12

    The naturally occurring M358R mutation of the plasma serpin α{sub 1}-proteinase inhibitor (API) changes both its cleavable reactive centre bond to Arg–Ser and the efficacy with which it inhibits different proteases, reducing the rate of inhibition of neutrophil elastase, and enhancing that of thrombin, factor XIa, and kallikrein, by several orders of magnitude. Although another plasma serpin with an Arg–Ser reactive centre, antithrombin (AT), has been shown to inhibit factor VIIa (FVIIa), no published data are available with respect to FVIIa inhibition by API M358R. Recombinant bacterially-expressed API M358R and plasma-derived AT were therefore compared using gel-based and kinetic assays of FVIIa integrity and activity. Under pseudo-first order conditions of excess serpin over protease, both AT and API M358R formed denaturation-resistant inhibitory complexes with FVIIa in reactions accelerated by TF; AT, but not API M358R, also required heparin for maximal activity. The second order rate constant for heparin-independent API M358R-mediated FVIIa inhibition was determined to be 7.8 ± 0.8 × 10{sup 2} M{sup −1}sec{sup −1}. We conclude that API M358R inhibits FVIIa by forming inhibitory complexes of the serpin type more rapidly than AT in the absence of heparin. The likely 20-fold excess of API M358R over AT in patient plasma during inflammation raises the possibility that it could contribute to the hemorrhagic tendencies manifested by rare individuals expressing this mutant serpin. - Highlights: • The inhibitory specificity of the serpin alpha-1-proteinase inhibitor (API) is sharply altered in the M358R variant. • API M358R forms denaturation-resistant complexes with coagulation factor VIIa at a rate accelerated by tissue factor but unaffected by heparin. • Complex formation was shown by gel-based assays and quantified kinetically by inhibition of FVIIa-dependent amidolysis.

  1. Are Proteinase 3 and Cathepsin C Enzymes Related to Pathogenesis of Periodontitis?

    Directory of Open Access Journals (Sweden)

    Oya Türkoğlu

    2014-01-01

    Full Text Available Aim. Cathepsin C is the activator of the polymorphonuclear leukocyte-derived proteinase 3, which contributes to inflammatory processes. The aim of the present study was to investigate gingival crevicular fluid (GCF proteinase 3 and cathepsin C levels in periodontal diseases. Design. Eighteen patients with chronic periodontitis (CP, 20 patients with generalized aggressive periodontitis (G-AgP, 20 patients with gingivitis, and 18 healthy subjects were included in the study. Periodontal parameters including probing depth, clinical attachment level, papilla bleeding index, and plaque index were assessed in all study subjects. GCF proteinase 3 and cathepsin C levels were analyzed by ELISA. Results. GCF proteinase 3 total amount was significantly higher in diseased groups compared to control group, after adjusting age P0.05. Periodontal parameters of sampling sites were positively correlated with GCF proteinase 3 total amounts P0.05. Conclusions. Elevated levels of GCF proteinase 3 in CP, G-AgP, and gingivitis might suggest that proteinase 3 plays a role during inflammatory periodontal events in host response. However, cathepsin C in GCF does not seem to have an effect on the pathogenesis of periodontal diseases.

  2. Purification and Characterization of an Extracellular Proteinase from Brevibacterium-Linens ATCC-9174

    DEFF Research Database (Denmark)

    Rattray, F P; Bockelmann, W; Fox, P F

    1995-01-01

    An extracellular serine proteinase from Brevibacterium linens ATCC 9174 was purified to homogeneity. pH and temperature optima were 8,5 and 50 degrees C, respectively. The results for the molecular mass of the proteinase were 56 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis...... and 126 kDa by gel filtration, indicating that the native enzyme exists as a dimer. Mg2+ and Ca2+ activated the proteinase, as did NaCl; however, Hg2+ Fe2+, and Zn2+ caused strong inhibition. The sequence of the first 20 N-terminal amino acids was NH2-Ala-Lys- Asn...

  3. Chemokine degradation by the Group A streptococcal serine proteinase ScpC can be reconstituted in vitro and requires two separate domains.

    Science.gov (United States)

    Fritzer, Andrea; Noiges, Birgit; Schweiger, Daniela; Rek, Angelika; Kungl, Andreas J; von Gabain, Alexander; Nagy, Eszter; Meinke, Andreas L

    2009-08-27

    Streptococcus pyogenes is one of the most common human pathogens and possesses diverse mechanisms to evade the human immune defence. One example of its immune evasion is the degradation of the chemokine IL (interleukin)-8 by ScpC, a serine proteinase that prevents the recruitment of neutrophils to an infection site. By applying the ANTIGENome technology and using human serum antibodies, we identified Spy0416, annotated as ScpC, as a prominent antigen that induces protective immune responses in animals. We demonstrate here for the first time that the recombinant form of Spy0416 is capable of IL-8 degradation in vitro in a concentration- and time-dependent manner. Mutations in the conserved amino acid residues of the catalytic triad of Spy0416 completely abolished in vitro activity. However, the isolated predicted proteinase domain does not exhibit IL-8-degrading activity, but is dependent on the presence of the C-terminal region of Spy0416. Binding to IL-8 is mainly mediated by the catalytic domain. However, the C-terminal region modulates substrate binding, indicating that the proteolytic activity is amenable to regulation via the non-catalytic regions. The specificity for human substrates is not restricted to IL-8, since we also detected in vitro protease activity for another CXC chemokine GRO-alpha (growth-related oncogene alpha), but not for NAP-2 (neutrophil-activating protein 2), SDF (stromal-cell-derived factor)-1alpha, PF-4 (platelet factor 4), I-TAC (interferon-gamma-inducible T-cell alpha-chemoattractant), IP-10 (interferon-gamma-inducible protein 10) and MCP-1 (monocyte chemoattractant protein 1). The degradation of two human CXC chemokines in vitro, the high sequence conservation, the immunogenicity of the protein in humans and the shown protection in animal studies suggest that Spy0416 is a promising vaccine candidate for the prevention of infections by S. pyogenes.

  4. Myogenic and nonmyogenic cells differentially express proteinases, Hsc/Hsp70, and BAG-1 during skeletal muscle regeneration.

    Science.gov (United States)

    Duguez, Stéphanie; Bihan, Marie-Catherine Le; Gouttefangeas, Dominique; Féasson, Léonard; Freyssenet, Damien

    2003-07-01

    Skeletal muscle has a remarkable capacity to regenerate after injury. To determine whether changes in the expression of proteinases, 73-kDa constitutive heat shock cognate protein (Hsc70) and stress-inducible 72-kDa heat shock protein (Hsp70) (Hsc/Hsp70), and Bcl-2-associated gene product-1 (BAG-1) contribute to the remodeling response of muscle tissue, tibialis anterior muscles of male Sprague-Dawley rats were injected with 0.75% bupivacaine and removed at 3, 5, 7, 10, 14, 21, or 35 days postinjection (n = 5-7/group). The immunohistochemical analysis of desmin, alpha-actin, and developmental/neonatal myosin heavy chain expressions indicated the presence of myoblasts (days 3-7), inflammatory cells (days 3-7), degenerating myofibers (days 3-7), regenerating myofibers (days 5-10), and growing mature myofibers (days 10-21) in regenerating muscles. Our biochemical analysis documented profound adaptations in proteolytic metabolism characterized by significant increases in the enzyme activities of matrix metalloproteinases 2 and 9 and plasminogen activators (days 3-14), calpains 1 and 2 (days 3-7), cathepsins B and L(days 3-10), and proteasome (days 3-14). Proteasome activity was strongly correlated with proliferating cell nuclear antigen protein level, suggesting that proteasome played a key role in myoblast proliferation. The expression pattern of BAG-1, a regulatory cofactor of Hsc/Hsp70 at the interface between protein folding and proteasomal proteolysis, did not corroborate the changes in proteasome enzyme activity, suggesting that BAG-1 may promote other functions, such as the folding capacity of Hsc/Hsp70. Altogether, the diversity of functions attributed to proteinases in the present study was strongly supported by the relative changes in the proportion of myogenic and nonmyogenic cells over the time course of regeneration.

  5. Coffee cysteine proteinases and related inhibitors with high expression during grain maturation and germination.

    Science.gov (United States)

    Lepelley, Maud; Amor, Mohamed Ben; Martineau, Nelly; Cheminade, Gerald; Caillet, Victoria; McCarthy, James

    2012-03-01

    Cysteine proteinases perform multiple functions in seeds, including participation in remodelling polypeptides and recycling amino acids during maturation and germination. Currently, few details exist concerning these genes and proteins in coffee. Furthermore, there is limited information on the cysteine proteinase inhibitors which influence the activities of these proteinases. Two cysteine proteinase (CP) and four cysteine proteinase inhibitor (CPI) gene sequences have been identified in coffee with significant expression during the maturation and germination of coffee grain. Detailed expression analysis of the cysteine proteinase genes CcCP1 and CcCP4 in Robusta using quantitative RT-PCR showed that these transcripts accumulate primarily during grain maturation and germination/post germination. The corresponding proteins were expressed in E. coli and purified, but only one, CcCP4, which has a KDDL/KDEL C-terminal sequence, was found to be active after a short acid treatment. QRT-PCR expression analysis of the four cysteine proteinase inhibitor genes in Robusta showed that CcCPI-1 is primarily expressed in developing and germinating grain and CcCPI-4 is very highly expressed during the late post germination period, as well as in mature, but not immature leaves. Transcripts corresponding to CcCPI-2 and CcCPI-3 were detected in most tissues examined at relatively similar, but generally low levels. Several cysteine proteinase and cysteine proteinase inhibitor genes with strong, relatively specific expression during coffee grain maturation and germination are presented. The temporal expression of the CcCP1 gene suggests it is involved in modifying proteins during late grain maturation and germination. The expression pattern of CcCP4, and its close identity with KDEL containing CP proteins, implies this proteinase may play a role in protein and/or cell remodelling during late grain germination, and that it is likely to play a strong role in the programmed cell death

  6. Coffee cysteine proteinases and related inhibitors with high expression during grain maturation and germination

    Directory of Open Access Journals (Sweden)

    Lepelley Maud

    2012-03-01

    Full Text Available Abstract Background Cysteine proteinases perform multiple functions in seeds, including participation in remodelling polypeptides and recycling amino acids during maturation and germination. Currently, few details exist concerning these genes and proteins in coffee. Furthermore, there is limited information on the cysteine proteinase inhibitors which influence the activities of these proteinases. Results Two cysteine proteinase (CP and four cysteine proteinase inhibitor (CPI gene sequences have been identified in coffee with significant expression during the maturation and germination of coffee grain. Detailed expression analysis of the cysteine proteinase genes CcCP1 and CcCP4 in Robusta using quantitative RT-PCR showed that these transcripts accumulate primarily during grain maturation and germination/post germination. The corresponding proteins were expressed in E. coli and purified, but only one, CcCP4, which has a KDDL/KDEL C-terminal sequence, was found to be active after a short acid treatment. QRT-PCR expression analysis of the four cysteine proteinase inhibitor genes in Robusta showed that CcCPI-1 is primarily expressed in developing and germinating grain and CcCPI-4 is very highly expressed during the late post germination period, as well as in mature, but not immature leaves. Transcripts corresponding to CcCPI-2 and CcCPI-3 were detected in most tissues examined at relatively similar, but generally low levels. Conclusions Several cysteine proteinase and cysteine proteinase inhibitor genes with strong, relatively specific expression during coffee grain maturation and germination are presented. The temporal expression of the CcCP1 gene suggests it is involved in modifying proteins during late grain maturation and germination. The expression pattern of CcCP4, and its close identity with KDEL containing CP proteins, implies this proteinase may play a role in protein and/or cell remodelling during late grain germination, and that it is

  7. Accumulation of a maize proteinase inhibitor in response to wounding and insect feeding, and characterization of its activity toward digestive proteinases of Spodoptera littoralis larvae.

    Science.gov (United States)

    Tamayo, M C; Rufat, M; Bravo, J M; San Segundo, B

    2000-06-01

    The mpi gene encodes a maize proteinase inhibitor (MPI) protein whose mRNA accumulates in response to mechanical wounding. In this study, mpi gene expression in response to different types of damage was investigated. In mechanically damaged leaves of maize (Zea mays L.), mpi mRNA accumulation was affected by the degree of damage inflicted on the leaf. Consecutive wounds resulted in higher levels of mpi transcripts. The MPI protein was expressed in Escherichia coli and purified. Polyclonal antibodies were then produced and used to study MPI accumulation in insect-wounded and mechanically wounded maize leaves. When larvae of the lepidopteran insect Spodoptera littoralis were fed on maize leaves, MPI accumulated in tissues adjacent to the wound site. The level of inhibitor accumulation was higher in leaves chewed by larvae than in leaves that had been damaged mechanically. Longer feeding periods also resulted in higher levels of MPI accumulation. Additionally, the inhibitory properties of MPI toward mammalian and insect digestive serine proteinases were determined. Contrary to the majority of the plant proteinase inhibitors described, MPI is an inhibitor of mammalian elastase that only weakly inhibits mammalian chymotrypsin. However, both elastase and chymotrypsin-like activities from the larval midgut of S. littoralis were effectively inhibited by MPI. We discuss these results with regard to the function and evolution of plant proteinase inhibitors. The availability of a plant proteinase inhibitor which is able to inhibit the two types of insect digestive proteinase, elastase and chymotrypsin, might be useful for engineering protection against lepidopteran insect pests in transgenic plants.

  8. Biodistribution, pharmacokinetics and toxicity of a Vasconcellea cundinamarcensis proteinase fraction with pharmacological activity

    Directory of Open Access Journals (Sweden)

    Fernanda O. Lemos

    Full Text Available Abstract Prior studies demonstrate that a proteinase fraction from Vasconcellea cundinamarcensis V.M. Badillo, Caricaceae, exhibits wound healing activity in gastric and cutaneous models and antitumoral/antimetastatic effects. Here, we present the toxicity, pharmacokinetics and biodistribution data for this proteinase fraction following a single dose into Swiss mice by i.v., s.c. or p.o. routes. The i.v. and s.c. toxicity assays demonstrate that proteinase fraction at ≤20 mg/kg is non-lethal after single injection, while parental administration (p.o. of ≤300 mg/kg does not cause death. Based on p.o. acute toxicity dose using Organisation for Economic Cooperation and Development protocols, proteinase fraction ranks as Class IV “harmful” substance. Proteinase fraction shows high uptake determined as Kp (distribution tissue/blood in organs linked to metabolism and excretion. Also, high bioavailability (≈100% was observed by s.c. administration. The blood contents following i.v. dose fits into a pharmacokinetic bi-compartmental model, consisting of high removal constants – kel 0.22 h−1 and kd 2.32 h−1and a half-life – t½ = 3.13 h. The Ames test of proteinase fraction (0.01–1% demonstrates absence of mutagenic activity. Likewise, genotoxic evaluation of proteinase fraction (5 or 10 mg/kg, i.p. shows no influence in micronuclei frequency. In conclusion, the acute doses for proteinase fraction lack mutagenic and genotoxic activity, clearing the way for clinical assays.

  9. Interpain A, a cysteine proteinase from Prevotella intermedia, inhibits complement by degrading complement factor C3.

    Science.gov (United States)

    Potempa, Michal; Potempa, Jan; Kantyka, Tomasz; Nguyen, Ky-Anh; Wawrzonek, Katarzyna; Manandhar, Surya P; Popadiak, Katarzyna; Riesbeck, Kristian; Eick, Sigrun; Blom, Anna M

    2009-02-01

    Periodontitis is an inflammatory disease of the supporting structures of the teeth caused by, among other pathogens, Prevotella intermedia. Many strains of P. intermedia are resistant to killing by the human complement system, which is present at up to 70% of serum concentration in gingival crevicular fluid. Incubation of human serum with recombinant cysteine protease of P. intermedia (interpain A) resulted in a drastic decrease in bactericidal activity of the serum. Furthermore, a clinical strain 59 expressing interpain A was more serum-resistant than another clinical strain 57, which did not express interpain A, as determined by Western blotting. Moreover, in the presence of the cysteine protease inhibitor E64, the killing of strain 59 by human serum was enhanced. Importantly, we found that the majority of P. intermedia strains isolated from chronic and aggressive periodontitis carry and express the interpain A gene. The protective effect of interpain A against serum bactericidal activity was found to be attributable to its ability to inhibit all three complement pathways through the efficient degradation of the alpha-chain of C3 -- the major complement factor common to all three pathways. P. intermedia has been known to co-aggregate with P. gingivalis, which produce gingipains to efficiently degrade complement factors. Here, interpain A was found to have a synergistic effect with gingipains on complement degradation. In addition, interpain A was able to activate the C1 complex in serum, causing deposition of C1q on inert and bacterial surfaces, which may be important at initial stages of infection when local inflammatory reaction may be beneficial for a pathogen. Taken together, the newly characterized interpain A proteinase appears to be an important virulence factor of P. intermedia.

  10. Interpain A, a cysteine proteinase from Prevotella intermedia, inhibits complement by degrading complement factor C3.

    Directory of Open Access Journals (Sweden)

    Michal Potempa

    2009-02-01

    Full Text Available Periodontitis is an inflammatory disease of the supporting structures of the teeth caused by, among other pathogens, Prevotella intermedia. Many strains of P. intermedia are resistant to killing by the human complement system, which is present at up to 70% of serum concentration in gingival crevicular fluid. Incubation of human serum with recombinant cysteine protease of P. intermedia (interpain A resulted in a drastic decrease in bactericidal activity of the serum. Furthermore, a clinical strain 59 expressing interpain A was more serum-resistant than another clinical strain 57, which did not express interpain A, as determined by Western blotting. Moreover, in the presence of the cysteine protease inhibitor E64, the killing of strain 59 by human serum was enhanced. Importantly, we found that the majority of P. intermedia strains isolated from chronic and aggressive periodontitis carry and express the interpain A gene. The protective effect of interpain A against serum bactericidal activity was found to be attributable to its ability to inhibit all three complement pathways through the efficient degradation of the alpha-chain of C3 -- the major complement factor common to all three pathways. P. intermedia has been known to co-aggregate with P. gingivalis, which produce gingipains to efficiently degrade complement factors. Here, interpain A was found to have a synergistic effect with gingipains on complement degradation. In addition, interpain A was able to activate the C1 complex in serum, causing deposition of C1q on inert and bacterial surfaces, which may be important at initial stages of infection when local inflammatory reaction may be beneficial for a pathogen. Taken together, the newly characterized interpain A proteinase appears to be an important virulence factor of P. intermedia.

  11. Electrophoretic analysis of the "cross-class" interaction between novel inhibitory serpin, squamous cell carcinoma antigen-1 and cysteine proteinases.

    Science.gov (United States)

    Nawata, S; Nakamura, K; Tanaka, T; Numa, F; Suminami, Y; Tsunaga, N; Kakegawa, H; Katunuma, N; Kato, H

    1997-05-01

    We investigated the "cross-class" interaction between cysteine proteinases and a novel inhibitory serpin, recombinant squamous cell carcinoma (rSCC) antigen-1, which inhibits a serine proteinase, chymotrypsin. rSCC antigen-1 inhibited the cysteine proteinases, papain, papaya proteinase IV and cathepsin L. Interestingly, although rSCC antigen-1 formed sodium dodecyl sulfate (SDS)- and heat-stable complexes with chymotrypsin, rSCC antigen-1 gave the 40 kDa fragment and small molecular mass peptide by incubation with papain without forming an SDS- and heat-stable complex. The cleavage was observed between the Gly353-Ser354 bond, indicating that rSCC antigen-1 interacts with cysteine proteinases not at the predicted reactive site P1-P1' portion (Ser354-Ser355), but at the Gly353-Ser354 of the P2-P1 portion. These findings promote understanding of the "suicide inhibition" mechanism of SCC antigen-1 against cysteine proteinases.

  12. Granulomatosis with polyangiitis (Wegener granulomatosis): A proteinase-3 driven disease?

    Science.gov (United States)

    Witko-Sarsat, Véronique; Thieblemont, Nathalie

    2018-03-01

    Granulomatosis with polyangiitis (GPA, Wegener granulomatosis) is a systemic autoimmune vasculitis that affects small arteries, arterioles, and capillaries, most notably in the kidneys and lungs. In this disease, proteinase-3 (PR3), produced by neutrophils, is targeted by antineutrophil cytoplasmic antibodies (ANCA). Recent work by our group has shown how PR3 impairs the resolution of inflammation and deregulates the immune system. Normally, the clearance of activated neutrophils triggers an anti-inflammatory, pro-resolution process. In patients with GPA, however, macrophages phagocytose apoptotic neutrophils then release massive amounts of pro-inflammatory mediators, notably interleukin-1, thereby generating a pro-inflammatory microenvironment conducive to autoimmunity. This deregulation of immune processes is accompanied with activation of plasmacytoid dendritic cells and with polarization of T-helper-2 (Th2), Th9, and Th17 cells. These recent data highlight the dual role of PR3, both auto-antigenic and auto-inflammatory, thus potentially opening up new therapeutic avenues. Copyright © 2017 Société française de rhumatologie. Published by Elsevier SAS. All rights reserved.

  13. Identification of Novel Placentally Expressed Aspartic Proteinase in Humans

    Directory of Open Access Journals (Sweden)

    Marta Majewska

    2017-06-01

    Full Text Available This study presents pioneering data concerning the human pregnancy-associated glycoprotein-Like family, identified in the genome, of the term placental transcriptome and proteome. RNA-seq allowed the identification of 1364 bp hPAG-L/pep cDNA with at least 56.5% homology with other aspartic proteinases (APs. In silico analyses revealed 388 amino acids (aa of full-length hPAG-L polypeptide precursor, with 15 aa-signal peptide, 47 aa-blocking peptide and 326 aa-mature protein, and two Asp residues (D, specific for a catalytic cleft of the APs (VVFDTGSSNLWV91-102 and AIVDTGTSLLTG274-285. Capillary sequencing identified 9330 bp of the hPAG-L gene (Gen Bank Acc. No. KX533473, composed of nine exons and eight introns. Heterologous Western blotting revealed the presence of one dominant 60 kDa isoform of the hPAG-L amongst cellular placental proteins. Detection with anti-pPAG-P and anti-Rec pPAG2 polyclonals allowed identification of the hPAG-L proteins located within regions of chorionic villi, especially within the syncytiotrophoblast of term singleton placentas. Our novel data extend the present knowledge about the human genome, as well as placental transcriptome and proteome during term pregnancy. Presumably, this may contribute to establishing a new diagnostic tool for examination of some disturbances during human pregnancy, as well as growing interest from both scientific and clinical perspectives.

  14. Saccharomyces cerevisiae proteinase A excretion and wine making.

    Science.gov (United States)

    Song, Lulu; Chen, Yefu; Du, Yongjing; Wang, Xibin; Guo, Xuewu; Dong, Jian; Xiao, Dongguang

    2017-11-09

    Proteinase A (PrA), the major protease in Saccharomyces cerevisiae, plays an essential role in zymogen activation, sporulation, and other physiological processes in vivo. The extracellular secretion of PrA often occurs during alcoholic fermentation, especially in the later stages when the yeast cells are under stress conditions, and affects the quality and safety of fermented products. Thus, the mechanism underlying PrA excretion must be explored to improve the quality and safety of fermented products. This paper briefly introduces the structure and physiological function of PrA. Two transport routes of PrA, namely, the Golgi-to-vacuole pathway and the constitutive Golgi-to-plasma membrane pathway, are also discussed. Moreover, the research history and developments on the mechanism of extracellular PrA secretion are described. In addition, it is briefly discussed that calcium homeostasis plays an important role in the secretory pathway of proteins, implying that the regulation of PrA delivery to the plasma membrane requires the involvement of calcium ion. Finally, this review focuses on the effects of PrA excretion on wine making (including Chinese rice wine, grape wine, and beer brewage) and presents strategies to control PrA excretion.

  15. Receptor-mediated antigen delivery into macrophages. Complexing antigen to alpha 2-macroglobulin enhances presentation to T cells.

    Science.gov (United States)

    Chu, C T; Pizzo, S V

    1993-01-01

    Macrophages secrete alpha 2-macroglobulin (alpha 2M), a protein that may facilitate early Ag handling. alpha 2M is able to entrap and form covalent linkages with diverse proteins during a transient proteinase-activated state. The resulting complexes are rapidly endocytosed after binding to high affinity receptors. Such a system could be capable of efficiently delivering a multitude of proteins to macrophages. We have used T hybridoma clones that respond only to hen egg lysozyme, in a MHC-restricted manner, to probe the effect of complex formation on Ag uptake and processing by murine macrophages. Radiolabeled lysozyme was internalized more rapidly and to a greater extent when bound to alpha 2M than when unbound. Macrophages pulsed with lysozyme-alpha 2M-elastase complexes required 200 to 250 times less Ag than those pulsed with free lysozyme to achieve effective presentation to T cells. Adding equimolar amounts of alpha 2M-elastase complexes, or of alpha 2M-methylamine, to free lysozyme had no effect on basal lysozyme presentation. Receptor-recognized forms of alpha 2M, but not lysozyme or BSA, competed effectively for both uptake and presentation of lysozyme-alpha 2M-elastase complexes. These results indicate that proteinase-activated alpha 2M can enhance Ag processing by carrying Ag into macrophages through a receptor-mediated process.

  16. Differential gene expression for suicide-substrate serine proteinase inhibitors (serpins) in vegetative and grain tissues of barley

    DEFF Research Database (Denmark)

    Roberts, T.H.; Marttila, S.; Rasmussen, S.K.

    2003-01-01

    Proteins of the serpin superfamily (similar to43 kDa) from mature cereal grains are in vitro suicide-substrate inhibitors of specific mammalian serine proteinases of the chymotrypsin family. However, unlike the 'standard-mechanism' serine proteinase inhibitors (<25 kDa), the biological functions ...... their irreversible inhibitory mechanism in the inhibition of exogenous proteinases capable of breaking down seed storage proteins, and in the defence of specific cell types in vegetative tissues....

  17. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); Xiao, Shaobo, E-mail: shaoboxiao@yahoo.com [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China)

    2010-08-13

    Research highlights: {yields} FMDV L{sup pro} inhibits poly(I:C)-induced IFN-{alpha}1/{beta} mRNA expression. {yields} L{sup pro} inhibits MDA5-mediated activation of the IFN-{alpha}1/{beta} promoter. {yields} L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes. {yields} L{sup pro} inhibits IFN-{alpha}1/{beta} promoter activation by decreasing IRF-3/7 in protein levels. {yields} The ability to process eIF-4G of L{sup pro} is not necessary to inhibit IFN-{alpha}1/{beta} activation. -- Abstract: The leader proteinase (L{sup pro}) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-{beta} (IFN-{beta}) antagonist that disrupts the integrity of transcription factor nuclear factor {kappa}B (NF-{kappa}B). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-{alpha}1/{beta} expression caused by L{sup pro} was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-{alpha}/{beta}. Furthermore, overexpression of L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L{sup pro} mutants indicated that the ability to process eIF-4G of L{sup pro} is not required for suppressing dsRNA-induced activation of the IFN-{alpha}1/{beta} promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-{kappa}B, L{sup pro} also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  18. Plant Proteinase Inhibitors in Therapeutics – Focus on Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Sandhya Srikanth

    2016-12-01

    Full Text Available Plants are known to have many secondary metabolites and phytochemical compounds which are highly explored at biochemical and molecular genetics level and exploited enormously in the human health care sector. However, there are other less explored small molecular weight proteins, which inhibit proteases/proteinases. Plants are good sources of protease inhibitors (PIs which protect them against diseases, insects, pests, and herbivores. In the past, proteinaceous PIs were considered primarily as protein-degrading enzymes. Nevertheless, this view has significantly changed and PIs are now treated as very important signaling molecules in many biological activities such as inflammation, apoptosis, blood clotting and hormone processing. In recent years, PIs have been examined extensively as therapeutic agents, primarily to deal with various human cancers. Interestingly, many plant-based PIs are also found to be effective against cardiovascular diseases, osteoporosis, inflammatory diseases and neurological disorders. Several plant PIs are under further evaluation in in vitro clinical trials. Among all types of PIs, Bowman-Birk inhibitors (BBI has been studied extensively in the treatment of many diseases, especially in the field of cancer prevention. So far, crops such as beans, potatoes, barley, squash, millet, wheat, buckwheat, groundnut, chickpea, pigeonpea, corn and pineapple have been identified as good sources of PIs. The PI content of such foods has a significant influence on human health disorders, particularly in the regions where people mostly depend on these kind of foods. These natural PIs vary in concentration, protease specificity, heat stability, and sometimes several PIs may be present in the same species or tissue. However, it is important to carry out individual studies to identify the potential effects of each PI on human health. PIs in plants make them incredible sources to determine novel PIs with specific pharmacological and

  19. Effect of pH on the production of alkaline proteinase by alkalophilic Bacillus sp

    International Nuclear Information System (INIS)

    Kitada, Makio; Horikoshi, Koki

    1976-01-01

    The effect of the pH of the medium on the microbial growth and alkaline proteinase production, and on the uptake of various substances by alkalophilic Bacillus sp. No.8-1 were studied to investigate the physiological properties of alkalophilic bacteria. Both the microbial growth and alkaline proteinase production by replacement culture were maximum between pH 9 and 10. The alkaline proteinase production sources were also effective for the production. The uptake of various substances such as glucose, acetate, amino acids, and uracil, necessary for proteinase production by this strain, was maximum between pH 9 and 10. The uptake of α-aminoisobutyric acid, a nonmetabolizable amino acid analogue, was also maximum at pH 10. The pH-dependence of these substance was not due to their ionic forms being affected by extracellular pH. It was concluded from above results that good production of alkaline proteinase in alkaline media was due to the active uptake of various nutrients in this culture condition. (auth.)

  20. Determination of germ tube, phospholipase, and proteinase production by bloodstream isolates of Candida albicans

    Directory of Open Access Journals (Sweden)

    Antonella Souza Mattei

    2013-06-01

    Full Text Available Introduction Candida albicans is a commensal and opportunistic agent that causes infection in immunocompromised individuals. Several attributes contribute to the virulence and pathogenicity of this yeast, including the production of germ tubes (GTs and extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate GT production and phospholipase and proteinase activities in bloodstream isolates of C. albicans. Methods One hundred fifty-three C. albicans isolates were obtained from blood samples and analyzed for GT, phospholipase, and proteinase production. The assays were performed in duplicate in egg yolk medium containing bovine serum albumin and human serum. Results Detectable amounts of proteinase were produced by 97% of the isolates, and 78% of the isolates produced phospholipase. GTs were produced by 95% of the isolates. A majority of the isolates exhibited low levels of phospholipase production and high levels of proteinase production. Conclusions Bloodstream isolates of C. albicans produce virulence factors such as GT and hydrolytic enzymes that enable them to cause infection under favorable conditions.

  1. The secreted cathepsin L-like proteinases of the trematode, Fasciola hepatica, contain 3-hydroxyproline residues.

    Science.gov (United States)

    Wijffels, G L; Panaccio, M; Salvatore, L; Wilson, L; Walker, I D; Spithill, T W

    1994-01-01

    The cysteine proteinases synthesized by the adult stage of the trematode Fasciola hepatica were found to be a very heterogeneous group of proteins as demonstrated by one- and two-dimensional gel analyses. N-terminal amino acid sequencing indicated the presence of at least two distinct gene products among the secreted cysteine proteinases. Enzymic studies and peptide sequence analysis of the excreted/secreted cysteine proteinases suggested a close relationship to the plant thiol cathepsins and the mammalian cathepsin L subfamily. The cloning of a representative cDNA for a putative Fasciola cathepsin confirmed similarities to the cathepsin L subfamily but revealed low identity with the cathepsin-like proteinases of the related trematode, Schistosoma, nematode cathepsins and the mammalian cathepsin B subfamily. Furthermore, peptide and protein sequencing revealed the modification of certain highly conserved prolines to unusual 3-hydroxyproline derivatives. This is the first report of modified prolines in any proteinase. This finding, as well as the high activities of these cathepsins at neutral to alkaline pH values, raises a number of questions as to the physiological function of these thiol cathepsins and their interaction with host tissues. Images Figure 1 Figure 2 Figure 3 Figure 6 PMID:8192668

  2. Kazal-type serine proteinase inhibitors in the midgut of Phlebotomus papatasi

    Directory of Open Access Journals (Sweden)

    Leah Theresa Sigle

    2013-09-01

    Full Text Available Sandflies (Diptera: Psychodidae are important disease vectors of parasites of the genus Leishmania, as well as bacteria and viruses. Following studies of the midgut transcriptome of Phlebotomus papatasi, the principal vector of Leishmania major, two non-classical Kazal-type serine proteinase inhibitors were identified (PpKzl1 and PpKzl2. Analyses of expression profiles indicated that PpKzl1 and PpKzl2 transcripts are both regulated by blood-feeding in the midgut of P. papatasi and are also expressed in males, larva and pupa. We expressed a recombinant PpKzl2 in a mammalian expression system (CHO-S free style cells that was applied to in vitro studies to assess serine proteinase inhibition. Recombinant PpKzl2 inhibited α-chymotrypsin to 9.4% residual activity and also inhibited α-thrombin and trypsin to 33.5% and 63.9% residual activity, suggesting that native PpKzl2 is an active serine proteinase inhibitor and likely involved in regulating digestive enzymes in the midgut. Early stages of Leishmania are susceptible to killing by digestive proteinases in the sandfly midgut. Thus, characterising serine proteinase inhibitors may provide new targets and strategies to prevent transmission of Leishmania.

  3. PAR-2, IL-4R, TGF-beta and TNF-alpha in bronchoalveolar lavage distinguishes extrinsic allergic alveolitis from sarcoidosis

    Czech Academy of Sciences Publication Activity Database

    Matěj, R.; Smětáková, M.; Vašáková, M.; Nováková, J.; Šterclová, M.; Kukal, J.; Olejár, Tomáš

    2014-01-01

    Roč. 8, č. 2 (2014), s. 533-538 ISSN 1792-0981 Institutional support: RVO:67985823 Keywords : sarcoidosis * extrinsic allergic alveolitis * interleukin 4 receptor * transforming growth factor beta * tumor necrosis factor alpha * proteinase activated receptor 2 Subject RIV: EC - Immunology Impact factor: 1.269, year: 2014

  4. Buffett's Alpha

    DEFF Research Database (Denmark)

    Frazzini, Andrea; Kabiller, David; Heje Pedersen, Lasse

    Berkshire Hathaway has realized a Sharpe ratio of 0.76, higher than any other stock or mutual fund with a history of more than 30 years, and Berkshire has a significant alpha to traditional risk factors. However, we find that the alpha becomes insignificant when controlling for exposures to Betting-Against-Beta...... in publicly traded stocks versus wholly-owned private companies, we find that the former performs the best, suggesting that Buffett's returns are more due to stock selection than to his effect on management. These results have broad implications for market efficiency and the implementability of academic...

  5. Coronavirus 3CL(pro) proteinase cleavage sites: Possible relevance to SARS virus pathology

    DEFF Research Database (Denmark)

    Kiemer, Lars; Lund, Ole; Brunak, Søren

    2004-01-01

    . Prompted by this, we set out to analyse and predict cleavage by the coronavirus main proteinase using computational methods. Results: We retrieved sequence data on seven fully sequenced coronaviruses and identified the main 3CL proteinase cleavage sites in polyproteins using alignments. A neural network...... which might be important to elucidate coronavirus pathology. Furthermore, the method might assist in design of proteinase inhibitors for treatment of SARS and possible future diseases caused by coronaviruses.......Background: Despite the passing of more than a year since the first outbreak of Severe Acute Respiratory Syndrome (SARS), efficient counter-measures are still few and many believe that reappearance of SARS, or a similar disease caused by a coronavirus, is not unlikely. For other virus families like...

  6. pH-dependent processing of yeast procarboxypeptidase Y by proteinase A in vivo and in vitro

    DEFF Research Database (Denmark)

    Sørensen, S O; van den Hazel, H B; Kielland-Brandt, Morten

    1994-01-01

    Carboxypeptidase Y is a vacuolar enzyme from Saccharomyces cerevisiae. It enters the vacuole as a zymogen, procarboxypeptidase Y, which is immediately processed in a reaction involving two endoproteases, proteinase A and proteinase B. We have investigated the in vitro activation of purified...

  7. Th1 Cell Development Induced by Cysteine Proteinases A and B in Localized Cutaneous Leishmaniasis Due to Leishmania guyanensis

    Science.gov (United States)

    Pascalis, Hervé; Lavergne, Anne; Bourreau, Eliane; Prévot-Linguet, Ghislaine; Kariminia, Amina; Pradinaud, Roger; Rafati, Sima; Launois, Pascal

    2003-01-01

    The cysteine proteinases CPA and CPB from Leishmania major induced Th1 responses in patients with leishmaniasis due to Leishmania guyanensis. Furthermore, cysteine proteinases induced neither interleukin 4 (IL-4) nor IL-13 and low levels of IL-10 in controls and patients. The results suggest that CPs would be quite good candidates for a vaccine against different Leishmania species. PMID:12704171

  8. Mechanisms of liver fibrosis associated with experimental Fasciola hepatica infection: roles of Fas2 proteinase and hepatic stellate cell activation.

    Science.gov (United States)

    Marcos, Luis A; Terashima, Angélica; Yi, Pedro; Andrade, Roy; Cubero, Francisco J; Albanis, Efsevia; Gotuzzo, Eduardo; Espinoza, Jose R; Friedman, Scott L

    2011-02-01

    We have evaluated the possible mechanisms of liver fibrosis caused by Fasciola hepatica in an animal model and in culture using immortalized human stellate cells. Liver biopsies of F. hepatica-infected rats were performed at wk 8 and 16. Serum-starved LX-2 cells, a human stellate cell line, were exposed to increasing concentrations of Fas2 antigen. The expression of key fibrosis-related genes was evaluated by qRT-PCR. There was a significant correlation between fibrogenic gene expression and both intensity and duration of infection. LX-2 cells exposed to Fas2 showed progressively increased expression of mRNAs for Collagen I, alpha-smooth muscle-actin, platelet-derived growth factor beta receptor, and tissue inhibitor of metalloproteinase II; inhibition of Fas2 cysteine proteinase activity by E-64 abrogated these increases, suggesting that the protease activity of Fas2 is involved in fibrogenic stimulation. In summary, F. hepatica infection is associated with up-regulation of mRNAs associated with hepatic fibrogenesis in vivo and in activated hepatic stellate cells.

  9. Inhibition of plasmin, urokinase, tissue plasminogen activator, and C1S by a myxoma virus serine proteinase inhibitor.

    Science.gov (United States)

    Lomas, D A; Evans, D L; Upton, C; McFadden, G; Carrell, R W

    1993-01-05

    The myxoma and malignant rabbit fibroma poxviruses are lethal tumorigenic viruses of rabbits whose virulence is modulated by the production of a virus-encoded secreted serine proteinase inhibitor, SERP-1. This viral protein was detected in medium harvested from myxoma and malignant rabbit fibroma virus-infected cells, and its inhibitory profile has been characterized by gel and kinetic analysis. SERP-1 forms complexes with and inhibits the human fibrinolytic enzymes plasmin, urokinase, and two-chain tissue-type plasminogen activator (association rate constants 3.4 x 10(4), 4.3 x 10(4), and 3.6 x 10(4) M-1 s-1 respectively). It is also able to inhibit C1S, the first enzyme in the complement cascade with an association rate constant which was unaffected by the addition of heparin (1.3 x 10(3) M-1 s-1). SERP-1 acts as a substrate for and is cleaved by thrombin, porcine trypsin, human neutrophil elastase, porcine pancreatic elastase, thermolysin, subtilisin, bovine alpha-chymotrypsin, and factor Xa. Incubation with kallikrein and cathepsin G had no effect. The structure of SERP-1 has been modeled on other members of the serpin family which revealed the characteristic serpin architecture apart from the absence of the D-helix. Structural analysis and kinetic assays demonstrate that the absence of this region does not prevent inhibitory activity and furthermore allow the identification of cysteine residues involved in internal and intermolecular disulfide bonding.

  10. Secretory leukocyte proteinase inhibitor-competent DNA deposits are potent stimulators of plasmacytoid dendritic cells

    DEFF Research Database (Denmark)

    Skrzeczynska-Moncznik, Joanna; Wlodarczyk, Agnieszka; Zabieglo, Katarzyna

    2012-01-01

    Secretory leukocyte proteinase inhibitor (SLPI) is a well-established inhibitor of serine proteases such as human neutrophil elastase (HNE) and a NF-κB regulatory agent in immune cells. In this paper, we report that SLPI plays a previously uncharacterized role in regulating activation of plasmacy......Secretory leukocyte proteinase inhibitor (SLPI) is a well-established inhibitor of serine proteases such as human neutrophil elastase (HNE) and a NF-κB regulatory agent in immune cells. In this paper, we report that SLPI plays a previously uncharacterized role in regulating activation...

  11. The cell envelope subtilisin-like proteinase is a virulence determinant for Streptococcus suis

    Directory of Open Access Journals (Sweden)

    Gottschalk Marcelo

    2010-02-01

    Full Text Available Abstract Background Streptococcus suis is a major swine pathogen and zoonotic agent that mainly causes septicemia, meningitis, and endocarditis. It has recently been suggested that proteinases produced by S. suis (serotype 2 are potential virulence determinants. In the present study, we screened a S. suis mutant library created by the insertion of Tn917 transposon in order to isolate a mutant deficient in a cell surface proteinase. We characterized the gene and assessed the proteinase for its potential as a virulence factor. Results Two mutants (G6G and M3G possessing a single Tn917 insertion were isolated. The affected gene coded for a protein (SSU0757 that shared a high degree of identity with Streptococccus thermophilus PrtS (95.9% and, to a lesser extent, with Streptococcus agalactiae CspA (49.5%, which are cell surface serine proteinases. The SSU0757 protein had a calculated molecular mass of 169.6 kDa and contained the catalytic triad characteristic of subtilisin family proteinases: motif I (Asp200, motif II (His239, and motif III (Ser568. SSU0757 also had the Gram-positive cell wall anchoring motif (Leu-Pro-X-Thr-Gly at the carboxy-terminus, which was followed by a hydrophobic domain. All the S. suis isolates tested, which belonged to different serotypes, possessed the gene encoding the SSU0757 protein. The two mutants devoid of subtilisin-like proteinase activity had longer generation times and were more susceptible to killing by whole blood than the wild-type parent strain P1/7. The virulence of the G6G and M3G mutants was compared to the wild-type strain in the CD1 mouse model. Significant differences in mortality rates were noted between the P1/7 group and the M3G and G6G groups (p Conclusion In summary, we identified a gene coding for a cell surface subtilisin-like serine proteinase that is widely distributed in S. suis. Evidences were brought for the involvement of this proteinase in S. suis virulence.

  12. Expression of human α1-proteinase inhibitor in Aspergillus niger

    Directory of Open Access Journals (Sweden)

    Punt Peter J

    2007-10-01

    Full Text Available Abstract Background Human α1-proteinase inhibitor (α1-PI, also known as antitrypsin, is the most abundant serine protease inhibitor (serpin in plasma. Its deficiency is associated with development of progressive, ultimately fatal emphysema. Currently in the United States, α1-PI is available for replacement therapy as an FDA licensed plasma-derived (pd product. However, the plasma source itself is limited; moreover, even with efficient viral inactivation steps used in manufacture of plasma products, the risk of contamination from emerging viruses may still exist. Therefore, recombinant α1-PI (r-α1-PI could provide an attractive alternative. Although r-α1-PI has been produced in several hosts, protein stability in vitro and rapid clearance from the circulation have been major issues, primarily due to absent or altered glycosylation. Results We have explored the possibility of expressing the gene for human α1-PI in the filamentous fungus Aspergillus niger (A. niger, a system reported to be capable of providing more "mammalian-like" glycosylation patterns to secretable proteins than commonly used yeast hosts. Our expression strategy was based on fusion of α1-PI with a strongly expressed, secreted leader protein (glucoamylase G2, separated by dibasic processing site (N-V-I-S-K-R that provides in vivo cleavage. SDS-PAGE, Western blot, ELISA, and α1-PI activity assays enabled us to select the transformant(s secreting a biologically active glycosylated r-α1-PI with yields of up to 12 mg/L. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS analysis further confirmed that molecular mass of the r-α1-PI was similar to that of the pd-α1-PI. In vitro stability of the r-α1-PI from A. niger was tested in comparison with pd-α1-PI reference and non-glycosylated human r-α1-PI from E. coli. Conclusion We examined the suitability of the filamentous fungus A. niger for the expression of the human gene for α1-PI, a medium size

  13. The aspartic proteinase family of three Phytophthora species

    Directory of Open Access Journals (Sweden)

    ten Have Arjen

    2011-05-01

    Full Text Available Abstract Background Phytophthora species are oomycete plant pathogens with such major social and economic impact that genome sequences have been determined for Phytophthora infestans, P. sojae and P. ramorum. Pepsin-like aspartic proteinases (APs are produced in a wide variety of species (from bacteria to humans and contain conserved motifs and landmark residues. APs fulfil critical roles in infectious organisms and their host cells. Annotation of Phytophthora APs would provide invaluable information for studies into their roles in the physiology of Phytophthora species and interactions with their hosts. Results Genomes of Phytophthora infestans, P. sojae and P. ramorum contain 11-12 genes encoding APs. Nine of the original gene models in the P. infestans database and several in P. sojae and P. ramorum (three and four, respectively were erroneous. Gene models were corrected on the basis of EST data, consistent positioning of introns between orthologues and conservation of hallmark motifs. Phylogenetic analysis resolved the Phytophthora APs into 5 clades. Of the 12 sub-families, several contained an unconventional architecture, as they either lacked a signal peptide or a propart region. Remarkably, almost all APs are predicted to be membrane-bound. Conclusions One of the twelve Phytophthora APs is an unprecedented fusion protein with a putative G-protein coupled receptor as the C-terminal partner. The others appear to be related to well-documented enzymes from other species, including a vacuolar enzyme that is encoded in every fungal genome sequenced to date. Unexpectedly, however, the oomycetes were found to have both active and probably-inactive forms of an AP similar to vertebrate BACE, the enzyme responsible for initiating the processing cascade that generates the Aβ peptide central to Alzheimer's Disease. The oomycetes also encode enzymes similar to plasmepsin V, a membrane-bound AP that cleaves effector proteins of the malaria parasite

  14. The aspartic proteinase family of three Phytophthora species

    Science.gov (United States)

    2011-01-01

    Background Phytophthora species are oomycete plant pathogens with such major social and economic impact that genome sequences have been determined for Phytophthora infestans, P. sojae and P. ramorum. Pepsin-like aspartic proteinases (APs) are produced in a wide variety of species (from bacteria to humans) and contain conserved motifs and landmark residues. APs fulfil critical roles in infectious organisms and their host cells. Annotation of Phytophthora APs would provide invaluable information for studies into their roles in the physiology of Phytophthora species and interactions with their hosts. Results Genomes of Phytophthora infestans, P. sojae and P. ramorum contain 11-12 genes encoding APs. Nine of the original gene models in the P. infestans database and several in P. sojae and P. ramorum (three and four, respectively) were erroneous. Gene models were corrected on the basis of EST data, consistent positioning of introns between orthologues and conservation of hallmark motifs. Phylogenetic analysis resolved the Phytophthora APs into 5 clades. Of the 12 sub-families, several contained an unconventional architecture, as they either lacked a signal peptide or a propart region. Remarkably, almost all APs are predicted to be membrane-bound. Conclusions One of the twelve Phytophthora APs is an unprecedented fusion protein with a putative G-protein coupled receptor as the C-terminal partner. The others appear to be related to well-documented enzymes from other species, including a vacuolar enzyme that is encoded in every fungal genome sequenced to date. Unexpectedly, however, the oomycetes were found to have both active and probably-inactive forms of an AP similar to vertebrate BACE, the enzyme responsible for initiating the processing cascade that generates the Aβ peptide central to Alzheimer's Disease. The oomycetes also encode enzymes similar to plasmepsin V, a membrane-bound AP that cleaves effector proteins of the malaria parasite Plasmodium falciparum during

  15. [Epizoologic studies of the detection of antibodies against Aujeszky's disease virus in sera and blood eluates of swine from Thailand using ELISA ("Enzygnost," Behring), serum neutralization test and "Aujeszky Latex Kit" (Iffa Merieux)].

    Science.gov (United States)

    Leamcharaskul, P; Renner-Müller, I C; Munz, E; Reimann, M

    1990-08-01

    The results of three tests for Aujeszky's disease were analysed and compared. The presence of Aujeszky's antibodies was determined by "Enzyme-linked-Immunosorbent-Assays" (ELISA, "Enzygnost"), Behring company, Marburg; "Serum-Neutralization-Tests" (SNT); and "Latex Agglutination-Tests" (LT, "Aujeszky-Latex-Kit"), Iffa Merieux company, Laupheim. Whole blood and sera samples were taken from 805 swine from 26 of Thailand's provinces. These samples were analysed to determine if eluates of whole blood on filter paper were as effective as corresponding sera samples in determining the presence of Aujeszky's disease antibodies. From a total of 805 samples, 26% of the serum and 18% of the blood eluate samples showed a positive result when tested by the ELISA method. Clearly, testing whole blood eluates provides results which are inferior to results from sera samples. Therefore the ELISA whole blood eluates test can only be recommended with reservations. Further testing was done on 645 serum samples using SNT. Samples tested were those which gave negative, suspicious, or weakly positive results when tested by ELISA. Using SNT, 23% of these showed a positive result. Many serum and blood eluate samples were also tested by LT. Most of these test samples were chosen because they were deemed suspicious. Suspicious samples were defined as those which had deviant test results. According to these results the sensitivity of LT was between the sensitivity of SNT and ELISA. Owner survey responses tended to state that few animals had been vaccinated. This coupled with the frequency of antibody occurrence proves the high rate of infection among Thailand's swine population.

  16. Random substitution of large parts of the propeptide of yeast proteinase A

    DEFF Research Database (Denmark)

    van den Hazel, H B; Kielland-Brandt, Morten; Winther, Jakob R.

    1995-01-01

    The yeast aspartic protease, proteinase A, has a 54 amino-acid propeptide, which is removed during activation of the zymogen in the vacuole. Apart from being involved inhibition/activation, the propeptide has been shown to be essential for formation of a stable active enzyme (van den Hazel, H. B...

  17. Subunit structure of karatasin, the proteinase isolated from Bromelia plumieri (karatas).

    Science.gov (United States)

    Montes, C; Amador, M; Cuevas, D; Cordoba, F

    1990-01-01

    Close to 15% of the karatasin proteinase activity in the fruit juice of Bromelia plumieri (karatas) is present outside dialysis Visking tubing in 7 days in 0.2 M acetate buffer (pH) 3.5 or 6.5) containing phenyl mercuric acetate. The small proteinase(s), distinct from the 85% activity in juice due to nondialysable karatasin with a reported Mr of 24,868, separates across Spectrapore (13 kDa) membranes but not across Spectrapore with 3.5 kDa average pore diameter. The dialyzed proteinase is named karatasin-D (K-D). Purified non-Dialysable karatasin can be dissociated to what seems to be K-D by incubation in a buffer solution, containing SDS and 2-mercaptoethanol with phenyl mercuric acetate, in dialysis experiments for 8 days at room temperature using Spectrapore 13 kDa tubing. Thus, native karatasin in B. plumieri fruit juice seem to be the result of association of 2 small molecular mass K-D subunits, linked together by disulfide bonds and electrostatic forces, in equilibrium with small amounts of free K-D molecules. The amino acid composition and partial sequence of karatasin up to the 14th position from the amino terminus have discrete analogies with papain and with stem bromelain.

  18. Secreted aspartate proteinases, a virulence factor of Candida spp.: Occurrence among clinical isolates

    Czech Academy of Sciences Publication Activity Database

    Hamal, P.; Dostál, Jiří; Raclavský, V.; Krylová, M.; Pichová, Iva; Hrušková-Heidingsfeldová, Olga

    2004-01-01

    Roč. 49, č. 4 (2004), s. 491-496 ISSN 0015-5632 R&D Projects: GA MZd NI6485 Institutional research plan: CEZ:AV0Z4055905 Keywords : Candida spp. * aspartate proteinases * RAPD typing Subject RIV: CE - Biochemistry Impact factor: 1.034, year: 2004

  19. The digestion of phagocytosed collagen is inhibited by the proteinase inhibitors leupeptin and E-64

    NARCIS (Netherlands)

    Everts, V.; Beertsen, W.; Tigchelaar-Gutter, W.

    1985-01-01

    Using morphometric methods the effects of the thiol-proteinase inhibitors leupeptin and E-64 on the digestion of intracytoplasmic collagen fibrils were studied in cultured mouse bone explants. Both drugs caused a dose-dependent increase of lysosomal structures containing cross-banded collagen

  20. In vivo and in vitro effect of Acacia nilotica seed proteinase inhibitors ...

    Indian Academy of Sciences (India)

    Acacia nilotica proteinase inhibitor (AnPI) was isolated by ammonium sulphate precipitation followed by chromatography on DEAE-Sephadex A-25 and resulted in a purification of 10.68-fold with a 19.5% yield. Electrophoretic analysis of purified AnPI protein resolved into a single band with molecular weight of ...

  1. Fasciola gigantica cathepsin L proteinase-based synthetic peptide for immunodiagnosis and prevention of sheep fasciolosis

    Czech Academy of Sciences Publication Activity Database

    Ježek, Jan; El Ridi, R.; Salah, M.; Wagih, A.; Aziz, H. W.; Tallima, H.; El Shafie, M. H.; Khalek, T. A.; Ammou, F. F. A.; Strongylis, C.; Moussis, V.; Tsikaris, V.

    2008-01-01

    Roč. 90, č. 3 (2008), s. 349-357 ISSN 0006-3525 Institutional research plan: CEZ:AV0Z40550506 Keywords : cathepsin L proteinase * peptides * sequential oligopeptide carriers * synthetic peptide vaccine * Fasciiola gigantica Subject RIV: CC - Organic Chemistry Impact factor: 2.823, year: 2008

  2. A triticale water-deficit-inducible phytocystatin inhibits endogenous cysteine proteinases in vitro.

    Science.gov (United States)

    Chojnacka, Magdalena; Szewińska, Joanna; Mielecki, Marcin; Nykiel, Małgorzata; Imai, Ryozo; Bielawski, Wiesław; Orzechowski, Sławomir

    2015-02-01

    Water-deficit is accompanied by an increase in proteolysis. Phytocystatins are plant inhibitors of cysteine proteinases that belong to the papain and legumain family. A cDNA encoding the protein inhibitor TrcC-8 was identified in the vegetative organs of triticale. In response to water-deficit, increases in the mRNA levels of TrcC-8 were observed in leaf and root tissues. Immunoblot analysis indicated that accumulation of the TrcC-8 protein occurred after 72h of water-deficit in the seedlings. Using recombinant protein, inhibitory activity of TrcC-8 against cysteine proteases from triticale and wheat tissues was analyzed. Under water-deficit conditions, there are increases in cysteine proteinase activities in both plant tissues. The cysteine proteinase activities were inhibited by addition of the recombinant TrcC-8 protein. These results suggest a potential role for the triticale phytocystatin in modulating cysteine proteinase activities during water-deficit conditions. Copyright © 2014 Elsevier GmbH. All rights reserved.

  3. Modeling the growth and proteinase A production in continuous cultures of recombinant Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Carlsen, Morten; Jochumsen, Kirsten Væver; Emborg, Claus

    1997-01-01

    Overexpression of the homologous protein proteinase A (PrA) in Saccharomyces cerevisiae has been achieved by inserting the PrA gene (PEP4) with its own promoter on a 2 mu multicopy plasmid. With this system the specific PrA production rate was found to be described well by a linear function...

  4. Proteinase K and the structure of PrPse: the good, the bad, and the ugly

    Science.gov (United States)

    Infectious proteins (prions) are, ironically, defined by their resistance to proteolytic digestion. A defining characteristic of the transmissible isoform of the prion protein (PrPSc) is its partial resistance to proteinase K (PK) digestion. Diagnosis of prion disease typically relies upon immunod...

  5. Hydrolytic activity of Virgibacillus sp. SK37, a starter culture of fish sauce fermentation, and its cell-bound proteinases.

    Science.gov (United States)

    Sinsuwan, Sornchai; Rodtong, Sureelak; Yongsawatdigul, Jirawat

    2012-08-01

    Fish sauce production relies on a natural fermentation process requiring 12-18 months for process completion. Virgibacillus sp. SK37 has been shown to be a potential strain for fish sauce acceleration. However, hydrolytic activity of proteinases bound at cell surface of this strain has not been well elucidated. Addition of 0.2 % CaCl(2) (w/w) in conjunction with starter cultures of Virgibacillus sp. SK 37 increased protein hydrolysis as measured by α-amino group content throughout fermentation (P bound proteinases from Virgibacillus sp. SK 37 were extracted into a free form by incubating the washed cells in Ca(2+)-free buffer at 37 °C for 2 h. Cell-bound proteinases revealed molecular mass of 19, 20, 22, 32, 34, and 44 kDa based on a synthetic peptide zymogram. The proteinases showed subtilisin-like serine characteristics with the highest activity at 50 °C and pH 8 and 11. Activity of the extracted proteinases increased ~4 times at ≥100 mM CaCl(2). In addition, CaCl(2) enhanced thermal stability of the extracted proteinases. Enzymes showed proteolytic activity in either the absence or presence of 10 and 25 % NaCl toward fish muscle, soy protein isolate, and casein substrates. Cell-bound proteinases were likely to play an important role in protein hydrolysis during fish sauce fermentation.

  6. pH-dependent processing of yeast procarboxypeptidase Y by proteinase A in vivo and in vitro

    DEFF Research Database (Denmark)

    Sørensen, S O; van den Hazel, H B; Kielland-Brandt, Morten

    1994-01-01

    procarboxypeptidase Y by purified proteinase A. This has identified two different processing intermediates; one active and one inactive. The intermediates define a 33 amino acid segment of the 91 amino acid propeptide as sufficient for maintaining the enzyme in an inactive state. The inactive intermediate...... activity. Efficient processing of procarboxypeptidase Y in the absence of proteinase B is dependent on acidic vacuolar pH, and the processing at neutral pH is slow and takes place in two steps similar to those identified in vitro.......Carboxypeptidase Y is a vacuolar enzyme from Saccharomyces cerevisiae. It enters the vacuole as a zymogen, procarboxypeptidase Y, which is immediately processed in a reaction involving two endoproteases, proteinase A and proteinase B. We have investigated the in vitro activation of purified...

  7. Binding properties of the regulatory domains in Manduca sexta hemolymph proteinase-14, an initiation enzyme of the prophenoloxidase activation system

    OpenAIRE

    Wang, Yang; Jiang, Haobo

    2009-01-01

    Pathogen recognition and rapid initiation of defense responses are essential for the survival of host insects. In Manduca sexta, hemolymph proteinase-14 precursor (proHP14) senses non-self presence and triggers a branched serine proteinase pathway which leads to prophenoloxidase activation and melanin formation around the invading organisms. To understand functions of individual domains in HP14, we have produced a series of HP14 domains and truncation mutants and studied their interactions wi...

  8. Effect of added proteinases and level of starter culture on the formation of biogenic amines in raw milk Manchego cheese.

    Science.gov (United States)

    Fernández-García, E; Tomillo, J; Núñez, M

    1999-11-15

    The influence of two proteinases (Bacillus subtilis neutral proteinase and Micrococcus sp. cysteine proteinase) and two starter culture levels (0.1% and 1%) on biogenic amine formation has been studied in raw ewes' milk Manchego cheese. Amino acid decarboxylating micro-organisms were determined on tyrosine enriched selective media. Biogenic amines were analysed by capillary electrophoresis in citrate buffer at pH 3.6. Addition of proteinases and level of starter culture did not influence the population of micro-organisms with amino acid decarboxylating activity, which represented on average 1% of the bacterial population in 30-day-old cheeses. Tyramine and histamine were detected in all batches of cheese from day 30. Concentrations of tyramine and histamine were higher in cheeses made from milk with neutral proteinase (up to 356 and 284 mg kg(-1), respectively, after 90 days) than in cheeses made from milk with cysteine proteinase (up to 269 and 189 mg kg(-1), respectively) or with no proteinase added (up to 305 and 226 mg kg(-1), respectively). Formation of tyramine and histamine was also favoured in cheeses made with 1% starter culture with respect to cheeses made with only 0.1% starter culture, probably due to the higher pH values of the former cheeses. After 90 days of ripening, concentrations of 10-20 mg kg(-1) phenylethylamine were observed in 9 of the 12 batches, and levels < 10 mg kg(-1) tryptamine were only detected in 3 batches, with no significant relationship between the concentration of these amines and proteinase addition or level of starter culture.

  9. Degradation of immunoglobulins, protease inhibitors, and interleukin-1 by a secretory proteinase of Acanthamoeba castellanii

    Science.gov (United States)

    Na, Byoung-Kuk; Cho, Jong-Hwa; Song, Chul-Yong; Kim, Tong-Soo

    2002-01-01

    The effect of a secretory proteinase from the pathogenic amoebae Acanthamoeba castellanii on host's defense-oriented or regulatory proteins such as immunoglobulins, interleukin-1, and protease inhibitors was investigated. The enzyme was found to degrade secretory immunoglobulin A (sIgA), IgG, and IgM. It also degraded interleukin-1α (IL-1α) and IL-1β. Its activity was not inhibited by endogenous protease inhibitors, such as α2-macroglobulin, α1-trypsin inhibitor, and α2-antiplasmin. Furthermore, the enzyme rapidly degraded those endogenous protease inhibitors as well. The degradation of host's defense-oriented or regulatory proteins by the Acanthamoeba proteinase suggested that the enzyme might be an important virulence factor in the pathogenesis of Acanthamoeba infection. PMID:12073735

  10. A preliminary neutron crystallographic study of proteinase K at pD 6.5

    Energy Technology Data Exchange (ETDEWEB)

    Gardberg, Anna S [ORNL; Blakeley, Matthew P. [Institut Laue-Langevin (ILL); Myles, Dean A A [ORNL

    2009-01-01

    AbstractA preliminary neutron crystallographic study of the proteolytic enzyme proteinase K is presented. Large hydrogenated crystals were prepared in deuterated crystallization buffer using the vapour-diffusion method. Data were collected to a resolution of 2.3 on the LADI-III diffractometer at the Institut Laue Langevin (ILL) in 2.5 days. The results demonstrate the feasibility of a full neutron crystallographic analysis of this structure aimed at providing relevant information on the location of H atoms, particularly at the active site. This information will contribute to further understanding of the molecular mechanisms underlying proteinase K's catalytic activity and to an enriched understanding of the subtilisin clan of serine proteases.

  11. Coulomb correction to elastic. alpha. -. alpha. scattering

    Energy Technology Data Exchange (ETDEWEB)

    Bera, P.K.; Jana, A.K.; Haque, N.; Talukdar, B. (Department of Physics, Visva-Bharati University, Santiniketan-731235, West Bengal, India (IN))

    1991-02-01

    The elastic {alpha}-{alpha} scattering is treated within the framework of a generalized phase-function method (GPFM). This generalization consists in absorbing the effect of Coulomb interaction in the comparison functions for developing the phase equation. Based on values of scattering phase shifts computed by the present method, it is concluded that the GPFM provides an uncomplicated approach to rigorous Coulomb correction in the {alpha}-{alpha} scattering.

  12. Novel Aggregation Properties of Candida albicans Secreted Aspartyl Proteinase Sap6 Mediate Virulence in Oral Candidiasis.

    Science.gov (United States)

    Kumar, Rohitashw; Saraswat, Darpan; Tati, Swetha; Edgerton, Mira

    2015-07-01

    Candida albicans, a commensal fungus of the oral microbiome, causes oral candidiasis in humans with localized or systemic immune deficiencies. Secreted aspartic proteinases (Saps) are a family of 10 related proteases and are virulence factors due to their proteolytic activity, as well as their roles in adherence and colonization of host tissues. We found that mice infected sublingually with C. albicans cells overexpressing Sap6 (SAP6 OE and a Δsap8 strain) had thicker fungal plaques and more severe oral infection, while infection with the Δsap6 strain was attenuated. These hypervirulent strains had highly aggregative colony structure in vitro and higher secreted proteinase activity; however, the levels of proteinase activity of C. albicans Saps did not uniformly match their abilities to damage cultured oral epithelial cells (SCC-15 cells). Hyphal induction in cells overexpressing Sap6 (SAP6 OE and Δsap8 cells) resulted in formation of large cell-cell aggregates. These aggregates could be produced in germinated wild-type cells by addition of native or heat-inactivated Sap6. Sap6 bound only to germinated cells and increased C. albicans adhesion to oral epithelial cells. The adhesion properties of Sap6 were lost upon deletion of its integrin-binding motif (RGD) and could be inhibited by addition of RGD peptide or anti-integrin antibodies. Thus, Sap6 (but not Sap5) has an alternative novel function in cell-cell aggregation, independent of its proteinase activity, to promote infection and virulence in oral candidiasis.

  13. Bioprocessing and immobilization of cell envelope proteinases from Lactobacillus delbrueckii subsp. lactis 313, for protein degradation

    OpenAIRE

    Agyei, Dominic

    2017-01-01

    Proteolytic enzymes are a useful class of biomolecules due to their ubiquity and the plethora of physiological roles they play in living systems. These enzymes are esponsible for the breakdown of proteins to peptides and have several applications in food, pharmaceuticals, diagnostics, photographic, waste treatments, bioremediation, and in the textile industry. Cell-envelope proteinases (CEPs) are a special class of industrially relevant extracellular proteolytic enzymes obtained from la...

  14. In vitro differential activity of phospholipases and acid proteinases of clinical isolates of Candida

    Directory of Open Access Journals (Sweden)

    Aurean D'Eça Júnior

    2011-06-01

    Full Text Available INTRODUCTION: Candida yeasts are commensals; however, if the balance of normal flora is disrupted or the immune defenses are compromised, Candida species can cause disease manifestations. Several attributes contribute to the virulence and pathogenicity of Candida, including the production of extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate the in vitro activity of phospholipases and acid proteinases in clinical isolates of Candida spp. METHODS: Eighty-two isolates from hospitalized patients collected from various sites of origin were analyzed. Phospholipase production was performed in egg yolk medium and the production of proteinase was verified in a medium containing bovine serum albumin. The study was performed in triplicate. RESULTS: Fifty-six (68.3% of isolates tested were phospholipase positive and 16 (44.4% were positive for proteinase activity. C. tropicalis was the species with the highest number of positive isolates for phospholipase (91.7%. Statistically significant differences were observed in relation to production of phospholipases among species (p<0,0001 and among the strains from different sites of origin (p=0.014. Regarding the production of acid protease, the isolates of C. parapsilosis tested presented a larger number of producers (69.2%. Among the species analyzed, the percentage of protease producing isolates did not differ statistically (χ2=1.9 p=0.5901 (χ2=1.9 p=0.5901. CONCLUSIONS: The majority of C. non-albicans and all C. albicans isolates were great producers of hydrolytic enzymes and, consequently, might be able to cause infection under favorable conditions.

  15. Intrahepatic expression of interferon alpha & interferon alpha ...

    African Journals Online (AJOL)

    kemrilib

    Alpha m-RNA while 30% only expressed Interferon Alpha Receptor m-RNA. Responders and non-responders to Interferon therapy ... expression of IFN Alpha Receptor mRNA. Regardless of the response to interferon, histological .... generation reverse hybridisation, line probe assay. (Inno-LiPA HCV II; Innogenetics, Ghent,.

  16. Implantation Serine Proteinases heterodimerize and are critical in hatching and implantation

    Directory of Open Access Journals (Sweden)

    Meng Guoliang

    2006-12-01

    Full Text Available Abstract Background We have recently reported the expression of murine Implantation Serine Proteinase genes in pre-implantation embryos (ISP1 and uterus (ISP1 and ISP2. These proteinases belong to the S1 proteinase family and are similar to mast cell tryptases, which function as multimers. Results Here, we report the purification and initial characterization of ISP1 and 2 with respect to their physico-chemical properties and physiological function. In addition to being co-expressed in uterus, we show that ISP1 and ISP2 are also co-expressed in the pre-implantation embryo. Together, they form a heterodimer with an approximate molecular weight of 63 kD. This complex is the active form of the enzyme, which we have further characterized as being trypsin-like, based on substrate and inhibitor specificities. In addition to having a role in embryo hatching and outgrowth, we demonstrate that ISP enzyme is localized to the site of embryo invasion during implantation and that its activity is important for successful implantation in vivo. Conclusion On the basis of similarities in structural, chemical, and functional properties, we suggest that this ISP enzyme complex represents the classical hatching enzyme, strypsin. Our results demonstrate a critical role for ISP in embryo hatching and implantation.

  17. Fibronectin-degrading activity of Trypanosoma cruzi cysteine proteinase plays a role in host cell invasion.

    Science.gov (United States)

    Maeda, Fernando Yukio; Cortez, Cristian; Izidoro, Mario Augusto; Juliano, Luiz; Yoshida, Nobuko

    2014-12-01

    Trypanosoma cruzi, the agent of Chagas disease, binds to diverse extracellular matrix proteins. Such an ability prevails in the parasite forms that circulate in the bloodstream and contributes to host cell invasion. Whether this also applies to the insect-stage metacyclic trypomastigotes, the developmental forms that initiate infection in the mammalian host, is not clear. Using T. cruzi CL strain metacyclic forms, we investigated whether fibronectin bound to the parasites and affected target cell invasion. Fibronectin present in cell culture medium bound to metacyclic forms and was digested by cruzipain, the major T. cruzi cysteine proteinase. G strain, with negligible cruzipain activity, displayed a minimal fibronectin-degrading effect. Binding to fibronectin was mediated by gp82, the metacyclic stage-specific surface molecule implicated in parasite internalization. When exogenous fibronectin was present at concentrations higher than cruzipain can properly digest, or fibronectin expression was stimulated by treatment of epithelial HeLa cells with transforming growth factor beta, the parasite invasion was reduced. Treatment of HeLa cells with purified recombinant cruzipain increased parasite internalization, whereas the treatment of parasites with cysteine proteinase inhibitor had the opposite effect. Metacyclic trypomastigote entry into HeLa cells was not affected by anti-β1 integrin antibody but was inhibited by anti-fibronectin antibody. Overall, our results have indicated that the cysteine proteinase of T. cruzi metacyclic forms, through its fibronectin-degrading activity, is implicated in host cell invasion. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  18. Cell-matrix interactions: focus on proteoglycan-proteinase interplay and pharmacological targeting in cancer.

    Science.gov (United States)

    Theocharis, Achilleas D; Gialeli, Chrisostomi; Bouris, Panagiotis; Giannopoulou, Efstathia; Skandalis, Spyros S; Aletras, Alexios J; Iozzo, Renato V; Karamanos, Nikos K

    2014-11-01

    Proteoglycans are major constituents of extracellular matrices, as well as cell surfaces and basement membranes. They play key roles in supporting the dynamic extracellular matrix by generating complex structural networks with other macromolecules and by regulating cellular phenotypes and signaling. It is becoming evident, however, that proteolytic enzymes are required partners for matrix remodeling and for modulating cell signaling via matrix constituents. Proteinases contribute to all stages of diseases, particularly cancer development and progression, and contextually participate in either the removal of damaged products or in the processing of matrix molecules and signaling receptors. The dynamic interplay between proteoglycans and proteolytic enzymes is a crucial biological step that contributes to the pathophysiology of cancer and inflammation. Moreover, proteoglycans are implicated in the expression and secretion of proteolytic enzymes and often modulate their activities. In this review, we describe the emerging biological roles of proteoglycans and proteinases, with a special emphasis on their complex interplay. We critically evaluate this important proteoglycan-proteinase interactome and discuss future challenges with respect to targeting this axis in the treatment of cancer. © 2014 FEBS.

  19. Snake venom serine proteinases specificity mapping by proteomic identification of cleavage sites.

    Science.gov (United States)

    Zelanis, André; Huesgen, Pitter F; Oliveira, Ana Karina; Tashima, Alexandre K; Serrano, Solange M T; Overall, Christopher M

    2015-01-15

    Many snake venom toxins are serine proteases but their specific in vivo targets are mostly unknown. Various act on components of the coagulation cascade, and fibrinolytic and kallikrein-kinin systems to trigger various pathological effects observed in the envenomation. Despite showing high similarity in terms of primary structure snake venom serine proteinases (SVSPs) show exquisite specificity towards macromolecular substrates. Therefore, the characterization of their peptide bond specificity is important for understanding the active site preference associated with effective proteolysis as well as for the design of peptide substrates and inhibitors. Bothrops jararaca contains various SVSPs among which Bothrops protease A is a specific fibrinogenolytic agent and PA-BJ is a platelet-activating enzyme. In this study we used proteome derived peptide libraries in the Proteomic Identification of protease Cleavage Sites (PICS) approach to explore the peptide bond specificity of Bothrops protease A and PA-BJ in order to determine their individual peptide cleavage sequences. A total of 371 cleavage sites (208 for Bothrops protease A and 163 for PA-BJ) were detected and both proteinases displayed a clear preference for arginine at the P1 position. Moreover, the analysis of the specificity profiles of Bothrops protease A and PA-BJ revealed subtle differences in the preferences along P6-P6', despite a common yet unusual preference for Pro at P2. Taken together, these results map the subsite specificity of both SVSPs and shed light in the functional differences between these proteinases. Proteolysis is key to various pathological effects observed upon envenomation by viperid snakes. The use of the Proteomic Identification of protease Cleavage Sites (PICS) approach for the easy mapping of proteinase subsite preferences at both the prime- and non-prime sides concurrently gives rise to a fresh understanding of the interaction of the snake venom serine proteinases with peptide and

  20. [Phospholipase and proteinase production by Malassezia pachydermatis isolated in dogs with and without otitis].

    Science.gov (United States)

    Ortiz, Gustavo; Martín, M Carmen; Carrillo-Muñoz, Alfonso J; Payá, M Jesús

    2013-01-01

    Malassezia pachydermatis is part of the skin microbiota of dogs and cats. M. pachydermatis has been associated with external otitis and seborrhoeic dermatitis, reported more often in dogs than in cats. When the physical, chemical or immunological mechanisms of the skin are altered, M. pachydermatis could act as a pathogen. Thus, several virulence factors, such as the ability to produce esterase, lipase, lipoxygenase, protease, chondroitin sulphatase, and hyaluronidase, have been studied. In the present study, we aim to identify the phospholipase activity measured at pH 6.3, and the proteinase activity measured at pH 6.3 and pH 6.8 (pH from ears of dogs with external otitis) of M. pachydermatis strains isolated from dogs with and without external otitis. The phospholipase activity was measured using a semi-quantitative method with egg yolk, and the proteinase activity with a semi-quantitative method using bovine serum albumin agar. The study was performed on 96 isolates of M. pachydermatis, 43 isolated from dogs without clinical symptoms of otitis, and 52 isolated from dogs with otitis. In our study, 75.8% of the isolates showed phospholipase activity at pH 6.3, and 81 and 97.9% of them showed proteinase activity measured at pH 6.3 and 6.8, respectively. A higher phospholipase activity was detected in strains isolated from dogs with otitis. The proteinase activity was increased at a pH of 6.8 (97.9%) in comparison to a pH of 6.3 (81%). Our results suggest that the phospholipase activity may play an important role in the invasion of host tissues in chronic canine otitis cases. The proteinase activity results obtained in this study suggest that a reduction in the pH of the treatment may improve its efficacy in the resolution of M. pachydermatis otitis. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  1. The H1 histone-specific proteinase is associated with nuclear matrix and stimulated by DNA containing breaks of denatured sites

    International Nuclear Information System (INIS)

    Gaziev, A.I.; Kutsyj, M.P.

    1988-01-01

    Discovery of proteinase in nuclear matrix specific of H1 histone and dependent presence of breaks or denatured sites in DNA permits to assume that the given enzyme, obviously, participates in replication and DNA repair, in regulation of genes expression. Removal of H1 histone by proteinase is, probably, necessary for procedure of these processes, and, obviously, this proteinase suffers conformational changes in the composition of the DNA-histone complex. H1 histone disintegration in nucleohistone containing damaged sites of DNA by specific proteinase, probably, represents one of the mechanisms for providing DNA repair in cells of higher organisms

  2. Kazal-type proteinase inhibitor from disk abalone (Haliotis discus discus): molecular characterization and transcriptional response upon immune stimulation.

    Science.gov (United States)

    Wickramaarachchi, W D Niroshana; De Zoysa, Mahanama; Whang, Ilson; Wan, Qiang; Lee, Jehee

    2013-09-01

    Proteinases and proteinase inhibitors are involved in several biological and physiological processes in all multicellular organisms. Proteinase inhibitors play a key role in regulating the activity of the respective proteinases. Among serine proteinase inhibitors, kazal-type proteinase inhibitors (KPIs) are widely found in mammals, avians, and a variety of invertebrates. In this study, we describe the identification of a kazal-type serine proteinase inhibitor (Ab-KPI) from the disk abalone, Haliotis discus discus, which is presumably involved in innate immunity. The full-length cDNA of Ab-KPI includes 600 bp nucleotides with an open reading frame (ORF) encoding a polypeptide of 143 amino acids. The deduced amino acid sequence of Ab-KPI contains a putative 17-amino acid signal peptide and two tandem kazal domains with high similarity to other kazal-type SPIs. Each kazal domain consists of reactive site (P1) residue containing a leucine (L), and a threonine (T) located in the second amino acid position after the second conserved cysteine of each domain. Temporal expression of Ab-KPI was assessed by real time quantitative PCR in hemocytes and mantle tissue following bacterial and viral hemorrhagic septicemia virus (VHSV) challenge, and tissue injury. At 6 h post-bacterial and -VHSV challenge, Ab-KPI expression in hemocytes was increased 14-fold and 4-fold, respectively, compared to control samples. The highest up-regulations upon tissue injury were shown at 9 h and 12 h in hemocytes and mantle, respectively. The transcriptional modulation of Ab-KPI following bacterial and viral challenges and tissue injury indicates that it might be involved in immune defense as well as wound healing process in abalone. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Comparison of self-processing of foot-and-mouth disease virus leader proteinase and porcine reproductive and respiratory syndrome virus leader proteinase nsp1α

    Energy Technology Data Exchange (ETDEWEB)

    Steinberger, Jutta [Max F. Perutz Laboratories, Medical University of Vienna, Department of Medical Biochemistry, Dr. Bohr-Gasse 9/3, A-1030 Vienna (Austria); Kontaxis, Georg [Max F. Perutz Laboratories, University of Vienna, Department of Structural and Computational Biology, Campus Vienna Biocenter 5, A-1030 Vienna (Austria); Rancan, Chiara [Helmholtz Zentrum München, Department of Gene Vectors, Haematologikum, Marchioninistrasse 25, D-81377 Munich (Germany); Skern, Tim, E-mail: timothy.skern@meduniwien.ac.at [Max F. Perutz Laboratories, Medical University of Vienna, Department of Medical Biochemistry, Dr. Bohr-Gasse 9/3, A-1030 Vienna (Austria)

    2013-09-01

    The foot-and-mouth disease virus leader proteinase (Lb{sup pro}) cleaves itself off the nascent viral polyprotein. NMR studies on the monomeric variant Lb{sup pro} L200F provide structural evidence for intramolecular self-processing. {sup 15}N-HSQC measurements of Lb{sup pro} L200F showed specifically shifted backbone signals in the active and substrate binding sites compared to the monomeric variant sLb{sup pro}, lacking six C-terminal residues. This indicates transient intramolecular interactions between the C-terminal extension (CTE) of one molecule and its own active site. Contrastingly, the porcine reproductive and respiratory syndrome virus (PRRSV) leader proteinase nsp1α, with a papain-like fold like Lb{sup pro}, stably binds its own CTE. Parts of the β-sheet domains but none of the α-helical domains of Lb{sup pro} and nsp1α superimpose; consequently, the α-helical domain of nsp1α is oriented differently relative to its β-sheet domain. This provides a large interaction surface for the CTE with the globular domain, stabilising the intramolecular complex. Consequently, self-processing inactivates nsp1α but not Lb{sup pro}. - Highlights: • We examine self-processing of the leader protease of foot-and-mouth disease virus. • NMR analysis strongly supports intramolecular self-processing. • Self-processing is a dynamic process with no stable complex. • Structural comparison with nsp1α of PRRSV which forms stable intramolecular complex. • Subdomain orientation explains differences in stability of intramolecular complexes.

  4. Comparison of self-processing of foot-and-mouth disease virus leader proteinase and porcine reproductive and respiratory syndrome virus leader proteinase nsp1α

    International Nuclear Information System (INIS)

    Steinberger, Jutta; Kontaxis, Georg; Rancan, Chiara; Skern, Tim

    2013-01-01

    The foot-and-mouth disease virus leader proteinase (Lb pro ) cleaves itself off the nascent viral polyprotein. NMR studies on the monomeric variant Lb pro L200F provide structural evidence for intramolecular self-processing. 15 N-HSQC measurements of Lb pro L200F showed specifically shifted backbone signals in the active and substrate binding sites compared to the monomeric variant sLb pro , lacking six C-terminal residues. This indicates transient intramolecular interactions between the C-terminal extension (CTE) of one molecule and its own active site. Contrastingly, the porcine reproductive and respiratory syndrome virus (PRRSV) leader proteinase nsp1α, with a papain-like fold like Lb pro , stably binds its own CTE. Parts of the β-sheet domains but none of the α-helical domains of Lb pro and nsp1α superimpose; consequently, the α-helical domain of nsp1α is oriented differently relative to its β-sheet domain. This provides a large interaction surface for the CTE with the globular domain, stabilising the intramolecular complex. Consequently, self-processing inactivates nsp1α but not Lb pro . - Highlights: • We examine self-processing of the leader protease of foot-and-mouth disease virus. • NMR analysis strongly supports intramolecular self-processing. • Self-processing is a dynamic process with no stable complex. • Structural comparison with nsp1α of PRRSV which forms stable intramolecular complex. • Subdomain orientation explains differences in stability of intramolecular complexes

  5. Purification, characterization and cloning of an aspartic proteinase inhibitor from squash phloem exudate.

    Science.gov (United States)

    Christeller, J T; Farley, P C; Ramsay, R J; Sullivan, P A; Laing, W A

    1998-05-15

    Phloem exudate from squash fruit contains heat-inactivated material which inhibits pepsin activity. This inhibitory activity was purified by mild acid treatment, chromatography on trypsin-agarose, Sephadex G-75 and reverse-phase HPLC, resulting in the elution of three peaks with pepsin-inhibitory activity. N-terminal sequencing indicated a common sequence of MGPGPAIGEVIG and the presence of minor species with seven- or two-amino-acid N-terminal extensions beyond this point. Microheterogeneity in this end sequence was exhibited within and between two preparations. Internal sequencing of a major peak after a trypsin digestion gave the sequence FYNVVVLEK. The common N-terminal sequence was used to design a degenerate primer for 3' rapid amplification of cDNA ends and cDNA clones encoding two isoforms of the inhibitor were obtained. The open reading frames of both cDNAs encoded proteins (96% identical) which contained the experimentally determined internal sequence. The amino acid content calculated from the predicted amino acid sequence was very similar to that measured by amino acid analysis of the purified inhibitor. The two predicted amino acid sequences (96 residues) had neither similarity to any other aspartic proteinase inhibitor nor similarity to any other protein. The inhibitors have a molecular mass of 10,552 Da, measured by matrix-assisted laser-desorption ionisation time-of-flight mass spectrometry and approximately 10,000 Da by SDS/PAGE, and behave as dimers of approximately 21,000 Da during chromatography on Superdex G-75 gel-filtration medium. The calculated molecular masses from the predicted amino acid sequences were 10,551 Da and 10,527 Da. The inhibitor was capable of inhibiting pepsin (Ki = 2 nM) and a secreted aspartic proteinase from the fungus Glomerella cingulata (Ki = 20 nM). The inhibitor, which is stable over acid and neutral pH, has been named squash aspartic proteinase inhibitor (SQAPI).

  6. Processing, activity, and inhibition of recombinant cyprosin, an aspartic proteinase from cardoon (Cynara cardunculus).

    Science.gov (United States)

    White, P C; Cordeiro, M C; Arnold, D; Brodelius, P E; Kay, J

    1999-06-11

    The cDNA encoding the precursor of an aspartic proteinase from the flowers of the cardoon, Cynara cardunculus, was expressed in Pichia pastoris, and the recombinant, mature cyprosin that accumulated in the culture medium was purified and characterized. The resultant mixture of microheterogeneous forms was shown to consist of glycosylated heavy chains (34 or 32 kDa) plus associated light chains with molecular weights in the region of 14,000-18,000, resulting from excision of most, but not all, of the 104 residues contributed by the unique region known as the plant specific insert. SDS-polyacrylamide gel electrophoresis under non-reducing conditions indicated that disulfide bonding held the heavy and light chains together in the heterodimeric enzyme forms. In contrast, when a construct was expressed in which the nucleotides encoding the 104 residues of the plant specific insert were deleted, the inactive, unprocessed precursor form (procyprosin) accumulated, indicating that the plant-specific insert has a role in ensuring that the nascent polypeptide is folded properly and rendered capable of being activated to generate mature, active proteinase. Kinetic parameters were derived for the hydrolysis of a synthetic peptide substrate by wild-type, recombinant cyprosin at a variety of pH and temperature values and the subsite requirements of the enzyme were mapped using a systematic series of synthetic inhibitors. The significance is discussed of the susceptibility of cyprosin to inhibitors of human immunodeficiency virus proteinase and particularly of renin, some of which were found to have subnanomolar potencies against the plant enzyme.

  7. Effect of oral antiseptic agents on phospholipase and proteinase enzymes of Candida albicans.

    Science.gov (United States)

    Uygun-Can, Banu; Kadir, Tanju; Gumru, Birsay

    2016-02-01

    Candida-associated denture stomatitis is the most prevalent form of oral candida infections among the denture wearers. Generally, antiseptic oral rinses used in the treatment of these infections are considered as an adjunct or alternative antifungal treatment. Studies have suggested that the intraoral concentrations of antiseptics decrease substantially to the sub-therapeutic levels on account of the dynamics of the oral cavity. This condition yields the question about the minimum antiseptic concentration that effect the character or pathogenesis of Candida during treatment. The extracellular phospholipase and proteinase enzymes of Candida albicans are regarded to have a crucial role in the pathogenesis of human fungal infections. Therefore, the aim of this study was to investigate the effect of different sub-therapeutic concentrations of chlorhexidine gluconate, hexetidine and triclosan on the production of these enzymes by C. albicans strains isolated from 20 patients with denture stomatitis. Phospholipase test was done by using Sabouraud dextrose agar with egg yolk, proteinase test was done by using bovine serum albumin agar. Phospholipase test was done by using Sabouraud dextrose agar with egg yolk, proteinase test was done by using bovine serum albumin agar. Exoenzyme production of 20 strains which were brief exposured to sub-therapeutic concentrations of three antiseptic agents decreased significantly compared with the strains that were not exposured with antiseptic values (pantiseptics (pantiseptic was compared, there were no significant differences between enzymatic activities (p>0.05). The results of this study show that sub-therapeutic levels of each antiseptic may modulate candidal exoenzyme production, consequently suppressing pathogenicity of C. albicans. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. X-ray, neutron and NMR studies of the catalytic mechanism of aspartic proteinases.

    Science.gov (United States)

    Coates, Leighton; Erskine, Peter T; Mall, Sanjay; Gill, Raj; Wood, Steve P; Myles, Dean A A; Cooper, Jonathan B

    2006-09-01

    Current proposals for the catalytic mechanism of aspartic proteinases are largely based on X-ray structures of bound oligopeptide inhibitors possessing non-hydrolysable analogues of the scissile peptide bond. Until recent years, the positions of protons on the catalytic aspartates and the ligand in these complexes had not been determined with certainty due to the inadequate resolution of these analyses. There has been much interest in locating the catalytic protons at the active site of aspartic proteinases since this has major implications for detailed understanding of the mechanism of action and the design of improved transition state mimics for therapeutic applications. In this review we discuss the results of studies which have shed light on the locations of protons at the catalytic centre. The first direct determination of the proton positions stemmed from neutron diffraction data collected from crystals of the fungal aspartic proteinase endothiapepsin bound to a transition state analogue (H261). The neutron structure of the complex at a resolution of 2.1 A provided evidence that Asp 215 is protonated and that Asp 32 is the negatively charged residue in the transition state complex. Atomic resolution X-ray studies of inhibitor complexes have corroborated this finding. A similar study of the native enzyme established that it, unexpectedly, has a dipeptide bound at the catalytic site which is consistent with classical reports of inhibition by short peptides and the ability of pepsins to catalyse transpeptidation reactions. Studies by NMR have confirmed the findings of low-barrier and single-well hydrogen bonds in the complexes with transition state analogues.

  9. Protective role of purified cysteine proteinases against Fasciola gigantica infection in experimental animals.

    Science.gov (United States)

    El-Ahwany, Eman; Rabia, Ibrahim; Nagy, Faten; Zoheiry, Mona; Diab, Tarek; Zada, Suher

    2012-03-01

    Fascioliasis is one of the public health problems in the world. Cysteine proteinases (CP) released by Fasciola gigantica play a key role in parasite feeding, migration through host tissues, and in immune evasion. There has been some evidence from several parasite systems that proteinases might have potential as protective antigens against parasitic infections. Cysteine proteinases were purified and tested in vaccine trials of sheep infected with the liver fluke. Multiple doses (2 mg of CP in Freund's adjuvant followed by 3 booster doses 1 mg each at 4 week intervals) were injected intramuscularly into sheep 1 week prior to infect orally with 300 F. gigantica metacercariae. All the sheep were humanely slaughtered 12 weeks after the first immunization. Changes in the worm burden, ova count, and humoral and cellular responses were evaluated. Significant reduction was observed in the worm burden (56.9%), bile egg count (70.7%), and fecel egg count (75.2%). Immunization with CP was also found to be associated with increases of total IgG, IgG(1), and IgG(2) (P<0.05). Data showed that the serum cytokine levels of pro-inflammatory cytokines, IL-12, IFN-γ, and TNF-α, revealed significant decreases (P<0.05). However, the anti-inflammatory cytokine levels, IL-10, TGF-β, and IL-6, showed significant increases (P<0.05). In conclusion, it has been found that CP released by F. gigantica are highly important candidates for a vaccine antigen because of their role in the fluke biology and host-parasite relationships.

  10. Two distinct phases of apoptosis in mammary gland involution: proteinase-independent and -dependent pathways

    Energy Technology Data Exchange (ETDEWEB)

    Lund, Leif R; Romer, John; Thomasset, Nicole; Solberg, Helene; Pyke, Charles; Bissell, Mina J; Dano, Keld; Werb, Zena

    1996-01-01

    Postlactational involution of the mammary gland is characterized by two distinct physiological events: apoptosis of the secretory, epithelial cells undergoing programmed cell death, and proteolytic degradation of the mammary gland basement membrane. We examined the spatial and temporal patterns of apoptotic cells in relation to those of proteinases during involution of the BALB/c mouse mammary gland. Apoptosis was almost absent during lactation but became evident at day 2 of involution, when {beta}-casein gene expression was still high. Apoptotic cells were then seen at least up to day 8 of involution, when {beta}-casein gene expression was being extinguished. Expression of sulfated glycoprotein-2 (SGP-2), interleukin-1{beta} converting enzyme (ICE) and tissue inhibitor of metalloproteinases-1 was upregulated at day 2, when apoptotic cells were seen initially. Expression of the matrix metalloproteinases gelatinase A and stromelysin-1 and the serine proteinase urokinase-type plasminogen activator, which was low during lactation, was strongly upregulated in parallel starting at day 4 after weaning, coinciding with start of the collapse of the lobulo-alveolar structures and the intensive tissue remodeling in involution. The major sites of mRNA synthesis for these proteinases were fibroblast-like cells in the periductal stroma and stromal cells surrounding the collapsed alveoli, suggesting that the degradative phase of involution is due to a specialized mesenchymal-epithelial interaction. To elucidate the functional role of these proteinases during involution, at the onset of weaning we treated mice systemically with the glucocorticoid hydrocortisone, which is known to inhibit mammary gland involution. Although the initial wave of apoptotic cells appeared in the lumina of the gland, the dramatic regression and tissue remodeling usually evident by day 5 was substantially inhibited by systemic treatment with hydrocortisone. mRNA and protein for gelatinase A, stromelysin

  11. Characterization of kininogenase activity of an acidic proteinase isolated from human kidney

    OpenAIRE

    Gomes, RAS; Juliano, L. [UNIFESP; Chagas, JR [UNIFESP; Hial, V

    1997-01-01

    An acidic proteinase was purified from human kidney cortex. the enzyme showed a molecular mass of 31 kDa by SDS-PAGE, 36 kDa by gel filtration, and isoelectric points of 5.2 and 6.1. the optimum pH for hydrolysis of bovine hemoglobin was about 3.5. Reverse-phase KPLC analysis of the incubation mixture of the enzyme with human plasma showed the presence of an active peptide on rat uterus muscle with the same retention time as the methionyl-lysyl-bradykinin (MLBK) standard. the specific activit...

  12. Purification, crystallization and preliminary crystallographic studies of a Kunitz-type proteinase inhibitor from tamarind (Tamarindus indica) seeds.

    Science.gov (United States)

    Patil, Dipak N; Chaudhry, Anshul; Sharma, Ashwani K; Tomar, Shailly; Kumar, Pravindra

    2009-07-01

    A Kunitz-type proteinase inhibitor has been purified from tamarind (Tamarindus indica) seeds. SDS-PAGE analysis of a purified sample showed a homogeneous band corresponding to a molecular weight of 21 kDa. The protein was identified as a Kunitz-type proteinase inhibitor based on N-terminal amino-acid sequence analysis. It was crystallized by the vapour-diffusion method using PEG 6000. The crystals belonged to the orthorhombic space group C222(1), with unit-cell parameters a = 37.2, b = 77.1, c = 129.1 A. Diffraction data were collected to a resolution of 2.7 A. Preliminary crystallographic analysis indicated the presence of one proteinase inhibitor molecule in the asymmetric unit, with a solvent content of 44%.

  13. Comparison of ACE inhibitory activity in skimmed goat and cow milk hydrolyzed by alcalase, flavourzyme, neutral protease and proteinase K

    Directory of Open Access Journals (Sweden)

    Bao Chunju

    2016-06-01

    Full Text Available Angiotensin I converting enzyme (ACE inhibitory peptides derived from milk proteins have obvious effect of lowering blood pressure, safe and non-toxic side effects. This study compared four commercial proteases, namely alcalase, flavourzyme, neutral protease and proteinase K for their ACE inhibitory activity in skimmed goat and cow milk and identified the best one with higher ACE inhibitory activity. The degree of hydrolysis (DH of alcalase and proteinase K were much higher than flavourzyme, neutral protease for both skimmed goat and cow milk. Alcalase was the best enzyme to produce ACE inhibitory peptides from goat milk, with the ACE inhibitory activity 95.31%, while proteinase K was the optimal protease for hydrolyzing cow milk, with 81.28% ACE inhibitory activity. Furthermore, no correlation was obtained between the ACE inhibitory activity and DH for both goat and cow milk.

  14. Expression of glutamate transporter, GABRA6, serine proteinase inhibitor 2 and low levels of glutamate and GABA in the brain of knock-out mouse for Canavan disease.

    Science.gov (United States)

    Surendran, Sankar; Rady, Peter L; Michals-Matalon, Kimberlee; Quast, Michael J; Rassin, David K; Campbell, Gerald A; Ezell, Ed L; Wei, Jingna; Tyring, Stephen K; Szucs, Sylvia; Matalon, Reuben

    2003-08-30

    Canavan disease (CD) is an autosomal recessive leukodystrophy characterized by spongy degeneration of the brain. The clinical features of CD are hypotonia, megalencephaly, and mental retardation leading to early death. While aspartoacylase (ASPA) activity increases with age in the wild type mouse brain, there is no ASPA activity in the CD mouse brain. So far ASPA deficiency and elevated NAA have been ascribed with the CD. Other factors affecting the brain that result from ASPA deficiency may lead pathophysiology of CD. The NMR spectra and amino acid analysis showed lower levels of glutamate and gamma-aminobutyric acid in the CD mouse brain compared to the wild type. Microarray gene expression on CD mouse brain showed glutamate transporter-EAAT4 and gamma-aminobutyric acid-A receptor, subunit alpha6 (GABRA6) were lower 9.7- and 119.1-fold, respectively. Serine proteinase inhibitor 2 (Spi2) was 29.9-fold higher in the CD mouse brain compared to the wild type. The decrease of GABRA6 and high expression of Spi2 in CD mouse brain were also confirmed by real-time RT-PCR. This first report showing abnormal expression of EAAT4, GABRA6, Spi2 combined with lower levels of glutamate and GABA are likely to be associated with the pathophysiology of CD.

  15. Functional characterization of single-domain cystatin-like cysteine proteinase inhibitors expressed by the trematode Fasciola hepatica.

    Science.gov (United States)

    Cancela, Martín; Corvo, Ileana; DA Silva, Edileuza; Teichmann, Aline; Roche, Leda; Díaz, Alvaro; Tort, José Fransisco; Ferreira, Henrique B; Zaha, Arnaldo

    2017-11-01

    Cystatins are small, phylogenetically conserved proteins that are tight-binding inhibitors of cysteine proteinases. The liver fluke Fasciola hepatica uses a diverse set of cysteine proteinases of the papain superfamily for host invasion, immune evasion and nutrition, but little is known about the regulation of these enzymes. The aim of this work is to characterize the cystatin repertoire of F. hepatica. For this purpose, we first surveyed the available sequence databases, identifying three different F. hepatica single-domain cystatins. In agreement with the in silico predictions, at least three small proteins with cysteine proteinase binding activity were identified. Phylogenetic analyses showed that the three cystatins (named FhStf-1, -2 and -3) are members of the I25A subfamily (stefins). Whereas FhStf-1 grouped with classical stefins, FhStf-2 and 3 fell in a divergent stefin subgroup unusually featuring signal peptides. Recombinant rFhStf-1, -2 and -3 had potent inhibitory activity against F. hepatica cathepsin L cysteine proteinases but differed in their capacity to inhibit mammalian cathepsin B, L and C. FhStf-1 was localized in the F. hepatica reproductive organs (testes and ovary), and at the surface lamella of the adult gut, where it may regulate cysteine proteinases related with reproduction and digestion, respectively. FhStf-1 was also detected among F. hepatica excretion-secretion (E/S) products of adult flukes. This suggests that it is secreted by non-classical secretory pathway and that it may interact with host lysosomal cysteine proteinases.

  16. Brewer's spent grain and corn steep liquor as alternative culture medium substrates for proteinase production by Streptomyces malaysiensis AMT-3.

    Science.gov (United States)

    do Nascimento, Rodrigo Pires; Junior, Nelson Alves; Coelho, Rosalie Reed Rodrigues

    2011-10-01

    Brewer's spent grain and corn steep liquor or yeast extract were used as the sole organic forms for proteinase production by Streptomyces malaysiensis in submerged fermentation. The influence of the C and N concentrations, as well as the incubation periods, were assessed. Eight proteolytic bands were detected through gelatin-gel-electrophoresis in the various extracts obtained from the different media and after different incubation periods, with apparent molecular masses of 20, 35, 43, 50, 70, 100, 116 and 212 kDa. The results obtained suggest an opportunity for exploring this alternative strategy for proteinases production by actinomycetes, using BSG and CSL as economically feasible substrates.

  17. Brewer's spent grain and corn steep liquor as alternative culture medium substrates for proteinase production by Streptomyces malaysiensis AMT-3

    Directory of Open Access Journals (Sweden)

    Rodrigo Pires do Nascimento

    2011-12-01

    Full Text Available Brewer's spent grain and corn steep liquor or yeast extract were used as the sole organic forms for proteinase production by Streptomyces malaysiensis in submerged fermentation. The influence of the C and N concentrations, as well as the incubation periods, were assessed. Eight proteolytic bands were detected through gelatin-gel-electrophoresis in the various extracts obtained from the different media and after different incubation periods, with apparent molecular masses of 20, 35, 43, 50, 70, 100, 116 and 212 kDa. The results obtained suggest an opportunity for exploring this alternative strategy for proteinases production by actinomycetes, using BSG and CSL as economically feasible substrates.

  18. Diversity in proteinase specificity of thermophilic lactobacilli as revealed by hydrolysis of dairy and vegetable proteins.

    Science.gov (United States)

    Pescuma, Micaela; Espeche Turbay, María Beatriz; Mozzi, Fernanda; Font de Valdez, Graciela; Savoy de Giori, Graciela; Hebert, Elvira María

    2013-09-01

    Ability of industrially relevant species of thermophilic lactobacilli strains to hydrolyze proteins from animal (caseins and β-lactoglobulin) and vegetable (soybean and wheat) sources, as well as influence of peptide content of growth medium on cell envelope-associated proteinase (CEP) activity, was evaluated. Lactobacillus delbrueckii subsp. lactis (CRL 581 and 654), L. delbrueckii subsp. bulgaricus (CRL 454 and 656), Lactobacillus acidophilus (CRL 636 and 1063), and Lactobacillus helveticus (CRL 1062 and 1177) were grown in a chemically defined medium supplemented or not with 1 % Casitone. All strains hydrolyzed mainly β-casein, while degradation of αs-caseins was strain dependent. Contrariwise, κ-Casein was poorly degraded by the studied lactobacilli. β-Lactoglobulin was mainly hydrolyzed by CRL 656, CRL 636, and CRL 1062 strains. The L. delbrueckii subsp. lactis strains, L. delbrueckii subsp. bulgaricus CRL 656, and L. helveticus CRL 1177 degraded gliadins in high extent, while the L. acidophilus and L. helveticus strains highly hydrolyzed soy proteins. Proteinase production was inhibited by Casitone, the most affected being the L. delbrueckii subsp. lactis species. This study highlights the importance of proteolytic diversity of lactobacilli for rational strain selection when formulating hydrolyzed dairy or vegetable food products.

  19. Modified TB rapid test by proteinase K for rapid diagnosis of pleural tuberculosis.

    Science.gov (United States)

    Yari, Shamsi; Hadizadeh Tasbiti, Alireza; Ghanei, Mostafa; Shokrgozar, Mohammad Ali; Fateh, Abolfazl; Yari, Fatemeh; Bahrmand, Ahmadreza

    2016-03-01

    The diagnosis of pleural tuberculosis continues to be a challenge due to the low sensitivity of traditional diagnostic methods. Better and more rapid tests are needed for diagnosis of pleural TB. In this study, pleural fluids were tested with rapid test to determine Mycobacterium tuberculosis (MTB antigen). Affinity chromatography was used to purify specific polyclonal antibodies against MTB antigen. Pleural samples after decontamination were treated with proteinase K. Rapid test for pleural fluids was prepared by specific antibody. Rapid test was performed on 85 pleural fluid patients. The patients had a mean age of 46.55 ± 15.96 years and 38 were men. The performance of rapid test, using proteinase K, was found to be the most impressive: sensitivity 93%, specificity 94%, PPV 90%, and NPV 96% compared with adenosine deaminase test (ADA), PCR, smear, and culture. The present study did demonstrate that modified TB rapid test can substantially improve the diagnosis of extrapulmonary TB. © 2015 APMIS. Published by John Wiley & Sons Ltd.

  20. A single amino acid substitution affects substrate specificity in cysteine proteinases from Fasciola hepatica.

    Science.gov (United States)

    Smooker, P. M.; Whisstock, J. C.; Irving, J. A.; Siyaguna, S.; Spithill, T. W.; Pike, R. N.

    2000-01-01

    The trematode Fasciola hepatica secretes a number of cathepsin L-like proteases that are proposed to be involved in feeding, migration, and immune evasion by the parasite. To date, six full cDNA sequences encoding cathepsin L preproproteins have been identified. Previous studies have demonstrated that one of these cathepsins (L2) is unusual in that it is able to cleave substrates with a proline in the P2 position, translating into an unusual ability (for a cysteine proteinase) to clot fibrinogen. In this study, we report the sequence of a novel cathepsin (L5) and compare the substrate specificity of a recombinant enzyme with that of recombinant cathepsin L2. Despite sharing 80% sequence identity with cathepsin L2, cathepsin L5 does not exhibit substantial catalytic activity against substrates containing proline in the P2 position. Molecular modeling studies suggested that a single amino acid change (L69Y) in the mature proteinases may account for the difference in specificity at the S2 subsite. Recombinant cathepsin L5/L69Y was expressed in yeast and a substantial increase in the ability of this variant to accommodate substrates with a proline residue in the P2 position was observed. Thus, we have identified a single amino acid substitution that can substantially influence the architecture of the S2 subsite of F. hepatica cathepsin L proteases. PMID:11206078

  1. Identification of Placental Aspartic Proteinase in the Eurasian Beaver (Castor fiber L.).

    Science.gov (United States)

    Lipka, Aleksandra; Panasiewicz, Grzegorz; Majewska, Marta; Paukszto, Lukasz; Bieniek-Kobuszewska, Martyna; Szafranska, Bozena

    2018-04-18

    Aspartic proteinases (AP) form a multigenic group widely distributed in various organisms and includes pepsins (pep), cathepsins D and E, pregnancy associated glycoproteins (PAGs) as well as plant, fungal, and retroviral proteinases. This study describes the transcript identification and expression localization of the AP within the discoid placenta of the Castor fiber . We identified 1257 bp of the AP cDNA sequence, encoding 391 amino acids (aa) of the polypeptide precursor composed of 16 aa signal peptide, 46 aa pro-piece, and 329 aa of the mature protein. Within the AP precursor, one site of potential N -glycosylation (NPS 119–121 ) and two Asp residues (D) specific for the catalytic cleft of AP were identified (VLFDTGSSNLWV 91–102 and GIVDTGTSLLTV 277–288 ). The highest homology of the identified placental AP nucleotide and aa sequence was to mouse pepsinogen C (75.8% and 70.1%, respectively). Identified AP also shared high homology with other superfamily members: PAGs, cathepsins, and napsins. The AP identified in this study was named as pepsinogen/PAG-Like (pep/PAG-L). Diversified pep/PAG-L protein profiles with a dominant 58 kDa isoform were identified. Immune reactive signals of the pep/PAG-L were localized within the trophectodermal cells of the beaver placenta. This is the first report describing the placental AP (pep/PAG-L) in the C. fiber .

  2. Intracellular localization of Treponema denticola chymotrypsin-like proteinase in chronic periodontitis

    Directory of Open Access Journals (Sweden)

    Emilia Marttila

    2014-07-01

    Full Text Available Treponema denticola is an important periodontal pathogen capable of tissue invasion. Its chymotrypsin-like proteinase (CTLP can degrade a number of basement membrane components in vitro, thus suggesting a contribution to tissue invasion by the spirochete. The aim of this study was to analyze the localization of CTLP in chronic periodontitis tissues ex vivo. A polyclonal antibody specific to T. denticola cell-bound CTLP was used to detect the spirochetes in the gingival tissues of patients with moderate to severe chronic periodontitis (n=25 by immunohistochemistry and periodic acid-Schiff staining (PAS. The presence of T. denticola in the periodontal tissue samples was analyzed by PCR. Periodontal tissue samples of 12 of the 25 patients were found to be positive for T. denticola by PCR. Moreover, CTLP could be detected in the periodontal tissues of all these patients by immunohistochemistry. In the epithelium, the CTLP was mostly intracellular. Typically, the positive staining could be seen throughout the whole depth of the epithelium. When detected extracellularly, CTLP was localized mainly as granular deposits. The connective tissue stained diffusely positive in four cases. The positive staining co-localized with the PAS stain in nine cases. T. denticola and its CTLP could be detected in diseased human periodontium both intra- and extracellularly. The granular staining pattern was suggestive of the presence of T. denticola bacteria, whereas the more diffused staining pattern was indicative of the recent presence of the bacterium and shedding of the cell-bound proteinase.

  3. TISSUE INHIBITOR OF METALLOPROTEINASE 1, MATRIX METALLOPROTEINASE 9, ALPHA-1 ANTITRYPSIN, METALLOTHIONEIN AND UROKINASE TYPE PLASMINOGEN ACTIVATOR RECEPTOR IN SKIN BIOPSIES FROM PATIENTS AFFECTED BY AUTOIMMUNE BLISTERING DISEASES

    Directory of Open Access Journals (Sweden)

    Ana Maria Abreu Velez

    2013-07-01

    Full Text Available Introduction: Proteinases and proteinase inhibitors have been described to play a role in autoimmune skin blistering diseases. We studied skin lesional biopsies from patients affected by several autoimmune skin blistering diseases for proteinases and proteinase inhibitors. Methods: We utilized immunohistochemistry to evaluate biopsies for alpha-1-antitrypsin, human matrix metalloproteinase 9 (MMP9, human tissue inhibitor of metalloproteinases 1 (TIMP-1, metallothionein and urokinase type plasminogen activator receptor (uPAR. We tested 30 patients affected by endemic pemphigus, 30 controls from the endemic area, and 15 normal controls. We also tested 30 biopsies from patients with bullous pemphigoid (BP, 20 with pemphigus vulgaris (PV, 8 with pemphigus foliaceus, and 14 with dermatitis herpetiformis (DH. Results: Contrary to findings in the current literature, most autoimmune skin blistering disease biopsies were negative for uPAR and MMP9. Only some chronic patients with El Bagre-EPF were positive to MMP9 in the dermis, in proximity to telocytes. TIMP-1 and metallothionein were positive in half of the biopsies from BP patients at the basement membrane of the skin, within several skin appendices, in areas of dermal blood vessel inflammation and within dermal mesenchymal-epithelial cell junctions.

  4. In vitro differential activity of phospholipases and acid proteinases of clinical isolates of Candida Atividade diferencial in vitro de fosfolipases e proteinases ácidas de isolados clínicos de Candida

    Directory of Open Access Journals (Sweden)

    Aurean D'Eça Júnior

    2011-06-01

    Full Text Available INTRODUCTION: Candida yeasts are commensals; however, if the balance of normal flora is disrupted or the immune defenses are compromised, Candida species can cause disease manifestations. Several attributes contribute to the virulence and pathogenicity of Candida, including the production of extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate the in vitro activity of phospholipases and acid proteinases in clinical isolates of Candida spp. METHODS: Eighty-two isolates from hospitalized patients collected from various sites of origin were analyzed. Phospholipase production was performed in egg yolk medium and the production of proteinase was verified in a medium containing bovine serum albumin. The study was performed in triplicate. RESULTS: Fifty-six (68.3% of isolates tested were phospholipase positive and 16 (44.4% were positive for proteinase activity. C. tropicalis was the species with the highest number of positive isolates for phospholipase (91.7%. Statistically significant differences were observed in relation to production of phospholipases among species (pINTRODUÇÃO: Candida são leveduras comensais, porém, se o equilíbrio da flora normal for interrompido ou as defesas imunitárias estiverem comprometidas, espécies de Candida podem causar manifestações de doença. Vários atributos contribuem na virulência e patogenicidade de Candida, inclusive a produção de enzimas extracelulares hidrolíticas, especialmente fosfolipases e proteinases. O objetivo deste estudo foi verificar a atividade in vitro de fosfolipases e proteinases ácidas em isolados clínicos de Candida spp. MÉTODOS: Oitenta e dois isolados provenientes de pacientes hospitalizados coletados a partir de sítios de origem diversos foram analisados. A produção de fosfolipase foi verificada em meio egg yolk e a de proteinase em meio contendo soro albumina bovina. O estudo foi feito em triplicata. RESULTADOS

  5. The determination of $\\alpha_s$ by the ALPHA collaboration

    CERN Document Server

    Bruno, Mattia

    2016-01-01

    We review the ALPHA collaboration strategy for obtaining the QCD coupling at high scale. In the three-flavor effective theory it avoids the use of perturbation theory at $\\alpha > 0.2$ and at the same time has the physical scales small compared to the cutoff $1/a$ in all stages of the computation. The result $\\Lambda_\\overline{MS}^{(3)}=332(14)$~MeV is translated to $\\alpha_\\overline{MS}(m_Z)=0.1179(10)(2)$ by use of (high order) perturbative relations between the effective theory couplings at the charm and beauty quark "thresholds". The error of this perturbative step is discussed and estimated as $0.0002$.

  6. Recombinant proteinase 3 (Wregener's antigen) expressed in Pichia pastoris is functionally active and is recognized by patient sera

    NARCIS (Netherlands)

    Harmsen, MC; Heeringa, P; VanderGeld, YM; Huitema, MG; Klimp, A; Tiran, A; Kallenberg, CGM

    1997-01-01

    The open reading frame of human proteinase 3 (PR3) without the prepro-peptide was cloned and expressed in Escherichia coli (rcPR3) and in Pichia pastoris (rpPR3). The 6-histidine tagged rpPR3 was efficiently secreted into culture supernatant from which it could be purified by immobilized metal

  7. The epidermal growth factor precursor in the rat kidney seems to be processed by an aprotinin sensitive proteinase

    DEFF Research Database (Denmark)

    Nexø, Ebba; Poulsen, Steen Seier; Raaberg, Lasse

    1992-01-01

    Epidermal growth factor (EGF) is synthesized as a membrane bound precursor in the rat kidney. The precursor seems to be processed by an aprotinin sensitive proteinase. Intravenous infusion of aprotinin reduces the urinary excretion of EGF by 85% and increases the amount of renal EGF. Kidney membr...

  8. Kinetic modelling of enzyme inactivation : kinetics of heat inactivation of the extracellular proteinase from Pseudomonas fluorescens 22F

    NARCIS (Netherlands)

    Schokker, E.P.

    1997-01-01

    The kinetics of heat inactivation of the extracellular proteinase from Pseudomonas fluorescens 22F was studied. It was established, by making use of kinetic modelling, that heat inactivation in the temperature range 35 - 70 °C was most likely caused

  9. Estimation of biofilm, proteinase & phospholipase production of the Candida species isolated from the oropharyngeal samples in HIV-infected patients

    Directory of Open Access Journals (Sweden)

    Vicky Lahkar

    2017-01-01

    Interpretation & conclusions: Although C. albicans was the most common Candida species identified in HIV positive patients, the emergence of NAC was of special concern. Virulence factors such as biofilms, proteinases and phospholipases were noted in both these groups. Further research is required for better understanding of the pathogenic role of Candida species so as to aid in therapeutic interventions.

  10. Rats and mice immunised with chimeric human/mouse proteinase 3 produce autoantibodies to mouse Pr3 and rat granulocytes

    NARCIS (Netherlands)

    van der Geld, Ymke M.; Hellmark, Thomas; Selga, Daina; Heeringa, Peter; Huitema, Minke G.; Limburg, Pieter C.; Kallenberg, Cees G. M.

    2007-01-01

    Aim: In this study, we employed chimeric human/ mouse Proteinase 3 ( PR3) proteins as tools to induce an autoantibody response to PR3 in rats and mice. Method: Rats and mice were immunised with recombinant human PR3 ( HPR3), recombinant murine PR3 ( mPR3), single chimeric human/ mouse PR3 ( HHm,

  11. Proteinases in excretory-secretory products of Toxocara canis second-stage larvae: zymography and modeling insights.

    Science.gov (United States)

    González-Páez, Gonzalo Ernesto; Alba-Hurtado, Fernando; García-Tovar, Carlos Gerardo; Argüello-García, Raúl

    2014-01-01

    Components released in excretory-secretory products of Toxocara canis larvae (TES) include phosphatidylethanolamine-binding proteins (TES26), mucins (TES120, MUC2-5), and C-type lectins (TES32, TES70) and their biochemical, immunological, and diagnostic properties have been extensively studied albeit proteinase activities towards physiological substrates are almost unknown. Proteolytic activities in TES samples were first analyzed by gel electrophoresis with gelatin as substrate. Major activities of ~400, 120, and 32 kDa in TES were relatively similar over a broad pH range (5.5-9.0) and all these were of the serine-type as leupeptin abolished gelatinolysis. Further, the ~400 kDa component degraded all physiological substrates tested (laminin, fibronectin, albumin, and goat IgG) and the 120 kDa component degraded albumin and goat IgG while proteinases of lower MW (45, 32, and 26 kDa) only degraded laminin and fibronectin, preferentially at alkaline pH (9.0). By protein modeling approaches using the known sequences of TES components, only TES26 and MUC4 displayed folding patterns significantly related to reference serine proteinases. These data suggest that most of serine proteinase activities secreted in vitro by infective larvae of T. canis have intriguing nature but otherwise help the parasite to affect multiple components of somatic organs and bodily fluids within the infected host.

  12. Extensive expansion of A1 family aspartic proteinases in fungi revealed by evolutionary analyses of 107 complete eukaryotic proteomes

    NARCIS (Netherlands)

    Revuelta, M.V.; Kan, van J.A.L.; Kay, J.; Have, ten A.

    2014-01-01

    The A1 family of eukaryotic aspartic proteinases (APs) forms one of the 16 AP families. Although one of the best characterized families, the recent increase in genome sequence data has revealed many fungal AP homologs with novel sequence characteristics. This study was performed to explore the

  13. Molecular cloning and functional characterisation of a cathepsin L-like proteinases from the fish kinetoplastid parasite Trypanosoma carassii

    NARCIS (Netherlands)

    Ruszczyk, A.; Forlenza, M.; Savelkoul, H.F.J.; Wiegertjes, G.F.

    2008-01-01

    Trypanosoma carassii is a fish kinetoplastid parasite that belongs to the family Trypanosomatida. In the present study we cloned a cathepsin L-like proteinase from T. carassii. The nucleotide sequence of 1371 bp translated into a preproprotein of 456 amino acids. The preproprotein contained the

  14. Lipases and proteinases in milk : occurrence, heat inactivation, and their importance for the keeping quality of milk products

    NARCIS (Netherlands)

    Driessen, F.M.

    1983-01-01

    The occurrence and heat inactivation of native and bacterial lipases and proteinases in milk were studied.

    Production of these enzymes by Gram-negative psychrotrophic bacteria in milk was found to take place towards the end of exponential growth and in the stationary growth

  15. Molecular basis of Colorado potato beetle adaptation to potato plant defence at the level of digestive cysteine proteinases

    NARCIS (Netherlands)

    Gruden, K.; Kuipers, A.G.J.; Guncar, G.; Slapar, N.; Strukelj, B.; Jongsma, M.A.

    2004-01-01

    Potato synthesises high levels of proteinase inhibitors in response to insect attack. This can adversely affect protein digestion in the insects, leading to reduced growth, delayed development and lowered fecundity. Colorado potato beetle overcomes this defence mechanism by changing the composition

  16. Whole body 3-methylhistidine production and proteinase activities in porcine muscle after protein-free feeding and realimentation.

    NARCIS (Netherlands)

    Hemel-Grooten, van den H.N.A.; Rathmacher, J.A.; Garssen, G.J.; Schreurs, V.V.A.M.; Verstegen, M.W.A.

    1998-01-01

    Whole body 3-methylhistidine (3MH) production rates and proteinase activities in porcine skeletal muscles were studied during a protein-free feeding period and subsequent realimentation. Out of 54 castrated male pigs (35 kg on day 0), six pigs were slaughtered on day 0, and 48 were randomly divided

  17. Characterization of extracellular polymeric matrix, and treatment of Fusobacterium nucleatum and Porphyromonas gingivalis biofilms with DNase I and proteinase K

    Directory of Open Access Journals (Sweden)

    Marwan Mansoor Ali Mohammed

    2013-01-01

    Full Text Available Background: Biofilms are organized communities of microorganisms embedded in a self-produced extracellular polymeric matrix (EPM, often with great phylogenetic variety. Bacteria in the subgingival biofilm are key factors that cause periodontal diseases; among these are the Gram-negative bacteria Fusobacterium nucleatum and Porphyromonas gingivalis. The objectives of this study were to characterize the major components of the EPM and to test the effect of deoxyribonuclease I (DNase I and proteinase K. Methods: F. nucleatum and P. gingivalis bacterial cells were grown in dynamic and static biofilm models. The effects of DNase I and proteinase K enzymes on the major components of the EPM were tested during biofilm formation and on mature biofilm. Confocal laser scanning microscopy was used in observing biofilm structure. Results: Proteins and carbohydrates were the major components of the biofilm matrix, and extracellular DNA (eDNA was also present. DNase I and proteinase K enzymes had little effect on biofilms in the conditions used. In the flow cell, F. nucleatum was able to grow in partially oxygenated conditions while P. gingivalis failed to form biofilm alone in similar conditions. F. nucleatum supported the growth of P. gingivalis when they were grown together as dual species biofilm. Conclusion: DNase I and proteinase K had little effect on the biofilm matrix in the conditions used. F. nucleatum formed biofilm easily and supported the growth of P. gingivalis, which preferred anaerobic conditions.

  18. The Contribution of Proteinase-Activated Receptors to Intracellular Signaling, Transcellular Transport and Autophagy in Alzheimer´s Disease

    Czech Academy of Sciences Publication Activity Database

    Matěj, R.; Rohan, Z.; Holada, K.; Olejár, Tomáš

    2015-01-01

    Roč. 12, č. 1 (2015), s. 2-12 ISSN 1567-2050 Institutional support: RVO:67985823 Keywords : Alzheimer ´s Disease * autophagy * proteinase-activated receptors Subject RIV: EA - Cell Biology Impact factor: 3.145, year: 2015

  19. Formation of biogenic amines in raw milk Hispánico cheese manufactured with proteinases and different levels of starter culture.

    Science.gov (United States)

    Fernandez-García, E; Tomillo, J; Nuñez, M

    2000-11-01

    Two proteinases, a neutral proteinase from Bacillus subtilis and a cysteine proteinase from Micrococcus sp., were used to accelerate the ripening process of raw cow's milk Hispánico cheese, a semihard variety. Two levels (0.1% and 1%) of a commercial starter culture containing Lactococcus lactis subsp. lactis and L. lactis subsp. cremoris were added for cheese manufacture. The influence of both factors, proteinase addition and level of starter culture, on the growth of amino acid-decarboxylating microorganisms and on the formation of biogenic amines during cheese ripening was investigated in duplicate experiments. The population of tyrosine decarboxylase-positive bacteria, which represented less than 1% of the total bacterial population in most cheese samples, and tyrosine decarboxylase-positive lactobacilli was not influenced by proteinase addition or level of starter culture. Tyramine was detected in all batches of cheese from day 30. Its concentration was significantly (P < 0.05) influenced by proteinase addition but not by the level of starter culture and increased with cheese age. After 90 days of ripening, 103 to 191 mg/kg of tyramine was found in the different cheese batches. Histamine was not detected until day 60 in cheese with neutral proteinase and 1% starter culture and until day 90 in the rest of the cheeses. The concentration of this amine did not exceed 20 mg/kg in any of the batches investigated. Phenylethylamine and tryptamine were not found in any of the samples.

  20. Processing of predicted substrates of fungal Kex2 proteinases from Candida albicans, C. glabrata, Saccharomyces cerevisiae and Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Bader Oliver

    2008-07-01

    Full Text Available Abstract Background Kexin-like proteinases are a subfamily of the subtilisin-like serine proteinases with multiple regulatory functions in eukaryotes. In the yeast Saccharomyces cerevisiae the Kex2 protein is biochemically well investigated, however, with the exception of a few well known proteins such as the α-pheromone precursors, killer toxin precursors and aspartic proteinase propeptides, very few substrates are known. Fungal kex2 deletion mutants display pleiotropic phenotypes that are thought to result from the failure to proteolytically activate such substrates. Results In this study we have aimed at providing an improved assembly of Kex2 target proteins to explain the phenotypes observed in fungal kex2 deletion mutants by in vitro digestion of recombinant substrates from Candida albicans and C. glabrata. We identified CaEce1, CA0365, one member of the Pry protein family and CaOps4-homolog proteins as novel Kex2 substrates. Conclusion Statistical analysis of the cleavage sites revealed extended subsite recognition of negatively charged residues in the P1', P2' and P4' positions, which is also reflected in construction of the respective binding pockets in the ScKex2 enzyme. Additionally, we provide evidence for the existence of structural constrains in potential substrates prohibiting proteolysis. Furthermore, by using purified Kex2 proteinases from S. cerevisiae, P. pastoris, C. albicans and C. glabrata, we show that while the substrate specificity is generally conserved between organisms, the proteinases are still distinct from each other and are likely to have additional unique substrate recognition.

  1. Identification of B cell recognized linear epitopes in a snake venom serine proteinase from the central American bushmaster Lachesis stenophrys.

    Science.gov (United States)

    Madrigal, M; Alape-Girón, A; Barboza-Arguedas, E; Aguilar-Ulloa, W; Flores-Díaz, M

    2017-12-15

    Snake venom serine proteinases are toxins that perturb hemostasis acting on proteins from the blood coagulation cascade, the fibrinolytic or the kallikrein-kinin system. Despite the relevance of these enzymes in envenomations by viper bites, the characterization of the antibody response to these toxins at the molecular level has not been previously addressed. In this work surface-located B cell recognized linear epitopes from a Lachesis stenophrys venom serine proteinase (UniProt accession number Q072L7) were predicted using an artificial neuronal network at the ABCpred server, the corresponding peptides were synthesized and their immunoreactivity was analyzed against a panel of experimental and therapeutic antivenoms. A molecular model of the L. stenophrys enzyme was built using as a template the structure of the D. acutus Dav-PA serine proteinase (Q9I8X1), which displays the highest degree of sequence similarity to the L. stenophrys enzyme among proteins of known 3D structure, and the surface-located epitopes were identified in the protein model using iCn3D. A total of 13 peptides corresponding to the surface exposed predicted epitopes from L. stenophrys serine proteinase were synthesized and, their reactivity with a rabbit antiserum against the recombinant enzyme and a panel of antivenoms was evaluated by a capture ELISA. Some of the epitopes recognized by monospecific and polyspecific antivenoms comprise sequences overlapping motifs conserved in viper venom serine proteinases. The identification and characterization of relevant epitopes recognized by B cells in snake venom toxins may provide valuable information for the preparation of immunogens that help in the production of improved therapeutic antivenoms. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Proteinase-activated receptor 2 is involved in the behavioural changes associated with sickness behaviour.

    Science.gov (United States)

    Abulkassim, Roua; Brett, Ros; MacKenzie, Scott M; Bushell, Trevor J

    2016-06-15

    Proteinase-activated receptor-2 (PAR2) is widely expressed in the CNS but whether it plays a key role in inflammation-related behavioural changes remains unknown. Hence, in the present study we have examined whether PAR2 contributes to behaviour associated with systemic inflammation using PAR2 transgenic mice. The onset of sickness behaviour was delayed and the recovery accelerated in PAR2(-/-) mice in the LPS-induced model of sickness behaviour. In contrast, PAR2 does not contribute to behaviour under normal conditions. In conclusion, these data suggest that PAR2 does not contribute to behaviour in the normal healthy brain but it plays a role in inflammation-related behavioural changes. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Antioxidant activity of bovine casein hydrolysates produced by Ficus carica L.-derived proteinase.

    Science.gov (United States)

    Di Pierro, Giovanna; O'Keeffe, Martina B; Poyarkov, Alexey; Lomolino, Giovanna; FitzGerald, Richard J

    2014-08-01

    A Ficus carica L. latex proteinase preparation was investigated for its ability to produce antioxidant hydrolysates/peptides from bovine casein (CN). The Oxygen Radical Absorbance Capacity (ORAC) values for NaCN and β-CN hydrolysates ranged from 0.06 to 0.18, and from 0.51 to 1.19μmol Trolox equivalents/mg freeze-dried sample, respectively. Gel permeation HPLC showed that the β-CN hydrolysate with a degree of hydrolysis of 21% had 65% of peptide material with a molecular mass <500Da. The RP-UPLC profiles also indicated that β-CN was substantially hydrolysed during the early stages of hydrolysis. Analysis of the 4h β-CN hydrolysate by LC-ESI-MS/MS allowed identification of 8 peptide sequences with potential antioxidant properties. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. Role of calcium-dependent proteinase in molt-induced claw muscle atrophy

    Energy Technology Data Exchange (ETDEWEB)

    Mykles, D.L.; Skinner, D.M.

    1984-01-01

    The claw closer muscle of the Bermuda land crab Gecarcinus lateralis undergoes a sequential atrophy and restoration during each intermolt cycle. Muscle protein decreases 40% during proecdysis and is restored following ecdysis. Amino acid incorporation into protein of postecdysial muscle is five times greater than that in anecdysial muscle. Since the rates of protein synthesis in anecdysial and proecdysial muscle are the same it appears that proecdysial muscle atrophy is caused primarily by an increase in protein degradation. A calcium-dependent proteinase (CDP) active at neutral pH has been implicated in the nonlysosomal hydrolysis of myofibrillar proteins. We have examined the role of a CDP in atrophy of the claw closer muscle. The many similarities between crustacean and vertebrate CDPs have established this crustacean system as a simple and convenient model for the role of Ca/sup 2 +/-dependent proteolysis in myofibrillar protein turnover and its manifestation in the structure of the sarcomere. 16 references, 8 figures. (ACR)

  5. Enhancement of native and phosphorylated TDP-43 immunoreactivity by proteinase K treatment following autoclave heating.

    Science.gov (United States)

    Mori, Fumiaki; Tanji, Kunikazu; Kakita, Akiyoshi; Takahashi, Hitoshi; Wakabayashi, Koichi

    2011-08-01

    TDP-43 is a major disease protein in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43 (FTLD-TDP). To evaluate the effectiveness of proteinase K (PK) treatment in antigen retrieval for native and phosphorylated TDP-43 protein, we examined the temporal cortex and spinal cord from patients with sporadic ALS and FTLD-TDP and control subjects. PK treatment following heat retrieval enhanced the immunoreactivity for native TDP-43 in controls as well as for native and phosphorylated TDP-43 in ALS and FTLD-TDP. A significant number of TDP-43-positive neuropil threads were demonstrated in lesions, in which routine immunohistochemistry revealed that the predominant inclusions are cytoplasmic. This retrieval method is the best of immunohistochemical techniques for demonstrating TDP-43 pathology, especially in the neuropil. © 2010 Japanese Society of Neuropathology.

  6. Lyman Alpha Control

    CERN Document Server

    Nielsen, Daniel Stefaniak

    2015-01-01

    This document gives an overview of how to operate the Lyman Alpha Control application written in LabVIEW along with things to watch out for. Overview of the LabVIEW code itself as well as the physical wiring of and connections from/to the NI PCI-6229 DAQ box is also included. The Lyman Alpha Control application is the interface between the ALPHA sequencer and the HighFinesse Wavelength Meter as well as the Lyman Alpha laser setup. The application measures the wavelength of the output light from the Lyman Alpha cavity through the Wavelength Meter. The application can use the Wavelength Meter’s PID capabilities to stabilize the Lyman Alpha laser output as well as switch between up to three frequencies.

  7. New ALPHA-2 magnet

    CERN Multimedia

    Anaïs Schaeffer

    2012-01-01

    On 21 June, members of the ALPHA collaboration celebrated the handover of the first solenoid designed for the ALPHA-2 experiment. The magnet has since been successfully installed and is working well.   Khalid Mansoor, Sumera Yamin and Jeffrey Hangst in front of the new ALPHA-2 solenoid. “This was the first of three identical solenoids that will be installed between now and September, as the rest of the ALPHA-2 device is installed and commissioned,” explains ALPHA spokesperson Jeffrey Hangst. “These magnets are designed to allow us to transfer particles - antiprotons, electrons and positrons - between various parts of the new ALPHA-2 device by controlling the transverse size of the particle bunch that is being transferred.” Sumera Yamin and Khalid Mansoor, two Pakistani scientists from the National Centre for Physics in Islamabad, came to CERN in February specifically to design and manufacture these magnets. “We had the chance to work on act...

  8. Application of Asian pumpkin (Cucurbita ficifolia) serine proteinase for production of biologically active peptides from casein.

    Science.gov (United States)

    Dąbrowska, Anna; Szołtysik, Marek; Babij, Konrad; Pokora, Marta; Zambrowicz, Aleksandra; Chrzanowska, Józefa

    2013-01-01

    The main objective of this study was to determine potential application of a serine proteinase derived from Asian pumpkin for obtaining biologically active peptides from casein. The course of casein hydrolysis by three doses of the enzyme (50, 150, 300 U/mg of protein) was monitored for 24 hours by the determinations of: hydrolysis degree DH (%), free amino group content (μmole Gly/g), RP HPLC peptide profiles and by polyacrylamide gel electrophoresis. In all hydrolyzates analyzed antioxidant activities were determined using three tests: the ability to reduce iron ions in FRAP test, the ability to scavenge free radicals in DPPH test, and Fe(2+) chelating activity. The antimicrobial activity of obtained peptide fractions was determined as the ability to inhibit the growth of Escherichia coli, Bacillus cereus and Pseudomonas fluorescens in a diffusion plate test. The deepest degradation, expressed as the DH [%] and the free amino group content (67% and 7528 µmole Gly/mg, respectively), was noted in samples hydrolyzed with 300 U/ml of enzyme for 24 hours, while in other samples the determined values were about three and two times lower. The results were in agreement with the peptide profiles obtained by RP HPLC. The highest antioxidative activities determined in all tests were seen for the casein hydrolysate obtained with 300 U/mg protein of serine proteinase after 24 h of reaction (2.15 µM Trolox/mg, 96.15 µg Fe(3+)/mg, 814.97 µg Fe(2+)/mg). Antimicrobial activity was presented in three preparations. In other samples no antimicrobial activity was detected.

  9. Metabolic mapping of proteinase activity with emphasis on in situ zymography of gelatinases: review and protocols.

    Science.gov (United States)

    Frederiks, Wilma M; Mook, Olaf R F

    2004-06-01

    Proteases are essential for protein catabolism, regulation of a wide range of biological processes, and in the pathogenesis of many diseases. Several techniques are available to localize activity of proteases in tissue sections or cell preparations. For localization of the activity of matrix metalloproteinases, in situ zymography was introduced some decades ago. The procedure is based on zymography using SDS polyacrylamide gels containing gelatin, casein, or fibrin as substrate. For in situ zymography, either a photographic emulsion containing gelatin or a fluorescence-labeled proteinaceous macromolecular substrate is brought into contact with a tissue section or cell preparation. After incubation, enzymatic activity is revealed as white spots in a dark background or as black spots in a fluorescent background. However, this approach does not allow precise localization of proteinase activity because of limited sensitivity. A major improvement in sensitivity was achieved with the introduction of dye-quenched (DQ-)gelatin, which is gelatin that is heavily labeled with FITC molecules so that its fluorescence is quenched. After cleavage of DQ-gelatin by gelatinolytic activity, fluorescent peptides are produced that are visible against a weakly fluorescent background. The incubation with DQ-gelatin can be combined with simultaneous immunohistochemical detection of a protein on the same section. To draw valid conclusions from the findings with in situ zymography, specific inhibitors need to be used and the technique has to be combined with immunohistochemistry and zymography. In that case, in situ zymography provides data that extend our understanding of the role of specific proteinases in various physiological and pathological conditions.

  10. Isolation and characterization of a proteinase K-sensitive PrPSc fraction.

    Science.gov (United States)

    Pastrana, Miguel A; Sajnani, Gustavo; Onisko, Bruce; Castilla, Joaquín; Morales, Rodrigo; Soto, Claudio; Requena, Jesús R

    2006-12-26

    Recent studies have shown that a sizable fraction of PrPSc present in prion-infected tissues is, contrary to previous conceptions, sensitive to digestion by proteinase K (PK). This finding has important implications in the context of diagnosis of prion disease, as PK has been extensively used in attempts to distinguish between PrPSc and PrPC. Even more importantly, PK-sensitive PrPSc (sPrPSc) might be essential to understand the process of conversion and aggregation of PrPC leading to infectivity. We have isolated a fraction of sPrPSc. This material was obtained by differential centrifugation at an intermediate speed of Syrian hamster PrPSc obtained through a conventional procedure based on ultracentrifugation in the presence of detergents. PK-sensitive PrPSc is completely degraded under standard conditions (50 mug/mL of proteinase K at 37 degrees C for 1 h) and can also be digested with trypsin. Centrifugation in a sucrose gradient showed sPrPSc to correspond to the lower molecular weight fractions of the continuous range of oligomers that constitute PrPSc. PK-sensitive PrPSc has the ability to convert PrPC into protease-resistant PrPSc, as assessed by the protein misfolding cyclic amplification assay (PMCA). Limited proteolysis of sPrPSc using trypsin allows for identification of regions that are particularly susceptible to digestion, i.e., are partially exposed and flexible; we have identified as such the regions around residues K110, R136, R151, K220, and R229. PK-sensitive PrPSc isolates should prove useful for structural studies to help understand fundamental issues of the molecular biology of PrPSc and in the quest to design tests to detect preclinical prion disease.

  11. Proteins of the kidney microvillar membrane. Aspartate aminopeptidase: purification by immunoadsorbent chromatography and properties of the detergent- and proteinase-solubilized forms

    DEFF Research Database (Denmark)

    Danielsen, Erik Michael; Norén, O; Sjöström, H

    1980-01-01

    revealed 1 g-atom of Ca/143000 g of protein. Two forms of the enzyme were purified: an amphipathic form solubilized from the membrane by Triton X-100 (detergent form) and a hydrophilic form released by incubation with trypsin (proteinase form). The detergent form exhibited charge-shift in crossed...... immunoelectrophoresis when anionic or cationic detergents were present. On gel filtration, mol.wts. of 350000--400000 and 270000 were calculated for the detergent and proteinase forms. Electron microscopy after negative staining of the proteinase form revealed a dimeric structure. Electrophoresis of either form...

  12. Alpha Shapes and Proteins

    DEFF Research Database (Denmark)

    Winter, Pawel; Sterner, Henrik; Sterner, Peter

    2009-01-01

    We provide a unified description of (weighted) alpha shapes, beta shapes and the corresponding simplicialcomplexes. We discuss their applicability to various protein-related problems. We also discuss filtrations of alpha shapes and touch upon related persistence issues.We claim that the full...... potential of alpha-shapes and related geometrical constructs in protein-related problems yet remains to be realized and verified. We suggest parallel algorithms for (weighted) alpha shapes, and we argue that future use of filtrations and kinetic variants for larger proteins will need such implementation....

  13. Targeted Alpha Therapy: From Alpha to Omega

    International Nuclear Information System (INIS)

    Allen, Barry J; Clarke, Raymond; Huang Chenyu

    2013-01-01

    This review covers the broad spectrum of Targeted Alpha Therapy (TAT) research in Australia; from in vitro and in vivo studies to clinical trials. The principle of tumour anti-vascular alpha therapy (TAVAT) is discussed in terms of its validation by Monte Carlo calculations of vascular models and the potential role of biological dosimetry is examined. Summmary of this review is as follows: 1. The essence of TAT 2. Therapeutic objectives 3. TAVAT and Monte Carlo microdosimetry 4. Biological dosimetry 5. Preclinical studies 6. Clinical trials 7. What next? 8. Obstacles. (author)

  14. Alpha-1-antitrypsin is produced by human neutrophil granulocytes and their precursors and liberated during granule exocytosis

    DEFF Research Database (Denmark)

    Clemmensen, Stine N; Jacobsen, Lars C; Rørvig, Sara

    2011-01-01

    Alpha-1-antitrypsin (A1AT) is an important inhibitor of neutrophil proteases including elastase, cathepsin G, and proteinase 3. Transcription profiling data suggest that A1AT is expressed by human neutrophil granulocytes during all developmental stages. A1AT has hitherto only been found associated....... Neutrophils from patients with A1AT-deficiency carrying the (PI)ZZ mutation in the A1AT gene appeared structurally and functionally normal, but A1AT produced in leukocytes of these patients lacked the ability to bind proteases efficiently. We conclude that A1AT generation and release from neutrophils add...

  15. Alpha1-antitrypsin deficiency

    DEFF Research Database (Denmark)

    Stolk, Jan; Seersholm, Niels; Kalsheker, Noor

    2006-01-01

    The Alpha One International Registry (AIR), a multinational research program focused on alpha1-antitrypsin (AAT) deficiency, was formed in response to a World Health Organization recommendation. Each of the nearly 20 participating countries maintains a national registry of patients with AAT defic...... epidemiology, inflammatory and signalling processes, therapeutic advances, and lung imaging techniques....

  16. Alpha clustering in nuclei

    International Nuclear Information System (INIS)

    Hodgson, P.E.

    1990-01-01

    The effects of nucleon clustering in nuclei are described, with reference to both nuclear structure and nuclear reactions, and the advantages of using the cluster formalism to describe a range of phenomena are discussed. It is shown that bound and scattering alpha-particle states can be described in a unified way using an energy-dependent alpha-nucleus potential. (author)

  17. Matriptase-2, a membrane-bound mosaic serine proteinase predominantly expressed in human liver and showing degrading activity against extracellular matrix proteins.

    Science.gov (United States)

    Velasco, Gloria; Cal, Santiago; Quesada, Victor; Sánchez, Luis M; López-Otín, Carlos

    2002-10-04

    We have identified and cloned a fetal liver cDNA encoding a new serine proteinase that has been called matriptase-2. This protein exhibits a domain organization similar to other members of an emerging family of membrane-bound serine proteinases known as type II transmembrane serine proteinases. Matriptase-2 contains a short cytoplasmic domain, a type II transmembrane sequence, a central region with several modular structural domains including two CUB (complement factor C1s/C1r, urchin embryonic growth factor, bone morphogenetic protein) domains and three low density lipoprotein receptor tandem repeats, and finally, a C-terminal catalytic domain with all typical features of serine proteinases. The human matriptase-2 gene maps to 22q12-q13, a location that differs from all type II transmembrane serine proteinase genes mapped to date. Immunofluorescence and Western blot analysis of COS-7 cells transfected with the isolated cDNA confirmed that matriptase-2 is anchored to the cell surface. Matriptase-2 was expressed in Escherichia coli, and the purified recombinant protein hydrolyzed synthetic substrates used for assaying serine proteinases and endogenous proteins such as type I collagen, fibronectin, and fibrinogen. Matriptase-2 could also activate single-chain urokinase plasminogen activator, albeit with low efficiency. These activities were abolished by inhibitors of serine proteinases but not by inhibitors of other classes of proteolytic enzymes. Northern blot analysis demonstrated that matriptase-2 transcripts are only detected at significant levels in both fetal and adult liver, suggesting that this novel serine proteinase may play a specialized role in matrix remodeling processes taking place in this tissue during development or in adult tissues.

  18. Genetics Home Reference: alpha thalassemia

    Science.gov (United States)

    ... Facebook Twitter Home Health Conditions Alpha thalassemia Alpha thalassemia Printable PDF Open All Close All Enable Javascript to view the expand/collapse boxes. Description Alpha thalassemia is a blood disorder that reduces the production ...

  19. Effects of endogenous cysteine proteinases on structures of collagen fibres from dermis of sea cucumber (Stichopus japonicus).

    Science.gov (United States)

    Liu, Yu-Xin; Zhou, Da-Yong; Ma, Dong-Dong; Liu, Zi-Qiang; Liu, Yan-Fei; Song, Liang; Dong, Xiu-Ping; Li, Dong-Mei; Zhu, Bei-Wei; Konno, Kunihiko; Shahidi, Fereidoon

    2017-10-01

    Autolysis of sea cucumber, caused by endogenous enzymes, leads to postharvest quality deterioration of sea cucumber. However, the effects of endogenous proteinases on structures of collagen fibres, the major biologically relevant substrates in the body wall of sea cucumber, are less clear. Collagen fibres were prepared from the dermis of sea cucumber (Stichopus japonicus), and the structural consequences of degradation of the collagen fibres caused by endogenous cysteine proteinases (ECP) from Stichopus japonicus were examined. Scanning electron microscopic images showed that ECP caused partial disaggregation of collagen fibres into collagen fibrils by disrupting interfibrillar proteoglycan bridges. Differential scanning calorimetry and Fourier transform infrared analysis revealed increased structural disorder of fibrillar collagen caused by ECP. SDS-PAGE and chemical analysis indicated that ECP can liberate glycosaminoglycan, hydroxyproline and collagen fragments from collagen fibres. Thus ECP can cause disintegration of collagen fibres by degrading interfibrillar proteoglycan bridges. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Orientation of the brush-border membranal proteinase which specifically splits the catalytic subunit of cAMP-dependent protein kinase.

    Science.gov (United States)

    De Jonge, H; Schmeeda, H; Shaltiel, S

    1987-12-15

    The active site of the rat intestinal brush-border membranal proteinase [Alhanaty E. and Shaltiel S. (1979) Biochem. Biophys. Res. Commun. 89, 323-332], which splits the catalytic subunit (C) of cAMP-dependent protein kinase with a remarkable specificity [Alhanaty E., Tauber-Finkelstein, M., Schmeeda, H. and Shaltiel, S. (1985) Curr. Topics Cell. Regul. 27, 267-277], is shown to face predominantly the cell exterior; vesicles prepared from these brush-borders (mostly sealed and right-side-out) fully express the proteinase activity as judged by the fact that there is no increase in activity upon rupture or solubilization of the vesicles. Although the brush-border vesicles contain a cAMP-dependent protein kinase, this membrane-bound kinase is not likely to be the physiological target of the proteinase, since it appears to have an intracellular orientation and, at least in the vesicles, to be inaccessible to the proteinase. It is, therefore, suggested that the physiological substrate of the proteinase might be either an extracellular cAMP-dependent protein kinase, which is lost (e.g. removed, inactivated or degraded) in the course of vesicle isolation, or a kinase domain in one of the family of proteins recently shown to have a considerable structural and conformational homology with C. Alternatively the physiological site of action of this kinase-splitting proteinase might be an intracellular organelle to which it is translocated by endocytosis.

  1. Analysis of green kiwi fruit (Actinidia deliciosa cv. Hayward) proteinases by two-dimensional zymography and direct identification of zymographic spots by mass spectrometry.

    Science.gov (United States)

    Larocca, Marilena; Rossano, Rocco; Riccio, Paolo

    2010-11-01

    Proteinases present in kiwi fruits are potentially allergenic enzymes belonging to the papain family of cysteine proteinases. Actinidin is a prominent kiwi enzyme. The study of kiwi proteinases is important for the follow-up of fruit maturation, a deeper insight in the allergenic properties of individual proteins, and the application of kiwi proteinases for meat tenderisation and other industrial purposes. Kiwi crude extracts were analysed by two-dimensional zymography on gelatin-containing gels. The digestion by the reactivated proteolytic enzymes after electrophoresis resulted in insights into kiwi proteinases. A mixture of several enzyme isotypes with the same pI but different molecular mass was observed. Clear spots, corresponding to the proteolytic activities, were excised, digested with trypsin, and submitted to MALDI-ToF mass spectrometry for protein identification. The most representative enzyme was actinidin. The innovative achievements of the present study are the: (1) two-dimensional zymographic map of kiwi gelatinases without the need for extensive purification; and (2) direct identification of proteinase isotypes by means of direct MALDI-ToF MS analysis of the zymographic spots. 2010 Society of Chemical Industry

  2. A secreted aspartic proteinase from Glomerella cingulata: purification of the enzyme and molecular cloning of the cDNA.

    Science.gov (United States)

    Clark, S J; Templeton, M D; Sullivan, P A

    1997-04-01

    A secreted aspartic proteinase from Glomerella cingulata (GcSAP) was purified to homogeneity by ion exchange chromatography. The enzyme has an M, of 36000 as estimated by SDS-PAGE, optimal activity from pH 3.5 to pH 4.0 and is inhibited by pepstatin. The N-terminal sequence, 23 residues long, was used to design a gene-specific primer. This was used in 3' RACE (rapid amplification of cDNA ends) PCR to amplify a 1.2 kb fragment of the gcsap cDNA. A second gene-specific primer was designed and used in 5' RACE PCR to clone the 5' region. This yielded a 600 bp DNA fragment and completed the open reading frame. The gcsap open reading frame encodes a protein with a 78 residue prepro-sequence typical of other fungal secreted aspartic proteinases. Based on the deduced sequence, the mature enzyme contains 329 amino acids and shows approximately 40% identity to other fungal aspartic proteinases. Subsequent cloning and sequencing of gcsap fragments obtained from PCR with genomic DNA revealed a 73 bp intron beginning at nt 728. Southern analyses at medium and high stringency indicated that G. cingulata possesses one gene for the secreted aspartic proteinase, and Northern blots indicated that gene expression was induced by exogenous protein and repressed by ammonium salts. GcSAP is a putative pathogenicity factor of G. cingulata, and it will now be possible to create SAP-mutants and assess the role GcSAP plays in pathogenicity.

  3. Saccharomyces cerevisiae can secrete Sapp1p proteinase of Candida parapsilosis but cannot use it for efficient nitrogen acquisition

    Czech Academy of Sciences Publication Activity Database

    Vinterová, Zuzana; Bauerová, Václava; Dostál, Jiří; Sychrová, Hana; Hrušková-Heidingsfeldová, Olga; Pichová, Iva

    2013-01-01

    Roč. 51, č. 3 (2013), s. 336-344 ISSN 1225-8873 R&D Projects: GA ČR GA310/09/1945; GA ČR GAP302/12/1151 Institutional support: RVO:61388963 ; RVO:67985823 Keywords : Candida parapsilosis * Saccharomyces cerevisiae * secreted aspartic proteinase * SAPP1 * nitrogen metabolism Subject RIV: EE - Microbiology, Virology; EE - Microbiology, Virology (FGU-C) Impact factor: 1.529, year: 2013

  4. Evidence for the presence of proteolytically active secreted aspartic proteinase 1 of Candida parapsilosis in the cell wall

    Czech Academy of Sciences Publication Activity Database

    Vinterová, Zuzana; Šanda, Miloslav; Dostál, Jiří; Hrušková-Heidingsfeldová, Olga; Pichová, Iva

    2011-01-01

    Roč. 20, č. 12 (2011), s. 2004-2012 ISSN 0961-8368 R&D Projects: GA MŠk(CZ) LC531; GA ČR GA310/09/1945 Institutional research plan: CEZ:AV0Z40550506 Keywords : Candida parapsilosis * secreted aspartic proteinases * Sapp1p * cell wall * biotin * proteolytic activity Subject RIV: CE - Biochemistry Impact factor: 2.798, year: 2011

  5. Aspartic proteinases are expressed in pitchers of the carnivorous plant Nepenthes alata Blanco.

    Science.gov (United States)

    An, Chung-Il; Fukusaki, Ei-ichiro; Kobayashi, Akio

    2002-03-01

    Carnivorous plants acquire significant amounts of nitrogen from insects. The tropical carnivorous plant Nepenthes accumulates acidic fluid containing aspartic proteinase (AP) in its trapping organs (pitchers), suggesting that the plant utilizes insect protein as a nitrogen source. Aspartic proteinases have been purified and characterized from sterile pitcher fluid of several species of Nepenthes; however, there is, as of yet, no information about sequence and expression of Nepenthes AP genes. To identify the pitcher AP, we cloned plant AP homologs from N. alata and examined their expressions. Five AP homologs ( NaAP1-NaAP5) were obtained by reverse transcription-polymerase chain reaction with degenerate primers designed for the conserved sequences of plant APs. Alignment of deduced amino acid sequences with other plant APs demonstrated that NaAP1-NaAP4 contained a plant-specific insert (PSI), a unique sequence of plant AP. However, NaAP5 did not possess the insert, and had a shorter sequence (by >100 amino acids) than the other APs. Northern analysis using a part of the coding region of NaAP1 as a probe showed that bands of approx. 1.8 kb corresponding to the sizes of NaAP1-NaAP4 mRNA were present in roots, stems, leaves, tendrils, and lower part of the pitchers, but a band of approx. 1.3 kb corresponding to the size of NaAP5 mRNA was not observed in any organs. In pitchers, highest expressions of NaAP1-NaAP4 were seen in the lower part of open pitchers containing natural prey, suggesting that the expressions of NaAP1-NaAP4 are coupled with prey capture. Transcripts of NaAP2 and NaAP4 were detected in the digestive glands, where AP secretion may occur. This result suggests that NaAP2 and NaAP4 are the possible APs secreted into the pitcher of N. alata.

  6. Proteinase-activated receptor 4 stimulation-induced epithelial-mesenchymal transition in alveolar epithelial cells

    Directory of Open Access Journals (Sweden)

    Araki Hiromasa

    2007-04-01

    Full Text Available Abstract Background Proteinase-activated receptors (PARs; PAR1–4 that can be activated by serine proteinases such as thrombin and neutrophil catepsin G are known to contribute to the pathogenesis of various pulmonary diseases including fibrosis. Among these PARs, especially PAR4, a newly identified subtype, is highly expressed in the lung. Here, we examined whether PAR4 stimulation plays a role in the formation of fibrotic response in the lung, through alveolar epithelial-mesenchymal transition (EMT which contributes to the increase in myofibroblast population. Methods EMT was assessed by measuring the changes in each specific cell markers, E-cadherin for epithelial cell, α-smooth muscle actin (α-SMA for myofibroblast, using primary cultured mouse alveolar epithelial cells and human lung carcinoma-derived alveolar epithelial cell line (A549 cells. Results Stimulation of PAR with thrombin (1 U/ml or a synthetic PAR4 agonist peptide (AYPGKF-NH2, 100 μM for 72 h induced morphological changes from cobblestone-like structure to elongated shape in primary cultured alveolar epithelial cells and A549 cells. In immunocytochemical analyses of these cells, such PAR4 stimulation decreased E-cadherin-like immunoreactivity and increased α-SMA-like immunoreactivity, as observed with a typical EMT-inducer, tumor growth factor-β (TGF-β. Western blot analyses of PAR4-stimulated A549 cells also showed similar changes in expression of these EMT-related marker proteins. Such PAR4-mediated changes were attenuated by inhibitors of epidermal growth factor receptor (EGFR kinase and Src. PAR4-mediated morphological changes in primary cultured alveolar epithelial cells were reduced in the presence of these inhibitors. PAR4 stimulation increased tyrosine phosphorylated EGFR or tyrosine phosphorylated Src level in A549 cells, and the former response being inhibited by Src inhibitor. Conclusion PAR4 stimulation of alveolar epithelial cells induced epithelial

  7. Staphylococcus aureus alpha-hemolysin activates the NLRP3-inflammasome in human and mouse monocytic cells.

    Directory of Open Access Journals (Sweden)

    Robin R Craven

    Full Text Available Community Acquired Methicillin Resistant Staphylococcus aureus (CA-MRSA causes severe necrotizing infections of the skin, soft tissues, and lungs. Staphylococcal alpha-hemolysin is an essential virulence factor in mouse models of CA-MRSA necrotizing pneumonia. S. aureus alpha-hemolysin has long been known to induce inflammatory signaling and cell death in host organisms, however the mechanism underlying these signaling events were not well understood. Using highly purified recombinant alpha-hemolysin, we now demonstrate that alpha-hemolysin activates the Nucleotide-binding domain and leucine-rich repeat containing gene family, pyrin domain containing 3 protein (NLRP3-inflammasome, a host inflammatory signaling complex involved in responses to pathogens and endogenous danger signals. Non-cytolytic mutant alpha-hemolysin molecules fail to elicit NLRP3-inflammasome signaling, demonstrating that the responses are not due to non-specific activation of this innate immune signaling system by bacterially derived proteins. In monocyte-derived cells from humans and mice, inflammasome assembly in response to alpha-hemolysin results in activation of the cysteine proteinase, caspase-1. We also show that inflammasome activation by alpha-hemolysin works in conjunction with signaling by other CA-MRSA-derived Pathogen Associated Molecular Patterns (PAMPs to induce secretion of pro-inflammatory cytokines IL-1beta and IL-18. Additionally, alpha-hemolysin induces cell death in these cells through an NLRP3-dependent program of cellular necrosis, resulting in the release of endogenous pro-inflammatory molecules, like the chromatin-associated protein, High-mobility group box 1 (HMGB1. These studies link the activity of a major S. aureus virulence factor to a specific host signaling pathway. The cellular events linked to inflammasome activity have clear relevance to the disease processes associated with CA-MRSA including tissue necrosis and inflammation.

  8. Staphylococcus aureus alpha-hemolysin activates the NLRP3-inflammasome in human and mouse monocytic cells.

    Science.gov (United States)

    Craven, Robin R; Gao, Xi; Allen, Irving C; Gris, Denis; Bubeck Wardenburg, Juliane; McElvania-Tekippe, Erin; Ting, Jenny P; Duncan, Joseph A

    2009-10-14

    Community Acquired Methicillin Resistant Staphylococcus aureus (CA-MRSA) causes severe necrotizing infections of the skin, soft tissues, and lungs. Staphylococcal alpha-hemolysin is an essential virulence factor in mouse models of CA-MRSA necrotizing pneumonia. S. aureus alpha-hemolysin has long been known to induce inflammatory signaling and cell death in host organisms, however the mechanism underlying these signaling events were not well understood. Using highly purified recombinant alpha-hemolysin, we now demonstrate that alpha-hemolysin activates the Nucleotide-binding domain and leucine-rich repeat containing gene family, pyrin domain containing 3 protein (NLRP3)-inflammasome, a host inflammatory signaling complex involved in responses to pathogens and endogenous danger signals. Non-cytolytic mutant alpha-hemolysin molecules fail to elicit NLRP3-inflammasome signaling, demonstrating that the responses are not due to non-specific activation of this innate immune signaling system by bacterially derived proteins. In monocyte-derived cells from humans and mice, inflammasome assembly in response to alpha-hemolysin results in activation of the cysteine proteinase, caspase-1. We also show that inflammasome activation by alpha-hemolysin works in conjunction with signaling by other CA-MRSA-derived Pathogen Associated Molecular Patterns (PAMPs) to induce secretion of pro-inflammatory cytokines IL-1beta and IL-18. Additionally, alpha-hemolysin induces cell death in these cells through an NLRP3-dependent program of cellular necrosis, resulting in the release of endogenous pro-inflammatory molecules, like the chromatin-associated protein, High-mobility group box 1 (HMGB1). These studies link the activity of a major S. aureus virulence factor to a specific host signaling pathway. The cellular events linked to inflammasome activity have clear relevance to the disease processes associated with CA-MRSA including tissue necrosis and inflammation.

  9. Functional Properties of a Cysteine Proteinase from Pineapple Fruit with Improved Resistance to Fungal Pathogens in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Wei Wang

    2014-02-01

    Full Text Available In plant cells, many cysteine proteinases (CPs are synthesized as precursors in the endoplasmic reticulum, and then are subject to post-translational modifications to form the active mature proteinases. They participate in various cellular and physiological functions. Here, AcCP2, a CP from pineapple fruit (Ananas comosus L. belonging to the C1A subfamily is analyzed based on the molecular modeling and homology alignment. Transcripts of AcCP2 can be detected in the different parts of fruits (particularly outer sarcocarps, and gradually increased during fruit development until maturity. To analyze the substrate specificity of AcCP2, the recombinant protein was overexpressed and purified from Pichia pastoris. The precursor of purified AcCP2 can be processed to a 25 kDa active form after acid treatment (pH 4.3. Its optimum proteolytic activity to Bz-Phe-Val-Arg-NH-Mec is at neutral pH. In addition, the overexpression of AcCP2 gene in Arabidopsis thaliana can improve the resistance to fungal pathogen of Botrytis cinerea. These data indicate that AcCP2 is a multifunctional proteinase, and its expression could cause fruit developmental characteristics of pineapple and resistance responses in transgenic Arabidopsis plants.

  10. Neutrophil proteinase 3 and dipeptidyl peptidase I (cathepsin C) as pharmacological targets in granulomatosis with polyangiitis (Wegener granulomatosis).

    Science.gov (United States)

    Korkmaz, Brice; Lesner, Adam; Letast, Stephanie; Mahdi, Yassir K; Jourdan, Marie-Lise; Dallet-Choisy, Sandrine; Marchand-Adam, Sylvain; Kellenberger, Christine; Viaud-Massuard, Marie-Claude; Jenne, Dieter E; Gauthier, Francis

    2013-07-01

    Neutrophils are among the first cells implicated in acute inflammation. Leaving the blood circulation, they quickly migrate through the interstitial space of tissues and liberate oxidants and other antimicrobial proteins together with serine proteinases. Neutrophil elastase, cathepsin G, proteinase 3 (PR3), and neutrophil serine protease 4 are four hematopoietic serine proteases activated by dipeptidyl peptidase I during neutrophil maturation and are mainly stored in cytoplasmic azurophilic granules. They regulate inflammatory and immune responses after their release from activated neutrophils at inflammatory sites. Membrane-bound PR3 (mbPR3) at the neutrophil surface is the prime antigenic target of antineutrophil cytoplasmic autoantibodies (ANCA) in granulomatosis with polyangiitis (GPA), a vasculitis of small blood vessels and granulomatous inflammation of the upper and/or lower respiratory tracts. The interaction of ANCA with mbPR3 results in excessive activation of neutrophils to produce reactive oxygen species and liberation of granular proteinases to the pericellular environment. In this review, we focus on PR3 and dipeptidyl peptidase I as attractive pharmacological targets whose inhibition is expected to attenuate autoimmune activation of neutrophils in GPA.

  11. Alpha Thalassemia (For Parents)

    Science.gov (United States)

    ... the body has a problem producing alpha globin Beta thalassemia : when the body has a problem producing beta ... Transfusion Blood Test: Hemoglobin Electrophoresis Sickle Cell Disease Beta Thalassemia Blood All About Genetics Prenatal Genetic Counseling Genetic ...

  12. A group-specific inhibitor of lysosomal cysteine proteinases selectively inhibits both proteolytic degradation and presentation of the antigen dinitrophenyl-poly-L-lysine by guinea pig accessory cells to T cells

    DEFF Research Database (Denmark)

    Buus, S; Werdelin, O

    1986-01-01

    A limited intralysosomal proteolytic degradation is probably a key event in the accessory cell processing of large protein antigens before their presentation to T cells. With the aid of highly specific inhibitors of proteinases, we have examined the role of proteolysis in the presentation...... of antigens by guinea pig accessory cells. The proteinase inhibitor benzyloxycarbonyl-phenylalanylalanine-diazomethyl-ketone, which selectively inhibits cysteine proteinases, was used to block this set of enzymes in cultured cells. We demonstrate that the selective inhibition of the cysteine proteinases...... inhibitor. Another inhibitor, pepstatin A, which selectively blocks aspartic proteinases, did not block the presentation of dinitrophenyl-poly-L-lysine. The results identify cysteine proteinases, probably lysosomal, as one of the groups of enzymes involved in antigen processing....

  13. Truncation of a P1 leader proteinase facilitates potyvirus replication in a non-permissive host.

    Science.gov (United States)

    Shan, Hongying; Pasin, Fabio; Tzanetakis, Ioannis E; Simón-Mateo, Carmen; García, Juan Antonio; Rodamilans, Bernardo

    2017-11-08

    The Potyviridae family is a major group of plant viruses that includes c. 200 species, most of which have narrow host ranges. The potyvirid P1 leader proteinase self-cleaves from the remainder of the viral polyprotein and shows large sequence variability linked to host adaptation. P1 proteins can be classified as Type A or Type B on the basis, amongst other things, of their dependence or not on a host factor to develop their protease activity. In this work, we studied Type A proteases from the Potyviridae family, characterizing their host factor requirements. Our in vitro cleavage analyses of potyvirid P1 proteases showed that the N-terminal domain is relevant for host factor interaction and suggested that the C-terminal domain is also involved. In the absence of plant factors, the N-terminal end of Plum pox virus P1 antagonizes protease self-processing. We performed extended deletion mutagenesis analysis to define the N-terminal antagonistic domain of P1. In viral infections, removal of the P1 protease antagonistic domain led to a gain-of-function phenotype, strongly increasing local infection in a non-permissive host. Altogether, our results shed new insights into the adaptation and evolution of potyvirids. © 2017 BSPP AND JOHN WILEY & SONS LTD.

  14. Proteinase K-containing lipid nanoparticles for therapeutic delivery of siRNA LOR-1284.

    Science.gov (United States)

    Kim, Dong Chul; Cho, Young Ah; Li, Hong; Yung, Bryant C; Lee, Robert J

    2014-07-01

    The objective of the present study was to develop an efficient delivery vehicle for siRNA LOR-1284 through incorporation of proteinase K (PrK) as a means of preventing siRNA degradation by serum nucleases. Lipid nanoparticle-PrK-siRNA (LN-PrK-siRNA) complexes were synthesized and characterized. siRNA complexed with PrK and liposomes composed of dimethyldioctadecyl ammonium bromide/cholesterol/Tween 80 (60:35:5 molar ratio) were investigated for down-regulation of R2 mRNA activity in KB human carcinoma cells. Treatment with LN-PrK-siRNA (30:0.3:1 molar ratio) significantly reduced levels of R2 mRNA compared to siRNA-liposomes without PrK in serum-containing medium. LN-PrK-siRNA complexes showed increased stability in serum and reduced toxicity in KB cells relative to LN-siRNA complexes. LN-PrK-siRNA complexes are promising delivery vehicles for siRNA. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  15. Blocking proteinase-activated receptor 2 alleviated neuropathic pain evoked by spinal cord injury.

    Science.gov (United States)

    Wei, H; Wei, Y; Tian, F; Niu, T; Yi, G

    2016-01-01

    Spinal cord injury (SCI) is an extremely serious type of physical trauma observed in clinics. Especially, neuropathic pain resulting from SCI has a lasting and significant impact on most aspects of daily life. Thus, a better understanding of the molecular pathways responsible for the cause of neuropathic pain observed in SCI is important to develop effectively therapeutic agents and treatment strategies. Proteinase-activated receptors (PARs) are a family member of G-protein-coupled receptors and are activated by a proteolytic mechanism. One of its subtypes PAR2 has been reported to be engaged in mechanical and thermal hyperalgesia. Thus, in this study we specifically examined the underlying mechanisms responsible for SCI evoked-neuropathic pain in a rat model. Overall, we demonstrated that SCI increases PAR2 and its downstream pathways TRPV1 and TRPA1 expression in the superficial dorsal horn of the spinal cord. Also, we showed that blocking spinal PAR2 by intrathecal injection of FSLLRY-NH2 significantly inhibits neuropathic pain responses induced by mechanical and thermal stimulation whereas FSLLRY-NH2 decreases the protein expression of TRPV1 and TRPA1 as well as the levels of substance P and calcitonin gene-related peptide. Results of this study have important implications, i.e. targeting one or more of these signaling molecules involved in activation of PAR2 and TRPV1/TRPA1 evoked by SCI may present new opportunities for treatment and management of neuropathic pain often observed in patients with SCI.

  16. Structural and functional characteristics of plant proteinase inhibitor-II (PI-II) family.

    Science.gov (United States)

    Rehman, Shazia; Aziz, Ejaz; Akhtar, Wasim; Ilyas, Muhammad; Mahmood, Tariq

    2017-05-01

    Plant proteinase inhibitor-II (PI-II) proteins are one of the promising defensive proteins that helped the plants to resist against different kinds of unfavorable conditions. Different roles for PI-II have been suggested such as regulation of endogenous proteases, modulation of plant growth and developmental processes and mediating stress responses. The basic knowledge on genetic and molecular diversity of these proteins has provided significant insight into their gene structure and evolutionary relationships in various members of this family. Phylogenetic comparisons of these family genes in different plants suggested that the high rate of retention of gene duplication and inhibitory domain multiplication may have resulted in the expansion and functional diversification of these proteins. Currently, a large number of transgenic plants expressing PI-II genes are being developed for enhancing the defensive capabilities against insects, bacteria and pathogenic fungi. Much emphasis is yet to be given to exploit this ever expanding repertoire of genes for improving abiotic stress resistance in transgenic crops. This review presents an overview about the current knowledge on PI-II family genes, their multifunctional role in plant defense and physiology with their potential applications in biotechnology.

  17. Molecular and biochemical characterization of hemoglobinase, a cysteine proteinase, in Paragonimus westermani

    Science.gov (United States)

    Choi, Joon-Hyuck; Lee, Jae-Hyuk; Yu, Hak-Sun; Jeong, Hae-Jin; Kim, Jin; Hong, Yeon-Chul; Chung, Dong-Il

    2006-01-01

    The mammalian trematode Paragonimus westermani is a typical digenetic parasite, which can cause paragonimiasis in humans. Host tissues and blood cells are important sources of nutrients for development, growth and reproduction of P. westermani. In this study, a cDNA clone encoding a 47 kDa hemoglobinase of P. westermani was characterized by sequencing analysis, and its localization was investigated immunohistochemically. The phylogenetic tree prepared based on the hemoglobinase gene showed high homology with hemoglobinases of Fasciola hepatica and Schistosoma spp. Moreover, recombinant P. westermani hemoglobinase degradaded human hemoglobin at acidic pH (from 3.0 to 5.5) and its activity was almost completely inhibited by E-64, a cysteine proteinase inhibitor. Immunohistochemical studies showed that P. westermani hemoglobinase was localized in the epithelium of the adult worm intestine implying that the protein has a specific function. These observations suggest that hemoglobinase may act as a digestive enzyme for acquisition of nutrients from host hemoglobin. Further investigations may provide insights into hemoglobin catabolism in P. westermani. PMID:16969056

  18. Purification, cloning and autoproteolytic processing of an aspartic proteinase from Centaurea calcitrapa.

    Science.gov (United States)

    Domingos, A; Cardoso, P C; Xue, Z T; Clemente, A; Brodelius, P E; Pais, M S

    2000-12-01

    Plant aspartic proteinases (APs) have been isolated from several seed and leaf sources but the only well characterized enzymes from flowers are cardosins and cyprosins from cardoon, Cynara cardunculus L. Here we report a full-length cDNA clone encoding an AP named cenprosin from the flowers of Centaurea calcitrapa L., a thistle related to cardoon. As found for all eukaryotic APs, the deduced primary sequence consists of a signal sequence, a propart and a mature enzyme. In addition, an internal sequence region of 104 residues typical only of plant APs (a plant-specific insert) is present in the primary structure. Northern analysis revealed that the strongest expression is in fresh flowers. The enzyme is also expressed in fairly high amounts in seeds and in leaves, a feature not detected for cardoon APs. The corresponding enzyme was purified in its precursor form from fresh flowers using ammonium-sulfate precipitation followed by ion-exchange and hydrophobic-interaction chromatography. The processing of the precursor into its mature form was studied in vitro. The enzyme underwent autocatalytic processing at pH 3.0 resulting in two chains of 16 and 30 kDa. When dried flowers were used as a starting material for purification, only 16- and 30-kDa chains were obtained, suggesting that autoproteolytic activation of procenprosin in vivo occurs mainly during drying of the flowers. This may indicate a specific degradative role for the enzyme during senescence of the flowers.

  19. Potato type I and II proteinase inhibitors: modulating plant physiology and host resistance.

    Science.gov (United States)

    Turra, David; Lorito, Matteo

    2011-08-01

    Serine protease inhibitors (PIs) are a large and complex group of plant proteins. Members of the potato type I (Pin1) and II (Pin2) proteinase inhibitor families are among the first and most extensively characterized plant PIs. Many insects and phytopathogenic microorganisms use intracellular and extracellular serine proteases playing important roles in pathogenesis. Plants, however, are able to fight these pathogens through the activation of an intricate defence system that leads to the accumulation of various PIs, including Pin1 and Pin2. Several transgenic plants over-expressing members of the Pin1 and Pin2 families have been obtained in the last twenty years and their enhanced defensive capabilities demonstrated against insects, fungi and bacteria. Furthermore, Pin1 and Pin2 genetically engineered plants showed altered regulation of different plant physiological processes (e.g., dehydratation response, programmed cell death, plant growth, trichome density and branching), supporting an endogenous role in various plant species in addition to the well established defensive one. This review summarizes the current knowledge about Pin1 and Pin2 structure, the role of these proteins in plant defence and physiology, and their potential exploitation in biotechnology.

  20. Circulating ADAM17 Level Reflects Disease Activity in Proteinase-3 ANCA-Associated Vasculitis

    Science.gov (United States)

    Bertram, Anna; Lovric, Svjetlana; Engel, Alissa; Beese, Michaela; Wyss, Kristin; Hertel, Barbara; Park, Joon-Keun; Becker, Jan U.; Kegel, Johanna; Haller, Hermann; Haubitz, Marion

    2015-01-01

    ANCA-associated vasculitides are characterized by inflammatory destruction of small vessels accompanied by enhanced cleavage of membrane-bound proteins. One of the main proteases responsible for ectodomain shedding is disintegrin and metalloproteinase domain-containing protein 17 (ADAM17). Given its potential role in aggravating vascular dysfunction, we examined the role of ADAM17 in active proteinase-3 (PR3)-positive ANCA-associated vasculitis (AAV). ADAM17 concentration was significantly increased in plasma samples from patients with active PR3-AAV compared with samples from patients in remission or from other controls with renal nonvascular diseases. Comparably, plasma levels of the ADAM17 substrate syndecan-1 were significantly enhanced in active AAV. We also observed that plasma-derived ADAM17 retained its specific proteolytic activity and was partly located on extracellular microparticles. Transcript levels of ADAM17 were increased in blood samples of patients with active AAV, but those of ADAM10 or tissue inhibitor of metalloproteinases 3, which inhibits ADAMs, were not. We also performed a microRNA (miR) screen and identified miR-634 as significantly upregulated in blood samples from patients with active AAV. In vitro, miR-634 mimics induced a proinflammatory phenotype in monocyte-derived macrophages, with enhanced expression and release of ADAM17 and IL-6. These data suggest that ADAM17 has a prominent role in AAV and might account for the vascular complications associated with this disease. PMID:25788529

  1. Conjugation of biogenic polyamine (putrescine) with proteinase K: Spectroscopic and theoretical insights.

    Science.gov (United States)

    Hosseini-Koupaei, Mansoore; Shareghi, Behzad; Saboury, Ali Akbar

    2017-05-01

    To understand the influence of polyamine on conformation, stabilization and function of proteins, we used multispectroscopic and simulation methods through structural, stability and kinetic measurements of proteinase K (PK) as a model enzyme combined with putrescine (Put). Structural variability of PK was investigated at different concentrations of Put, using circular dichroism, spectrofluorescence and UV-vis measurements. The secondary structure of PK was changed through β-sheet to α-helix switch induced by Put. Spontaneity of the PK-Put complexation, through hydrogen and van der Waals interactions, altered the microenvironment of aromatic residues due to the exposure of them to the solvent. UV-vis measurement also supported the secondary and tertiary structure alteration of PK as a function of Put concentration. Analysis of kinetic parameters and stability studies revealed that Put could act as an enhancer of activity and stabilizer of PK. Our experiments showed that stability and activity changes of enzyme were closely associated to the conformational alterations of enzyme. The molecular simulation results also demonstrated that Put could spontaneously bind and alter the structure of PK, thereby confirming the experimental results. Overall, the results showed that Put could bind to PK and improve its stability and activity, thereby promising various biotechnological and industrial applications. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Computational study of some benzamidine-based inhibitors of thrombin-like snake venom proteinases

    Science.gov (United States)

    Henriques, Elsa S.; Nascimento, Marco A. C.; Ramos, Maria João

    Pit viper venoms contain a number of serine proteinases that, despite their observed coagulant thrombin-like action in vitro, exhibit a paradoxical benign defibrinogenating (anticoagulant) action in vivo, with clinical applications in preventing thrombi and improved blood circulation. Considering that several benzamidine-based inhibitors, some highly selective to thrombin, also inhibit the enzymatic activity of such venombins, the modeling of their enzyme-inhibitor interactions could provide valuable information on the topological factors that determine the divergences in activity. The first step, and the object of the present study, was to derive the necessary set of parameters, consistent with the CHARMM force field, and to perform molecular dynamics (MD) simulations on a few selected representatives of the inhibitors in question under physiological conditions. Bonding and van der Waals parameters were derived by analogy to similar ones in the existing force field. Net atomic charges were obtained with a restrained fitting to the molecular electrostatic potential generated at B3LYP/6-31G(d) level. The parameters were refined to reproduce the available experimental geometries and crystal data, and the MD simulations of the free inhibitors in aqueous solution at 298 K provided an insightful description of their available conformational space.

  3. Single-Step Purification and Characterization of A Recombinant Serine Proteinase Inhibitor from Transgenic Plants.

    Science.gov (United States)

    Jha, Shweta; Agarwal, Saurabh; Sanyal, Indraneel; Amla, D V

    2016-05-01

    Expression of recombinant therapeutic proteins in transgenic plants has a tremendous impact on safe and economical production of biomolecules for biopharmaceutical industry. The major limitation in their production is downstream processing of recombinant protein to obtain higher yield and purity of the final product. In this study, a simple and rapid process has been developed for purification of therapeutic recombinant α1-proteinase inhibitor (rα1-PI) from transgenic tomato plants, which is an abundant serine protease inhibitor in human serum and chiefly inhibits the activity of neutrophil elastase in lungs. We have expressed rα1-PI with modified synthetic gene in transgenic tomato plants at a very high level (≃3.2 % of total soluble protein). The heterologous protein was extracted with (NH4)2SO4 precipitation, followed by chromatographic separation on different matrices. However, only immunoaffinity chromatography resulted into homogenous preparation of rα1-PI with 54 % recovery. The plant-purified rα1-PI showed molecular mass and structural conformation comparable to native serum α1-PI, as shown by mass spectrometry and optical spectroscopy. The results of elastase inhibition assay revealed biological activity of the purified rα1-PI protein. This work demonstrates a simple and efficient one-step purification of rα1-PI from transgenic plants, which is an essential prerequisite for further therapeutic development.

  4. Treponema denticola chymotrypsin-like proteinase (CTLP) integrates spirochaetes within oral microbial communities

    Science.gov (United States)

    Cogoni, Valentina; Morgan-Smith, Alex; Fenno, J. Christopher; Dymock, David

    2012-01-01

    Treponema denticola is found ubiquitously in the human oral cavity and is mainly associated with bacterial communities implicated in the establishment and development of periodontal disease. The ability to become integrated within biofilm communities is crucial to the growth and survival of oral bacteria, and involves inter-bacterial coaggregation, metabolic cooperation, and synergy against host defences. In this article we show that the chymotrypsin-like proteinase (CTLP), found within a high-molecular-mass complex on the cell surface, mediates adherence of T. denticola to other potential periodontal pathogens, Porphyromonas gingivalis, Fusobacterium nucleatum, Prevotella intermedia and Parvimonas micra. Proteolytic activity per se did not appear to be required for the interactions, and expression of the major outer-sheath protein (Msp) was not necessary, except for binding Parv. micra. Biofilms of densely packed cells and matrix, up to 40 µm in depth, were formed between T. denticola and P. gingivalis on salivary pellicle, with T. denticola cells enriched in the upper layers. Expression of CTLP, but not Msp, was critical for dual-species biofilm formation with P. gingivalis. T. denticola did not form dual-species biofilms with any of the other three periodontal bacterial species under various conditions. Synergy between T. denticola and P. gingivalis was also shown by increased inhibition of blood clotting, which was CTLP-dependent. The results demonstrate the critical role of CTLP in interactions of T. denticola with other oral micro-organisms, leading to synergy in microbial community development and host tissue pathogenesis. PMID:22313692

  5. Proteinase K and the structure of PrPSc: The good, the bad and the ugly.

    Science.gov (United States)

    Silva, Christopher J; Vázquez-Fernández, Ester; Onisko, Bruce; Requena, Jesús R

    2015-09-02

    Infectious proteins (prions) are, ironically, defined by their resistance to proteolytic digestion. A defining characteristic of the transmissible isoform of the prion protein (PrP(Sc)) is its partial resistance to proteinase K (PK) digestion. Diagnosis of prion disease typically relies upon immunodetection of PK-digested PrP(Sc) by Western blot, ELISA or immunohistochemical detection. PK digestion has also been used to detect differences in prion strains. Thus, PK has been a crucial tool to detect and, thereby, control the spread of prions. PK has also been used as a tool to probe the structure of PrP(Sc). Mass spectrometry and antibodies have been used to identify PK cleavage sites in PrP(Sc). These results have been used to identify the more accessible, flexible stretches connecting the β-strand components in PrP(Sc). These data, combined with physical constraints imposed by spectroscopic results, were used to propose a qualitative model for the structure of PrP(Sc). Assuming that PrP(Sc) is a four rung β-solenoid, we have threaded the PrP sequence to satisfy the PK proteolysis data and other experimental constraints. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Stable and Simple Immobilization of Proteinase K Inside Glass Tubes and Microfluidic Channels.

    Science.gov (United States)

    Küchler, Andreas; Bleich, Julian N; Sebastian, Bernhard; Dittrich, Petra S; Walde, Peter

    2015-11-25

    Engyodontium album proteinase K (proK) is widely used for degrading proteinaceous impurities during the isolation of nucleic acids from biological samples, or in proteomics and prion research. Toward applications of proK in flow reactors, a simple method for the stable immobilization of proK inside glass micropipette tubes was developed. The immobilization of the enzyme was achieved by adsorption of a dendronized polymer-enzyme conjugate from aqueous solution. This conjugate was first synthesized from a polycationic dendronized polymer (denpol) and proK and consisted, on average, of 2000 denpol repeating units and 140 proK molecules, which were attached along the denpol chain via stable bis-aryl hydrazone bonds. Although the immobilization of proK inside the tube was based on nonspecific, noncovalent interactions only, the immobilized proK did not leak from the tube and remained active during prolonged storage at 4 °C and during continuous operation at 25 °C and pH = 7.0. The procedure developed was successfully applied for the immobilization of proK on a glass/PDMS (polydimethylsiloxane) microchip, which is a requirement for applications in the field of proK-based protein analysis with such type of microfluidic devices.

  7. ALPHA-2: the sequel

    CERN Multimedia

    Katarina Anthony

    2012-01-01

    While many experiments are methodically planning for intense works over the long shutdown, there is one experiment that is already working at full steam: ALPHA-2. Its final components arrived last month and will completely replace the previous ALPHA set-up. Unlike its predecessor, this next generation experiment has been specifically designed to measure the properties of antimatter.   The ALPHA team lower the new superconducting solenoid magnet into place. The ALPHA collaboration is working at full speed to complete the ALPHA-2 set-up for mid-November – this will give them a few weeks of running before the AD shutdown on 17 December. “We really want to get some experience with this device this year so that, if we need to make any changes, we will have time during the long shutdown in which to make them,” says Jeffrey Hangst, ALPHA spokesperson. “Rather than starting the 2014 run in the commissioning stage, we will be up and running from the get go.&...

  8. NMR analysis of the interaction of picornaviral proteinases Lb and 2A with their substrate eukaryotic initiation factor 4GII.

    Science.gov (United States)

    Aumayr, Martina; Fedosyuk, Sofiya; Ruzicska, Katharina; Sousa-Blin, Carla; Kontaxis, Georg; Skern, Tim

    2015-12-01

    Messenger RNA is recruited to the eukaryotic ribosome by a complex including the eukaryotic initiation factor (eIF) 4E (the cap-binding protein), the scaffold protein eIF4G and the RNA helicase eIF4A. To shut-off host-cell protein synthesis, eIF4G is cleaved during picornaviral infection by a virally encoded proteinase; the structural basis of this reaction and its stimulation by eIF4E is unclear. We have structurally and biochemically investigated the interaction of purified foot-and-mouth disease virus (FMDV) leader proteinase (Lb(pro)), human rhinovirus 2 (HRV2) 2A proteinase (2A(pro)) and coxsackievirus B4 (CVB4) 2A(pro) with purified eIF4GII, eIF4E and the eIF4GII/eIF4E complex. Using nuclear magnetic resonance (NMR), we completed (13)C/(15) N sequential backbone assignment of human eIF4GII residues 551-745 and examined their binding to murine eIF4E. eIF4GII551-745 is intrinsically unstructured and remains so when bound to eIF4E. NMR and biophysical techniques for determining stoichiometry and binding constants revealed that the papain-like Lb(pro) only forms a stable complex with eIF4GII(551-745) in the presence of eIF4E, with KD values in the low nanomolar range; Lb(pro) contacts both eIF4GII and eIF4E. Furthermore, the unrelated chymotrypsin-like 2A(pro) from HRV2 and CVB4 also build a stable complex with eIF4GII/eIF4E, but with K(D) values in the low micromolar range. The HRV2 enzyme also forms a stable complex with eIF4E; however, none of the proteinases tested complex stably with eIF4GII alone. Thus, these three picornaviral proteinases have independently evolved to establish distinct triangular heterotrimeric protein complexes that may actively target ribosomes involved in mRNA recruitment to ensure efficient host cell shut-off. © 2015 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

  9. Expression of triplicated and quadruplicated alpha globin genes in sheep.

    Science.gov (United States)

    Vestri, R; Pieragostini, E; Yang, F; di Gregorio, P; Rando, A; Masina, P

    1991-01-01

    In the sheep alpha alpha alpha globin gene haplotype, the three genes display from the 5' to the 3' end the percentage efficiencies of about 30:14:6, as indicated by the amounts of the three types of alpha chain produced in the alpha alpha alpha/alpha alpha alpha homozygotes. The 3' gene in the alpha alpha alpha alpha haplotype appears to have an efficiency around 1%, as suggested by analysis of one quadruple alpha homozygote. Moreover, the total outputs of the alpha alpha alpha as well as of the alpha alpha alpha alpha haplotypes do not substantially differ from that of the common alpha alpha haplotype.

  10. Monte Carlo alpha calculation

    Energy Technology Data Exchange (ETDEWEB)

    Brockway, D.; Soran, P.; Whalen, P.

    1985-01-01

    A Monte Carlo algorithm to efficiently calculate static alpha eigenvalues, N = ne/sup ..cap alpha..t/, for supercritical systems has been developed and tested. A direct Monte Carlo approach to calculating a static alpha is to simply follow the buildup in time of neutrons in a supercritical system and evaluate the logarithmic derivative of the neutron population with respect to time. This procedure is expensive, and the solution is very noisy and almost useless for a system near critical. The modified approach is to convert the time-dependent problem to a static ..cap alpha../sup -/eigenvalue problem and regress ..cap alpha.. on solutions of a/sup -/ k/sup -/eigenvalue problem. In practice, this procedure is much more efficient than the direct calculation, and produces much more accurate results. Because the Monte Carlo codes are intrinsically three-dimensional and use elaborate continuous-energy cross sections, this technique is now used as a standard for evaluating other calculational techniques in odd geometries or with group cross sections.

  11. Crystal structure of the alkaline proteinase Savinase from Bacillus lentus at 1.4 A resolution.

    Science.gov (United States)

    Betzel, C; Klupsch, S; Papendorf, G; Hastrup, S; Branner, S; Wilson, K S

    1992-01-20

    Savinase (EC3.4.21.14) is secreted by the alkalophilic bacterium Bacillus lentus and is a representative of that subgroup of subtilisin enzymes with maximum stability in the pH range 7 to 10 and high activity in the range 8 to 12. It is therefore of major industrial importance for use in detergents. The crystal structure of the native form of Savinase has been refined using X-ray diffraction data to 1.4 A resolution. The starting model was that of subtilisin Carlsberg. A comparison to the structures of the closely related subtilisins Carlsberg and BPN' and to the more distant thermitase and proteinase K is presented. The structure of Savinase is very similar to those of homologous Bacillus subtilisins. There are two calcium ions in the structure, equivalent to the strong and the weak calcium-binding sites in subtilisin Carlsberg and subtilisin BPN', well known for their stabilizing effect on the subtilisins. The structure of Savinase shows novel features that can be related to its stability and activity. The relatively high number of salt bridges in Savinase is likely to contribute to its high thermal stability. The non-conservative substitutions and deletions in the hydrophobic binding pocket S1 result in the most significant structural differences from the other subtilisins. The different composition of the S1 binding loop as well as the more hydrophobic character of the substrate-binding region probably contribute to the alkaline activity profile of the enzyme. The model of Savinase contains 1880 protein atoms, 159 water molecules and two calcium ions. The crystallographic R-factor [formula; see text].

  12. Mandatory role of proteinase-activated receptor 1 in experimental bladder inflammation

    Directory of Open Access Journals (Sweden)

    Davis Carole A

    2007-03-01

    Full Text Available Abstract Background In general, inflammation plays a role in most bladder pathologies and represents a defense reaction to injury that often times is two edged. In particular, bladder neurogenic inflammation involves the participation of mast cells and sensory nerves. Increased mast cell numbers and tryptase release represent one of the prevalent etiologic theories for interstitial cystitis and other urinary bladder inflammatory conditions. The activity of mast cell-derived tryptase as well as thrombin is significantly increased during inflammation. Those enzymes activate specific G-protein coupled proteinase-activated receptors (PARs. Four PARs have been cloned so far, and not only are all four receptors highly expressed in different cell types of the mouse urinary bladder, but their expression is altered during experimental bladder inflammation. We hypothesize that PARs may link mast cell-derived proteases to bladder inflammation and, therefore, play a fundamental role in the pathogenesis of cystitis. Results Here, we demonstrate that in addition to the mouse urinary bladder, all four PA receptors are also expressed in the J82 human urothelial cell line. Intravesical administration of PAR-activating peptides in mice leads to an inflammatory reaction characterized by edema and granulocyte infiltration. Moreover, the inflammatory response to intravesical instillation of known pro-inflammatory stimuli such as E. coli lipopolysaccharide (LPS, substance P, and antigen was strongly attenuated by PAR1-, and to a lesser extent, by PAR2-deficiency. Conclusion Our results reveal an overriding participation of PAR1 in bladder inflammation, provide a working model for the involvement of downstream signaling, and evoke testable hypotheses regarding the role of PARs in bladder inflammation. It remains to be determined whether or not mechanisms targeting PAR1 gene silencing or PAR1 blockade will ameliorate the clinical manifestations of cystitis.

  13. Correlation between culture medium pH, extracellular proteinase activity, and cell growth of Candida albicans in insoluble stratum corneum-supplemented media.

    Science.gov (United States)

    Tsuboi, R; Matsuda, K; Ko, I J; Ogawa, H

    1989-01-01

    Candida albicans produces a major extracellular proteinase whose activities are observed only in weakly acidic pH. However, in affected lesions, a variety of pH conditions exist, including neutral pH. To verify the pathological importance of the extracellular proteinase, the correlation between culture medium pH, extracellular proteinase activity, and cell growth of C. albicans was followed for 3 weeks with unbuffered and insoluble stratum corneum-supplemented liquid media. Each medium pH, initially adjusted within a range of pH 3-7 by the addition of sodium hydroxide or hydrochloric acid solution, was acidified, and a subsequent high proteolytic activity and rapid fungal growth were observed. After full fungal growth, neutralization of each medium to pH 7 and reduction of proteinase activity occurred. Results from a glucose addition experiment suggest that acidification of each medium was produced by the acid formation from glucose and neutralization by the exhaustion of glucose and increase of ammonia from denatured stratum corneum. These data suggest that extracellular proteinase from C. albicans could act as a virulence factor under a wide range of pH conditions by the acidification of the environmental pH close to the organism.

  14. Monte Carlo alpha deposition

    International Nuclear Information System (INIS)

    Talley, T.L.; Evans, F.

    1988-01-01

    Prior work demonstrated the importance of nuclear scattering to fusion product energy deposition in hot plasmas. This suggests careful examination of nuclear physics details in burning plasma simulations. An existing Monte Carlo fast ion transport code is being expanded to be a test bed for this examination. An initial extension, the energy deposition of fast alpha particles in a hot deuterium plasma, is reported. The deposition times and deposition ranges are modified by allowing nuclear scattering. Up to 10% of the initial alpha particle energy is carried to greater ranges and times by the more mobile recoil deuterons. 4 refs., 5 figs., 2 tabs

  15. Buffett’s Alpha

    DEFF Research Database (Denmark)

    Frazzini, Andrea; Kabiller, David; Heje Pedersen, Lasse

    Berkshire Hathaway has realized a Sharpe ratio of 0.76, higher than any other stock or mutual fund with a history of more than 30 years, and Berkshire has a significant alpha to traditional risk factors. However, we find that the alpha becomes insignificant when controlling for exposures to Betting......-Against-Beta and Quality-Minus-Junk factors. Further, we estimate that Buffett’s leverage is about 1.6-to-1 on average. Buffett’s returns appear to be neither luck nor magic, but, rather, reward for the use of leverage combined with a focus on cheap, safe, quality stocks. Decomposing Berkshires’ portfolio into ownership...

  16. Inducible expression of a fusion gene encoding two proteinase inhibitors leads to insect and pathogen resistance in transgenic rice.

    Science.gov (United States)

    Quilis, Jordi; López-García, Belén; Meynard, Donaldo; Guiderdoni, Emmanuel; San Segundo, Blanca

    2014-04-01

    Plant proteinase inhibitors (PIs) are considered as candidates for increased insect resistance in transgenic plants. Insect adaptation to PI ingestion might, however, compromise the benefits received by transgenic expression of PIs. In this study, the maize proteinase inhibitor (MPI), an inhibitor of insect serine proteinases, and the potato carboxypeptidase inhibitor (PCI) were fused into a single open reading frame and introduced into rice plants. The two PIs were linked using either the processing site of the Bacillus thuringiensis Cry1B precursor protein or the 2A sequence from the foot-and-mouth disease virus (FMDV). Expression of each fusion gene was driven by the wound- and pathogen-inducible mpi promoter. The mpi-pci fusion gene was stably inherited for at least three generations with no penalty on plant phenotype. An important reduction in larval weight of Chilo suppressalis fed on mpi-pci rice, compared with larvae fed on wild-type plants, was observed. Expression of the mpi-pci fusion gene confers resistance to C. suppressalis (striped stem borer), one of the most important insect pest of rice. The mpi-pci expression systems described may represent a suitable strategy for insect pest control, better than strategies based on the use of single PI genes, by preventing insect adaptive responses. The rice plants expressing the mpi-pci fusion gene also showed enhanced resistance to infection by the fungus Magnaporthe oryzae, the causal agent of the rice blast disease. Our results illustrate the usefulness of the inducible expression of the mpi-pci fusion gene for dual resistance against insects and pathogens in rice plants. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  17. Binding properties of the regulatory domains in Manduca sexta hemolymph proteinase-14, an initiation enzyme of the prophenoloxidase activation system.

    Science.gov (United States)

    Wang, Yang; Jiang, Haobo

    2010-03-01

    Pathogen recognition and rapid initiation of defense responses are essential for the survival of host insects. In Manduca sexta, hemolymph proteinase-14 precursor (proHP14) senses non-self presence and triggers a branched serine proteinase pathway which leads to prophenoloxidase activation and melanin formation around the invading organisms. To understand functions of individual domains in HP14, we have produced a series of HP14 domains and truncation mutants and studied their interactions with microbial polysaccharides and beta-1,3-glucan recognition protein-1 (betaGRP1)-a biosensor for fungal and bacterial infection. These include: the low-density lipoprotein receptor class A repeats 1-5 (LDL(1-5)), Sushi domain, Wonton domain, and proteinase catalytic domain of HP14, as well as proHP14 missing 1-4 LDL repeats (DeltaLDL(1), DeltaLDL(12), DeltaLDL(1-3) and DeltaLDL(1-4)). LDL(1-5), Sushi, and Wonton domains specifically recognized Lys-type PG, whereas the latter two also bound betaGRP1. Wonton in addition bound to lipopolysaccharide (LPS), lipoteichoic acid (LTA), and meso-diaminopimelic acid (DAP)-type peptidoglycan (PG). The four N-terminally truncated proHP14 (DeltaL(x)) further confirmed specific interactions with LPS, LTA, DAP-PG, Lys-PG, laminarin, and betaGRP1. These binding data suggest a broad specificity of proHP14 in pattern recognition. Its role in mediating immune responses is anticipated to be influenced by other plasma factors and surface structures of invading pathogens. Copyright 2009 Elsevier Ltd. All rights reserved.

  18. The role of the proteinase inhibitor ovorubin in apple snail eggs resembles plant embryo defense against predation.

    Directory of Open Access Journals (Sweden)

    Marcos Sebastián Dreon

    Full Text Available BACKGROUND: Fieldwork has thoroughly established that most eggs are intensely predated. Among the few exceptions are the aerial egg clutches from the aquatic snail Pomacea canaliculata which have virtually no predators. Its defenses are advertised by the pigmented ovorubin perivitellin providing a conspicuous reddish coloration. The nature of the defense however, was not clear, except for a screening for defenses that identified a neurotoxic perivitellin with lethal effect on rodents. Ovorubin is a proteinase inhibitor (PI whose role to protect against pathogens was taken for granted, according to the prevailing assumption. Through biochemical, biophysical and feeding experiments we studied the proteinase inhibitor function of ovorubin in egg defenses. METHODOLOGY/PRINCIPAL FINDINGS: Mass spectrometry sequencing indicated ovorubin belongs to the Kunitz-type serine proteinase inhibitor family. It specifically binds trypsin as determined by small angle X-ray scattering (SAXS and cross-linking studies but, in contrast to the classical assumption, it does not prevent bacterial growth. Ovorubin was found extremely resistant to in vitro gastrointestinal proteolysis. Moreover feeding studies showed that ovorubin ingestion diminishes growth rate in rats indicating that this highly stable PI is capable of surviving passage through the gastrointestinal tract in a biologically active form. CONCLUSIONS: To our knowledge, this is the first direct evidence of the interaction of an egg PI with a digestive protease of potential predators, limiting predator's ability to digest egg nutrients. This role has not been reported in the animal kingdom but it is similar to plant defenses against herbivory. Further, this would be the only defense model with no trade-offs between conspicuousness and noxiousness by encoding into the same molecule both the aposematic warning signal and an antinutritive/antidigestive defense. These defenses, combined with a neurotoxin and

  19. The Role of the Proteinase Inhibitor Ovorubin in Apple Snail Eggs Resembles Plant Embryo Defense against Predation

    Science.gov (United States)

    Dreon, Marcos Sebastián; Ituarte, Santiago; Heras, Horacio

    2010-01-01

    Background Fieldwork has thoroughly established that most eggs are intensely predated. Among the few exceptions are the aerial egg clutches from the aquatic snail Pomacea canaliculata which have virtually no predators. Its defenses are advertised by the pigmented ovorubin perivitellin providing a conspicuous reddish coloration. The nature of the defense however, was not clear, except for a screening for defenses that identified a neurotoxic perivitellin with lethal effect on rodents. Ovorubin is a proteinase inhibitor (PI) whose role to protect against pathogens was taken for granted, according to the prevailing assumption. Through biochemical, biophysical and feeding experiments we studied the proteinase inhibitor function of ovorubin in egg defenses. Methodology/Principal Findings Mass spectrometry sequencing indicated ovorubin belongs to the Kunitz-type serine proteinase inhibitor family. It specifically binds trypsin as determined by small angle X-ray scattering (SAXS) and cross-linking studies but, in contrast to the classical assumption, it does not prevent bacterial growth. Ovorubin was found extremely resistant to in vitro gastrointestinal proteolysis. Moreover feeding studies showed that ovorubin ingestion diminishes growth rate in rats indicating that this highly stable PI is capable of surviving passage through the gastrointestinal tract in a biologically active form. Conclusions To our knowledge, this is the first direct evidence of the interaction of an egg PI with a digestive protease of potential predators, limiting predator's ability to digest egg nutrients. This role has not been reported in the animal kingdom but it is similar to plant defenses against herbivory. Further, this would be the only defense model with no trade-offs between conspicuousness and noxiousness by encoding into the same molecule both the aposematic warning signal and an antinutritive/antidigestive defense. These defenses, combined with a neurotoxin and probably unpalatable

  20. Proteinase-Activated Receptor 2 Is a Novel Regulator of TGF-β Signaling in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    David Witte

    2016-11-01

    Full Text Available TGF-β has a dual role in tumorigenesis, acting as a tumor suppressor in normal cells and in the early stages of tumor development while promoting carcinogenesis and metastasis in advanced tumor stages. The final outcome of the TGF-β response is determined by cell-autonomous mechanisms and genetic alterations such as genomic instability and somatic mutations, but also by a plethora of external signals derived from the tumor microenvironment, such as cell-to-cell interactions, growth factors and extracellular matrix proteins and proteolytic enzymes. Serine proteinases mediate their cellular effects via activation of proteinase-activated receptors (PARs, a subclass of G protein-coupled receptors that are activated by proteolytic cleavage. We have recently identified PAR2 as a factor required for TGF-β1-dependent cell motility in ductal pancreatic adenocarcinoma (PDAC cells. In this article, we review what is known on the TGF-β-PAR2 signaling crosstalk and its relevance for tumor growth and metastasis. Since PAR2 is activated through various serine proteinases, it may couple TGF-β signaling to a diverse range of other physiological processes, such as local inflammation, systemic coagulation or pathogen infection. Moreover, since PAR2 controls expression of the TGF-β type I receptor ALK5, PAR2 may also impact signaling by other TGF-β superfamily members that signal through ALK5, such as myostatin and GDF15/MIC-1. If so, PAR2 could represent a molecular linker between PDAC development and cancer-related cachexia.

  1. Identification of a novel SERPINA-1 mutation causing alpha-1 antitrypsin deficiency in a patient with severe bronchiectasis and pulmonary embolism

    Directory of Open Access Journals (Sweden)

    Milger K

    2015-05-01

    Full Text Available Katrin Milger,1 Lesca Miriam Holdt,2 Daniel Teupser,2 Rudolf Maria Huber,1 Jürgen Behr,1 Nikolaus Kneidinger1 1Department of Internal Medicine V, University of Munich, Comprehensive Pneumology Center, Member of the German Center for Lung Research, 2Institute of Laboratory Medicine, University of Munich, Munich, Germany Abstract: Deficiency in the serine protease inhibitor, alpha-1 antitrypsin (AAT, is known to cause emphysema and liver disease. Other manifestations, including airway disease or skin disorders, have also been described. A 44-year-old woman presented to our emergency department with dyspnea and respiratory insufficiency. She had never smoked, and had been diagnosed with COPD 9 years earlier. Three months previously, she had suffered a pulmonary embolism. Chest computed tomography scan revealed severe cystic bronchiectasis with destruction of the lung parenchyma. The sweat test was normal and there was no evidence of the cystic fibrosis transmembrane conductance regulator (CFTR mutation. Capillary zone electrophoresis showed a decrease of alpha-1 globin band and AAT levels were below the quantification limit (<25 mg/dL. No S or Z mutation was identified, but sequencing analysis found a homozygous cytosine and adenine (CA insertion in exon 2 of the SERPINA-1 gene, probably leading to a dysfunctional protein (PI Null/Null. This mutation has not been previously identified. The atypical presentation of the patient, with severe cystic bronchiectasis, highlights AAT deficiency as a differential diagnosis in bronchiectasis. Further, awareness should be raised regarding a possible increased risk of thromboembolism associated with AAT deficiency. Keywords: alpha-1 antitrypsin deficiency, bronchiectasis, SERPINA-1 mutation, pulmonary embolism

  2. Classification of Lactococcus lactis cell envelope proteinase based on gene sequencing, peptides formed after hydrolysis of milk, and computer modeling

    DEFF Research Database (Denmark)

    Børsting, Mette Winther; Qvist, K.B.; Brockmann, E.

    2015-01-01

    Lactococcus lactis strains depend on a proteolytic system for growth in milk to release essential AA from casein. The cleavage specificities of the cell envelope proteinase (CEP) can vary between strains and environments and whether the enzyme is released or bound to the cell wall. Thirty-eight Lc....... lactis strains were grouped according to their CEP AA sequences and according to identified peptides after hydrolysis of milk. Finally, AA positions in the substrate binding region were suggested by the use of a new CEP template based on Streptococcus C5a CEP. Aligning the CEP AA sequences of 38 strains...

  3. Case Study - Alpha

    Directory of Open Access Journals (Sweden)

    Stephen Leybourne

    2016-11-01

    Full Text Available This case study was developed from an actual scenario by Dr. Steve Leybourne of Boston University.  The case documents the historical evolution of an organization, and has been used successfully in courses dealing with organizational and cultural change, and the utilization of ‘soft skills’ in project-based management. This is a short case, ideal for classroom use and discussion.  The issues are easily accessible to students, and there is a single wide ranging question that allows for the inclusion of many issues surrounding strategic decision-making, and behavioural and cultural change. Alpha was one of the earlier companies in the USA to invest in large, edge-of-town superstores, with plentiful free vehicle parking, selling food and related household products. Alpha was created in the 1950s as a subsidiary of a major publicly quoted retail group.  It started business by opening a string of very large discount stores in converted industrial and warehouse premises in the south of the United States. In the early days shoppers were offered a limited range of very competitively priced products. When Alpha went public in 1981 it was the fourth largest food retailer in the US, selling an ever-widening range of food and non-food products.  Its success continued to be based on high volume, low margins and good value for money, under the slogan of ‘Alpha Price.’

  4. Alpha-mannosidosis

    DEFF Research Database (Denmark)

    Borgwardt, Line; Stensland, Hilde Monica Frostad Riise; Olsen, Klaus Juul

    2015-01-01

    of the three subgroups of genotype/subcellular localisation and the clinical and biochemical data were done to investigate the potential relationship between genotype and phenotype in alpha-mannosidosis. Statistical analyses were performed using the SPSS software. Analyses of covariance were performed...

  5. Active trafficking of alpha 1 antitrypsin across the lung endothelium.

    Directory of Open Access Journals (Sweden)

    Angelia D Lockett

    Full Text Available The homeostatic lung protective effects of alpha-1 antitrypsin (A1AT may require the transport of circulating proteinase inhibitor across an intact lung endothelial barrier. We hypothesized that uninjured pulmonary endothelial cells transport A1AT to lung epithelial cells. Purified human A1AT was rapidly taken up by confluent primary rat pulmonary endothelial cell monolayers, was secreted extracellularly, both apically and basolaterally, and was taken up by adjacent rat lung epithelial cells co-cultured on polarized transwells. Similarly, polarized primary human lung epithelial cells took up basolaterally-, but not apically-supplied A1AT, followed by apical secretion. Evidence of A1AT transcytosis across lung microcirculation was confirmed in vivo by two-photon intravital microscopy in mice. Time-lapse confocal microscopy indicated that A1AT co-localized with Golgi in the endothelium whilst inhibition of the classical secretory pathway with tunicamycin significantly increased intracellular retention of A1AT. However, inhibition of Golgi secretion promoted non-classical A1AT secretion, associated with microparticle release. Polymerized A1AT or A1AT supplied to endothelial cells exposed to soluble cigarette smoke extract had decreased transcytosis. These results suggest previously unappreciated pathways of A1AT bidirectional uptake and secretion from lung endothelial cells towards the alveolar epithelium and airspaces. A1AT trafficking may determine its functional bioavailablity in the lung, which could be impaired in individuals exposed to smoking or in those with A1AT deficiency.

  6. Ixodes scapularis tick serine proteinase inhibitor (serpin gene family; annotation and transcriptional analysis

    Directory of Open Access Journals (Sweden)

    Chalaire Katelyn C

    2009-05-01

    Full Text Available Abstract Background Serine proteinase inhibitors (Serpins are a large superfamily of structurally related, but functionally diverse proteins that control essential proteolytic pathways in most branches of life. Given their importance in the biology of many organisms, the concept that ticks might utilize serpins to evade host defenses and immunizing against or disrupting their functions as targets for tick control is an appealing option. Results A sequence homology search strategy has allowed us to identify at least 45 tick serpin genes in the Ixodes scapularis genome that are structurally segregated into 32 intronless and 13 intron-containing genes. Nine of the intron-containing serpins occur in a cluster of 11 genes that span 170 kb of DNA sequence. Based on consensus amino acid residues in the reactive center loop (RCL and signal peptide scanning, 93% are putatively inhibitory while 82% are putatively extracellular. Among the 11 different amino acid residues that are predicted at the P1 sites, 16 sequences possess basic amino acid (R/K residues. Temporal and spatial expression analyses revealed that 40 of the 45 serpins are differentially expressed in salivary glands (SG and/or midguts (MG of unfed and partially fed ticks. Ten of the 38 serpin genes were expressed from six to 24 hrs of feeding while six and fives genes each are predominantly or exclusively expressed in either MG and SG respectively. Conclusion Given the diversity among tick species, sizes of tick serpin families are likely to be variable. However this study provides insight on the potential sizes of serpin protein families in ticks. Ticks must overcome inflammation, complement activation and blood coagulation to complete feeding. Since these pathways are regulated by serpins that have basic residues at their P1 sites, we speculate that I. scapularis may utilize some of the serpins reported in this study to manipulate host defense. We have discussed our data in the context of

  7. Allosteric modulation of proteinase 3 activity by anti-neutrophil cytoplasmic antibodies in granulomatosis with polyangiitis.

    Science.gov (United States)

    Hinkofer, Lisa C; Hummel, Amber M; Stone, John H; Hoffman, Gary S; Merkel, Peter A; Spiera, E Robert F; St Clair, William; McCune, Joseph W; Davis, John C; Specks, Ulrich; Jenne, Dieter E

    2015-05-01

    Anti-neutrophil cytoplasmic antibodies (ANCA) with proteinase 3 (PR3) specificity are a useful laboratory biomarker for the diagnosis of Granulomatosis with Polyangiitis (GPA) and are believed to be implicated in the pathogenesis. It has been repeatedly suggested that disease activity of GPA is more closely related to the appearance and rise of PR3-inhibiting ANCA than to an increase of total ANCA. Previous studies on a limited number of patient samples, however, have yielded inconclusive results. To overcome the previous methodological limitations, we established a new ultrasensitive method to quantify the inhibitory capacity of PR3-ANCA using small volumes of plasma from patients with GPA. A large collection of longitudinally-collected samples from the Wegener Granulomatosis Etanercept Trial (WGET) became available to us to determine the functional effects of ANCA on PR3 in comparison to clinical disease manifestations. In these patient samples we not only detected PR3-ANCA with inhibitory capacity, but also PR3-ANCA with enhancing effects on PR3 activity. However no correlation of these activity-modulating PR3-ANCA with disease activity at either the time of enrollment or over the course of disease was found. Only patients with pulmonary involvement, especially patients with nodule formation in the respiratory tract, showed a slight, but not significant, decrease of inhibitory capacity. Epitope mapping of the activity-modulating PR3-ANCA revealed a binding on the active site surface of PR3. Yet these ANCA were able to bind to PR3 with an occupied active site cleft, indicating an allosteric mechanism of inhibition. The recently described signal ratio between the MCPR3-3 and MCPR3-2 capture ELISA was consistent with the binding of activity-modulating ANCA to the active site surface. Evidence for a shared epitope between activity-modulating PR3-ANCA and MCPR3-7, however, was very limited, suggesting that a majority of PR3-ANCA species do not inhibit PR3 by the same

  8. Demystifying AlphaGo Zero as AlphaGo GAN

    OpenAIRE

    Dong, Xiao; Wu, Jiasong; Zhou, Ling

    2017-01-01

    The astonishing success of AlphaGo Zero\\cite{Silver_AlphaGo} invokes a worldwide discussion of the future of our human society with a mixed mood of hope, anxiousness, excitement and fear. We try to dymystify AlphaGo Zero by a qualitative analysis to indicate that AlphaGo Zero can be understood as a specially structured GAN system which is expected to possess an inherent good convergence property. Thus we deduct the success of AlphaGo Zero may not be a sign of a new generation of AI.

  9. $\\alpha$-Representation for QCD

    OpenAIRE

    Tuan, Richard Hong

    1998-01-01

    An $\\alpha$-parameter representation is derived for gauge field theories.It involves, relative to a scalar field theory, only constants and derivatives with respect to the $\\alpha$-parameters. Simple rules are given to obtain the $\\alpha$-representation for a Feynman graph with an arbitrary number of loops in gauge theories in the Feynman gauge.

  10. Alpha Theta Meditation: Phenomenological, neurophysiologic ...

    African Journals Online (AJOL)

    Alpha Theta Meditation: Phenomenological, neurophysiologic, mindfulness, mood, health and sport implications. ... the single alpha theta meditation was associated with elevated alpha and theta activity, as well as decrease in negative mood perceptions, especially with regard to anxiety, sadness and confusion scores.

  11. Detergent-Stable Salt-Activated Proteinases from Virgibacillus halodenitrificans SK1-3-7 Isolated from Fish Sauce Fermentation.

    Science.gov (United States)

    Montriwong, Aungkawipa; Rodtong, Sureelak; Yongsawatdigul, Jirawat

    2015-05-01

    The NaCl-activated and detergent-stable proteinases from Virgibacillus halodenitrificans SK1-3-7 isolated from fish sauce fermentation were purified and characterized. The enzymes with molecular masses of 20 and 36 kDa showed caseinolytic activity on a zymogram. Optimum azocaseinolytic activity was at 60 °C and pH 9. The proteolytic activity increased in the presence of 10 mM CaCl2 and 0.5 M NaCl and showed high stability at 0-2 M NaCl. The enzymes were stable at pH 4-10 and 10-50 °C. The enzymes preferably hydrolyzed Suc-Ala-Ala-Pro-Phe-pNA and were completely inhibited by phenylmethanesulfonyl fluoride (PMSF), showing subtilisin-like characteristics. Activity and stability remained high in the presence of H2O2 and various surfactants. The enzymes exhibited high stability (>95%) in various organic solvents (DMSO, butanol, ethanol, 2-propanol, and acetonitrile) at concentration of 50%. The V. halodenitrificans SK1-3-7 proteinases showed potential as a biocatalyst in aqueous-organic solvent systems and as an additive in laundry detergent.

  12. Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper

    Directory of Open Access Journals (Sweden)

    A.V Pérez

    2008-01-01

    Full Text Available A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for approximately 0.13% of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 µg and fibrinogen (minimum coagulant dose = 4.2 µg in vitro, and promotes defibrin(ogenation in vivo (minimum defibrin(ogenating dose = 1.0 µg. In addition, when injected intravenously in mice at doses of 5 and 10 µg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the `gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

  13. Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper.

    Science.gov (United States)

    Pérez, A V; Rucavado, A; Sanz, L; Calvete, J J; Gutiérrez, J M

    2008-01-01

    A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for approximately 0.13% of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 microg) and fibrinogen (minimum coagulant dose = 4.2 microg) in vitro, and promotes defibrin(ogen)ation in vivo (minimum defibrin(ogen)ating dose = 1.0 microg). In addition, when injected intravenously in mice at doses of 5 and 10 microg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the ;gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

  14. Molecular karyotype and chromosomal localization of genes encoding ß-tubulin, cysteine proteinase, hsp 70 and actin in Trypanosoma rangeli

    Directory of Open Access Journals (Sweden)

    CB Toaldo

    2001-01-01

    Full Text Available The molecular karyotype of nine Trypanosoma rangeli strains was analyzed by contour-clamped homogeneous electric field electrophoresis, followed by the chromosomal localization of ß-tubulin, cysteine proteinase, 70 kDa heat shock protein (hsp 70 and actin genes. The T. rangeli strains were isolated from either insects or mammals from El Salvador, Honduras, Venezuela, Colombia, Panama and southern Brazil. Also, T. cruzi CL-Brener clone was included for comparison. Despite the great similarity observed among strains from Brazil, the molecular karyotype of all T. rangeli strains analyzed revealed extensive chromosome polymorphism. In addition, it was possible to distinguish T. rangeli from T. cruzi by the chromosomal DNA electrophoresis pattern. The localization of ß-tubulin genes revealed differences among T. rangeli strains and confirmed the similarity between the isolates from Brazil. Hybridization assays using probes directed to the cysteine proteinase, hsp 70 and actin genes discriminated T. rangeli from T. cruzi, proving that these genes are useful molecular markers for the differential diagnosis between these two species. Numerical analysis based on the molecular karyotype data revealed a high degree of polymorphism among T. rangeli strains isolated from southern Brazil and strains isolated from Central and the northern South America. The T. cruzi reference strain was not clustered with any T. rangeli strain.

  15. Assessing Proteinase K Resistance of Fish Prion Proteins in a Scrapie-Infected Mouse Neuroblastoma Cell Line

    Directory of Open Access Journals (Sweden)

    Evgenia Salta

    2014-11-01

    Full Text Available The key event in prion pathogenesis is the structural conversion of the normal cellular protein, PrPC, into an aberrant and partially proteinase K resistant isoform, PrPSc. Since the minimum requirement for a prion disease phenotype is the expression of endogenous PrP in the host, species carrying orthologue prion genes, such as fish, could in theory support prion pathogenesis. Our previous work has demonstrated the development of abnormal protein deposition in sea bream brain, following oral challenge of the fish with natural prion infectious material. In this study, we used a prion-infected mouse neuroblastoma cell line for the expression of three different mature fish PrP proteins and the evaluation of the resistance of the exogenously expressed proteins to proteinase K treatment (PK, as an indicator of a possible prion conversion. No evidence of resistance to PK was detected for any of the studied recombinant proteins. Although not indicative of an absolute inability of the fish PrPs to structurally convert to pathogenic isoforms, the absence of PK-resistance may be due to supramolecular and conformational differences between the mammalian and piscine PrPs.

  16. Glucose-6-phosphate isomerase is an endogenous inhibitor to myofibril-bound serine proteinase of crucian carp (Carassius auratus).

    Science.gov (United States)

    Sun, Le-Chang; Zhou, Li-Gen; Du, Cui-Hong; Cai, Qiu-Feng; Hara, Kenji; Su, Wen-Jin; Cao, Min-Jie

    2009-06-24

    Glucose-6-phosphate isomerase (GPI) was purified to homogeneity from the skeletal muscle of crucian carp ( Carassius auratus ) by ammonium sulfate fractionation, column chromatographies of Q-Sepharose, SP-Sepharose, and Superdex 200 with a yield of 8.0%, and purification folds of 468. The molecular mass of GPI was 120 kDa as estimated by gel filtration, while on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), two subunits (55 and 65 kDa) were identified, suggesting that it is a heterodimer. Interestingly, GPI revealed specific inhibitory activity toward a myofibril-bound serine proteinase (MBSP) from crucian carp, while no inhibitory activity was identified toward other serine proteinases, such as white croaker MBSP and crucian carp trypsin. Kinetic analysis showed that GPI is a competitive inhibitor toward MBSP, and the K(i) was 0.32 microM. Our present results indicated that the multifunctional protein GPI is an endogenous inhibitor to MBSP and may play a significant role in the regulation of muscular protein metabolism in vivo.

  17. Rapid Characterization of Insulin Modifications and Sequence Variations by Proteinase K Digestion and UHPLC-ESI-MS

    Science.gov (United States)

    Yang, Rong-Sheng; Tang, Weijuan; Sheng, Huaming; Meng, Fanyu

    2018-01-01

    Discovery of novel insulin analogs as therapeutics has remained an active area of research. Compared with native human insulin, insulin analog molecules normally incorporate either covalent modifications or amino acid sequence variations. From the drug discovery and development perspective, methods for efficient and detailed characterization of these primary structural changes are very important. In this report, we demonstrate that proteinase K digestion coupled with UPLC-ESI-MS analysis provides a simple and rapid approach to characterize the modifications and sequence variations of insulin molecules. A commercially available proteinase K digestion kit was used to process recombinant human insulin (RHI), insulin glargine, and fluorescein isothiocynate-labeled recombinant human insulin (FITC-RHI) samples. The LC-MS data clearly showed that RHI and insulin glargine samples can be differentiated, and the FITC modifications in all three amine sites of the RHI molecule are well characterized. The end-to-end experiment and data interpretation was achieved within 60 min. This approach is fast and simple, and can be easily implemented in early drug discovery laboratories to facilitate research on more advanced insulin therapeutics. [Figure not available: see fulltext.

  18. Alpha scintillation radon counting

    International Nuclear Information System (INIS)

    Lucas, H.F. Jr.

    1977-01-01

    Radon counting chambers which utilize the alpha-scintillation properties of silver activated zinc sulfide are simple to construct, have a high efficiency, and, with proper design, may be relatively insensitive to variations in the pressure or purity of the counter filling. Chambers which were constructed from glass, metal, or plastic in a wide variety of shapes and sizes were evaluated for the accuracy and the precision of the radon counting. The principles affecting the alpha-scintillation radon counting chamber design and an analytic system suitable for a large scale study of the 222 Rn and 226 Ra content of either air or other environmental samples are described. Particular note is taken of those factors which affect the accuracy and the precision of the method for monitoring radioactivity around uranium mines

  19. Combining Alphas via Bounded Regression

    Directory of Open Access Journals (Sweden)

    Zura Kakushadze

    2015-11-01

    Full Text Available We give an explicit algorithm and source code for combining alpha streams via bounded regression. In practical applications, typically, there is insufficient history to compute a sample covariance matrix (SCM for a large number of alphas. To compute alpha allocation weights, one then resorts to (weighted regression over SCM principal components. Regression often produces alpha weights with insufficient diversification and/or skewed distribution against, e.g., turnover. This can be rectified by imposing bounds on alpha weights within the regression procedure. Bounded regression can also be applied to stock and other asset portfolio construction. We discuss illustrative examples.

  20. Interleukin-21, B cell activating factor and unmethylated CPG oligodeoxynucleotides synergize in promoting anti-proteinase 3 autoantibody production in vitro

    NARCIS (Netherlands)

    Lepse, Nikola; Land, Judith; Rutgers, Abraham; Kallenberg, Cornelis; Stegeman, Coen A.; Heeringa, Peter; Abdulahad, Wayel H.

    2012-01-01

    Background/Purpose: Anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitides (AAV) are characterized by the presence of circulating autoantibodies that are often directed against proteinase 3 (PR3). Although the mechanisms that lead to ANCA production in AAV are not clear, bacterial

  1. The propeptide is required for in vivo formation of stable active yeast proteinase A and can function even when not covalently linked to the mature region

    DEFF Research Database (Denmark)

    van den Hazel, H B; Kielland-Brandt, Morten; Winther, Jakob R.

    1993-01-01

    The PEP4-encoded aspartate protease proteinase A from Saccharomyces cerevisiae is synthesized as a zymogen (Ammerer, G., Hunter, C. P., Rothman, J. H., Saari, G. C., Valls, L. A., and Stevens, T. H. (1986) Mol. Cell. Biol. 6, 2490-2499; Woolford, C. A., Daniels, L. B., Park, F. J., Jones, E. W., ...

  2. Characterisation of cysteine proteinases responsible for digestive proteolysis in guts of larval Western corn rootworm (Diabrotica virgifera) by expression in the yeast Pichia pastoris

    NARCIS (Netherlands)

    Bown, D.P.; Wilkinson, H.S.; Jongsma, M.A.; Gatehouse, J.A.

    2004-01-01

    Cysteine proteinases are the major class of enzymes responsible for digestive proteolysis in western corn rootworm (Diabrotica virgifera), a serious pest of maize. A larval gut extract hydrolysed typical cathepsin substrates, such as Z-phe-arg-AMC and Z-arg-arg-AMC, and hydrolysis was inhibited by

  3. Cysteine proteinase inhibitor cystatin C in squamous cell carcinoma of the head and neck: relation to prognosis.

    Science.gov (United States)

    Strojan, P; Oblak, I; Svetic, B; Smid, L; Kos, J

    2004-05-17

    To determine the role of the cysteine proteinase inhibitor cystatin C in the invasive behavior of squamous cell carcinoma of the head and neck (SCCHN), Cystatin C protein level was measured in 82 pairs of primary tumour tissue and adjacent noncancerous mucosa, using the enzyme-linked immunosorbent assay. The median level of cystatin C in tumour tissue was 1.18 times lower than that in corresponding mucosa (P=0.031). In normal mucosa samples, the cystatin C level was influenced by the site of sampling: it was lower in nonlaryngeal tissue samples (oral cavity, oro- or hypopharynx) than in laryngeal samples (P=0.004). The tumour cystatin C level correlated inversely with pN-stage (P=0.047), whereas a trend of lower cystatin C levels was observed in the group with extranodal tumour extension compared to those with no extranodal spread (P=0.069). In univariate analysis, the patients with low tumour cystatin C levels exhibited poor disease-free survival (DFS, P=0.013) and disease-specific survival (DSS, P=0.013). In multivariate analysis, the most powerful predictor of survival was pN-stage (DFS: P=0.040, HR 2.78; DSS: P=0.011, HR 4.36,), followed by the cystatin C level (DFS: P=0.043, HR 0.22; DSS: P=0.067, HR 0.25). When comparing the prognostic strength of cystatin C to that of stefin A, another cysteine proteinase inhibitor, which emerged as the most significant prognosticator for survival in our previous study analysing the same cohort of patients, stefin A proved to be significantly more reliable predictor for both DFS and DSS than cystatin C. Our results indicate that cystatin C is implicated in the invasive behavior of SCCHN, and that there are variations in regulation of proteolytic pathways under nonmalignant conditions, inherent to individual subsites inside the upper aerodigestive tract. The correlation between high cystatin C levels and improved survival concurs with the concept of the protective role of high levels of cysteine proteinase inhibitors in

  4. Treatment of alpha bearing wastes

    International Nuclear Information System (INIS)

    1988-01-01

    This report deals with the current state of the art of alpha waste treatment, which is an integral part of the overall nuclear waste management system. The International Atomic Energy Agency (IAEA) defines alpha bearing waste as 'waste containing one or more alpha emitting radionuclides, usually actinides, in quantities above acceptable limits'. The limits are established by national regulatory bodies. The limits above which wastes are considered as alpha contaminated refer to the concentrations of alpha emitters that need special consideration for occupational exposures and/or potential safety, health, or environmental impact during one or more steps from generation through disposal. Owing to the widespread use of waste segregation by source - that is, based upon the 'suspect origin' of the material - significant volumes of waste are being handled as alpha contaminated which, in fact, do not require such consideration by reason of risk or environmental concern. The quantification of de minimis concepts by national regulatory bodies could largely contribute to the safe reduction of waste volumes and associated costs. Other factors which could significantly contribute to the reduction of alpha waste arisings are an increased application of assaying and sorting, instrumentation and the use of feedback mechanisms to control or modify the processes which generate these wastes. Alpha bearing wastes are generated during fabrication and reprocessing of nuclear fuels, decommissioning of alpha contaminated facilities, and other activities. Most alpha wastes are contact handled, but a small portion may require shielding or remote handling because of high levels of neutron (n), beta (β), or gamma (γ) emissions associated with the waste material. This report describes the sources and characteristics of alpha wastes and strategies for alpha waste management. General descriptions of treatment processes for solid and liquid alpha wastes are included. 71 refs, 14 figs, 9 tabs

  5. The alpha effect

    International Nuclear Information System (INIS)

    Anon.

    1982-01-01

    Much of the recent interest in RAM system reliability stems from concern over alpha particle soft error rates reported for the initial 64 k RAMs. With increasing memory density likely in the next few years the problem of soft errors is rearing its head again. A few years ago ITT carried out experiments on 16k RAMs and found no significant problems. However, recent tests have shown a raise in the number of soft errors with 64k RAMs, and the launch of 256k and 512k memories is likely to make the problem acute

  6. Alpha-mannosidosis

    Directory of Open Access Journals (Sweden)

    Nilssen Øivind

    2008-07-01

    Full Text Available Abstract Alpha-mannosidosis is an inherited lysosomal storage disorder characterized by immune deficiency, facial and skeletal abnormalities, hearing impairment, and intellectual disability. It occurs in approximately 1 of 500,000 live births. The children are often born apparently normal, and their condition worsens progressively. Some children are born with ankle equinus or develop hydrocephalus in the first year of life. Main features are immune deficiency (manifested by recurrent infections, especially in the first decade of life, skeletal abnormalities (mild-to-moderate dysostosis multiplex, scoliosis and deformation of the sternum, hearing impairment (moderate-to-severe sensorineural hearing loss, gradual impairment of mental functions and speech, and often, periods of psychosis. Associated motor function disturbances include muscular weakness, joint abnormalities and ataxia. The facial trait include large head with prominent forehead, rounded eyebrows, flattened nasal bridge, macroglossia, widely spaced teeth, and prognathism. Slight strabismus is common. The clinical variability is significant, representing a continuum in severity. The disorder is caused by lysosomal alpha-mannosidase deficiency. Alpha-mannosidosis is inherited in an autosomal recessive fashion and is caused by mutations in the MAN2B1 gene located on chromosome 19 (19 p13.2-q12. Diagnosis is made by measuring acid alpha-mannosidase activity in leukocytes or other nucleated cells and can be confirmed by genetic testing. Elevated urinary secretion of mannose-rich oligosaccharides is suggestive, but not diagnostic. Differential diagnoses are mainly the other lysosomal storage diseases like the mucopolysaccharidoses. Genetic counseling should be given to explain the nature of the disease and to detect carriers. Antenatal diagnosis is possible, based on both biochemical and genetic methods. The management should be pro-active, preventing complications and treating

  7. Alpha activity measurement with lsc

    International Nuclear Information System (INIS)

    Dobrin, R. I.; Dulama, C. N.; Ciocirlan, C. N.; Toma, A.; Stoica, S. M.; Valeca, M.

    2013-01-01

    Recently, we showed that the alpha activity in liquid samples can be measured using a liquid scintillation analyzer without alpha/beta discrimination capability. The purpose of this work was to evaluate the performances of the method and to optimize the procedure of the sample preparation. A series of tests was performed to validate the procedure of alpha emitting radionuclides extraction in aqueous samples with Actinide Resin, especially regarding to the contact time required to extract all alpha nuclides. The main conclusions were that a minimum 18 hours stirring time is needed to achieve a percent recovery of the alpha nuclides grater than 90% and that the counting efficiency of alphas measurements with LSC is nearly 100%. (authors)

  8. 18F-FDG PET/CT findings preceded elevation of serum proteinase 3 antineutrophil cytoplasmic antibodies in Wegener granulomatosis.

    Science.gov (United States)

    Ito, Kimiteru; Minamimoto, Ryogo; Yamashita, Hiroyuki; Morooka, Miyako; Okasaki, Momoko; Mimori, Akio; Kubota, Kazuo

    2014-01-01

    A 67-year-old woman underwent F-FDG PET/CT after developing a fever of unknown origin. PET/CT revealed intensive FDG uptake at the nasal and lung lesions. On the laboratory data, serum myeloperoxidase antineutrophil cytoplasmic antibodies (ANCA) titer was elevated, although serum directed against proteinase 3 (PR3) ANCA titer was within normal limits. One month after treatment, follow-up PET/CT revealed decreased FDG uptake at the lesions. One year later, serum PR3-ANCA titer elevated, which finally led to a diagnosis of Wegener granulomatosis (WG). WG lesions may be detected earlier by FDG PET/CT than by serum PR3-ANCA titers.

  9. The urinary excretion of epidermal growth factor in the rat is reduced by aprotinin, a proteinase inhibitor

    DEFF Research Database (Denmark)

    Jørgensen, P E; Raaberg, Lasse; Poulsen, Steen Seier

    1990-01-01

    of immunoreactive EGF in the kidney tissue is increased after aprotinin administration (median amount 0.11 pmol EGF/mg protein versus less than 0.04 pmol EGF/mg protein, P less than 0.001). Neither the creatinine clearance, the total urinary protein output, nor the volume of urine produced was affected by aprotinin....... in vivo is processed by an aprotinin inhibitable proteinase. EGF is produced in the kidneys as a precursor with a molecular weight of approximately 130 kDa. In rat urine, nanomolar amounts of 6 kDa EGF are excreted per 24 h together with small amounts of high molecular weight forms of EGF. During i...

  10. Synthesis of the proteinase inhibitor LEKTI domain 6 by the fragment condensation method and regioselective disulfide bond formation.

    Science.gov (United States)

    Vasileiou, Zoe; Barlos, Kostas K; Gatos, Dimitrios; Adermann, Knut; Deraison, Celine; Barlos, Kleomenis

    2010-01-01

    Proteinase inhibitors are of high pharmaceutical interest and are drug candidates for a variety of indications. Specific kallikrein inhibitors are important for their antitumor activity and their potential application to the treatment of skin diseases. In this study we describe the synthesis of domain 6 of the kallikrein inhibitor Lympho-Epithilial Kazal-Type Inhibitor (LEKTI) by the fragment condensation method and site-directed cystine bridge formation. To obtain the linear LEKTI precursor, the condensation was best performed in solution, coupling the protected fragment 1-22 to 23-68. This method yielded LEKTI domain 6 of high purity and equipotent to the recombinantly produced peptide. (c) 2010 Wiley Periodicals, Inc.

  11. Induction of protective immunity in cattle against infection with Fasciola hepatica by vaccination with cathepsin L proteinases and with hemoglobin.

    Science.gov (United States)

    Dalton, J P; McGonigle, S; Rolph, T P; Andrews, S J

    1996-01-01

    Two cathepsin L proteinases, cathepsin L1 and cathepsin L2, secreted by liver flukes may be involved in tissue penetration, nutrition, and protection from immune attack. To ascertain the immunoprophylactic potential of these proteinases, and of another molecule, liver fluke hemoglobin (Hb), we performed vaccine trials in cattle. In the first vaccine trial various doses of cathepsin L1 were tested. The mean protection level obtained was 53.7%. In a second vaccine trial cathepsin L1 and Hb elicited 42.5 and 43.8% protection levels, respectively, while a combination of the two molecules induced a significantly higher level of protection (51.9%). Cathepsin L2 was not examined alone; however, vaccination of cattle with a combination of cathepsin L2 and Hb elicited the highest level of protection (72.4%). The animals that received cathepsin L1-Hb or cathepsin L2-Hb showed reduced liver damage as assessed by serum glutamic dehydrogenase and gamma-glutamyl transferase levels. Furthermore, a reduced viability was observed for fluke eggs recovered from all vaccine groups. This anti-embryonation effect of vaccination was particularly evident in the group that received cathepsin L2-Hb where >98% of the eggs recovered did not embryonate to miracidia. Although all vaccine preparations induced high antibody titers which were boosted following the challenge infection, there was no correlation between antibody titers and protection. The results of these trials demonstrate that cathepsin Ls and Hb could form the basis of a molecular vaccine that would not only reduce parasite burden but would also prevent transmission of liver fluke disease. PMID:8945548

  12. The 3D structure and function of digestive cathepsin L-like proteinases of Tenebrio molitor larval midgut.

    Science.gov (United States)

    Beton, Daniela; Guzzo, Cristiane R; Ribeiro, Alberto F; Farah, Chuck S; Terra, Walter R

    2012-09-01

    Cathepsin L-like proteinases (CAL) are major digestive proteinases in the beetle Tenebrio molitor. Procathepsin Ls 2 (pCAL2) and 3 (pCAL3) were expressed as recombinant proteins in Escherichia coli, purified and activated under acidic conditions. Immunoblot analyses of different T. molitor larval tissues demonstrated that a polyclonal antibody to pCAL3 recognized pCAL3 and cathepsin L 3 (CAL3) only in the anterior two-thirds of midgut tissue and midgut luminal contents of T. molitor larvae. Furthermore, immunocytolocalization data indicated that pCAL3 occurs in secretory vesicles and microvilli in anterior midgut. Therefore CAL3, like cathepsin L 2 (CAL2), is a digestive enzyme secreted by T. molitor anterior midgut. CAL3 hydrolyses Z-FR-MCA and Z-RR-MCA (typical cathepsin substrates), whereas CAL2 hydrolyses only Z-FR-MCA. Active site mutants (pCAL2C25S and pCAL3C26S) were constructed by replacing the catalytic cysteine with serine to prevent autocatalytic processing. Recombinant pCAL2 and pCAL3 mutants (pCAL2C25S and pCAL3C26S) were prepared, crystallized and their 3D structures determined at 1.85 and 2.1 Å, respectively. While the overall structure of these enzymes is similar to other members of the papain superfamily, structural differences in the S2 subsite explain their substrate specificities. The data also supported models for CAL trafficking to lysosomes and to secretory vesicles to be discharged into midgut contents. Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Patterns and determinants of acute psychiatric readmissions | Behr ...

    African Journals Online (AJOL)

    Objectives. Deinstitutionalisation and shortage of psychiatric beds worldwide has led to extensive research into the risk factors and interventions associated with rapid and recurrent admission to hospital. Little research of this nature has taken place in South Africa, particularly with regard to acute hospital admissions.

  14. Patterns and determinants of acute psychiatric readmissions | Behr ...

    African Journals Online (AJOL)

    Little research of this nature has taken place in South Africa, particularly with regard to acute hospital admissions. This study attempted primarily to assess the effect of length of stay and administration of depot antipsychotics in hospital on time to readmission. Design. A retrospective cohort of 180 admissions was followed up ...

  15. Molecular characterization of alpha 1- and alpha 2-adrenoceptors.

    Science.gov (United States)

    Harrison, J K; Pearson, W R; Lynch, K R

    1991-02-01

    Three 'alpha 1-adrenoceptors' and three 'alpha 2-adrenoceptors' have now been cloned. How closely do these receptors match the native receptors that have been identified pharmacologically? What are the properties of these receptors, and how do they relate to other members of the cationic amine receptor family? Kevin Lynch and his colleagues discuss these questions in this review.

  16. Alpha wastes treatment

    International Nuclear Information System (INIS)

    Thouvenot, P.

    2000-01-01

    Alter 2004, the alpha wastes issued from the Commissariat a l'Energie Atomique installations will be sent to the CEDRA plant. The aims of this installation are decontamination and wastes storage. Because of recent environmental regulations concerning ozone layer depletion, the use of CFC 113 in the decontamination unit, as previously planned, is impossible. Two alternatives processes are studied: the AVD process and an aqueous process including surfactants. Best formulations for both processes are defined issuing degreasing kinetics. It is observed that a good degreasing efficiency is linked to a good decontamination efficiency. Best results are obtained with the aqueous process. Furthermore, from the point of view of an existing waste treatment unit, the aqueous process turns out to be more suitable than the AVD process. (author)

  17. Mind Your p's and Alphas.

    Science.gov (United States)

    Stallings, William M.

    In the educational research literature alpha, the a priori level of significance, and p, the a posteriori probability of obtaining a test statistic of at least a certain value when the null hypothesis is true, are often confused. Explanations for this confusion are offered. Paradoxically, alpha retains a prominent place in textbook discussions of…

  18. Summary of Alpha Particle Transport

    Energy Technology Data Exchange (ETDEWEB)

    Medley, S.S.; White, R.B.; Zweben, S.J.

    1998-08-19

    This paper summarizes the talks on alpha particle transport which were presented at the 5th International Atomic Energy Agency's Technical Committee Meeting on "Alpha Particles in Fusion Research" held at the Joint European Torus, England in September 1997.

  19. Proteinaceous alpha-araylase inhibitors

    DEFF Research Database (Denmark)

    Svensson, Birte; Fukuda, Kenji; Nielsen, P.K.

    2004-01-01

    Proteins that inhibit alpha-amylases have been isolated from plants and microorganisms. These inhibitors can have natural roles in the control of endogenous a-amylase activity or in defence against pathogens and pests; certain inhibitors are reported to be antinutritional factors. The alpha-amylase...... inhibitors belong to seven different protein structural families, most of which also contain evolutionary related proteins without inhibitory activity. Two families include bifunctional inhibitors acting both on alpha-amylases and proteases. High-resolution structures are available of target alpha-amylases...... in complex with inhibitors from five families. These structures indicate major diversity but also some similarity in the structural basis of alpha-amylase inhibition. Mutational analysis of the mechanism of inhibition was performed in a few cases and various protein engineering and biotechnological...

  20. A group-specific inhibitor of lysosomal cysteine proteinases selectively inhibits both proteolytic degradation and presentation of the antigen dinitrophenyl-poly-L-lysine by guinea pig accessory cells to T cells

    DEFF Research Database (Denmark)

    Buus, S; Werdelin, O

    1986-01-01

    A limited intralysosomal proteolytic degradation is probably a key event in the accessory cell processing of large protein antigens before their presentation to T cells. With the aid of highly specific inhibitors of proteinases, we have examined the role of proteolysis in the presentation...... of antigens by guinea pig accessory cells. The proteinase inhibitor benzyloxycarbonyl-phenylalanylalanine-diazomethyl-ketone, which selectively inhibits cysteine proteinases, was used to block this set of enzymes in cultured cells. We demonstrate that the selective inhibition of the cysteine proteinases...... of antigen-presenting cells causes a profound inhibition of both the proteolytic degradation and the presentation of the synthetic antigen dinitrophenyl-poly-L-lysine. In contrast, the presentation of another synthetic antigen, the copolymer of L-glutamic acid and L-alanine, was enhanced by the same...

  1. ALPHA freezes antiprotons

    CERN Multimedia

    CERN Bulletin

    2010-01-01

    Laboratories like CERN can routinely produce many different types of antiparticles. In 1995, the PS210 experiment formed the first antihydrogen atoms and a few years later, in 2002, ATRAP and ATHENA were already able to produce several thousand of them. However, no experiment in the world has succeeded in ‘trapping’ these anti-atoms in order to study them. This is the goal of the ALPHA experiment, which has recently managed to cool down the antiprotons to just a few Kelvin. This represents a major step towards trapping the anti-atom, thus opening a new avenue into the investigation of antimatter properties.   Members of the ALPHA collaboration working on the apparatus in the Antiproton Decelerator experimental hall at CERN. Just like the atom, the anti-atom is neutral. Unlike the atom, the anti-atom is made up of antiprotons (as opposed to protons in the atom) and positrons (as opposed to electrons). In order to thoroughly study the properties of the anti-atoms, scien...

  2. Alpha-1-antitrypsin deficiency.

    Science.gov (United States)

    Bals, Robert

    2010-10-01

    Alpha-1-antitrypsin deficiency (AATD) is a rare genetic disorder associated with the development of liver and lung disease. AAT is a 52-kD glycoprotein, produced mainly by hepatocytes and secreted into the blood. Agglomeration of the AAT-protein in hepatocytes can result in liver disease. Exposure to smoke is the major risk factor for the development of lung disease characterised as early chronic obstructive lung disease (COPD). Diagnosis is based on the analysis of the AAT genotype and phenotype. The measurement of the AAT serum level is useful as screening test. Liver biopsy is not necessary to establish the diagnosis. Therapy for AAT-related liver disease is supportive, a specific therapy is not available. AATD is a rare condition (1:5000-10000) and, as a consequence, data and information on diagnosis and treatment are not easily accessible. This chapter provides a comprehensive overview on AATD, covering basic biology, diagnostic and therapeutic approaches. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Measurement of $\\alpha_{s}$ with Radiative Hadronic Events

    CERN Document Server

    Abbiendi, G; Åkesson, P F; Alexander, G; Anagnostou, G; Anderson, K J; Asai, S; Axen, D; Bailey, I; Barberio, E; Barillari, T; Barlow, R J; Batley, R J; Bechtle, P; Behnke, T; Bell, K W; Bell, P J; Bella, G; Bellerive, A; Benelli, G; Bethke, S; Biebel, O; Boeriu, O; Bock, P; Boutemeur, M; Braibant, S; Brown, R M; Burckhart, H J; Campana, S; Capiluppi, P; Carnegie, R K; Carter, A A; Carter, J R; Chang, C Y; Charlton, D G; Ciocca, C; Csilling, A; Cuffiani, M; Dado, S; Dallavalle, M; de Roeck, A; De Wolf, E A; Desch, K; Dienes, B; Dubbert, J; Duchovni, E; Duckeck, G; Duerdoth, I P; Etzion, E; Fabbri, F; Ferrari, P; Fiedler, F; Fleck, I; Ford, M; Frey, A; Gagnon, P; Gary, J W; Geich-Gimbel, C; Giacomelli, G; Giacomelli, P; Giunta, M; Goldberg, J; Gross, E; Grunhaus, J; Gruwé, M; Sen-Gupta, A; Hajdu, C; Hamann, M; Hanson, G G; Harel, A; Hauschild, M; Hawkes, C M; Hawkings, R; Herten, G; Heuer, R D; Hill, J C; Horváth, D; Igo-Kemenes, P; Ishii, K; Jeremie, H; Jovanovic, P; Junk, T R; Kanzaki, J; Karlen, D; Kawagoe, K; Kawamoto, T; Keeler, R K; Kellogg, R G; Kennedy, B W; Kluth, S; Kobayashi, T; Kobel, M; Komamiya, S; Kramer, T; Krasznahorkays, A Jr; Krieger, P; Von Krogh, J; Kühl, T; Kupper, M; Lafferty, G D; Landsman, H; Lanske, D; Lellouch, D; Letts, J; Levinson, L; Lillich, J; Lloyd, S L; Loebinger, F K; Lü, J; Ludwig, A; Ludwig, J; Mader, W; Marcellini, S; Martin, A J; Mashimo, T; Mättig, P; McKenna, J; McPherson, R A; Meijers, F; Menges, W; Merritt, F S; Mes, H; Meyer, N; Michelini, A; Mihara, S; Mikenberg, G; Miller, D J; Mohr, W; Mori, T; Mutter, A; Nagai, K; Nakamura, I; Nanjo, H; Neal, H A; O'Neale, S W; Oh, A; Oreglia, M J; Orito, S; Pahl, C; Pásztor, G; Pater, J R; Pilcher, J E; Pinfold, J L; Plane, D E; Pooth, O; Przybycien, M; Quadt, A; Rabbertz, K; Rembser, C; Renkel, P; Roney, J M; Rossi, A M; Rozen, Y; Runge, K; Sachs, K; Saeki, T; Sarkisyan-Grinbaum, E; Schaile, A D; Schaile, O; Scharff-Hansen, P; Schiecks, J; Schörner-Sadenius, T; Schröder, M; Schumacher, M; Seuster, R; Shears, T G; Shen, B C; Sherwood, P; Skuja, A; Smith, A M; Sobie, R J; Söldner-Rembold, S; Spanó, F; Stahl, A; Strom, D; Ströhmer, R; Tarem, S; Tasevsky, M; Teuscher, R; Thomson, M A; Torrence, E; Toya, D; Trigger, I; Trócsányi, Z L; Tsur, E; Turner-Watson, M F; Ueda, I; Ujvári, B; Vollmer, C F; Vannerem, P; Vertesi, R; Verzocchi, M; Voss, H; Vossebeld, J; Ward, C P; Ward, D R; Watkins, P M; Watson, A T; Watson, N K; Wells, P S; Wengler, T; Wermes, N; Wilson, G W; Wilson, J A; Wolf, G; Wyatt, T R; Yamashita, S; Zer-Zion, D; Zivkovic, L

    2008-01-01

    Hadronic final states with a hard isolated photon are studied using data taken at centre-of-mass energies around the mass of the Z0 boson with the OPAL detector at LEP. The strong coupling alpha S is extracted by comparing data and QCD predictions for event shape observables at average reduced centre-of-mass energies ranging from 24 GeV to 78 GeV, and the energy dependence of alpha S is studied. Our results are consistent with the running of alpha S as predicted by QCD and show that within the uncertainties of our analysis event shapes in hadronic Z0 decays with hard and isolated photon radiation can be described by QCD at reduced centre-of-mass energies. Combining all values from different event shape observables and energies gives alpha S (Mz)=0.1182 pm 0.0015(stat.) pm 0.0101(syst.).

  4. Effect Of Five Proteases Including Alcalase, Flavourzyme, Papain, Proteinase K And Trypsin On Antioxidative Activities Of Casein Hydrolysate From Goat Milk

    OpenAIRE

    Shu Guowei; Zhang Qian; Chen He; Wan Hongchang; Li Hong

    2015-01-01

    Oxidation was related to the pathogenesis of human diseases. Adequate intake of antioxidant activity of food can reduce the levels of free radicals, prevent lipid peroxidation, and help the body against diseases. In the paper, casein from goat milk was hydrolyzed by five commercial proteases, namely, Alcalase, flavourzyme, papain, proteinase K and trypsin. The antioxidant activities of casein hydrolysates were assessed by evaluating hydrolysis degree, DPPH radical-scavenging activity, metal-c...

  5. What Powers Lyman alpha Blobs?

    OpenAIRE

    Ao, Y.; Matsuda, Y.; Beelen, A.; Henkel, C.; Cen, R.; De Breuck, C.; Francis, P.; Kovacs, A.; Lagache, G.; Lehnert, M.; Mao, M.; Menten, K. M.; Norris, R.; Omont, A.; Tatemastu, K.

    2015-01-01

    Lyman alpha blobs (LABs) are spatially extended lyman alpha nebulae seen at high redshift. The origin of Lyman alpha emission in the LABs is still unclear and under debate. To study their heating mechanism(s), we present Australia Telescope Compact Array (ATCA) observations of the 20 cm radio emission and Herschel PACS and SPIRE measurements of the far-infrared (FIR) emission towards the four LABs in the protocluster J2143-4423 at z=2.38. Among the four LABs, B6 and B7 are detected in the rad...

  6. Alpha heating in toroidal devices

    Energy Technology Data Exchange (ETDEWEB)

    Miley, G.H.

    1978-01-01

    Ignition (or near-ignition) by alpha heating is a key objective for the achievement of economic fusion reactors. While good confinement of high-energy alphas appears possible in larger reactors, near-term tokamak-type ignition experiments as well as some concepts for small reactors (e.g., the Field-Reversed Mirror or FRM) potentially face marginal situations. Consequently, there is a strong motivation to develop methods to evaluate alpha losses and heating profiles in some detail. Such studies for a TFTR-size tokamak and for a small FRM are described here.

  7. Investigations on the activation of recombinant microbial pro-transglutaminase: in contrast to proteinase K, dispase removes the histidine-tag.

    Science.gov (United States)

    Sommer, Christian; Hertel, Thomas C; Schmelzer, Christian E H; Pietzsch, Markus

    2012-02-01

    In order to produce recombinant microbial transglutaminase (rMTG) which is free of the activating protease, dispase was used to activate the pro-rMTG followed by immobilized metal affinity chromatography (IMAC). As shown by MALDI-MS, the dispase does not only cleave the pro-sequence, but unfortunately also cleaves within the C-terminal histidine-tag. Hence, the active rMTG cannot properly bind to the IMAC material. As an alternative, proteinase K was investigated. This protease was successfully applied for the activation of purified pro-rMTG either as free or immobilized enzyme and the free enzyme was also applicable directly in the crude cell extract of E. coli. Thus, it enables a simple two-step activation/purification procedure resulting in protease-free and almost pure transglutaminase preparations. The protocol has been successfully applied to both, wild-type transglutaminase of Streptomyces mobaraensis as well as to the highly active variant S2P. Proteinase K activates the pro-rMTG without unwanted degradation of the histidine-tag. It turned out to be very important to inhibit proteinase K activity, e.g., by PMSF, prior to protein separation by SDS-PAGE.

  8. High-resolution X-ray study of the effects of deuteration on crystal growth and the crystal structure of proteinase K

    International Nuclear Information System (INIS)

    Chatake, Toshiyuki; Ishikawa, Takuya; Yanagisawa, Yasuhide; Yamada, Taro; Tanaka, Ichiro; Fujiwara, Satoru; Morimoro, Yukio

    2011-01-01

    A high-resolution X-ray crystallographic study of the effects of solvent deuteration on the crystallization of proteinase K shows negligibly small degradations of the crystals owing to solvent deuteration and small structural differences between nondeuterated and deuterated crystals of proteinase K. Deuteration of macromolecules is an important technique in neutron protein crystallography. Solvent deuteration of protein crystals is carried out by replacing water (H 2 O) with heavy water (D 2 O) prior to neutron diffraction experiments in order to diminish background noise. The effects of solvent deuteration on the crystallization of proteinase K (PK) with polyethylene glycol as a precipitant were investigated using high-resolution X-ray crystallography. In previous studies, eight NO 3 − anions were included in the PK crystal unit cell grown in NaNO 3 solution. In this study, however, the PK crystal structure did not contain NO 3 − anions; consequently, distortions of amino acids arising from the presence of NO 3 − anions were avoided in the present crystal structures. High-resolution (1.1 Å) X-ray diffraction studies showed that the degradation of PK crystals induced by solvent deuteration was so small that this degradation would be negligible for the purpose of neutron protein crystallography experiments at medium resolution. Comparison of the nonhydrogen structures of nondeuterated and deuterated crystal structures demonstrated very small structural differences. Moreover, a positive correlation between the root-mean-squared differences and B factors indicated that no systematic difference existed

  9. Effect Of Five Proteases Including Alcalase, Flavourzyme, Papain, Proteinase K And Trypsin On Antioxidative Activities Of Casein Hydrolysate From Goat Milk

    Directory of Open Access Journals (Sweden)

    Shu Guowei

    2015-12-01

    Full Text Available Oxidation was related to the pathogenesis of human diseases. Adequate intake of antioxidant activity of food can reduce the levels of free radicals, prevent lipid peroxidation, and help the body against diseases. In the paper, casein from goat milk was hydrolyzed by five commercial proteases, namely, Alcalase, flavourzyme, papain, proteinase K and trypsin. The antioxidant activities of casein hydrolysates were assessed by evaluating hydrolysis degree, DPPH radical-scavenging activity, metal-chelating activity and superoxide radical scavenging activity. The results showed as follows: the DH of proteinase K, Alcalase, and trypsin were higher significantly than those of papain and flavourzyme. The Fe2+-chelation activity and superoxide radical scavenging activity of casein hydrolysates from goat milk by Alcalase was higher than the others, the DPPH scavenging activities of casein hydrolysates by Alcalase and papain was higher than the others and the DPPH scavenging activities by Alcalase and papain had no significant diffierence (p<0.05, so the optimal proteinase for hydrolysis casein from goat milk to produce antioxidant peptide was Alcalase.

  10. The role of salt bridges on the temperature adaptation of aqualysin I, a thermostable subtilisin-like proteinase.

    Science.gov (United States)

    Jónsdóttir, Lilja B; Ellertsson, Brynjar Ö; Invernizzi, Gaetano; Magnúsdóttir, Manuela; Thorbjarnardóttir, Sigríður H; Papaleo, Elena; Kristjánsson, Magnús M

    2014-12-01

    Differences in salt bridges are believed to be a structural hallmark of homologous enzymes from differently temperature-adapted organisms. Nevertheless, the role of salt bridges on structural stability is still controversial. While it is clear that most buried salt bridges can have a functional or structural role, the same cannot be firmly stated for ion pairs that are exposed on the protein surface. Salt bridges, found in X-ray structures, may not be stably formed in solution as a result of high flexibility or high desolvation penalty. More studies are thus needed to clarify the picture on salt bridges and temperature adaptation. We contribute here to this scenario by combining atomistic simulations and experimental mutagenesis of eight mutant variants of aqualysin I, a thermophilic subtilisin-like proteinase, in which the residues involved in salt bridges and not conserved in a psychrophilic homolog were systematically mutated. We evaluated the effects of those mutations on thermal stability and on the kinetic parameters. Overall, we show here that only few key charged residues involved in salt bridges really contribute to the enzyme thermal stability. This is especially true when they are organized in networks, as here attested by the D17N mutation, which has the most remarkable effect on stability. Other mutations had smaller effects on the properties of the enzyme indicating that most of the isolated salt bridges are not a distinctive trait related to the enhanced thermal stability of the thermophilic subtilase. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Optimization of conditions to extract high quality DNA for PCR analysis from whole blood using SDS-proteinase K method

    Directory of Open Access Journals (Sweden)

    Wajhul Qamar

    2017-11-01

    Full Text Available In case of studies associated with human genetics, genomics, and pharmacogenetics the genomic DNA is extracted from the buccal cells, whole blood etc. Several methods are exploited by the researchers to extract DNA from the whole blood. One of these methods, which utilizes cell lysis and proteolytic properties of sodium dodecyl sulfate (SDS and proteinase K respectively, might also be called SDS-PK method. It does not include any hazardous chemicals such as phenol or chloroform and is inexpensive. However, several researchers report the same method with different formulas and conditions. During our experiments with whole blood DNA extraction we experienced problems such as protein contamination, DNA purity and yield when followed some SDS-PK protocols reported elsewhere. A260/A280 and A260/A230 ratios along with PCR amplification give a clear idea about the procedure that was followed to extract the DNA. In an effort to increase the DNA purity from human whole blood, we pointed out some steps of the protocol that play a crucial role in determining the extraction of high quality DNA.

  12. Distinct proteinase 3-induced cytokine patterns in Wegener´s granulomatosis, Churg-Strauss syndrome, and healthy controls.

    Science.gov (United States)

    Fagin, Ursula; Csernok, Elena; Müller, Antje; Pitann, Silke; Fazio, Juliane; Krause, Kristina; Bremer, Philip; Wipfler-Freissmuth, Edith; Moosig, Frank; Gross, Wolfgang L; Lamprecht, Peter

    2011-01-01

    To analyse whether a specific cytokine pattern is elicited in response to the autoantigen proteinase 3 (PR3) in active Wegener's granulomatosis (WG). Six-colour flow cytometry was used to analyse cytokine production and surface markers of the total CD4+ T-cell population ex vivo and in PR3-stimulated T-cell lines of patients with active PR3-ANCA-positive WG, PR3-ANCA-negative Churg-Strauss syndrome (CSS), and healthy controls (HC). The cytokine response of the total PB CD4+ T cell population was skewed towards distinct pro-inflammatory cytokine patterns in WG (Th1-type) and CSS (Th17, Th1-/Th2-type). Th2-type as well as Th17 cell populations including Th17/Th1, Th17/Th2 and Th22 cells were elicited in response to PR3 stimulation in WG. In contrast, CSS patients displayed a Th2-type dominated response following PR3 stimulation. These data suggest that the cytokine response of the total CD4+ T-cell population and PR3-specific cells is influenced by the underlying disorder.

  13. Proteinase activated receptor 1 mediated fibrosis in a mouse model of liver injury: a role for bone marrow derived macrophages.

    Directory of Open Access Journals (Sweden)

    Yiannis N Kallis

    Full Text Available Liver fibrosis results from the co-ordinated actions of myofibroblasts and macrophages, a proportion of which are of bone marrow origin. The functional effect of such bone marrow-derived cells on liver fibrosis is unclear. We examine whether changing bone marrow genotype can down-regulate the liver's fibrotic response to injury and investigate mechanisms involved. Proteinase activated receptor 1 (PAR1 is up-regulated in fibrotic liver disease in humans, and deficiency of PAR1 is associated with reduced liver fibrosis in rodent models. In this study, recipient mice received bone marrow transplantation from PAR1-deficient or wild-type donors prior to carbon tetrachloride-induced liver fibrosis. Bone marrow transplantation alone from PAR1-deficient mice was able to confer significant reductions in hepatic collagen content and activated myofibroblast expansion on wild-type recipients. This effect was associated with a decrease in hepatic scar-associated macrophages and a reduction in macrophage recruitment from the bone marrow. In vitro, PAR1 signalling on bone marrow-derived macrophages directly induced their chemotaxis but did not stimulate proliferation. These data suggest that the bone marrow can modulate the fibrotic response of the liver to recurrent injury. PAR1 signalling can contribute to this response by mechanisms that include the regulation of macrophage recruitment.

  14. Anti-proteinase 3 anti-neutrophil cytoplasm autoantibodies recapitulate systemic vasculitis in mice with a humanized immune system.

    LENUS (Irish Health Repository)

    Little, Mark A

    2012-01-01

    Evidence is lacking for direct pathogenicity of human anti-proteinase-3 (PR3) antibodies in development of systemic vasculitis and granulomatosis with polyangiitis (GPA, Wegener\\'s granulomatosis). Progress in study of these antibodies in rodents has been hampered by lack of PR3 expression on murine neutrophils, and by different Fc-receptor affinities for IgG across species. Therefore, we tested whether human anti-PR3 antibodies can induce acute vasculitis in mice with a human immune system. Chimeric mice were generated by injecting human haematopoietic stem cells into irradiated NOD-scid-IL2Rγ⁻\\/⁻ mice. Matched chimera mice were treated with human IgG from patients with: anti-PR3 positive renal and lung vasculitis; patients with non-vasculitic renal disease; or healthy controls. Six-days later, 39% of anti-PR3 treated mice had haematuria, compared with none of controls. There was punctate bleeding on the surface of lungs of anti-PR3 treated animals, with histological evidence of vasculitis and haemorrhage. Anti-PR3 treated mice had mild pauci-immune proliferative glomerulonephritis, with infiltration of human and mouse leukocytes. In 3 mice (17%) more severe glomerular injury was present. There were no glomerular changes in controls. Human IgG from patients with anti-PR3 autoantibodies is therefore pathogenic. This model of anti-PR3 antibody-mediated vasculitis may be useful in dissecting mechanisms of microvascular injury.

  15. Anti-proteinase 3 anti-neutrophil cytoplasm autoantibodies recapitulate systemic vasculitis in mice with a humanized immune system.

    Directory of Open Access Journals (Sweden)

    Mark A Little

    Full Text Available Evidence is lacking for direct pathogenicity of human anti-proteinase-3 (PR3 antibodies in development of systemic vasculitis and granulomatosis with polyangiitis (GPA, Wegener's granulomatosis. Progress in study of these antibodies in rodents has been hampered by lack of PR3 expression on murine neutrophils, and by different Fc-receptor affinities for IgG across species. Therefore, we tested whether human anti-PR3 antibodies can induce acute vasculitis in mice with a human immune system. Chimeric mice were generated by injecting human haematopoietic stem cells into irradiated NOD-scid-IL2Rγ⁻/⁻ mice. Matched chimera mice were treated with human IgG from patients with: anti-PR3 positive renal and lung vasculitis; patients with non-vasculitic renal disease; or healthy controls. Six-days later, 39% of anti-PR3 treated mice had haematuria, compared with none of controls. There was punctate bleeding on the surface of lungs of anti-PR3 treated animals, with histological evidence of vasculitis and haemorrhage. Anti-PR3 treated mice had mild pauci-immune proliferative glomerulonephritis, with infiltration of human and mouse leukocytes. In 3 mice (17% more severe glomerular injury was present. There were no glomerular changes in controls. Human IgG from patients with anti-PR3 autoantibodies is therefore pathogenic. This model of anti-PR3 antibody-mediated vasculitis may be useful in dissecting mechanisms of microvascular injury.

  16. Putrescine-dependent re-localization of TvCP39, a cysteine proteinase involved in Trichomonas vaginalis cytotoxicity.

    Directory of Open Access Journals (Sweden)

    Bertha Isabel Carvajal-Gamez

    Full Text Available Polyamines are involved in the regulation of some Trichomonas vaginalis virulence factors such as the transcript, proteolytic activity, and cytotoxicity of TvCP65, a cysteine proteinase (CP involved in the trichomonal cytotoxicity. In this work, we reported the putrescine effect on TvCP39, other CP that also participate in the trichomonal cytotoxicity. Parasites treated with 1,4-diamino-2-butanone (DAB (an inhibitor of putrescine biosynthesis, diminished the amount and proteolytic activity of TvCP39 as compared with untreated parasites. Inhibition of putrescine biosynthesis also reduced ∼ 80% the tvcp39 mRNA levels according to RT-PCR and qRT-PCR assays. Additionally, actinomycin D-treatment showed that the tvcp39 mRNA half-life decreased in the absence of putrescine. However, this reduction was restored by exogenous putrescine addition, suggesting that putrescine is necessary for tvcp39 mRNA stability. TvCP39 was localized in the cytoplasm but, in DAB treated parasites transferred into exogenous putrescine culture media, TvCP39 was re-localized to the nucleus and nuclear periphery of trichomonads. Interestingly, the amount and proteolytic activity of TvCP39 was recovered as well as the tvcp39 mRNA levels were restored when putrescine exogenous was added to the DAB-treated parasites. In conclusion, our data show that putrescine regulate the TvCP39 expression, protein amount, proteolytic activity, and cellular localization.

  17. Putrescine-dependent re-localization of TvCP39, a cysteine proteinase involved in Trichomonas vaginalis cytotoxicity.

    Science.gov (United States)

    Carvajal-Gamez, Bertha Isabel; Quintas-Granados, Laura Itzel; Arroyo, Rossana; Vázquez-Carrillo, Laura Isabel; Ramón-Luing, Lucero De los Angeles; Carrillo-Tapia, Eduardo; Alvarez-Sánchez, María Elizbeth

    2014-01-01

    Polyamines are involved in the regulation of some Trichomonas vaginalis virulence factors such as the transcript, proteolytic activity, and cytotoxicity of TvCP65, a cysteine proteinase (CP) involved in the trichomonal cytotoxicity. In this work, we reported the putrescine effect on TvCP39, other CP that also participate in the trichomonal cytotoxicity. Parasites treated with 1,4-diamino-2-butanone (DAB) (an inhibitor of putrescine biosynthesis), diminished the amount and proteolytic activity of TvCP39 as compared with untreated parasites. Inhibition of putrescine biosynthesis also reduced ∼ 80% the tvcp39 mRNA levels according to RT-PCR and qRT-PCR assays. Additionally, actinomycin D-treatment showed that the tvcp39 mRNA half-life decreased in the absence of putrescine. However, this reduction was restored by exogenous putrescine addition, suggesting that putrescine is necessary for tvcp39 mRNA stability. TvCP39 was localized in the cytoplasm but, in DAB treated parasites transferred into exogenous putrescine culture media, TvCP39 was re-localized to the nucleus and nuclear periphery of trichomonads. Interestingly, the amount and proteolytic activity of TvCP39 was recovered as well as the tvcp39 mRNA levels were restored when putrescine exogenous was added to the DAB-treated parasites. In conclusion, our data show that putrescine regulate the TvCP39 expression, protein amount, proteolytic activity, and cellular localization.

  18. Comparison of clinical and environmental isolates of Acanthamoeba based on morphology, protease and gelatinase activity, and the cysteine proteinase gene

    Directory of Open Access Journals (Sweden)

    Gil M. Penuliar

    2010-06-01

    Full Text Available Acanthamoeba spp. are opportunistic pathogens that cause amebic keratitis and granulomatous amebic encephalitis in man. Recent attempts to correlate pathogenicity with species have been proven difficult due to inconsistencies in morphology-based classification. The objectives of this study were: (1 to compare clinical and environmental isolates based on morphology, protease and gelatinase activity, and the cysteine proteinase (CP gene, and (2 to determine whether these features can be used to differentiate the isolates. Results show some degree of variation in trophozoite and cyst morphology. Zymography, demonstrated gross differences in banding patterns, and the protease activity of clinical isolates was greater than the environmental isolates (p-value < 0.01. Amplification of the CP gene yielded two bands in the environmental isolates, approximately 755 bp and 440 bp in length. In contrast, only one band, either the 755 bp or 440 bp band was amplified in the clinical isolates. The results confirmed the limitations of morphology in differentiating Acanthamoeba species, and suggest that zymography, protease activity, and detection of the CP gene are useful reference tests to distinguish pathogenic from non-pathogenic isolates.

  19. A proteinase inhibitor from Caesalpinia echinata (pau-brasil) seeds for plasma kallikrein, plasmin and factor XIIa.

    Science.gov (United States)

    Cruz-Silva, Ilana; Gozzo, Andrezza J; Nunes, Viviane A; Carmona, Adriana K; Faljoni-Alario, Adelaide; Oliva, Maria Luiza V; Sampaio, Misako U; Sampaio, Claudio A M; Araujo, Mariana S

    2004-11-01

    Caesalpinia echinata is a tree belonging to the Leguminosae family. The red color of the trunk, looking like burning wood ('brasa' in Portuguese), is the origin of the name Brazil. Seeds of leguminous plants contain high amounts of serine proteinase inhibitors that can affect different biological processes. Here we show that a protein isolated from seeds of C. echinata is able to inhibit enzymes that participate in blood coagulation and fibrinolysis. This inhibitor (CeKI) was purified to homogeneity by ion exchange and reversed-phase chromatography. SDS-PAGE indicated a single polypeptide chain with a molecular mass of 20 kDa. CeKI inhibits human plasma kallikrein ( K i =3.1 nM), plasmin ( K i =0.18 nM), factor XIIa ( K i =0.18 nM), trypsin ( K i =21.5 nM) and factor Xa ( K i =0.49 mM). CeKI inhibited kinin release from highmolecular- mass kininogen by kallikrein in vitro . The N-terminal sequence, determined by automatic Edman degradation, identified the inhibitor as a member of the Kunitz family. The secondary structure, determined by circular dichroism, is mainly a random coil followed by beta-sheet structure. The action of CeKI on enzymes of the blood-clotting intrinsic pathway was confirmed by prolongation of the activated partial thromboplastin time.

  20. Classification of alpha 1-adrenoceptor subtypes

    NARCIS (Netherlands)

    Michel, M. C.; Kenny, B.; Schwinn, D. A.

    1995-01-01

    Two alpha 1-adrenoceptor subtypes (alpha 1A and alpha 1B) have been detected in various tissues by pharmacological techniques, and three distinct cDNAs encoding alpha 1-adrenoceptor subtypes have been cloned. The profile of an increasing number of subtype-selective compounds at cloned and endogenous

  1. Triplication of alpha-globin genes is responsible for unusual alpha 113Leu/alpha 113His-globin chain ratios in sheep.

    Science.gov (United States)

    Vestri, R; Masina, P; Rando, A; Testa, A; Di Gregorio, P

    1987-10-01

    By investigations at the DNA and protein level, it has been shown that in sheep a previously detected, presumed quantitative allele of the II alpha 113His gene, displaying a reduced efficiency (called the II alpha 113His decreases gene), is carried by a chromosome bearing three alpha-globin loci. In particular, five sheep having an alpha 113Leu/alpha 113His-chain ratio of about 13:1 (13:1 phenotype) possessed the -I alpha 113Leu-II alpha 113Leu-/-I alpha 113Leu-II alpha 113Leu-III alpha 113His decreases genotype. One sheep showing a alpha 113Leu/alpha 113His-chain ratio of about 3:1 (3:1 phenotype) had the -I alpha 113Leu-II alpha 113His-/-I alpha 113Leu-II alpha 113Leu-III alpha 113His decreases genotype, while one sheep having a chain ratio of about 6:1 (6:1 phenotype) carried the -I alpha 113Leu-II alpha 113Leu-II alpha 113His decreases-/-I alpha 113Leu-II alpha 113Leu-III alpha 113His decreases genotype. Nineteen sheep, displaying the common phenotypes, all possessed the alpha alpha/alpha alpha gene arrangement. Furthermore, the possible location of the gene with reduced efficiency and the expression of the three genes in the triple alpha-globin loci chromosome are discussed.

  2. Contribution to the study of the alpha-alpha interaction

    International Nuclear Information System (INIS)

    Darriulat, Pierre

    1965-01-01

    The new variable energy cyclotron at Berkeley that can accelerate an alpha beam up to an energy of 130 MeV and the mass production of lithium diffused junctions have enabled us to perform 2 series of measurement, in the first one we use alpha beams with an energy ranging between 50 and 120 MeV to study alpha-alpha forces in the second one we use the flexibility of the variable energy cyclotron the resonances around 40 MeV, region that can not yet be reached by tandem accelerators. This work is divided into 6 chapters. The first chapter is dedicated to the formalism of partial wave analysis and the theory of the compound nucleus. In the second chapter the author presents the 88 cyclotron at Berkeley and the diffusion chamber, the alpha detectors are lithium diffused junctions made of silicon. The third chapter deals with the experimental methods used and the issue of the reduction of the volume of data. In the fourth chapter the results obtained in the upper part of the energy range are described in terms of complex shifts that allow the description of the α-α interaction at high energy. The very low impact parameter has enabled us to find 2 new components (l=6 and l=8) of the rotational spectrum and to define a more accurate phenomenological potential. The fifth chapter is dedicated to the narrow resonances we have found between 12 and 27 MeV. We present in the last chapter a calculation of the binding energy of C 12 in which we have considered the 12 C nucleus as formed by 3 alpha particles interacting with each other through the phenomenological potential defined above

  3. Workshop on Precision Measurements of $\\alpha_s$

    Energy Technology Data Exchange (ETDEWEB)

    Bethke, Siegfried; /Munich, Max Planck Inst.; Hoang, Andre H.; /Vienna U.; Kluth, Stefan; /Munich, Max Planck Inst.; Schieck, Jochen; /Munich U.; Stewart, Iain W.; Aoki, S.; Beneke, M.; Bethke, S.; Blumlein, J.; Brambilla, N.; Brodsky, S.; /MIT, LNS

    2011-10-01

    These are the proceedings of the Workshop on Precision Measurements of {alpha}{sub s} held at the Max-Planck-Institute for Physics, Munich, February 9-11, 2011. The workshop explored in depth the determination of {alpha}{sub s}(m{sub Z}) in the {ovr MS} scheme from the key categories where high precision measurements are currently being made, including DIS and global PDF fits, {tau}-decays, electro-weak precision observables and Z-decays, event-shapes, and lattice QCD. These proceedings contain a short summary contribution from the speakers, as well as the lists of authors, conveners, participants, and talks.

  4. Functional proteomic of Matrix Metallo-proteinases (MMP) dedicated to the detection of active forms of MMP in complex proteome

    International Nuclear Information System (INIS)

    David, A.

    2007-07-01

    The Matrix Metallo-proteinases (M.M.P.) represent a family of Zinc dependent extracellular proteinases able to cleave collectively all the proteins constituting the extracellular matrix. Currently, 23 human M.M.P. have been identified and are characterized by their sequence in amino-acids and their highly conserved 3 D structure. These enzymes are expressed constitutively during the tissue remodeling process. Their over-expression in various diseases tightly related to inflammatory processes (arthritis, emphysema, cancer) described M.M.P. as choice therapeutic targets. However, as the tissue remodeling implicates modification of cellular contacts, M.M.P. appear currently as proteins involved in signalling pathways. Recent works demonstrating that M.M.P. are able to cleave substrates, which are different than proteins constituting the extracellular matrix, reinforce this vision. In order to identify the individual role and the protein expression level of M.M.P. in pathological context, we developed a new technique of functional proteomics dedicated to the detection of active forms of M.M.P. in tumour samples. This technique relied on the development of a new photoaffinity probe, based on the structure of a potent phosphinic inhibitor of M.M.P., allowing targeting and isolating active forms of M.M.P. by photoaffinity labelling. Furthermore, as the new developed probe incorporated a radioactive element, photoaffinity labelling permitted to radiolabel the targeted proteins. This probe demonstrated in vitro its remarkable ability to covalently modify the h M.M.P.-12, with a singular cross-linking yield, determined at 42 %, displaying an extremely sensitive detection (2.5 fmoles of h M.M.P.-12). When added to complex proteome, the photoaffinity probe presents the same sensibility of detection for the h M.M.P.-12 (5 fmoles); importantly, in this case, h M.M.P.-12 represents only 0.001 % of the totality of the proteins present in the sample. Moreover, this technique allows

  5. Redundant signaling mechanisms contribute to the vasodilatory response of the afferent arteriole to proteinase-activated receptor-2.

    Science.gov (United States)

    Wang, Xuemei; Hollenberg, Morley D; Loutzenhiser, Rodger

    2005-01-01

    We previously demonstrated that stimulation of proteinase-activated receptor-2 (PAR-2) by SLIGRL-NH(2) elicits afferent arteriolar vasodilation, in part, by elaborating nitric oxide (NO), suggesting an endothelium-dependent mechanism (Trottier G, Hollenberg M, Wang X, Gui Y, Loutzenhiser K, and Loutzenhiser R. Am J Physiol Renal Physiol 282: F891-F897, 2002). In the present study, we characterized the NO-independent component of this response, using the in vitro perfused hydronephrotic rat kidney. SLIGRL-NH(2) (10 mumol/l) dilated afferent arterioles preconstricted with ANG II, and the initial transient component of this response was resistant to NO synthase (NOS) and cyclooxygenase inhibition. This NO-independent response was not prevented by treatment with 10 nmol/l charybdotoxin and 1 mumol/l apamin, a manipulation that prevents the endothelium-derived hyperpolarizing factor (EDHF)-like response of the afferent arteriole to acetylcholine, nor was it blocked by the addition of 1 mmol/l tetraethylammonium (TEA) or 50 mumol/l 17-octadecynoic acid, treatments that block the EDHF-like response to bradykinin. To determine whether the PAR-2 response additionally involves the electrogenic Na(+)-K(+)-ATPase, responses were evaluated in the presence of 3 mmol/l ouabain. In this setting, SLIGRL-NH(2) induced a biphasic dilation in control and a transient response after NOS inhibition. The latter was not prevented by charybdotoxin plus apamin or by TEA alone but was abolished by combined treatment with charybdotoxin, apamin, and TEA. This treatment did not prevent the NO-dependent dilation evoked in the absence of NOS inhibition. Our findings indicate a remarkable redundancy in the signaling cascade mediating PAR-2 -induced afferent arteriolar vasodilation, suggesting an importance in settings such as inflamation or ischemia, in which vascular mechanisms might be impaired and the PAR system is thought to be activated.

  6. JMJD3 is involved in neutrophil membrane proteinase 3 overexpression during the hyperinflammatory response in early sepsis.

    Science.gov (United States)

    Chen, Yang; Liu, Zhaojun; Pan, Tingting; Chen, Erzhen; Mao, Enqiang; Chen, Ying; Tan, Ruoming; Wang, Xiaoli; Tian, Rui; Liu, Jialin; Qu, Hongping

    2018-04-02

    Excessive production of pro-inflammatory cytokines in early sepsis causes high early mortality rates. Membrane proteinase 3 (mPR3) expression on neutrophils plays a critical role in pro-inflammatory cytokine production. However, the mechanism underlying mPR3 overexpression in early sepsis is unknown. Here, we explored mPR3 expression in early sepsis and its regulatory mechanism. Thirty-two patients with sepsis and 20 healthy controls were prospectively enrolled. On day 1 after the onset of sepsis, mPR3 and jumonji domain-containing protein D3 (JMJD3) expression levels were measured in peripheral blood neutrophils. Lipopolysaccharide (LPS) was employed to induce JMJD3 expression in vitro, and GSK-J4 was used to inhibit JMJD3. Neutrophils were divided into four groups, control, LPS, LPS + GSK-J4, and GSK-J4, and cultured with THP-1 cells respectively. Plasma and culture supernatant cytokine levels were measured by enzyme-linked immunosorbent assays. Neutrophil mPR3 levels were significantly higher in patients with early sepsis than in healthy controls. Plasma cytokine (IL-1β and TNF-α) levels were increased in patients with sepsis exhibiting high mPR3 expression. Additionally, JMJD3 expression levels in neutrophils were increased in early sepsis. In vitro, both mPR3 on neutrophils and IL-1β in culture supernatants increased in response to LPS stimulation. Neutrophil mPR3 expression and IL-1β levels were significantly reduced by GSK-J4 in cells treated with LPS. IL-1β level was significantly higher in LPS-stimulated co-culture supernatants than in the corresponding individual cultured cells. Thus, our results suggest that JMJD3 contributes to the high expression of neutrophil mPR3, which promotes the production of proinflammatory IL-1β in early sepsis. Copyright © 2018. Published by Elsevier B.V.

  7. Neutrophil elastase induces inflammation and pain in mouse knee joints via activation of proteinase-activated receptor-2.

    Science.gov (United States)

    Muley, Milind M; Reid, Allison R; Botz, Bálint; Bölcskei, Kata; Helyes, Zsuzsanna; McDougall, Jason J

    2016-02-01

    Neutrophil elastase plays a crucial role in arthritis. Here, its potential in triggering joint inflammation and pain was assessed, and whether these effects were mediated by proteinase-activated receptor-2 (PAR2). Neutrophil elastase (5 μg) was injected into the knee joints of mice and changes in blood perfusion, leukocyte kinetics and paw withdrawal threshold were assessed. Similar experiments were performed in animals pretreated with the neutrophil elastase inhibitor sivelestat, the PAR2 antagonist GB83, the p44/42 MAPK inhibitor U0126 and in PAR2 receptor knockout (KO) mice. Neutrophil elastase activity was also evaluated in arthritic joints by fluorescent imaging and sivelestat was assessed for anti-inflammatory and analgesic properties. Intra-articular injection of neutrophil elastase caused an increase in blood perfusion, leukocyte kinetics and a decrease in paw withdrawal threshold. Sivelestat treatment suppressed this effect. The PAR2 antagonist GB83 reversed neutrophil elastase-induced synovitis and pain and these responses were also attenuated in PAR2 KO mice. The MAPK inhibitor U0126 also blocked neutrophil elastase-induced inflammation and pain. Active neutrophil elastase was increased in acutely inflamed knees as shown by an activatable fluorescent probe. Sivelestat appeared to reduce neutrophil elastase activity, but had only a moderate anti-inflammatory effect in this model. Neutrophil elastase induced acute inflammation and pain in knee joints of mice. These changes are PAR2-dependent and appear to involve activation of a p44/42 MAPK pathway. Blocking neutrophil elastase, PAR2 and p44/42 MAPK activity can reduce inflammation and pain, suggesting their utility as therapeutic targets. © 2015 The British Pharmacological Society.

  8. A double WAP domain-containing protein Es-DWD1 from Eriocheir sinensis exhibits antimicrobial and proteinase inhibitory activities.

    Directory of Open Access Journals (Sweden)

    Shuang Li

    Full Text Available Whey acidic proteins (WAP belong to a large gene family of antibacterial peptides, which are critical in the host immune response against microbial invasion. The common feature of these proteins is a single WAP domain maintained by at least one four-disulfide core (4-DSC structure rich in cysteine residues. In this study, a double WAP domain (DWD-containing protein, Es-DWD1, was first cloned from the Chinese mitten crab (Eriocheirsinensis. The full-length Es-DWD1cDNA was 1193 bp, including a 411 bp open reading frame (ORF encoding 136 amino acids with a signal peptide of 22 amino acids in the N-terminus. A comparison with other reported invertebrate and vertebrate sequences revealed the presence of WAP domains characteristic of WAP superfamilies. As determined by quantitative real-time RT-PCR, Es-DWD1 transcripts were ubiquitously expressed in all tissues, but it was up-regulated in hemocytes post-challenge with pathogen-associated molecular patterns (PAMPs. The mature recombinant Es-DWD1 (rEs-DWD1 protein exhibited different binding activities to bacteria and fungus. Moreover, rEs-DWD1 could exert agglutination activities against Bacillus subtilis and Pichiapastoris and demonstrated inhibitory activities against the growth of Staphylococcus aureus, Aeromonas hydrophila and P. pastoris. Furthermore, rEs-DWD1 showed a specific protease inhibitory activity in B. subtilis. Coating of rEs-DWD1 onto agarose beads enhanced encapsulation of the beads by crab hemocytes. Collectively, the results suggest that Es-DWD1 is a double WAP domain containing protein with antimicrobial and proteinase inhibitory activities, which play significant roles in the immunity of crustaceans.

  9. Conditioning of alpha bearing wastes

    International Nuclear Information System (INIS)

    1991-01-01

    Alpha bearing wastes are generated during the reprocessing of spent fuel, mixed oxide fuel fabrication, decommissioning and other activities. The safe and effective management of these wastes is of particular importance owing to the radiotoxicity and long lived characteristics of certain transuranic (TRU) elements. The management of alpha bearing wastes involves a number of stages which include collection, characterization, segregation, treatment, conditioning, transport, storage and disposal. This report describes the currently available matrices and technologies for the conditioning of alpha wastes and relates them to their compatibility with the other stages of the waste management process. The selection of a specific immobilization process is dependent on the waste treatment state and the subsequent handling, transport, storage and disposal requirements. The overall objectives of immobilization are similar for all waste producers and processors, which are to produce: (a) Waste forms with sufficient mechanical, physical and chemical stability to satisfy all stages of handling, transport and storage (referred to as the short term requirements), and (b) Waste forms which will satisfy disposal requirements and inhibit the release of radionuclides to the biosphere (referred to as the long term requirements). Cement and bitumen processes have already been successfully applied to alpha waste conditioning on the industrial scale in many of the IAEA Member States. Cement systems based on BFS and pozzolanic cements have emerged as the principal encapsulation matrices for the full range of alpha bearing wastes. Alternative technologies, such as polymers and ceramics, are being developed for specific waste streams but are unlikely to meet widespread application owing to cost and process complexity. The merits of alpha waste conditioning are improved performance in transport, storage and disposal combined with enhanced public perception of waste management operations. These

  10. "Purification and evaluation of somatic, excretory-secretory and Cysteine proteinase antigens of Fasciola Hepatica using IgG-ELISA in diagnosing Fascioliasis "

    Directory of Open Access Journals (Sweden)

    "Rokni MB

    2001-08-01

    Full Text Available Fasciolosis, or liver fluke disease, caused by parasites of the genus Fasciola is emerging as an important disease in man and animals, in the world and Iran, particularly in nortern parts. The economical losses in domestic animals are considerable. In the recent decade there were two major outbreaks of human fasciolosis in the Caspian region, northern part of Iran with 7000-10000 infected cases. Sicne it is impossible to diagnose fasciolosis in acute phase using coprological methods and even in chronic phases its sensitivity is low, evaluating and establishing a reliable and cost-effetive test is indispensable and notewortly.In the present survey, we produced and examined the sensitivity and specificity of liver fluke homogenate (LFH , excretory-secetory (ES and cysteine proteinase (CP antigens of F. hepatica using IgG-ELISA test. A 25-27 kilo Dalton coomassie blue-stained band was observed and using of specific inhibitors indicated that this antigen belongs to the class of cysteine proteinase. The sensitivity of LFH, ES and CP antigen in IgG-ELISa was 100% for each, while their specificity was 97.8%, 98.8% and 98.8% respectively. There was a significant difference in mean OD values between cases of proven fasciolosis and other true negative cases, including healthy control individuals and patients with other parasitic diseases.This present report is the first to demonstrate the purification and evaluation of F. hepatica cysteine proteinase antigen by IgG-ELISA test for the diagnosis of fasciolosis in Iran. In conclusion, the IgG-ELISa using ES and CP show high sensitivity and specificity and would be a valuable tool to diagnose human fasciolosis in Iran, particularly in endemic areas.

  11. Negative Effects of a Nonhost Proteinase Inhibitor of ~19.8 kDa from Madhuca indica Seeds on Developmental Physiology of Helicoverpa armigera (Hübner

    Directory of Open Access Journals (Sweden)

    Farrukh Jamal

    2014-01-01

    Full Text Available An affinity purified trypsin inhibitor from the seed flour extracts of Madhuca indica (MiTI on denaturing polyacrylamide gel electrophoresis showed that MiTI consisted of a single polypeptide chain with molecular mass of ~19.8 kDa. MiTI inhibited the total proteolytic and trypsin-like activities of the midgut proteinases of Helicoverpa armigera larvae by 87.51% and 76.12%, respectively, at concentration of 5 µg/mL with an IC50 of 1.75 µg/mL against trypsin like midgut proteinases. The enzyme kinetic studies demonstrated that MiTI is a competitive inhibitor with a Ki value of 4.1×10−10 M for Helicoverpa trypsin like midgut proteinases. In vivo experiments with different concentrations of MiTI in artificial diet (0.5, 1.0, and 1.5% w/w showed an effective downfall in the larval body weight and an increase in larval mortality. The concentration of MiTI in the artificial diet to cause 50% mortality (LD50 of larvae was 1.5% w/w and that to cause reduction in mass of larvae by 50% (ED50 was 1.0% w/w. Nutritional indices observations suggest the toxic and adverse effects of MiTI on the growth and development of H. armigera larvae. The results suggest a strong bioinsecticidal potential of affinity purified MiTI which can be exploited in insect pest management of crop plants.

  12. Improved Alpha Testing Using Hashed Sampling.

    Science.gov (United States)

    Wyman, Chris; McGuire, Morgan

    2017-08-14

    We further describe and analyze the idea of hashed alpha testing from Wyman and McGuire [1], which builds on stochastic alpha testing and simplifies stochastic transparency. Typically, alpha testing provides a simple mechanism to mask out complex silhouettes using simple proxy geometry with applied alpha textures. While widely used, alpha testing has a long-standing problem: geometry can disappear entirely as alpha mapped polygons recede with distance. As foveated rendering for virtual reality spreads, this problem worsens as peripheral minification and prefiltering introduce this problem on nearby objects.

  13. Disruption of each of the secreted aspartyl proteinase genes SAP1, SAP2, and SAP3 of Candida albicans attenuates virulence.

    OpenAIRE

    Hube, B; Sanglard, D; Odds, F C; Hess, D; Monod, M; Schäfer, W; Brown, A J; Gow, N A

    1997-01-01

    Secreted aspartyl proteinases (Saps), encoded by a gene family with at least nine members (SAP1 to SAP9), are one of the most discussed virulence factors produced by the human pathogen Candida albicans. In order to study the role of each Sap isoenzyme in pathogenicity, we have constructed strains which harbor mutations at selected SAP genes. SAP1, SAP2, and SAP3, which are regulated differentially in vitro, were mutated by targeted gene disruption. The growth rates of all homozygous null muta...

  14. Immunological cross-reactivity of the major allergen from perennial ryegrass (Lolium perenne), Lol p I, and the cysteine proteinase, bromelain.

    Science.gov (United States)

    Pike, R N; Bagarozzi, D; Travis, J

    1997-04-01

    Antibodies prepared in rabbits against the major allergen from ryegrass (Lolium perenne), Lol p I, cross-reacted with the cysteine proteinase bromelain from pineapple and vice versa. Deglycosylation of the proteins showed that the cross-reaction was based on recognition of the carbohydrate moiety of the allergen, but for bromelain the cross-reaction was most likely due to a combination of factors. The results indicate that the carbohydrate residues from these allergens play an important role in cross-reactions found between them and possibly those from other species.

  15. Perspectives of digestive pest control with proteinase inhibitors that mainly affect the trypsin-like activity of Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae

    Directory of Open Access Journals (Sweden)

    M.E. Pereira

    2005-11-01

    Full Text Available The present study describes the main characteristics of the proteolytic activities of the velvetbean caterpillar, Anticarsia gemmatalis Hübner, and their sensitivity to proteinase inhibitors and activators. Midguts of last instar larvae reared on an artificial diet were homogenized in 0.15 M NaCl and centrifuged at 14,000 g for 10 min at 4ºC and the supernatants were used in enzymatic assays at 30ºC, pH 10.0. Basal total proteolytic activity (azocasein hydrolysis was 1.14 ± 0.15 absorbance variation min-1 mg protein-1, at 420 nm; basal trypsin-like activity (N-benzoyl-L-arginine-p-nitroanilide, BApNA, hydrolysis was 0.217 ± 0.02 mmol p-nitroaniline min-1 mg protein-1. The maximum proteolytic activities were observed at pH 10.5 using azocasein and at pH 10.0 using BApNA, this pH being identical to the midgut pH of 10.0. The maximum trypsin-like activity occurred at 50ºC, a temperature that reduces enzyme stability to 80 and 60% of the original, when pre-incubated for 5 and 30 min, respectively. Phenylmethylsulfonyl fluoride inhibited the proteolytic activities with an IC50 of 0.39 mM for azocasein hydrolysis and of 1.35 mM for BApNA hydrolysis. Benzamidine inhibited the hydrolysis with an IC50 of 0.69 and 0.076 mM for azocasein and BApNA, respectively. The absence of cysteine-proteinases is indicated by the fact that 2-mercaptoethanol and L-cysteine did not increase the rate of azocasein hydrolysis. These results demonstrate the presence of serine-proteinases and the predominance of trypsin-like activity in the midgut of Lepidoptera insects, now also detected in A. gemmatalis, and suggest this enzyme as a major target for pest control based on disruption of protein metabolism using proteinase inhibitors.

  16. Proteins of the kidney microvillar membrane. Aspartate aminopeptidase: purification by immunoadsorbent chromatography and properties of the detergent- and proteinase-solubilized forms

    DEFF Research Database (Denmark)

    Danielsen, Erik Michael; Norén, O; Sjöström, H

    1980-01-01

    Aminopeptidase A (aspartate aminopeptidase, EC 3.4.11.7) was purified 2000-fold from pig kidney cortex. The essential step in the purification was chromatography on an immunoadsorbent column prepared from a rabbit antiserum raised against pig intestinal aminopeptidase A. Glutamyl and aspartyl...... revealed 1 g-atom of Ca/143000 g of protein. Two forms of the enzyme were purified: an amphipathic form solubilized from the membrane by Triton X-100 (detergent form) and a hydrophilic form released by incubation with trypsin (proteinase form). The detergent form exhibited charge-shift in crossed...... protein....

  17. Nonenzymatic palmitoylation at Cys 3 causes extra-activation of the alpha-subunit of the stimulatory GTP-binding protein Gs.

    Science.gov (United States)

    Mollner, S; Ferreira, P; Beck, K; Pfeuffer, T

    1998-10-01

    Treatment of crude stimulatory GTP-binding protein of adenylyl cyclase (Gs) from turkey erythrocyte membranes with hydroxylamine results in twofold enhancement of adenylyl cyclase activity following reconstitution with adenylyl cyclase type V expressed in Spodoptera frugiperda cells (Sf9) cells. Enhancement by hydroxylamine of immunoaffinity purified Gs was still 1.5-fold, while that of Gs purified according to the multiple-step procedure by Northup, J. K., Sternweis, P. C., Smigel, M. D., Schleifer, L. S., Ross, E. M. & Gilman, A. G. (Proc. Natl Acad. Sci. USA 78, 6516-6520, 1980) was close to unity. The alpha-subunit of the stimulatory GTP-binding protein expressed in Escherichia coli (r(alpha)s), likewise, failed to show an effect by hydroxylamine. Surprisingly, guanosine 5'-O-(3-thiotriphosphate) (GTP[S])-liganded r(alpha)s, treated with palmitoyl-CoA for 14 h at 4 degrees C resulted in sixfold enhancement of reconstitutive activity. In contrast, that of the GDP-liganded r(alpha)s(beta)gamma heterotrimer was not improved by palmitoylation and consecutive activation with GTP[S], although incorporation of [3H]palmitate into momomer and heterotrimer was identical. While adenylyl cyclase type-V activity reconstituted by r(alpha)s x GTP[S] was not influenced by betagamma-subunits, that activated by palmitoylated r(alpha)s x GTP[S] was considerably inhibited, suggesting a higher affinity of palmitoylated r(alpha)s for betagamma-subunits. On treatment of either form with the proteinase Lys-C, less than 25% of the label was found in a stable M(r) 38000 fragment with intact C terminus, but lacking the N-terminal portion. Absence of the latter did not affect activation by r(alpha)s, but caused a >90% loss of extra-activation by palmitoylated r(alpha)s. The results also indicate that nonenzymatic, much in the same way as physiological enzymatic, palmitoylation of alpha(s) occurs predominantly on Cys 3.

  18. ALPHA,·ANTITRYPSIN DEFICIENCY*

    African Journals Online (AJOL)

    1971-02-06

    Feb 6, 1971 ... Lieberman," in fact, found that 15·2% of 66 patients hospitalized with pulmonary emphysema had heterozygous alpha,-antitrypsin deficiency. The over-all incidence of the deficiency was 25'8% in this group. Of patients under the age of 50 years, 47·8% had deficient levels. If such observations are confirmed ...

  19. Alpha sources deposit by sublimation

    International Nuclear Information System (INIS)

    Amoudry, F.; Eloy, J.F.

    1983-09-01

    We studied and realized a device able to perform some very thin substracts used for alpha spectrometry measurements. Sources are prepared by sublimation of the sample in a vacuum container. The energy required for this sublimation is furnished by a laser beam [fr

  20. Bothrops asper metalloproteinase BaP1 is inhibited by alpha(2)-macroglobulin and mouse serum and does not induce systemic hemorrhage or coagulopathy.

    Science.gov (United States)

    Escalante, Teresa; Rucavado, Alexandra; Kamiguti, Aura S; Theakston, R David G; Gutiérrez, José María

    2004-02-01

    The ability of the P-I metalloproteinase BaP1, isolated from the venom of the snake Bothrops asper, to induce systemic bleeding, thrombocytopenia and defibrinogenation was assessed in an experimental mouse model. Intravenous administration of BaP1 caused neither systemic bleeding nor any evidence of pathology in lungs, kidneys, liver, heart and brain. Moreover, there were no alterations in the whole blood clotting time or in platelet numbers. In addition, BaP1 did not inhibit collagen-induced platelet aggregation in vitro. Proteolytic and hemorrhagic activities of BaP1 were readily inhibited by the plasma proteinase inhibitor, alpha(2)-macroglobulin, and normal mouse serum also inhibited hemorrhage. Such inhibition may explain why BaP1 induces multiple local tissue-damaging effects, but is largely devoid of systemic toxicity.

  1. The role of proteases, endoplasmic reticulum stress and SERPINA1 heterozygosity in lung disease and alpha-1 anti-trypsin deficiency.

    LENUS (Irish Health Repository)

    Greene, Catherine M

    2012-02-01

    The serine proteinase inhibitor alpha-1 anti-trypsin (AAT) provides an antiprotease protective screen throughout the body. Mutations in the AAT gene (SERPINA1) that lead to deficiency in AAT are associated with chronic obstructive pulmonary diseases. The Z mutation encodes a misfolded variant of AAT that is not secreted effectively and accumulates intracellularly in the endoplasmic reticulum of hepatocytes and other AAT-producing cells. Until recently, it was thought that loss of antiprotease function was the major cause of ZAAT-related lung disease. However, the contribution of gain-of-function effects is now being recognized. Here we describe how both loss- and gain-of-function effects can contribute to ZAAT-related lung disease. In addition, we explore how SERPINA1 heterozygosity could contribute to smoking-induced chronic obstructive pulmonary diseases and consider the consequences.

  2. The effect of a metalloproteinase inhibitor (GI5402) on tumor necrosis factor-alpha (TNF-alpha) and TNF-alpha receptors during human endotoxemia

    NARCIS (Netherlands)

    Dekkers, P. E.; Lauw, F. N.; ten Hove, T.; te Velde, A. A.; Lumley, P.; Becherer, D.; van Deventer, S. J.; van der Poll, T.

    1999-01-01

    Tumor necrosis factor-alpha (TNF-alpha) is released from the cell surface by cleavage of pro-TNF-alpha by metalloproteinases (MPs). In cell cultures, inhibition of MPs has been found not only to reduce the release of TNF-alpha, but also to enhance the surface expression of TNF-alpha and TNF-alpha

  3. Mathematical model for the homeostasis of alpha-macroglobulins in the rat.

    Science.gov (United States)

    Aguirre, M C; Armendariz, M; Lupo, M; Rigalli, A

    2011-11-01

    Alpha-macroglobulins (AM) are proteins that inactivate proteinases. Sodium monofluorophosphate (MFP) binds to AM and transiently changes AM plasma levels. As a consequence MFP is useful to modify AM homeostasis. A mathematical model to study the homeostasis of AM is proposed in this paper. The model describes changes in plasma concentration of AM, MFP concentration in the gastrointestinal tract, MFP plasma concentration, plasma concentration of AMMFP and includes rate constants of the processes involved in AM homeostasis. Estimation of the rate constants values was achieved using experimental and mathematical resources. The homeostasis of AM after an oral dose of 80 μmol of MFP was analyzed with a simulation tool. Experimental conditions that modify the homeostasis of AM had been simulated and validated using specific drugs that change some parameter of the system. The mathematical model describes accurately the behavior of the biological model. The results allow concluding that the simplifications made did not underestimate the main processes involved in the homeostasis and, also that the assumptions made were correct. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Diabetic retinopathy: could the alpha-1 antitrypsin be a therapeutic option?

    Directory of Open Access Journals (Sweden)

    Gustavo Ortiz

    2014-01-01

    Full Text Available Diabetic retinopathy is one of the most important causes of blindness. The underlying mechanisms of this disease include inflammatory changes and remodeling processes of the extracellular-matrix (ECM leading to pericyte and vascular endothelial cell damage that affects the retinal circulation. In turn, this causes hypoxia leading to release of vascular endothelial growth factor (VEGF to induce the angiogenesis process. Alpha-1 antitrypsin (AAT is the most important circulating inhibitor of serine proteases (SERPIN. Its targets include elastase, plasmin, thrombin, trypsin, chymotrypsin, proteinase 3 (PR-3 and plasminogen activator (PAI. AAT modulates the effect of protease-activated receptors (PARs during inflammatory responses. Plasma levels of AAT can increase 4-fold during acute inflammation then is so-called acute phase protein (APPs. Individuals with low serum levels of AAT could develop disease in lung, liver and pancreas. AAT is involved in extracellular matrix remodeling and inflammation, particularly migration and chemotaxis of neutrophils. It can also suppress nitric oxide (NO by nitric oxide sintase (NOS inhibition. AAT binds their targets in an irreversible way resulting in product degradation. The aim of this review is to focus on the points of contact between multiple factors involved in diabetic retinopathy and AAT resembling pleiotropic effects that might be beneficial.

  5. Calibration of sources for alpha spectroscopy systems

    International Nuclear Information System (INIS)

    Freitas, I.S.M.; Goncalez, O.L.

    1992-01-01

    This paper describes the calibration methodology for measuring the total alpha activity of plane and thin sources with the Alpha Spectrometer for Silicon Detector in the Nuclear Measures and Dosimetry laboratory at IEAv/CTA. (author)

  6. Cysteine proteinases from papaya (Carica papaya) in the treatment of experimental Trichuris suis infection in pigs: two randomized controlled trials.

    Science.gov (United States)

    Levecke, Bruno; Buttle, David J; Behnke, Jerzy M; Duce, Ian R; Vercruysse, Jozef

    2014-05-30

    Cysteine proteinases (CPs) from papaya (Carica papaya) possess anthelmintic properties against human soil-transmitted helminths (STH, Ascaris lumbricoides, Trichuris trichiura and hookworm), but there is a lack of supportive and up-to-date efficacy data. We therefore conducted two randomized controlled trials in pigs to assess the efficacy of papaya CPs against experimental infections with T. suis. First, we assessed efficacy by means of egg (ERR) and adult worm reduction rate (WRR) of a single-oral dose of 450 μmol active CPs (CP450) against low (inoculum of 300 eggs) and high (inoculum of 3,000 eggs) intensity T. suis infections and compared the efficacy with those obtained after a single-oral dose of 400 mg albendazole (ALB). In the second trial, we determined and compared the efficacy of a series of CP doses (45 [CP45], 115 [CP115], 225 [CP225], and 450 [CP450] μmol) against high intensity infections. CP450 was highly efficacious against both levels of infection intensity, resulting in ERR and WRR of more than 97%. For both levels of infection intensity, CP450 was significantly more efficacious compared to ALB by means of WRR (low infection intensity: 99.0% vs. 39.0%; high infection intensity; 97.4% vs. 23.2%). When the efficacy was assessed by ERR, a significant difference was only observed for high intensity infections, CP450 being more efficacious than ALB (98.9% vs. 59.0%). For low infection intensities, there was no significant difference in ERR between CP450 (98.3%) and ALB (64.4%). The efficacy of CPs increased as a function of increasing dose. When determined by ERR, the efficacy ranged from 2.1% for CP45 to 99.2% for CP450. For WRR the results varied from -14.0% to 99.0%, respectively. Pairwise comparison revealed a significant difference in ERR and WRR only between CP45 and CP450, the latter being more efficacious. A single dose of 450 μmol CPs provided greater efficacy against T. suis infections in pigs than a single-oral dose of 400 mg ALB

  7. Co-expression of two genes, a chitinase (chit42) and proteinase (prb1), implicated in mycoparasitism by Trichoderma hamatum.

    Science.gov (United States)

    Steyaert, Johanna M; Stewart, Alison; Jaspers, Marlene V; Carpenter, Margaret; Ridgway, Hayley J

    2004-01-01

    Mycoparasitism of fungal plant pathogens by Trichoderma species is a complex process that involves the production and coordinated secretion of cell-wall degrading enzymes. Genes implicated in mycoparasitism by Trichoderma atroviride contain motifs in the promoter region, designated MYRE1-MYRE4, that are proposed to act as binding sites for a global inducer of the mycoparasitic response. The aim of our study was to establish whether these motifs also were present in Trichoderma hamatum and whether the presence of these motifs could predict co-expression when T. hamatum was confronted by a pathogen. Using a combination of targeted, degenerate and inverse PCR, homologues of the mycoparasitism-related genes ech42 (chit42), prb1 and lam1.3 (xbg1.3-110), which encode an endochitinase, proteinase, and β-1,3-glucanase, respectively, were cloned and sequenced from T. hamatum. Alignment of the promoter regions of the three genes revealed identical regions in the chit42 and prb1 promoters, which were 6-9 base pairs in length and conserved in position. Specifically, the regulator y motifs MYRE1-MYRE4 were fully conserved, together with a fifth motif, identified by this research. A substrate assay designed to investigate the response of these genes from T. harzianum and T. hamatum to a simple carbon source (glycerol) showed that, in contrast to chit42 and prb1, xbg1.3-110 was not expressed. Further comparison of the expression patterns of these three genes between T. harzianum and T. hamatum using the glycerol substrate assay showed that no chit42 or prb1 expression could be detected in T. harzianum when it was grown under the same conditions as T. hamatum. This showed that the response of these genes to glycerol was species specific and that a single expression pattern for these genes was not common to all Trichoderma species. Confrontation assays were used to investigate the response of the three T. hamatum genes to the more complex substrate posed by the fungal pathogen

  8. Challenges in pKa Predictions for Proteins: The case of Asp213 in Human Proteinase 3

    Science.gov (United States)

    Hajjar, Eric; Dejaegere, Annick; Reuter, Nathalie

    2009-09-01

    Knowledge of the protonation states of the ionizable residues in an enzyme is a prerequisite to an accurate description of its structure and mechanism. In practice, the use of the inappropriate protonation state for an amino acid in a molecular modeling computation (e.g., molecular dynamics simulation) is likely to lead to unrealistic results. Although methods using solvers of the linearized Poisson-Boltzmann equation have proven to yield accurate pKa predictions, they bear a number of limitations. They are quite demanding in terms of computational power and are sensitive to representation of the charges and their position (force field and protein conformation). Moreover they depend on the choice of a dielectric constant for the protein interior. In this manuscript, we describe the difficulties met when trying to predict the protonation state of a buried amino acid, located in a protein for which very little biochemical data is available. Such a case is highly representative of the challenges faced in theoretical biology studies. Proteinase 3 (PR3) is an enzyme involved in proteolytic events associated with inflammation. It is a potential target in the development of new anti-inflammatory therapeutic strategies. We report the results of pKa predictions of the aspartic acid 213 of PR3 with a FDPB solver. We probed the influence of the choice of the dielectric constant for the protein interior ɛp and the benefits of conformational sampling by molecular dynamics (MD) on the pKa prediction of this carboxylate group. Using only the FDPB calculations, we could not conclude on the protonation state of Asp213. MD simulations confronted to knowledge of the ligand-binding and reaction mechanism led us to decide on a protonated form of this aspartic acid. We also demonstrate that the use of the wrong protonation state leads to an unreliable structural model for PR3. pKa prediction with a fast empirical method yielded a pKa of 8.4 for Asp213, which is in agreement with our

  9. Enzyme replacement therapy for alpha-mannosidosis

    DEFF Research Database (Denmark)

    Borgwardt, Line Gutte; Dali, Christine I.; Fogh, J

    2013-01-01

    Alpha-mannosidosis (OMIM 248500) is a rare lysosomal storage disease (LSD) caused by alpha-mannosidase deficiency. Manifestations include intellectual disabilities, facial characteristics and hearing impairment. A recombinant human alpha-mannosidase (rhLAMAN) has been developed for weekly intrave...... intravenous enzyme replacement therapy (ERT). We present the preliminary data after 12 months of treatment....

  10. THE ALPHA/BETA-HYDROLASE FOLD

    NARCIS (Netherlands)

    OLLIS, DL; CHEAH, E; CYGLER, M; FROLOW, F; FRANKEN, SM; HAREL, M; REMINGTON, SJ; SILMAN, [No Value; SCHRAG, J; SUSSMAN, JL; VERSCHUEREN, KHG; GOLDMAN, A

    We have identified a new protein fold-the alpha/beta-hydrolase fold-that is common to several hydrolytic enzymes of widely differing phylogenetic origin and catalytic function. The core of each enzyme is similar: an alpha/beta-sheet, not barrel, of eight beta-sheets connected by alpha-helices. These

  11. Electrophoretic analysis of proteinases in sodium dodecyl sulfate-polyacrylamide gels containing copolymerized radiolabeled protein substrates: Application to proenkephalin processing enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Irvine, J.W.; Roberts, S.F.; Lindberg, I. (Louisiana State Univ. Medical Center, New Orleans (USA))

    1990-10-01

    A novel method is described for the zymographic analysis of proteinases in sodium dodecyl sulfate-polyacrylamide gels containing copolymerized radiolabeled protein substrates such as ({sup 35}S)methionine-labeled proenkephalin or {sup 125}I-labeled proinsulin. After electrophoresis the enzyme is reactivated and cleaves the radiolabeled in situ substrate into smaller peptides. These small peptides are able to diffuse out of the gel, leaving clear areas against a dark background when visualized by autoradiography. The technique can be used to detect as little as 200 fg of trypsin using only 50 ng (1.25 microCi) of ({sup 35}S)proenkephalin. Soluble- and membrane-bound adrenal trypsin-like enzyme were isolated from bovine adrenal chromaffin granules. Both proteinases cleaved ({sup 35}S)methionine-labeled proenkephalin but not {sup 125}I-labeled proinsulin. Moreover, both had a Mr of approximately 30,000. The potential of this technique for general use is discussed. An additional method using the synthetic fluorogenic substrate t-butoxycarbonyl Glu-Lys-Lys aminomethylcoumarin is also described.

  12. Assessment of cathepsin D and L-like proteinases of poultry red mite, Dermanyssus gallinae (De Geer), as potential vaccine antigens.

    Science.gov (United States)

    Bartley, Kathryn; Huntley, John F; Wright, Harry W; Nath, Mintu; Nisbet, Alasdair J

    2012-05-01

    Vaccination is a feasible strategy for controlling the haematophagous poultry red mite Dermanyssus gallinae. A cDNA library enriched for genes upregulated after feeding was created to identify potential vaccine antigens. From this library, a gene (Dg-CatD-1) encoding a 383 amino acid protein (Dg-CatD-1) with homology to cathepsin D lysosomal aspartyl proteinases was identified as a potential vaccine candidate. A second gene (Dg-CatL-1) encoding a 341 amino acid protein (Dg-CatL-1) with homology to cathepsin L cysteine proteinases was also selected for further study. IgY obtained from naturally infested hens failed to detect Dg-CatD-1 suggesting that it is a concealed antigen. Conversely, Dg-CatL-1 was detected by IgY derived from natural-infestation, indicating that infested hens are exposed to Dg-CatL-1. Mortality rates 120 h after mites had been fed anti-Dg-CatD-1 were significantly higher than those fed control IgY (PFCatD-1 and anti-Dg-CatL-1 IgY had 4·42 and 2·13 times higher risks of dying compared with controls (PFCatD-1 and L-1 both have potential as vaccine antigens as part of a multi-component vaccine and have the potential to be improved as vaccine antigens using alternative expression systems.

  13. Nature of the pygmy dipole resonance in Ce-140 studied in (alpha, alpha 'gamma) experiments

    NARCIS (Netherlands)

    Savran, D.; Babilon, M.; van den Berg, A.M.; Harakeh, M.N.; Hasper, J.; Matic, A.; Wörtche, H.J.; Zilges, A.

    2006-01-01

    A concentration of electric-dipole excitations below the particle threshold, which is frequently denoted as the pygmy dipole resonance, has been studied in the semimagic nucleus Ce-140 in (alpha, alpha(')gamma) experiments at E-alpha=136 MeV. The technique of alpha-gamma coincidence experiments

  14. Alternative splicing of T cell receptor (TCR) alpha chain transcripts containing V alpha 1 or V alpha 14 elements.

    Science.gov (United States)

    Mahotka, C; Hansen-Hagge, T E; Bartram, C R

    1995-10-01

    Human acute lymphoblastic leukemia cell lines represent valuable tools to investigate distinct steps of the complex regulatory pathways underlying T cell receptor recombination and expression. A case in point are V delta 2D delta 3 and subsequent V delta 2D delta 3J alpha rearrangements observed in human leukemic pre-B cells as well as in normal lymphopoiesis. The functional expression of these unusual (VD) delta (JC) alpha hybrids is almost exclusively prevented by alternative splicing events. In this report we show that alternative splicing at cryptic splice donor sites within V elements is not a unique feature of hybrid TCR delta/alpha transcripts. Among seven V alpha families analyzed by RT-PCR, alternatively spliced products were observed in TCR alpha recombinations containing V alpha 1 or V alpha 14 elements. In contrast to normal peripheral blood cells and thymocytes, the leukemia cell line JM expressing functional V alpha 1J alpha 3C alpha transcripts lacked evidence of aberrant TCR alpha RNA species.

  15. Alpha particles detection in nitrocellulose

    International Nuclear Information System (INIS)

    Romero C, M.

    1976-01-01

    The method for the manufacturing of the detection films follows these steps: preparation of the mass which includes nitrocellulose in the form of cotton as raw material ethyl acetate, cellosolve acetate, isopropyl and butyl alcohols as solvents and dioctyl phtalate as plasticiser; dilution of the paste; pouring of the diluted mass; and drying of the detection films. The results obtained experimentally are: The determination of the development times of the different thicknesses of the manufactured films. Response linearity of the detectors, variation of the number of tracks according to the distance of the source to the detector. Sizes of the diameter of the tracks depending of the distance detector-alpha emmission source. As a conclusion we can say the the nitrocellulose detectors are specific for alpha radiation; the more effective thicknesses in uranium prospecting works were those of 60 microns, since for the laboratory works the thicknesses of 30 to 40 microns were the ideal; the development technique of the detection films is simple and cheap and can be realized even in another place than the laboratory; this way of the manufacturing of nitrocellulose detection film sensitive to alpha nuclear radiation is open to future research. (author)

  16. Measurement and analysis of $\\alpha$ particle induced reactions on yttrium

    CERN Document Server

    Singh, N L; Chintalapudi, S N

    2000-01-01

    Excitation functions for /sup 89/Y[( alpha ,3n); ( alpha ,4n); ( alpha , p3n); ( alpha , alpha n); ( alpha , alpha 2n)] reactions were measured up to 50 MeV using stacked foil activation technique and HPGe gamma ray spectroscopy method. The experimental data were compared with calculations considering equilibrium as well as preequilibrium reactions according to the hybrid model of Blann (ALICE/90). For ( alpha , xnyp) type of reactions, the precompound contributions are described by the model. There seems to be indications of direct inelastic scattering effects in ( alpha , alpha xn) type of reactions. To the best of our knowledge, the excitation functions for ( alpha ,4n), ( alpha , p3n), ( alpha , alpha n) and ( alpha , alpha 2n) reactions were measured for the first time. (23 refs).

  17. Measurement and analysis of alpha particle induced reactions on yttrium

    Energy Technology Data Exchange (ETDEWEB)

    Singh, N.L.; Gadkari, M.S. [Baroda Univ. (India). Dept. of Physics; Chintalapudi, S.N. [IUC-DAEF Calcutta Centre, Calcutta (India)

    2000-05-01

    Excitation functions for {sup 89}Y[({alpha},3n);({alpha},4n);({alpha},p3n);({alpha},{alpha}n);({alpha},{alpha}2n)] reactions were measured up to 50 MeV using stacked foil activation technique and HPGe gamma ray spectroscopy method. The experimental data were compared with calculations considering equilibrium as well as preequilibrium reactions according to the hybrid model of Blann (ALICE/90). For ({alpha},xnyp) type of reactions, the precompound contributions are described by the model. There seems to be indications of direct inelastic scattering effects in ({alpha},{alpha}xn) type of reactions. To the best of our knowledge, the excitation functions for ({alpha},4n), ({alpha},p3n), ({alpha},{alpha}n) and ({alpha},{alpha}2n) reactions were measured for the first time. (orig.)

  18. Interferon Alpha in Systemic Lupus Erythematosus

    Directory of Open Access Journals (Sweden)

    Timothy B. Niewold

    2010-01-01

    Full Text Available The pleiotropic cytokine interferon alpha is involved in multiple aspects of lupus etiology and pathogenesis. Interferon alpha is important under normal circumstances for antiviral responses and immune activation. However, heightened levels of serum interferon alpha and expression of interferon response genes are common in lupus patients. Lupus-associated autoantibodies can drive the production of interferon alpha and heightened levels of interferon interfere with immune regulation. Several genes in the pathways leading to interferon production or signaling are associated with risk for lupus. Clinical and cellular manifestations of excess interferon alpha in lupus combined with the genetic risk factors associated with interferon make this cytokine a rare bridge between genetic risk and phenotypic effects. Interferon alpha influences the clinical picture of lupus and may represent a therapeutic target. This paper provides an overview of the cellular, genetic, and clinical aspects of interferon alpha in lupus.

  19. Novel alpha-mannosidase inhibitors; Nye alfa-mannosidaseinhibitorer

    Energy Technology Data Exchange (ETDEWEB)

    Farr, R.A.; Kang, M.S.; Peet, N.P.; Sunkara, S.P.

    1997-05-20

    [4S-(4{alpha}, 4a{beta}, 6{alpha}, 7{alpha}, 7a{alpha})]-Octahydro-1H-1-pyridine-4,5,6,7-tetrols and [4R-(4{alpha}, 4a{alpha}, 5{alpha}, 6{beta}, 7{beta}, 7a{beta})]-octahydro-1H-1-pyridine-4,5,6,7-tetrols are useful inhibitors of alpha-mannosidase and are useful immunostimulants, chemoprotective and radioprotective agents and antimetastatic agents.

  20. Activator protein 2alpha mediates parathyroid TGF-alpha self-induction in secondary hyperparathyroidism.

    Science.gov (United States)

    Arcidiacono, Maria Vittoria; Cozzolino, Mario; Spiegel, Noah; Tokumoto, Masanori; Yang, Jing; Lu, Yan; Sato, Tetsuhiko; Lomonte, Carlo; Basile, Carlo; Slatopolsky, Eduardo; Dusso, Adriana S

    2008-10-01

    In secondary hyperparathyroidism, enhanced expression of TGF-alpha in the parathyroid leads to its own upregulation, generating a feed-forward loop for TGF-alpha activation of its receptor, EGFR receptor (EGFR), which promotes parathyroid hyperplasia. These studies examined the role of activator protein 2alpha (AP2), an inducer of TGF-alpha gene transcription, in the upregulation of parathyroid TGF-alpha in secondary hyperparathyroidism. In rat and human secondary hyperparathyroidism, parathyroid AP2 expression strongly correlated with TGF-alpha levels and with the rate of parathyroid growth, as expected. Furthermore, the increases in rat parathyroid content of AP2 and its binding to a consensus AP2 DNA sequence preceded the increase in TGF-alpha induced by high dietary phosphate. More significant, in A431 cells, which provide a model of enhanced TGF-alpha and TGF-alpha self-induction, mutating the core AP2 site of the human TGF-alpha promoter markedly impaired promoter activity induced by endogenous or exogenous TGF-alpha. Important for therapy, in five-sixths nephrectomized rats fed high-phosphate diets, inhibition of parathyroid TGF-alpha self-induction using erlotinib, a highly specific inhibitor of TGF-alpha/EGFR-driven signals, reduced AP2 expression dosage dependently. This suggests that the increases in parathyroid AP2 occur downstream of EGFR activation by TGF-alpha and are required for TGF-alpha self-induction. Indeed, in A431 cells, erlotinib inhibition of TGF-alpha self-induction caused parallel reductions in AP2 expression and nuclear localization, as well as TGF-alpha mRNA and protein levels. In summary, increased AP2 expression and transcriptional activity at the TGF-alpha promoter determine the severity of the hyperplasia driven by parathyroid TGF-alpha self-upregulation in secondary hyperparathyroidism.

  1. The impact of single nucleotide polymorphism in monomeric alpha-amylase inhibitor genes from wild emmer wheat, primarily from Israel and Golan

    Directory of Open Access Journals (Sweden)

    Yan Ze-Hong

    2010-06-01

    Full Text Available Abstract Background Various enzyme inhibitors act on key insect gut digestive hydrolases, including alpha-amylases and proteinases. Alpha-amylase inhibitors have been widely investigated for their possible use in strengthening a plant's defense against insects that are highly dependent on starch as an energy source. We attempted to unravel the diversity of monomeric alpha-amylase inhibitor genes of Israeli and Golan Heights' wild emmer wheat with different ecological factors (e.g., geography, water, and temperature. Population methods that analyze the nature and frequency of allele diversity within a species and the codon analysis method (comparing patterns of synonymous and non-synonymous changes in protein coding sequences were used to detect natural selection. Results Three hundred and forty-eight sequences encoding monomeric alpha-amylase inhibitors (WMAI were obtained from 14 populations of wild emmer wheat. The frequency of SNPs in WMAI genes was 1 out of 16.3 bases, where 28 SNPs were detected in the coding sequence. The results of purifying and the positive selection hypothesis (p Conclusions Great diversity at the WMAI locus, both between and within populations, was detected in the populations of wild emmer wheat. It was revealed that WMAI were naturally selected for across populations by a ratio of dN/dS as expected. Ecological factors, singly or in combination, explained a significant proportion of the variations in the SNPs. A sharp genetic divergence over very short geographic distances compared to a small genetic divergence between large geographic distances also suggested that the SNPs were subjected to natural selection, and ecological factors had an important evolutionary role in polymorphisms at this locus. According to population and codon analysis, these results suggested that monomeric alpha-amylase inhibitors are adaptively selected under different environmental conditions.

  2. Leishmania mortality in sand fly blood meal is not species-specific and does not result from direct effect of proteinases.

    Science.gov (United States)

    Pruzinova, Katerina; Sadlova, Jovana; Myskova, Jitka; Lestinova, Tereza; Janda, Jozef; Volf, Petr

    2018-01-15

    Leishmania development in sand flies is confined to the alimentary tract and is closely connected with blood meal digestion. Previously, it has been published that activities of sand fly midgut proteases are harmful to Leishmania, especially to amastigote-promastigote transition forms. However, our experiments with various Leishmania-sand fly pairs gave quite opposite results. We evaluated the effect of semi-digested midgut content on different life stages of Leishmania donovani and Leishmania major in vitro. Various morphological forms of parasites, including macrophage-derived amastigotes and transition forms, were incubated 2 h with midguts dissected at various intervals (6-72 h) post-blood meal or with commercially available proteinase, and their viability was determined using flow cytometry. In parallel, using amastigote-initiated experimental infections, we compared development of L. donovani in sand flies that are either susceptible (Phlebotomus argentipes and P. orientalis) or refractory (P. papatasi and Sergentomyia schwetzi) to this parasite. In vitro, sand fly midgut homogenates affected L. major and L. donovani in a similar way; in all sand fly species, the most significant mortality effect was observed by the end of the blood meal digestion process. Surprisingly, the most susceptible Leishmania stages were promastigotes, while mortality of transforming parasites and amastigotes was significantly lower. Parasites were also susceptible to killing by rabbit blood in combination with proteinase, but resistant to proteinase itself. In vivo, L. donovani developed late-stage infections in both natural vectors; in P. argentipes the development was much faster than in P. orientalis. On the other hand, in refractory species P. papatasi and S. schwetzi, promastigotes survived activity of digestive enzymes but were lost during defecation. We demonstrated that Leishmania transition forms are more resistant to the killing effect of semi-digested blood meal than 24

  3. Alpha particle studies during JET DT experiments

    International Nuclear Information System (INIS)

    1999-01-01

    The 1997 DT experiment (DTE1) at the Joint European Torus included studies of the behaviour of alpha particles in high temperature plasmas. Clear alpha particle heating was observed in a series of otherwise similar 10MW hot-ion H-modes by scanning the DT mixture from 0%T to 93%T. Maxima in central temperature and energy content were obtained which corresponded with the maximum in fusion yield. Alfven Eigenmodes (AEs) have been detected in JET, driven by NBI or ICRH fast ions. However, in agreement with theory, no AE activity was observed in DT plasmas which could be attributed to alpha particle drive, except in the afterglow of some Optimised Shear pulses. Ion Cyclotron Emission (ICE) was detected at harmonics of the alpha particle cyclotron frequency at the outer edge of the plasma. The ICE is interpreted as being close to magnetoacoustic cyclotron instability, driven by inverted alpha distributions at the plasma edge. The high-energy neutral particle spectra showed features, which are ascribed to a mixture of alphas, neutralised by helium-like impurities, and deuterons, born from elastic collisions with alpha particles and neutralised by hydrogen-like impurities. The results of all these studies are consistent with classical alpha particle trapping and slowing-down. Future DT experiments will aim to increase alpha particle pressure, so interactions with plasma instabilities can be studied. The measurement of knock-on neutral triton spectra offers a clean way to determine confined alpha densities in these future experiments. (author)

  4. Alpha particle studies during JET DT experiments

    International Nuclear Information System (INIS)

    2001-01-01

    The 1997 DT experiment (DTE1) at the Joint European Torus included studies of the behaviour of alpha particles in high temperature plasmas. Clear alpha particle heating was observed in a series of otherwise similar 10MW hot-ion H-modes by scanning the DT mixture from 0%T to 93%T. Maxima in central temperature and energy content were obtained which corresponded with the maximum in fusion yield. Alfven Eigenmodes (AEs) have been detected in JET, driven by NBI or ICRH fast ions. However, in agreement with theory, no AE activity was observed in DT plasmas which could be attributed to alpha particle drive, except in the afterglow of some Optimised Shear pulses. Ion Cyclotron Emission (ICE) was detected at harmonics of the alpha particle cyclotron frequency at the outer edge of the plasma. The ICE is interpreted as being close to magnetoacoustic cyclotron instability, driven by inverted alpha distributions at the plasma edge. The high-energy neutral particle spectra showed features, which are ascribed to a mixture of alphas, neutralised by helium-like impurities, and deuterons, born from elastic collisions with alpha particles and neutralised by hydrogen-like impurities. The results of all these studies are consistent with classical alpha particle trapping and slowing-down. Future DT experiments will aim to increase alpha particle pressure, so interactions with plasma instabilities can be studied. The measurement of knock-on neutral triton spectra offers a clean way to determine confined alpha densities in these future experiments. (author)

  5. The Alpha Magnetic Spectrometer (AMS)

    CERN Document Server

    Alcaraz, J; Ambrosi, G; Anderhub, H; Ao, L; Arefev, A; Azzarello, P; Babucci, E; Baldini, L; Basile, M; Barancourt, D; Barão, F; Barbier, G; Barreira, G; Battiston, R; Becker, R; Becker, U; Bellagamba, L; Bene, P; Berdugo, J; Berges, P; Bertucci, B; Biland, A; Bizzaglia, S; Blasko, S; Bölla, G; Boschini, M; Bourquin, Maurice; Brocco, L; Bruni, G; Buénerd, M; Burger, J D; Burger, W J; Cai, X D; Camps, C; Cannarsa, P; Capell, M; Casadei, D; Casaus, J; Castellini, G; Cecchi, C; Chang, Y H; Chen, H F; Chen, H S; Chen, Z G; Chernoplekov, N A; Tzi Hong Chiueh; Chuang, Y L; Cindolo, F; Commichau, V; Contin, A; Crespo, P; Cristinziani, M; Cunha, J P D; Dai, T S; Deus, J D; Dinu, N; Djambazov, L; Dantone, I; Dong, Z R; Emonet, P; Engelberg, J; Eppling, F J; Eronen, T; Esposito, G; Extermann, P; Favier, Jean; Fiandrini, E; Fisher, P H; Flügge, G; Fouque, N; Galaktionov, Yu; Gervasi, M; Giusti, P; Grandi, D; Grimm, O; Gu, W Q; Hangarter, K; Hasan, A; Hermel, V; Hofer, H; Huang, M A; Hungerford, W; Ionica, M; Ionica, R; Jongmanns, M; Karlamaa, K; Karpinski, W; Kenney, G; Kenny, J; Kim, W; Klimentov, A; Kossakowski, R; Koutsenko, V F; Kraeber, M; Laborie, G; Laitinen, T; Lamanna, G; Laurenti, G; Lebedev, A; Lee, S C; Levi, G; Levchenko, P M; Liu, C L; Liu, H T; Lopes, I; Lu, G; Lü, Y S; Lübelsmeyer, K; Luckey, D; Lustermann, W; Maña, C; Margotti, A; Mayet, F; McNeil, R R; Meillon, B; Menichelli, M; Mihul, A; Mourao, A; Mujunen, A; Palmonari, F; Papi, A; Park, I H; Pauluzzi, M; Pauss, Felicitas; Perrin, E; Pesci, A; Pevsner, A; Pimenta, M; Plyaskin, V; Pozhidaev, V; Postolache, V; Produit, N; Rancoita, P G; Rapin, D; Raupach, F; Ren, D; Ren, Z; Ribordy, M; Richeux, J P; Riihonen, E; Ritakari, J; Röser, U; Roissin, C; Sagdeev, R; Sartorelli, G; Schwering, G; Scolieri, G; Seo, E S; Shoutko, V; Shoumilov, E; Siedling, R; Son, D; Song, T; Steuer, M; Sun, G S; Suter, H; Tang, X W; Ting, Samuel C C; Ting, S M; Tornikoski, M; Torsti, J; Ulbricht, J; Urpo, S; Usoskin, I; Valtonen, E; Vandenhirtz, J; Velcea, F; Velikhov, E P; Verlaat, B; Vetlitskii, I; Vezzu, F; Vialle, J P; Viertel, Gert M; Vitè, Davide F; Gunten, H V; Wallraff, W; Wang, B C; Wang, J Z; Wang, Y H; Wiik, K; Williams, C; Wu, S X; Xia, P C; Yan, J L; Yan, L G; Yang, C G; Yang, M; Ye, S W; Yeh, P; Xu, Z Z; Zhang, H Y; Zhang, Z P; Zhao, D X; Zhu, G Y; Zhu, W Z; Zhuang, H L; Zichichi, A; Zimmermann, B

    2002-01-01

    The Alpha Magnetic Spectrometer (AMS) is a large acceptance (0.65 sr m sup 2) detector designed to operate in the International Space Station (ISS) for three years. The purposes of the experiment are to search for cosmic antimatter and dark matter and to study the composition and energy spectrum of the primary cosmic rays. A 'scaled-down' version has been flown on the Space Shuttle Discovery for 10 days in June 1998. The complete AMS is programmed for installation on the ISS in October 2003 for an operational period of 3 yr. This contribution reports on the experimental configuration that will be installed on the ISS.

  6. Targeted alpha therapy for cancer

    Energy Technology Data Exchange (ETDEWEB)

    Allen, Barry J [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Raja, Chand [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Rizvi, Syed [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Li Yong [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Tsui, Wendy [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Zhang, David [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Song, Emma [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Qu, C F [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Kearsley, John [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Graham, Peter [Centre for Experimental Radiation Oncology, St George Cancer Care Centre, Gray St, Kogarah 2217, NSW (Australia); Thompson, John [Sydney Melanoma Unit, Royal Prince Alfred Hospital, Camperdown 2050 NSW (Australia)

    2004-08-21

    Targeted alpha therapy (TAT) offers the potential to inhibit the growth of micrometastases by selectively killing isolated and preangiogenic clusters of cancer cells. The practicality and efficacy of TAT is tested by in vitro and in vivo studies in melanoma, leukaemia, colorectal, breast and prostate cancers, and by a phase 1 trial of intralesional TAT for melanoma. The alpha-emitting radioisotope used is Bi-213, which is eluted from the Ac-225 generator and chelated to a cancer specific monoclonal antibody (mab) or protein (e.g. plasminogen activator inhibitor-2 PAI2) to form the alpha-conjugate (AC). Stable alpha-ACs have been produced which have been tested for specificity and cytotoxicity in vitro against melanoma (9.2.27 mab), leukaemia (WM60), colorectal (C30.6), breast (PAI2, herceptin), ovarian (PAI2, herceptin, C595), prostate (PAI2, J591) and pancreatic (PAI2, C595) cancers. Subcutaneous inoculation of 1-1.5 million human cancer cells into the flanks of nude mice causes tumours to grow in all mice. Tumour growth is compared for untreated controls, nonspecific AC and specific AC, for local (subcutaneous) and systemic (tail vein or intraperitoneal) injection models. The {sup 213}Bi-9.2.27 AC is injected into secondary skin melanomas in stage 4 patients in a dose escalation study to determine the effective tolerance dose, and to measure kinematics to obtain the equivalent dose to organs. In vitro studies show that TAT is one to two orders of magnitude more cytotoxic to targeted cells than non-specific ACs, specific beta emitting conjugates or free isotopes. In vivo local TAT at 2 days post-inoculation completely prevents tumour formation for all cancers tested so far. Intra-lesional TAT can completely regress advanced sc melanoma but is less successful for breast and prostate cancers. Systemic TAT inhibits the growth of sc melanoma xenografts and gives almost complete control of breast and prostate cancer tumour growth. Intralesional doses up to 450 {mu

  7. The Alpha Magnetic Spectrometer (ams)

    Science.gov (United States)

    Ionica, Maria

    2004-01-01

    The Alpha Magnetic Spectrometer (AMS), once installed on the International Space Station will provide precise measurements of the cosmic ray spectra up to TeV energy range, and will search for cosmological antimatter and missing matter. A prototype version of the detector was operated successfully on the space shuttle Discovery in June 1998 (STS-91). Here we briefly report on the design of the AMS apparatus and present the results of the measurements of the fluxes of proton, electron, positron and helium from the STS-91 flight.

  8. Crossing symmetry in Alpha space

    CERN Multimedia

    CERN. Geneva

    2017-01-01

    The conformal bootstrap program aims to catalog all conformal field theories (second-order phase transitions) in D dimensions. Despite its ambitious scope much progress has been made over the past decade, e.g. in computing critical exponents for the 3D O(N) models to high precision. At this stage, analytic methods to explore the CFT landscape are not as well developed. In this talk I will describe a new mathematical framework for the bootstrap known as "alpha space", which reduces crossing symmetry to a set of integral equations. Based on arXiv:1702.08471 (with Balt van Rees) and arXiv:1703.08159.

  9. The Alpha Magnetic Spectrometer (AMS)

    International Nuclear Information System (INIS)

    Alcaraz, J.; Alpat, B.; Ambrosi, G.; Anderhub, H.; Ao, L.; Arefiev, A.; Azzarello, P.; Babucci, E.; Baldini, L.; Basile, M.; Barancourt, D.; Barao, F.; Barbier, G.; Barreira, G.; Battiston, R.; Becker, R.; Becker, U.; Bellagamba, L.; Bene, P.; Berdugo, J.; Berges, P.; Bertucci, B.; Biland, A.; Bizzaglia, S.; Blasko, S.; Boella, G.; Boschini, M.; Bourquin, M.; Brocco, L.; Bruni, G.; Buenerd, M.; Burger, J.D.; Burger, W.J.; Cai, X.D.; Camps, C.; Cannarsa, P.; Capell, M.; Casadei, D.; Casaus, J.; Castellini, G.; Cecchi, C.; Chang, Y.H.; Chen, H.F.; Chen, H.S.; Chen, Z.G.; Chernoplekov, N.A.; Chiueh, T.H.; Chuang, Y.L.; Cindolo, F.; Commichau, V.; Contin, A.; Crespo, P.; Cristinziani, M.; Cunha, J.P. da; Dai, T.S.; Deus, J.D.; Dinu, N.; Djambazov, L.; DAntone, I.; Dong, Z.R.; Emonet, P.; Engelberg, J.; Eppling, F.J.; Eronen, T.; Esposito, G.; Extermann, P.; Favier, J.; Fiandrini, E.; Fisher, P.H.; Fluegge, G.; Fouque, N.; Galaktionov, Yu.; Gervasi, M.; Giusti, P.; Grandi, D.; Grimm, O.; Gu, W.Q.; Hangarter, K.; Hasan, A.; Hermel, V.; Hofer, H.; Huang, M.A.; Hungerford, W.; Ionica, M.; Ionica, R.; Jongmanns, M.; Karlamaa, K.; Karpinski, W.; Kenney, G.; Kenny, J.; Kim, W.; Klimentov, A.; Kossakowski, R.; Koutsenko, V.; Kraeber, M.; Laborie, G.; Laitinen, T.; Lamanna, G.; Laurenti, G.; Lebedev, A.; Lee, S.C.; Levi, G.; Levtchenko, P.; Liu, C.L.; Liu, H.T.; Lopes, I.; Lu, G.; Lu, Y.S.; Luebelsmeyer, K.; Luckey, D.; Lustermann, W.; Mana, C.; Margotti, A.; Mayet, F.; McNeil, R.R.; Meillon, B.; Menichelli, M.; Mihul, A.; Mourao, A.; Mujunen, A.; Palmonari, F.; Papi, A.; Park, I.H.; Pauluzzi, M.; Pauss, F.; Perrin, E.; Pesci, A.; Pevsner, A.; Pimenta, M.; Plyaskin, V.; Pojidaev, V.; Postolache, V.; Produit, N.; Rancoita, P.G.; Rapin, D.; Raupach, F.; Ren, D.; Ren, Z.; Ribordy, M.; Richeux, J.P.; Riihonen, E.; Ritakari, J.; Roeser, U.; Roissin, C.; Sagdeev, R.; Sartorelli, G.; Schultz von Dratzig, A.; Schwering, G.; Scolieri, G.; Seo, E.S.; Shoutko, V.; Shoumilov, E.; Siedling, R.; Son, D.; Song, T.; Steuer, M.; Sun, G.S.; Suter, H.; Tang, X.W.; Ting, S.C.C.Samuel C.C.; Ting, S.M.; Tornikoski, M.; Torsti, J.; Tr umper, J.; Ulbricht, J.; Urpo, S.; Usoskin, I.; Valtonen, E.; Vandenhirtz, J.; Velcea, F.; Velikhov, E.; Verlaat, B.; Vetlitsky, I.; Vezzu, F.; Vialle, J.P.; Viertel, G.; Vite, D.; Gunten, H. Von; Wicki, S.W.S. Waldmeier; Wallraff, W.; Wang, B.C.; Wang, J.Z.; Wang, Y.H.; Wiik, K.; Williams, C.; Wu, S.X.; Xia, P.C.; Yan, J.L.; Yan, L.G.; Yang, C.G.; Yang, M.; Ye, S.W.; Yeh, P.; Xu, Z.Z.; Zhang, H.Y.; Zhang, Z.P.; Zhao, D.X.; Zhu, G.Y.; Zhu, W.Z.; Zhuang, H.L.; Zichichi, A.; Zimmermann, B.

    2002-01-01

    The Alpha Magnetic Spectrometer (AMS) is a large acceptance (0.65 sr m 2 ) detector designed to operate in the International Space Station (ISS) for three years. The purposes of the experiment are to search for cosmic antimatter and dark matter and to study the composition and energy spectrum of the primary cosmic rays. A 'scaled-down' version has been flown on the Space Shuttle Discovery for 10 days in June 1998. The complete AMS is programmed for installation on the ISS in October 2003 for an operational period of 3 yr. This contribution reports on the experimental configuration that will be installed on the ISS

  10. PGF2alpha induced differential expression of genes involved in turnover of extracellular matrix in rat decidual cells

    Directory of Open Access Journals (Sweden)

    Callegari Eduardo A

    2005-01-01

    Full Text Available Abstract In the rat, the decidual tissue is an important component for maternal recognition of pregnancy. Decidualization can be induced by either the implantation of the blastocyst or by artificial stimuli. The process of decidua formation or decidualization, is characterized by growth and differentiation of endometrial stromal cells. Prostaglandin F2alpha (PGF2α has been shown to be involved in inhibition of implantation, alteration of embryo development, induction of luteal regression, and the mediation of pregnancy loss induced by microorganism infections. In order to establish a direct role for PGF2α in decidual function, we have evaluated its effects on the expression of an extensive array of genes using primary decidual cell culture. Upon treatment with PGF2α sixty genes were significantly down-regulated whereas only six genes were up-regulated (from a total of 1176 genes studied. Interestingly, the majority of the genes inhibited by PGF2α are either directly or indirectly involved in the turnover of the extracellular matrix (ECM. Genes such as gelatinase A (MMP2, cathepsin L, tissue inhibitor metalloproteinases 2 (TIMP2 and 3 (TIMP3, plasminogen activator inhibitor1 (PAI1, tissue type plasminogen activator (tPA, urokinase plasminogen activator (tPA, endothelin 1, calponin, carboxypeptidase D and calponin acidic were down regulated. The opposite effect was observed for prostromelysin 53 kDa (proMMP3, plasma proteinase I alpha and alpha 1 antiproteinase, all of which were significantly up-regulated by PGF2α. The results strongly suggest that the abortificient role of elevated levels of PGF2α after implantation is due, in large part, to inhibition of genes involved in the normal turnover of the extracellular matrix necessary for decidual formation.

  11. Assessing the level of matrix metal proteinases 1,8,9, their tissue inhibitor, type I, in cases of odontogenic phlegmons.

    Science.gov (United States)

    Markelova, E V; Romanchuk, A L; Prosekova, E V; Krasnikov, V E; Beniova, S N

    2017-01-01

    The article considers the measured values of the level of MMP-1, MMP-8 and MMP-9, and of their tissue inhibitor Type I (TIMP-1) in the blood serum and mixed saliva samples of 78 patients (31 women - 36.2 %, 47 men - 63.8 %) suffering from odontogenic phlegmons in the oral and maxillofacial region. The study indicators were assessed through the enzyme-linked immunosorbent assay using diagnostic sets RandD Diagnostics Inc. (USA). An uncontrolled hyperactivation of metal proteinases as one of the components of the systemic inflammatory response in case of phlegmon-related complications in the oral and maxillofacial region, as well as development of the sepsis syndrome were studied and it was determined that it results in distortion of the processes of reparative hystogeny and compel us to employ new approaches to the treatment of this type of patients (Tab. 2, Fig. 1, Ref. 13).

  12. Expression of cysteine proteinase type I and II of Leishmania infantum and their recognition by sera during canine and human visceral leishmaniasis.

    Science.gov (United States)

    Rafati, Sima; Nakhaee, Alireza; Taheri, Tahere; Ghashghaii, Andishe; Salmanian, Ali Hatef; Jimenez, Maribel; Mohebali, Mehdi; Masina, Slavica; Fasel, Nicolas

    2003-01-01

    In this study, the mature domains of type I (CPB) and type II (CPA) cysteine proteinases (CPs) of Leishmania infantum were expressed and their immunogenic properties defined using sera from active and recovered cases of human visceral leishmaniasis and sera from infected dogs. Immunoblotting and ELISA analysis indicated that a freeze/thaw extract of parasite antigens showed similar and intensive recognition in both active cases of human and dog sera but lower recognition in recovered human individuals. The total IgG of actively infected human sera was higher than in recovered cases when rCPs were used as antigen. In contrast to dog sera, both active and recovered human cases have higher recognition toward rCPB than rCPA. Furthermore, the asymptomatic dogs in contrast to the symptomatic cases exhibited specific lymphocyte proliferation to both crude antigens and rCPs.

  13. Prostanoid-dependent bladder pain caused by proteinase-activated receptor-2 activation in mice: Involvement of TRPV1 and T-type Ca2+ channels

    Directory of Open Access Journals (Sweden)

    Maho Tsubota

    2018-01-01

    Full Text Available We studied the pronociceptive role of proteinase-activated receptor-2 (PAR2 in mouse bladder. In female mice, intravesical infusion of the PAR2-activating peptide, SLIGRL-amide (SL, caused delayed mechanical hypersensitivity in the lower abdomen, namely ‘referred hyperalgesia’, 6–24 h after the administration. The PAR2-triggered referred hyperalgesia was prevented by indomethacin or a selective TRPV1 blocker, and restored by a T-type Ca2+ channel blocker. In human urothelial T24 cells, SL caused delayed prostaglandin E2 production and COX-2 upregulation. Our data suggest that luminal PAR2 stimulation in the bladder causes prostanoid-dependent referred hyperalgesia in mice, which involves the activation of TRPV1 and T-type Ca2+ channels.

  14. Immune response of rats vaccinated orally with various plant-expressed recombinant cysteine proteinase constructs when challenged with Fasciola hepatica metacercariae.

    Science.gov (United States)

    Kesik-Brodacka, Malgorzata; Lipiec, Agnieszka; Kozak Ljunggren, Monika; Jedlina, Luiza; Miedzinska, Katarzyna; Mikolajczak, Magdalena; Plucienniczak, Andrzej; Legocki, Andrzej B; Wedrychowicz, Halina

    2017-03-01

    Cysteine proteinases of Fasciola hepatica are important candidates for vaccine antigens because of their role in fluke biology and host-parasite relationships. In our previous experiments, we found that a recombinant cysteine proteinase cloned from adult F. hepatica (CPFhW) can protect rats against liver fluke infections when it is administered intramuscularly or intranasally in the form of cDNA. We also observed considerable protection upon challenge following mucosal vaccination with inclusion bodies containing recombinant CPFhW produced in Escherichia coli. In this study, we explore oral vaccination, which may be the desired method of delivery and is potentially capable of preventing infections at the site of helminth entry. To provide antigen encapsulation and to protect the vaccine antigen from degradation in the intestinal tract, transgenic plant-based systems are used. In the present study, we aimed to evaluate the protective ability of mucosal vaccinations of 12-week-old rats with CPFhW produced in a transgenic-plant-based system. To avoid inducing tolerance and to maximise the immune response induced by oral immunisation, we used the hepatitis B virus (HBV) core protein (HBcAg) as a carrier. Animals were immunised with two doses of the antigen and challenged with 25 or 30 metacercariae of F. hepatica. We obtained substantial protection after oral administration of the plant-produced hybrids of CPFhW and HBcAg. The highest level of protection (65.4%) was observed in animals immunised with transgenic plants expressing the mature CPFhW enzyme flanked by Gly-rich linkers and inserted into c/e1 epitope of truncated HBcAg. The immunised rats showed clear IgG1 and IgM responses to CPFhW for 4 consecutive weeks after the challenge.

  15. Quantum Estimates of Alpha Emitter Life Time

    Directory of Open Access Journals (Sweden)

    B. Santoso

    2006-01-01

    Full Text Available Quantum estimates of several alpha radioactive life time have been made using the probability of quantum tunneling through the nuclear potential barrier. It is assumed that for a given nucleus with mass number A and isotopic number Z, there exists an alpha particle moving freely back and forth in the nucleus with mass and isotopic numbers A -4 and Z-2. If the probability of penetrating the nuclear potential barrier is Τ, then after N times (N=1/Τ hitting the barrier an alpha particle is emitted. To obtain the elapsed time for emitting an alpha particle requires N times τ0, where τ0 is the time travel for alpha across the nuclear diameter, which is dependent on alpha energy. It is assumed here that this kinetic energy is the same as the emitted energy. The emitting alpha kinetic energies here are calculated by the difference of the masses of the parent and daughter nuclei and the alpha particles. They are in closed agreement with the experimental observations. While the alpha radioactive life time are not the same order of magnitudes but give the same linearity on the logarithmic scale as function of the inverse square root of energy.

  16. Sterically hindered C(alpha, alpha)-disubstituted alpha-amino acids: synthesis from alpha-nitroacetate and incorporation into peptides.

    Science.gov (United States)

    Fu, Y; Hammarström, L G; Miller, T J; Fronczek, F R; McLaughlin, M L; Hammer, R P

    2001-10-19

    The preparation of sterically hindered and polyfunctional C(alpha,alpha)-disubstituted alpha-amino acids (alpha alpha AAs) via alkylation of ethyl nitroacetate and transformation into derivatives ready for incorporation into peptides are described. Treatment of ethyl nitroacetate with N,N-diisopropylethylamine (DIEA) in the presence of a catalytic amount of tetraalkylammonium salt, followed by the addition of an activated alkyl halide or Michael acceptor, gives the doubly C-alkylated product in good to excellent yields. Selective nitro reduction with Zn in acetic acid or hydrogen over Raney Ni gives the corresponding amino ester that, upon saponification, can be protected with the fluorenylmethyloxycarbonyl (Fmoc) group. The first synthesis of an orthogonally protected, tetrafunctional C(alpha,alpha)-disubstituted analogue of aspartic acid, 2,2-bis(tert-butylcarboxymethyl)glycine (Bcmg), is described. Also, the sterically demanding C(alpha,alpha)-dibenzylglycine (Dbg) has been incorporated into a peptide using solid-phase synthesis. It was found that once sterically congested Dbg is at the peptide N-terminus, further chain extension becomes very difficult using uronium or phosphonium salts (PyAOP, PyAOP/HOAt, HATU). However, preformed amino acid symmetrical anhydride couples to N-terminal Dbg in almost quantitative yield in nonpolar solvent (dichloroethane-DMF, 9:1).

  17. Hypothalamic PGC-1 alpha Protects Against High-Fat Diet Exposure by Regulating ER alpha

    NARCIS (Netherlands)

    Morselli, Eugenia; Fuente-Martin, Esther; Finan, Brian; Kim, Min; Frank, Aaron; Garcia-Caceres, Cristina; Navas, Carlos Rodriguez; Gordillo, Ruth; Neinast, Michael; Kalainayakan, Sarada P.; Li, Dan L.; Gao, Yuanqing; Yi, Chun-Xia; Hahner, Lisa; Palmer, Biff F.; Tschöp, Matthias H.; Clegg, Deborah J.

    2014-01-01

    High-fat diets (HFDs) lead to obesity and inflammation in the central nervous system (CNS). Estrogens and estrogen receptor alpha (ER alpha) protect premenopausal females from the metabolic complications of inflammation and obesity-related disease. Here, we demonstrate that hypothalamic PGC-1 alpha

  18. Tumor necrosis factor alpha (TNF alpha) in human skin : A comparison of different antibodies for immunohistochemistry

    NARCIS (Netherlands)

    van der Laan, N; de Leij, LFMH; Buurman, W; Timens, W; ten Duis, HJ

    Conflicting results have been reported regarding the localization and presence of TNF alpha in normal human skin, To study TNF alpha expression, we tested a panel of antibodies directed against human TNF alpha, First, antibodies were tested for immunoreactivity on cytospots of isolated

  19. Study of the pygmy dipole resonance in Mo-94 using the (alpha, alpha ' gamma) coincidence technique

    NARCIS (Netherlands)

    Derya, V.; Endres, J.; Elvers, M.; Harakeh, M. N.; Pietralla, N.; Romig, C.; Savran, D.; Scheck, M.; Siebenhuehner, F.; Stoica, V. I.; Wortche, H. J.; Zilges, A.

    2013-01-01

    The (alpha, alpha' gamma) reaction at E-alpha = 136 MeV was used to study the electric dipole response in the open-shell vibrational nucleus Mo-94 below the neutron-separation threshold. The coincidence experiment has been performed at the Kernfysisch Versneller Instituut in Groningen, The

  20. Utilização da fração semipurificada da proteinase do Trypanosoma cruzi no imunodiagnóstico da doença de Chagas The use of a semipurified fraction of Trypanosoma cruzi proteinase in immunodiagnosis of Chagas' disease

    Directory of Open Access Journals (Sweden)

    Ajax Mercês Atta

    1984-12-01

    Full Text Available Foram sensibilizadas hemácias humanas 0 Rh negativo com a fração semipurificada (Fp da proteinase do Trypanosoma cruzi, e testadas quanto a antigenicidade com soros de pacientes portadores de tripanossomíase americana crônica e de outras doenças parasitárias não relacionadas. Reações de hemaglutinação positivas foram observadas com os soros de pacientes chagásicos e com alguns soros de indivíduos portadores de leishmaniose cutaneo-mucosa. Não foram observadas reações cruzadas com os soros de pacientes portadores de leishmaniose visceral, malária, toxoplasmose, sífilis, esquistossomose e mononucleose. Os resultados obtidos são favoráveis ao emprego desta fração antigênica em testes de imunodiagnóstico da tripanossomíase americana.Group 0 Rh negative human erytrocytes were coated with the semipurified fraction of T. cruzi proteinase and tested with sera both from patients with chagas' disease and from others with unrelated parasitic diseases. Positive haemagglutination reactions were only observed with the sera from the former and with that from two patients with mucocutaneous leishmaniasis. No crossed reactions were observed with visceral leishmaniasis, malaria, toxoplasmosis syphilis, schistosomiasis or mononucleosis sera. Results suggest that this purified fraction can be used in immunodiagnosis of American Trypanosomiasis.

  1. Psychiatric Symptoms in Alpha-Mannosidosis

    Science.gov (United States)

    Malm, D.; Pantel, J.; Linaker, O. M.

    2005-01-01

    Alpha-mannosidosis is characterized by mild to moderate intellectual disability (ID), moderate to severe neurosensory hearing loss, frequent infections, psychomotor disturbances and skeletal dysmorphism. For the first time, a panel of nine alpha-mannosidosis patients with psychiatric symptoms is presented. The clinical picture has several…

  2. Coefficient Alpha Bootstrap Confidence Interval under Nonnormality

    Science.gov (United States)

    Padilla, Miguel A.; Divers, Jasmin; Newton, Matthew

    2012-01-01

    Three different bootstrap methods for estimating confidence intervals (CIs) for coefficient alpha were investigated. In addition, the bootstrap methods were compared with the most promising coefficient alpha CI estimation methods reported in the literature. The CI methods were assessed through a Monte Carlo simulation utilizing conditions…

  3. Alpha Shape Topology of the Cosmic Web

    NARCIS (Netherlands)

    Weygaert, Rien van de; Platen, Erwin; Vegter, Gert; Eldering, Bob; Kruithof, Nico

    2010-01-01

    We study the topology of the Megaparsec Cosmic Web on the basis of the Alpha Shapes of the galaxy distribution. The simplicial complexes of the alpha shapes are used to determine the set of Betti numbers (βk, k = 1, . . . , D), which represent a complete characterization of the topology of a

  4. Central and peripheral alpha-adrenoceptors

    NARCIS (Netherlands)

    van Zwieten, P A; van Meel, J. C. A.; de Jonge, A; Wilffert, B; Timmermans, P B

    1982-01-01

    The recent interest in the characterization and functional, role of alpha-adrenoceptors has prompted us to study the following different, although interdigitated, lines of research: (a) The functional role of calcium ions in the process of vasoconstriction, induced by alpha 2-adrenoceptor

  5. Producing alpha-olefins using polyketide synthases

    Energy Technology Data Exchange (ETDEWEB)

    Fortman, Jeffrey L.; Katz, Leonard; Steen, Eric J.; Keasling, Jay D.

    2018-01-02

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing an .alpha.-olefin, such as 1-hexene or butadiene. The present invention also provides for a host cell comprising the PKS and when cultured produces the .alpha.-olefin.

  6. Bayesian Meta-Analysis of Coefficient Alpha

    Science.gov (United States)

    Brannick, Michael T.; Zhang, Nanhua

    2013-01-01

    The current paper describes and illustrates a Bayesian approach to the meta-analysis of coefficient alpha. Alpha is the most commonly used estimate of the reliability or consistency (freedom from measurement error) for educational and psychological measures. The conventional approach to meta-analysis uses inverse variance weights to combine…

  7. ALPHA experiment facility and Prof. Jeffrey Hangst.

    CERN Multimedia

    Maximilien Brice

    2010-01-01

    Picture 01-07: General views of the ALPHA experiment Picture 5: Andrea Gutierrez, a PhD student from UBC, transfers liquid helium from a storage dewar into the cryostat containing the superconducting magnetic trap used by the ALPHA experiment.Picture 08-11: Jeffery Hangst, spokesperson for ALPHA Picture 12: The ALPHA silicon detector, which surrounds the trapping resion and is used for imaging antiproton annihilations (Credit University of Liverpool) Picture 13: Untrapped antihydrogen atoms annihilating on the inner surface of the ALPHA trap. These are measured by the ALPHA annihilation detector. The events are concentrated at the electrode radius of about 22.3 mm. The coordinates are defined in the Nature article, Figure 1b. Picture 14: The electrodes (gold) for the ALPHA Penning trap being inserted into the vacuum chamber and cryostat assembly. This is the trap used to combine or "mix" positrons and antiprotons to make antihydrogen. (Credit: Niels Madsen ALPHA/Swansea.) Picture 15: Top, a diagram of the...

  8. Alpha-1 Antitrypsin Deficiency (Inherited Emphysema)

    Science.gov (United States)

    ... antitrypsin inactivates elastase once it has finished its job. Without alpha 1 antitrypsin, elastase can destroy the air sacs of the lung. How is the diagnosis made? Because Alpha-1 related disease is COPD, the diagnosis is made by the same methods. Your doctor may have you do a number ...

  9. A stable lipid-induced aggregate of alpha-synuclein

    NARCIS (Netherlands)

    Drescher, Malte; van Rooijen, Bart D; Veldhuis, Gertjan; Subramaniam, Vinod; Huber, Martina

    2010-01-01

    The Parkinson's disease-related protein alpha-Synuclein (alphaS) is a 140 residue intrinsically disordered protein. Its membrane-binding properties are thought to be relevant for its physiological or pathologic activity. Here, the interaction of alphaS with POPG

  10. Improved peak shape fitting in alpha spectra.

    Science.gov (United States)

    Pommé, S; Caro Marroyo, B

    2015-02-01

    Peak overlap is a recurrent issue in alpha-particle spectrometry, not only in routine analyses but also in the high-resolution spectra from which reference values for alpha emission probabilities are derived. In this work, improved peak shape formulae are presented for the deconvolution of alpha-particle spectra. They have been implemented as fit functions in a spreadsheet application and optimum fit parameters were searched with built-in optimisation routines. Deconvolution results are shown for a few challenging spectra with high statistical precision. The algorithm outperforms the best available routines for high-resolution spectrometry, which may facilitate a more reliable determination of alpha emission probabilities in the future. It is also applicable to alpha spectra with inferior energy resolution. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  11. Technical Basis for the Use of Alpha Absorption Corrections on RCF Gross Alpha Data

    International Nuclear Information System (INIS)

    Ceffalo, G.M.

    1999-01-01

    This document provides the supporting data and rationale for making absorption corrections to gross alpha data to correct alpha data for loss due to absorption in the sample matrix. For some time there has been concern that the gross alpha data produced by the Environmental Restoration Contractor Radiological Counting Facility, particularly gross alpha analysis on soils, has been biased toward low results, as no correction for self-absorption was applied to the counting data. The process was investigated, and a new methodology for alpha self-absorption has been developed

  12. Elimination of alpha-gal xenoreactive epitope: alpha-galactosidase treatment of porcine heart valves.

    Science.gov (United States)

    Choi, Sun-Young; Jeong, Hee-Jin; Lim, Hong-Gook; Park, Seong-Sik; Kim, Soo-Hwan; Kim, Yong Jin

    2012-05-01

    Porcine heart valves are among the most widely used tissue valves in clinical heart valve implantation. However, immunologic responses have been implicated as potential causes of the limited durability of xenograft heart valves. The study aim was to determine the effectiveness of alpha-galactosidase treatment used to degrade the major xenoreactive antigens found in xenograft heart valves. Fresh porcine heart valves and pericardium treated with alpha-galactosidase were studied to evaluate the xenoreactive galactose (alpha1,3) galactose (alpha-gal) antigen. Removal of the alpha-gal epitope from the porcine heart valve was monitored via 3,3'-diaminobenzidine staining intensity, while the removal of alpha-gal from N-glycans on porcine heart valves treated with recombinant alpha-galactosidase was determined either qualitatively or quantitatively by mass fingerprinting using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The porcine pericardium was used for monitoring the change in mechanical properties after alpha-galactosidase treatment. In addition, the biomechanical modification property of collagen fiber rearrangement on tissue was assessed using transmission electron microscopy (TEM). Following a 24-h incubation at pH 7.2, 4 degrees C, employing 0.1 U/ml of Bacteroides thetaiotaomicron-derived recombinant alpha-galactosidase, the enzyme effectively removed the alpha-gal epitopes expressed on porcine heart valves. The identification type of alpha-gal N-glycan on fresh aortic valve, aortic wall, pulmonary valve, and pulmonary wall was 7.1%, 10.3%, 6% and 8%, respectively. In the presence of alpha-galactosidase treatment, alpha-gal-containing N-glycans were converted into alpha-gal-negative N-glycans. Likewise, alpha-gal-containing N-glycans were not detected when MALDI-TOF MS quantitative analysis was used. Furthermore, no significant difference was observed in the mechanical properties and findings from TEM in alpha

  13. Three complement-like repeats compose the complete alpha2-macroglobulin binding site in the second ligand binding cluster of the low density lipoprotein receptor-related protein.

    Science.gov (United States)

    Dolmer, Klavs; Gettins, Peter G W

    2006-11-10

    Given the importance of the low density lipoprotein receptor-related protein (LRP) as an essential endocytosis and signaling receptor for many protein ligands, and of alpha2-macroglobulin (alpha2M)-proteinase complexes as one such set of ligands, an understanding of the specificity of their interaction with LRP is an important goal. A starting point is the known role of the 138-residue receptor binding domain (RBD) in binding to LRP. Previous studies have localized high affinity alpha2M binding to the eight complement repeat (CR)-containing cluster 2 of LRP. In the present study we have identified the minimum CR domains that constitute the full binding site for RBD and, hence, for alpha2M on LRP. We report on the ability of the triple construct of CR3-4-5 to bind RBD with an affinity (Kd = 130 nM) the same as for isolated RBD to intact LRP. This Kd is 30-fold smaller than for RBD to CR5-6-7, demonstrating the specificity of the interaction with CR3-4-5. Binding requires previously identified critical lysine residues but is almost pH-independent within the range of pH values encountered between extracellular and internal compartments, consistent with an earlier proposed model of intracellular ligand displacement by intramolecular YWTD domains. The present findings suggest a model to explain the ability of LRP to bind a wide range of structurally unrelated ligands in which a nonspecific ligand interaction with the acidic region present in most CR domains is augmented by interactions with other CR surface residues that are unique to a particular CR cluster.

  14. Regions involved in binding of urokinase-type-1 inhibitor complex and pro-urokinase to the endocytic alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein. Evidence that the urokinase receptor protects pro-urokinase against binding to the endocytic receptor.

    Science.gov (United States)

    Nykjaer, A; Kjøller, L; Cohen, R L; Lawrence, D A; Garni-Wagner, B A; Todd, R F; van Zonneveld, A J; Gliemann, J; Andreasen, P A

    1994-10-14

    The alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2MR/LRP) binds several ligands, including complex between the two chain urokinase-type plasminogen activator (uPA) and type-1 plasminogen activator inhibitor (PAI-1), and the single chain zymogen pro-urokinase (pro-uPA). We have used truncated variants of uPA and PAI-1 as well as Fab fragments of monoclonal antibodies with known epitopes to identify regions in the uPA.PAI-1 complex and in pro-uPA involved in binding to alpha 2MR/LRP.uPA.PAI-1 complex bound with high affinity (EC50 about 0.4 nM) via contacts in the PAI-1 moiety as well as the uPA serine proteinase domain and the uPAA chain. Pro-uPA bound with lower affinity (EC50 about 10 nM), and efficient binding to alpha 2MR/LRP was dependent on contact with both the A chain and the serine proteinase domain. We analyzed the effect of complex formation with the urokinase receptor since this is the primary target for binding of uPA.PAI-1 and pro-uPA at the cell surface, and since it has been demonstrated that urokinase receptor-bound uPA.PAI-1 complex is internalized following interaction with alpha 2 MR/LRP (Nykjaer, A., Petersen, C. M., Møller, B., Jensen, P.H., Moestrup, S.K., Holtet, T.L., Etzerodt M., Thøgersen, H.C., Munch, M., Andreasen, P.A., and Gliemann, J. (1992) J. Biol. Chem. 267, 14543-14546). Soluble recombinant urokinase receptor blocked the binding of pro-uPA to alpha 2MR/LRP but caused only a slight reduction in the affinity for binding of uPA.PAI-1. Moreover, pro-uPA bound to the urokinase receptor at the cell surface was not internalized and degraded unless activated to uPA and complexed with PAI-1. We conclude that pro-uPA is protected against degradation via alpha 2MR/LRP when bound to uPAR due to shielding of a binding contact in the A chain, whereas the affinity of uPAR-bound uPA.PAI-1 complex for binding to alpha 2MR/LRP remains sufficient to allow rapid internalization and degradation.

  15. Determination of alpha_s using jet cross section parameterizations at hadron colliders

    CERN Document Server

    Stenzel, H

    2001-01-01

    Precise measurements of the single inclusive jet cross section have been performed by the TEVATRON experiments and will be provided by the LHC experiments extending to larger values of transverse energy. Theoretical predictions of this observable at NLO in perturbative QCD depend both on the PDF parameterization set and on the value of the strong coupling constant alpha_s. In this paper the dependence of the jet cross section on alpha_s is investigated. A method is presented to extract alpha_s(E_T) from a cross section measurement based on a parameterization of the alpha_s dependence. Systematic uncertainties and the E_T-range of applicability are discussed. A comparative study is performed between the case of ppbar at sqrt{s}=1.8 TeV (TEVATRON) and pp scattering at sqrt{s}=14 TeV (LHC).

  16. Mapping High-Velocity H-alpha and Lyman-alpha Emission from Supernova 1987A

    Science.gov (United States)

    France, Kevin; McCray, Richard; Fransson, Claes; Larsson, Josefin; Frank, Kari A.; Burrows, David N.; Challis, Peter; Kirshner, Robert P.; Chevalier, Roger A.; Garnavich, Peter; hide

    2015-01-01

    We present new Hubble Space Telescope images of high-velocity H-alpha and Lyman-alpha emission in the outer debris of SN 1987A. The H-alpha images are dominated by emission from hydrogen atoms crossing the reverse shock. For the first time we observe emission from the reverse shock surface well above and below the equatorial ring, suggesting a bipolar or conical structure perpendicular to the ring plane. Using the H-alpha imaging, we measure the mass flux of hydrogen atoms crossing the reverse shock front, in the velocity intervals (-7,500 < V(sub obs) < -2,800 km/s) and (1,000 < V(sub obs) < 7,500 km/s), ?M(sub H) = 1.2 × 10(exp -3) M/ y. We also present the first Lyman-alpha imaging of the whole remnant and new Chandra X-ray observations. Comparing the spatial distribution of the Lyman-alpha and X-ray emission, we observe that the majority of the high-velocity Lyman-alpha emission originates interior to the equatorial ring. The observed Lyman-alpha/H-alpha photon ratio, R(L-alpha/H-alpha) approx. = 17, is significantly higher than the theoretically predicted ratio of approx. = 5 for neutral atoms crossing the reverse shock front. We attribute this excess to Lyman-alpha emission produced by X-ray heating of the outer debris. The spatial orientation of the Lyman-alpha and X-ray emission suggests that X-ray heating of the outer debris is the dominant Lyman-alpha production mechanism in SN 1987A at this phase in its evolution.

  17. Direct binding of syndecan-4 cytoplasmic domain to the catalytic domain of protein kinase C alpha (PKC alpha) increases focal adhesion localization of PKC alpha

    DEFF Research Database (Denmark)

    Lim, Ssang-Taek; Longley, Robert L; Couchman, John R

    2003-01-01

    alpha. Full-length PKC alpha weakly interacted with 4V by yeast two-hybrid assays, but PKC alpha constructs that lack the pseudosubstrate region or constructs of the whole catalytic domain interacted more strongly. A mutated 4V sequence (4V(YF): LGKKPIFKK) did not interact with PKC alpha, indicating...

  18. Increased virulence and competitive advantage of a/alpha over a/a or alpha/alpha offspring conserves the mating system of Candida albicans.

    Science.gov (United States)

    Lockhart, Shawn R; Wu, Wei; Radke, Joshua B; Zhao, Rui; Soll, David R

    2005-04-01

    The majority of Candida albicans strains in nature are a/alpha and must undergo homozygosis to a/a or alpha/alpha to mate. Here we have used a mouse model for systemic infection to test the hypothesis that a/alpha strains predominate in nature because they have a competitive advantage over a/a and alpha/alpha offspring in colonizing hosts. Single-strain injection experiments revealed that a/alpha strains were far more virulent than either their a/a or alpha/alpha offspring. When equal numbers of parent a/alpha and offspring a/a or alpha/alpha cells were co-injected, a/alpha always exhibited a competitive advantage at the time of extreme host morbidity or death. When equal numbers of an engineered a/a/alpha2 strain and its isogenic a/a parent strain were co-injected, the a/a/alpha2 strain exhibited a competitive advantage at the time of host morbidity or death, suggesting that the genotype of the mating-type (MTL) locus, not associated genes on chromosome 5, provides a competitive advantage. We therefore propose that heterozygosity at the MTL locus not only represses white-opaque switching and genes involved in the mating process, but also affects virulence, providing a competitive advantage to the a/alpha genotype that conserves the mating system of C. albicans in nature.

  19. Alpha intrusion on ovenight polysomnogram

    Directory of Open Access Journals (Sweden)

    Nahapetian R

    2014-06-01

    Full Text Available No abstract available. Article truncated after 150 words. A 30 year-old Army veteran with a past medical history significant for chronic lumbar back pain stemming from a fall-from-height injury sustained in 2006 was referred to the sleep laboratory for evaluation of chronic fatigue and excessive daytime hypersomnolence. His Epworth sleepiness scale score was 16. He denied a history of snoring and witnessed apnea. Body Mass Index (BMI was 25.7 kg/m2. His main sleep related complaints were frequent nocturnal arousals, poor sleep quality, un-refreshing sleep, prolonged latency to sleep onset, and nightmares. An In-lab attended diagnostic polysomnogram was performed. Sleep efficiency was reduced (73% and overall arousal index was not significantly elevated (3.2 events/hour. The sleep study showed rapid eye movement (REM related sleep disordered breathing that did not meet diagnostic criteria for sleep apnea. There was no evidence for period limb movement disorder. However, the study was significant for alpha wave intrusion in stage N2 non-REM and stage ...

  20. Diabetes and alpha lipoic acid

    Directory of Open Access Journals (Sweden)

    Issy eLaher

    2011-11-01

    Full Text Available Diabetes mellitus is a multi-faceted metabolic disorder where there is increased oxidative stress that contributes to the pathogenesis of this debilitating disease. This has prompted several investigations into the use of antioxidants as a complementary therapeutic approach. Alpha lipoic acid, a naturally occurring dithiol compound which plays an essential role in mitochondrial bioenergetic reactions, has gained considerable attention as an antioxidant for use in managing diabetic complications. Lipoic acid quenches reactive oxygen species, chelates metal ions, and reduces the oxidized forms of other antioxidants such as vitamin C, vitamin E and glutathione. It also boosts antioxidant defense system through Nrf2-mediated antioxidant gene expression and by modulation of peroxisome proliferator activated receptors-regulated genes. ALA inhibits nuclear factor kappa B and activates AMPK in skeletal muscles, which in turn have a plethora of metabolic consequences. These diverse actions suggest that a lipoic acid acts by multiple mechanisms, many of which have only been uncovered recently. In this review we briefly summarize the known biochemical properties of lipoic acid and then discussed the oxidative mechanisms implicated in diabetic complications and the mechanisms by which lipoic acid may ameliorate these reactions. The findings of some of the clinical trials in which lipoic acid administration has been tested in diabetic patients during the last 10 years are summarized. It appears that the clearest benefit of lipoic acid supplementation is in patients with diabetic neuropathy.

  1. Diagnostics for PLX-alpha

    Science.gov (United States)

    Gilmore, Mark; Hsu, Scott

    2015-11-01

    The goal of the Plasma Liner eXperiment PLX-alpha at Los Alamos National Laboratory is to establish the viability of creating a spherically imploding plasma liner for MIF and HED applications, using a spherical array of supersonic plasma jets launched by innovative contoured-gap coaxial plasma guns. PLX- α experiments will focus in particular on establishing the ram pressure and uniformity scalings of partial and fully spherical plasma liners. In order to characterize these parameters experimentally, a suite of diagnostics is planned, including multi-camera fast imaging, a 16-channel visible interferometer (upgraded from 8 channels) with reconfigurable, fiber-coupled front end, and visible and VUV high-resolution and survey spectroscopy. Tomographic reconstruction and data fusion techniques will be used in conjunction with interferometry, imaging, and synthetic diagnostics from modeling to characterize liner uniformity in 3D. Diagnostic and data analysis design, implementation, and status will be presented. Supported by the Advanced Research Projects Agency - Energy - U.S. Department of Energy.

  2. Effects of alpha particles on zebrafish embryos

    International Nuclear Information System (INIS)

    Yum, E.H.W.; Choi, V.W.Y.; Yu, K.N.; Li, V.W.T.; Cheng, S.H.

    2008-01-01

    Full text: Ionizing radiation such as X-ray and alpha particles can damage cellular macromolecules, which can lead to DNA single- and double-strand breaks. In the present work, we studied the effects of alpha particles on dechorionated zebrafish embryos. Thin polyallyldiglycol carbonate (PADC) films with a thickness of 16 μm were prepared from commercially available PADC films (with thickness of 100 μm) by chemical etching and used as support substrates for holding zebrafish embryos for alpha-particle irradiation. These films recorded alpha-particle hit positions, quantified the number and energy of alpha particles actually incident on the embryo cells, and thus enabled the calculation of the dose absorbed by the embryo cells. Irradiation was made at 1.25 hours post fertilization (hpf) with various absorbed dose. TdT-mediated dUTP Nick-End Labeling (TUNEL) assay was performed on the embryos at different time stages after irradiation. Marked apoptosis was detected only in embryos at earlier time stages. The results showed that DNA double-strand break during zebrafish embryogenesis can be induced by alpha-particle irradiation, which suggests that zebrafish is a potential model for assessing the effects of alpha-particle radiation

  3. Cortical Alpha Activity in Schizoaffective Patients

    Directory of Open Access Journals (Sweden)

    Mahdi Moeini

    2017-02-01

    Full Text Available Objective: Electrophysiological studies have identified abnormal oscillatory activities in the cerebral cortex in schizophrenia and mood disorders. Biological and pathophysiological evidence suggests specific deficits in serotonin (5-HT receptor function in schizoaffective disorder (SA, a clinical syndrome with characteristics of both schizophrenia and bipolar disorder. This study investigated alpha oscillations in patients with SA.Method: Electroencephalography was used to measure ongoing and evoked alpha oscillations in 38 adults meeting Diagnostic and Statistical Manual of Mental Disorders–Fourth Edition (DSM–IV criteria for SA, and in 39 healthy controls.Results: Spontaneous alpha power of the participants with SA was significantly lower than that of healthy participants [F (1, 75 = 8.81, P < 0.01]. Evoked alpha activity was also decreased in SA compared to controls [F (1, 75 = 5.67, P = 0.025].Conclusion: A strong reduction of alpha power in the posterior regions may reflect abnormality in the thalamocortical circuits. It is shown that hypoxia and reduced cerebral blood flow is associated with reduced alpha activity among different regions of the brain. Therefore, it can be concluded that greatly decreased alpha activity, particularly in centro-parietal and occipital regions, is related to SA symptoms such as hallucinations.

  4. Lucid dreaming and alpha activity: a preliminary report.

    Science.gov (United States)

    Ogilvie, R D; Hunt, H T; Tyson, P D; Lucescu, M L; Jeakins, D B

    1982-12-01

    10 good dream recallers spent 2 nights in the sleep lab during which they were awakened 4 times per night from REM sleep, twice during their highest alpha activity in REM, and twice during low REM alpha. 5 were given alpha feedback training prior to sleep onset. Arousals from high alpha REM sleep yielded significantly higher lucidity ratings. Alpha feedback had no effect upon lucidity or REM alpha levels. Similarities between lucid dreams and meditative phenomena are discussed.

  5. The impact of NaCl substitution with KCl on proteinase activities cell-free extract and cell-free supernatant at different pH levels and salt concentrations: Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus.

    Science.gov (United States)

    Ayyash, M M; Sherkat, F; Shah, N P

    2012-08-01

    The effect of NaCl substitution with KCl at different pH levels (6.0, 5.5, and 5.0) and salt concentrations on proteinase activities of cell-free and supernatant of Lactobacillus delbrueckii ssp. bulgaricus 11824 (L. bulgaricus) and Streptococcus thermophilus MS (ST) was investigated. MRS broths were separately mixed with 4 salt treatments (NaCl only, 1NaCl:1KCl, 1NaCl:3KCl, and KCl only) at 2 different concentrations (5% and 10%) and incubated at 37 °C for 22 h. The cell pellets were used to prepare proteinase of cell-free extract and the cell-free supernatants were used as source of extracellular proteinases. The proteolytic activities and protein contents of both fractions were determined. The supernatants after incubation of both fractions with 3 milk caseins (α-, β-, κ-casein) were subjected to angiotensin-converting-enzyme inhibitory (ACE-inhibitory) activity and proteolytic activity by ortho-phthalaldehyde (OPA) method. Significant differences were observed in ACE-inhibitory activities and proteolytic (OPA) between salt treatments of cell-free extract and cell-free supernatant of L. bulgaricus and S. thermophilus at same salt concentration and same pH level. There was a significant effect of pH level and salt treatments interaction on ACE-inhibitory activity, OPA activity and azocasein activity. To reduce sodium concentration in cheese by substituting of NaCl with KCl, it was important to study the effect on starter culture proteinases which play a vital role in ripening and texture profile of cheese. © 2012 Institute of Food Technologists®

  6. Anomalous atomic volume of alpha-Pu

    DEFF Research Database (Denmark)

    Kollar, J.; Vitos, Levente; Skriver, Hans Lomholt

    1997-01-01

    We have performed full charge-density calculations for the equilibrium atomic volumes of the alpha-phase light actinide metals using the local density approximation (LDA) and the generalized gradient approximation (GGA). The average deviation between the experimental and the GGA atomic radii is 1.......3%. The comparison between the LDA and GGA results show that the anomalously large atomic volume of alpha-Pu relative to alpha-Np can be ascribed to exchange-correlation effects connected with the presence of low coordinated sites in the structure where the f electrons are close to the onset of localization...

  7. Alpha spectral analysis via artificial neural networks

    International Nuclear Information System (INIS)

    Kangas, L.J.; Hashem, S.; Keller, P.E.; Kouzes, R.T.; Troyer, G.L.

    1994-10-01

    An artificial neural network system that assigns quality factors to alpha particle energy spectra is discussed. The alpha energy spectra are used to detect plutonium contamination in the work environment. The quality factors represent the levels of spectral degradation caused by miscalibration and foreign matter affecting the instruments. A set of spectra was labeled with a quality factor by an expert and used in training the artificial neural network expert system. The investigation shows that the expert knowledge of alpha spectra quality factors can be transferred to an ANN system

  8. Measurements of $\\alpha_s$ in $pp$ Collisions at the LHC

    CERN Document Server

    Warburton, Andreas; The ATLAS collaboration

    2015-01-01

    The coupling of the strong force, $\\alpha_s$, is deemed to be a fundamental parameter of Nature, and, beyond the quark masses, constitutes the only free parameter in the QCD Lagrangian. Provided is an overview of CERN Large Hadron Collider (LHC) measurements of $\\alpha_s(M_Z)$ evaluated at the $Z$-boson mass and of the running of $\\alpha_s(Q)$ as a function of energy-momentum transfer $Q$. The measurements were performed by the ATLAS and CMS Collaborations using proton-proton ($pp$) collisions with centre-of-mass energies of 7 TeV and data samples with time-integrated luminosities of up to 5 fb$^{-1}$. Four different categories of observable were used in the described extractions of $\\alpha_s$: inclusive jet cross sections, 3-jet to 2-jet inclusive cross-section ratios, 3-jet mass cross sections, and top-quark pair production cross sections. These results, which include the first NNLO measurement of $\\alpha_s$ at a hadron collider and the first determinations of $\\alpha_s$ at energy scales above 1 TeV, are co...

  9. A Quantitative Electrochemiluminescence Assay for Clostridium perfringens alpha toxin

    National Research Council Canada - National Science Library

    Merrill, Gerald A; Rivera, Victor R; Neal, Dwayne D; Young, Charles; Poli, Mark A

    2006-01-01

    .... Biotinylated antibodies to C. perfringens alpha toxin bound to streptavidin paramagnetic beads specifically immunoadsorbed soluble sample alpha toxin which subsequently selectively immunoadsorbed ruthenium (Ru...

  10. The effect of NaCl substitution with KCl on proteinase activities of cell-free extract and cell-free supernatant at different pH levels and salt concentrations: Lactobacillus acidophilus and Lactobacillus casei.

    Science.gov (United States)

    Ayyash, M M; Sherkat, F; Shah, N P

    2013-01-01

    The aim of this study was to investigate the effect of substitution of NaCl with KCl at different pH levels and salt concentrations on proteinase activity of cell-free extract and cell-free supernatant of the probiotics Lactobacillus acidophilus and Lactobacillus casei. de Man, Rogosa, and Sharpe broth aliquots were mixed with 2 pure salts (NaCl and KCl) and 2 salt concentrations at 2 concentration levels (5 and 10%), inoculated with Lactobacillus acidophilus or Lactobacillus casei, and incubated aerobically at 37°C for 22 h. The cultures were then centrifuged at 4,000×g for 30 min, and the collected cell pellets were used to prepare cell-wall proteinases and the supernatants used as a source of supernatant (extracellular) proteinases. The proteolytic activity and protein content of both portions were determined. After incubation of both portions with 3 milk caseins (α-, β-, κ-casein), the supernatants were individually subjected to analysis of angiotensin-converting enzyme (ACE)-inhibitory activity and proteolytic activity using the o-phthalaldehyde method. Significant differences were observed in ACE-inhibitory and proteolytic activities between salt substitution treatments of cell-free extract and cell-free supernatant from both probiotic strains at the same salt concentration and pH level. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  11. Neutron-induced alpha radiography

    International Nuclear Information System (INIS)

    Pereira, Marco Antonio Stanojev

    2008-01-01

    A new radiography technique to inspect thin samples was developed. Low energy alpha particles, generated by a boron based screen under thermal neutron irradiation, are used as penetrating radiation. The solid state nuclear track detector CR-39 has been used to register the image. The interaction of the α - particles with the CR-39 gives rise to damages which under an adequate chemical etching became tracks the basic units forming the image. A digital system was developed for data acquisition and data analysis as well as for image processing. The irradiation and etching conditions to obtain the best radiography are 1,3 hours and 25 minutes at 70 deg C respectively. For such conditions samples having 10 μm in thickness can be inspected with a spatial resolution of 32 μm. The use of the digital system has reduced the time spent for data acquisition and data analysis and has improved the radiography image visualization. Furthermore, by using the digital system, it was possible to study several new parameters regarding the tracks which are very important to understand and study the image formation theory in solid state nuclear track detectors, the one used in this thesis. Some radiography images are also shown which demonstrate the potential of the proposed radiography technique. When compared with the other radiography techniques already in use to inspect thin samples, the present one developed in the present paper allows a smaller time to obtain the image, it is not necessary to handle liquid radioactive substances, the detector is insensitive to β, γ, X-ray and visible light. (author)

  12. Liquid scintillation alpha particle spectrometry. Progress report

    International Nuclear Information System (INIS)

    Bell, L.L.; Hakooz, S.A.; Johnson, L.O.; Nieschmidt, E.B.; Meikrantz, D.H.

    1979-12-01

    Objective to develop a technique whereby Pu may be put into solution, extracted by solvent extraction into a suitable extractive scintillant and subsequently counted. Presented here are results of attempts to separate beta and alpha activities through pulse shape discrimination. A qualitative discussion is given which yields alpha particle peak widths, resolution and response. The detection efficiency for alpha particles in a liquid scintillant is 100%. Present detection sensitivities of the equipment being used are: 4.5 x 10 -6 μCi (100 s), 1.2 x 10 -6 μCi (1000 s), and 4.0 x 10 -7 μCi (10,000 s) at the 3 sigma level. The detectability of a particular alpha-emitting species is strongly dependent upon the population of other species. The ability to discriminate depends upon the system resolution. 14 figures, 2 tables

  13. NEW APPROACHES TO CONFINED ALPHA DIAGNOSTICS

    Energy Technology Data Exchange (ETDEWEB)

    FISHER,R.K

    2004-04-01

    Three new approaches to obtain information on the confined fast alphas in International Thermonuclear Experimental Reactor (ITER) are proposed. The first technique measures the energetic charge exchange (CX) neutrals that result from the alpha collision-induced knock-on fuel ion tails undergoing electron capture on the MeV D neutral beams planned for heating and current drive. The second technique measures the energetic knock-on neutron tail due to alphas using the lengths of the proton recoil tracks produced by neutron collisions in nuclear emulsions. The range of the 14 to 20 MeV recoil protons increases by {approx}140 microns per MeV. The third approach would measure the CX helium neutrals resulting from confined alphas capturing two electrons in the ablation cloud surrounding a dense gas jet that has been proposed for disruption mitigation in ITER.

  14. NEW APPROACHES TO CONFINED ALPHA DIAGNOSTICS

    International Nuclear Information System (INIS)

    FISHER, R.K.

    2004-01-01

    Three new approaches to obtain information on the confined fast alphas in International Thermonuclear Experimental Reactor (ITER) are proposed. The first technique measures the energetic charge exchange (CX) neutrals that result from the alpha collision-induced knock-on fuel ion tails undergoing electron capture on the MeV D neutral beams planned for heating and current drive. The second technique measures the energetic knock-on neutron tail due to alphas using the lengths of the proton recoil tracks produced by neutron collisions in nuclear emulsions. The range of the 14 to 20 MeV recoil protons increases by ∼140 microns per MeV. The third approach would measure the CX helium neutrals resulting from confined alphas capturing two electrons in the ablation cloud surrounding a dense gas jet that has been proposed for disruption mitigation in ITER

  15. Strichartz estimates on $alpha$-modulation spaces

    Directory of Open Access Journals (Sweden)

    Weichao Guo

    2013-05-01

    Full Text Available In this article, we consider some dispersive equations, including Schrodinger equations, nonelliptic Schrodinger equations, and wave equations. We develop some Strichartz estimates in the frame of alpha-modulation spaces.

  16. Targeted alpha therapy: Applications and current status

    Energy Technology Data Exchange (ETDEWEB)

    Bruchertseifer, Frank, E-mail: frank.bruchertseifer@ec.europa.eu [European Commission, Joint Research Centre, Karlsruhe (Germany)

    2017-07-01

    Full text: The field of targeted alpha therapy has been developed rapidly in the last decade. Besides {sup 223}Ra, {sup 211}At and {sup 212}Pb/{sup 212}Bi the alpha emitters {sup 225}Ac and {sup 213}Bi are promising therapeutic radionuclides for application in targeted alpha therapy of cancer and infectious diseases. The presentation will give a short overview about the current clinical treatments with alpha emitting radionuclides and will place an emphasis on the most promising clinical testing of peptides and antibodies labelled with {sup 225}Ac and {sup 213}Bi for treatment of metastatic castration-resistant prostate cancer patients with glioma and glioblastoma multiform, PSMA-positive tumor phenotype and bladder carcinoma in situ. (author)

  17. Calibration of alpha surface contamination monitor

    International Nuclear Information System (INIS)

    Freitas, I.S.M. de; Goncalez, O.L.

    1990-01-01

    In this work, the results, as well as the methodology, of the calibration of an alpha surface contamination monitor are presented. The calibration factors are obtained by least-squares fitting with effective variance. (author)

  18. Solar Imagery - Chromosphere - H-Alpha

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Collection includes a variety of H-alpha photographic datasets contributed by a number of national and private solar observatories located worldwide. Solar...

  19. Energy dependence of event shapes and of $\\alpha_s$ at LEP 2

    CERN Document Server

    Abreu, P; Adye, T; Adzic, P; Albrecht, Z; Alderweireld, T; Alekseev, G D; Alemany, R; Allmendinger, T; Allport, P P; Almehed, S; Amaldi, Ugo; Amapane, N; Amato, S; Anassontzis, E G; Andersson, P; Andreazza, A; Andringa, S; Antilogus, P; Apel, W D; Arnoud, Y; Åsman, B; Augustin, J E; Augustinus, A; Baillon, Paul; Bambade, P; Barão, F; Barbiellini, Guido; Barbier, R; Bardin, Dimitri Yuri; Barker, G; Baroncelli, A; Battaglia, Marco; Baubillier, M; Becks, K H; Begalli, M; Behrmann, A; Beillière, P; Belokopytov, Yu A; Belous, K S; Benekos, N C; Benvenuti, Alberto C; Bérat, C; Berggren, M; Bertini, D; Bertrand, D; Besançon, M; Bianchi, F; Bigi, M; Bilenky, S M; Bizouard, M A; Bloch, D; Blom, H M; Bonesini, M; Bonivento, W; Boonekamp, M; Booth, P S L; Borgland, A W; Borisov, G; Bosio, C; Botner, O; Boudinov, E; Bouquet, B; Bourdarios, C; Bowcock, T J V; Boyko, I; Bozovic, I; Bozzo, M; Branchini, P; Brenke, T; Brenner, R A; Brückman, P; Brunet, J M; Bugge, L; Buran, T; Burgsmüller, T; Buschbeck, Brigitte; Buschmann, P; Cabrera, S; Caccia, M; Calvi, M; Camporesi, T; Canale, V; Carena, F; Carroll, L; Caso, Carlo; Castillo-Gimenez, M V; Cattai, A; Cavallo, F R; Chabaud, V; Chapkin, M M; Charpentier, P; Chaussard, L; Checchia, P; Chelkov, G A; Chierici, R; Chliapnikov, P V; Chochula, P; Chorowicz, V; Chudoba, J; Cieslik, K; Collins, P; Contri, R; Cortina, E; Cosme, G; Cossutti, F; Cowell, J H; Crawley, H B; Crennell, D J; Crépé, S; Crosetti, G; Cuevas-Maestro, J; Czellar, S; Davenport, Martyn; Da Silva, W; Deghorain, A; Della Ricca, G; Delpierre, P A; Demaria, N; De Angelis, A; de Boer, Wim; De Clercq, C; De Lotto, B; De Min, A; De Paula, L S; Dijkstra, H; Di Ciaccio, Lucia; Dolbeau, J; Doroba, K; Dracos, M; Drees, J; Dris, M; Duperrin, A; Durand, J D; Eigen, G; Ekelöf, T J C; Ekspong, Gösta; Ellert, M; Elsing, M; Engel, J P; Erzen, B; Espirito-Santo, M C; Falk, E; Fanourakis, G K; Fassouliotis, D; Fayot, J; Feindt, Michael; Fenyuk, A; Ferrari, P; Ferrer, A; Ferrer-Ribas, E; Ferro, F; Fichet, S; Firestone, A; Flagmeyer, U; Föth, H; Fokitis, E; Fontanelli, F; Franek, B J; Frodesen, A G; Frühwirth, R; Fulda-Quenzer, F; Fuster, J A; Galloni, A; Gamba, D; Gamblin, S; Gandelman, M; García, C; Gaspar, C; Gaspar, M; Gasparini, U; Gavillet, P; Gazis, E N; Gelé, D; Ghodbane, N; Gil, I; Glege, F; Gokieli, R; Golob, B; Gómez-Ceballos, G; Gonçalves, P; González-Caballero, I; Gopal, Gian P; Gorn, L; Górski, M; Guz, Yu; Gracco, Valerio; Grahl, J; Graziani, E; Green, C; Grimm, H J; Gris, P; Grosdidier, G; Grzelak, K; Günther, M; Guy, J; Hahn, F; Hahn, S; Haider, S; Hallgren, A; Hamacher, K; Hansen, J; Harris, F J; Hedberg, V; Heising, S; Hernández, J J; Herquet, P; Herr, H; Hessing, T L; Heuser, J M; Higón, E; Holmgren, S O; Holt, P J; Hoorelbeke, S; Houlden, M A; Hrubec, Josef; Huet, K; Hughes, G J; Hultqvist, K; Jackson, J N; Jacobsson, R; Jalocha, P; Janik, R; Jarlskog, C; Jarlskog, G; Jarry, P; Jean-Marie, B; Johansson, E K; Jönsson, P E; Joram, C; Juillot, P; Kapusta, F; Karafasoulis, K; Katsanevas, S; Katsoufis, E C; Keränen, R; Kersevan, Borut P; Khomenko, B A; Khovanskii, N N; Kiiskinen, A P; King, B J; Kinvig, A; Kjaer, N J; Klapp, O; Klein, H; Kluit, P M; Kokkinias, P; Koratzinos, M; Kostyukhin, V; Kourkoumelis, C; Kuznetsov, O; Krammer, Manfred; Kriznic, E; Krstic, J; Krumshtein, Z; Kubinec, P; Kurowska, J; Kurvinen, K L; Lamsa, J; Lane, D W; Langefeld, P; Lapin, V; Laugier, J P; Lauhakangas, R; Leder, Gerhard; Ledroit, F; Lefébure, V; Leinonen, L; Leisos, A; Leitner, R; Lemonne, J; Lenzen, Georg; Lepeltier, V; Lesiak, T; Lethuillier, M; Libby, J; Liko, D; Lipniacka, A; Lippi, I; Lörstad, B; Loken, J G; Lopes, J H; López, J M; López-Fernandez, R; Loukas, D; Lutz, P; Lyons, L; MacNaughton, J N; Mahon, J R; Maio, A; Malek, A; Malmgren, T G M; Maltezos, S; Malychev, V; Mandl, F; Marco, J; Marco, R P; Maréchal, B; Margoni, M; Marin, J C; Mariotti, C; Markou, A; Martínez-Rivero, C; Martínez-Vidal, F; Martí i García, S; Mastroyiannopoulos, N; Matorras, F; Matteuzzi, C; Matthiae, Giorgio; Masik, J; Mazzucato, F; Mazzucato, M; McCubbin, M L; McKay, R; McNulty, R; McPherson, G; Meroni, C; Meyer, W T; Migliore, E; Mirabito, L; Mitaroff, Winfried A; Mjörnmark, U; Moa, T; Moch, M; Møller, R; Mönig, K; Monge, M R; Moreau, X; Morettini, P; Morton, G A; Müller, U; Münich, K; Mulders, M; Mulet-Marquis, C; Muresan, R; Murray, W J; Muryn, B; Myatt, Gerald; Myklebust, T; Naraghi, F; Nassiakou, M; Navarria, Francesco Luigi; Navas, S; Nawrocki, K; Negri, P; Némécek, S; Neufeld, N; Neumeister, N; Nicolaidou, R; Nielsen, B S; Nikolenko, M; Nomokonov, V P; Normand, Ainsley; Nygren, A; Obraztsov, V F; Olshevskii, A G; Onofre, A; Orava, Risto; Orazi, G; Österberg, K; Ouraou, A; Paganoni, M; Paiano, S; Pain, R; Paiva, R; Palacios, J; Palka, H; Papadopoulou, T D; Papageorgiou, K; Pape, L; Parkes, C; Parodi, F; Parzefall, U; Passeri, A; Passon, O; Pegoraro, M; Peralta, L; Pernicka, Manfred; Perrotta, A; Petridou, C; Petrolini, A; Phillips, H T; Pierre, F; Pimenta, M; Piotto, E; Podobnik, T; Pol, M E; Polok, G; Poropat, P; Pozdnyakov, V; Privitera, P; Pukhaeva, N; Pullia, Antonio; Radojicic, D; Ragazzi, S; Rahmani, H; Ratoff, P N; Read, A L; Rebecchi, P; Redaelli, N G; Regler, Meinhard; Reid, D; Reinhardt, R; Renton, P B; Resvanis, L K; Richard, F; Rídky, J; Rinaudo, G; Røhne, O M; Romero, A; Ronchese, P; Rosenberg, E I; Rosinsky, P; Roudeau, Patrick; Rovelli, T; Royon, C; Ruhlmann-Kleider, V; Ruiz, A; Saarikko, H; Sacquin, Yu; Sadovskii, A; Sajot, G; Salt, J; Sampsonidis, D; Sannino, M; Schneider, H; Schwemling, P; Schwering, B; Schwickerath, U; Schyns, M A E; Scuri, F; Seager, P; Sedykh, Yu; Segar, A M; Sekulin, R L; Shellard, R C; Sheridan, A; Siebel, M; Simard, L C; Simonetto, F; Sissakian, A N; Smadja, G; Smirnov, N; Smirnova, O G; Smith, G R; Sopczak, André; Sosnowski, R; Spassoff, Tz; Spiriti, E; Sponholz, P; Squarcia, S; Stanescu, C; Stanic, S; Stevenson, K; Stocchi, A; Strub, R; Stugu, B; Szczekowski, M; Szeptycka, M; Tabarelli de Fatis, T; Tegenfeldt, F; Terranova, F; Thomas, J; Timmermans, J; Tinti, N; Tkatchev, L G; Todorova-Nová, S; Tomaradze, A G; Tomé, B; Tonazzo, A; Tortora, L; Tranströmer, G; Treille, D; Tristram, G; Trochimczuk, M; Troncon, C; Tsirou, A L; Turluer, M L; Tyapkin, I A; Tzamarias, S; Ullaland, O; Uvarov, V; Valenti, G; Vallazza, E; Van der Velde, C; van Apeldoorn, G W; van Dam, P; Van Doninck, W K; Van Eldik, J; Van Lysebetten, A; Van Vulpen, I B; Vassilopoulos, N; Vegni, G; Ventura, L; Venus, W A; Verbeure, F; Verlato, M; Vertogradov, L S; Verzi, V; Vilanova, D; Vitale, L; Vlasov, E; Vodopyanov, A S; Vollmer, C F; Voulgaris, G; Vrba, V; Wahlen, H; Walck, C; Weiser, C; Wicke, D; Wickens, J H; Wilkinson, G R; Winter, M; Witek, M; Wolf, G; Yi, J; Yushchenko, O P; Zaitsev, A; Zalewska-Bak, A; Zalewski, Piotr; Zavrtanik, D; Zevgolatakos, E; Zimin, N I; Zucchelli, G C; Zumerle, G

    1999-01-01

    Infrared and collinear safe event shape distributions and their mean values are determined using the data taken at ve di erent centre of mass energies above $M_Z$ with the DELPHI detector at LEP. From the event shapes, the strong coupling $\\alpha_s$ is extracted in $O(\\alpha^2_s)$, NLLA and a combined scheme using hadronisation corrections evaluated with fragmentation model generators as well as using an analytical power ansatz. Comparing these measurements to those obtained at MZ, the energy dependence (running) of $\\alpha_s$ is accessible. The logarithmic energy slope of the inverse strong coupling is measured to be $d\\alpha_{s}^{-1}/d log(E_{cm}) = 1.39 \\pm 0.34(stat) \\pm 0.17(syst)$, in good agreement with the QCD expectation of 1.27.

  20. Influence of fast alpha diffusion and thermal alpha buildup on tokamak reactor performance

    International Nuclear Information System (INIS)

    Uckan, N.A.; Tolliver, J.S.; Houlberg, W.A.; Attenberger, S.E.

    1988-01-01

    The effect of fast alpha diffusion and thermal alpha accumulation on the confinement capability of a candidate Engineering Test Reactor plasma (Tokamak Ignition/Burn Experimental Reactor) in achieving ignition and steady-state driven operation has been assessed using both global and 1-1/2-dimensional transport models. Estimates are made of the threshold for radial diffusion of fast alphas and thermal alpha buildup. It is shown that a relatively low level of radial transport, when combined with large gradients in the fast alpha density, leads to a significant radial flow with a deleterious effect on plasma performance. Similarly, modest levels of thermal alpha concentration significantly influence the ignition and steady-state burn capability

  1. Measurement of the angle alpha at BABAR

    Energy Technology Data Exchange (ETDEWEB)

    Perez, A.; /Orsay, LAL

    2009-06-25

    The authors present recent measurements of the CKM angle {alpha} using data collected by the BABAR detector at the PEP-II asymmetric-energy e{sup +}e{sup -} collider at the SLAC National Accelerator Laboratory, operating at the {Upsilon}(4S) resonance. They present constraints on {alpha} from B {yields} {pi}{pi}, B {yields} {rho}{rho} and B {yields} {rho}{pi} decays.

  2. Improved peak shape fitting in alpha spectra

    OpenAIRE

    POMME Stefaan; CARO MARROYO BELEN

    2014-01-01

    Peak overlap is a recurrent issue ina lpha-particle spectrometry, not only in routine analyses but also in the high-resolution spectra from which reference values for alpha emission probabilities are derived. In this work, improved peak shape formulae are presented for the deconvolution of alpha-particle spectra. They have been implemented as fit functions in a spreadsheet application and optimum fit parameters were searched with built-in optimisation routines. Deconvolution results are shown...

  3. Conceptual design report for alpha waste incinerator

    International Nuclear Information System (INIS)

    1979-04-01

    The Alpha Waste Incinerator, a new facility in the SRP H-Area, will process transuranic or alpha-contaminated combustible solid wastes. It will seal the radioactive ash and scrubbing salt residues in cans for interim storage in drums on site burial ground pads. This report includes objectives, project estimate, schedule, standards and criteria, excluded costs, safety evaluation, energy consumption, environmental assessment, and key drawings

  4. Beteigeuze (Alpha Orionis) und Mintaka (Delta Orionis)

    Science.gov (United States)

    Vollmann, Wolfgang

    2013-02-01

    Magnitude measures transformed to Johnson V of Alpha Orionis (Betelgeuse) and Delta Orionis with a wide-angle lens and DSLR are presented and discussed. Alpha Orionis light changes are shown clearly. The primary and secondary eclipses of Delta Orionis with amplitudes of 0.12 and 0.05 mag respectively are clearly recorded. They occur near phase 0.00 and 0.50 respectively of current elements from VSX (2).

  5. Considering the determination of an alpha value

    International Nuclear Information System (INIS)

    Lorenz, B.

    1987-01-01

    Following an outline of the most important international methods of evaluating an alpha value (the monetary equivalent of one man-sievert) and an approach deemed suitable for use in the GDR, it is recommended that alpha be taken as 30,000 Mark per man-sievert in national cost-benefit analyses. This value should be revisited every five to ten years. (author)

  6. Alpha-root Processes for Derivatives pricing

    OpenAIRE

    Balakrishna, BS

    2010-01-01

    A class of mean reverting positive stochastic processes driven by alpha-stable distributions, referred to here as alpha-root processes in analogy to the square root process (Cox-Ingersoll-Ross process), is a subclass of affine processes, in particular continuous state branching processes with immigration (CBI processes). Being affine, they provide semi-analytical results for the implied term structures as well as for the characteristic exponents for their associated distributions. Their use h...

  7. Self-assembling, dynamic alphaPNAs

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2009-01-01

    In the recent report published in Science, Ghadiri and coworkers describe dynamic tPNAs, alphaPNA derivatives with a nucleobase attached via a thioester bond that are a step forward toward self-repairing and replicating molecules.......In the recent report published in Science, Ghadiri and coworkers describe dynamic tPNAs, alphaPNA derivatives with a nucleobase attached via a thioester bond that are a step forward toward self-repairing and replicating molecules....

  8. Alpha-emitters for medical therapy workshop

    International Nuclear Information System (INIS)

    Feinendegen, L.E.; McClure, J.J.

    1996-01-01

    A workshop on ''Alpha-Emitters for Medical Therapy'' was held May 30-31, 1996 in Denver Colorado to identify research goals and potential clinical needs for applying alpha-particle emitters and to provide DOE with sufficient information for future planning. The workshop was attended by 36 participants representing radiooncology, nuclear medicine, immunotherapy, radiobiology, molecular biology, biochemistry, radiopharmaceutical chemistry, dosimetry, and physics. This report provides a summary of the key points and recommendations arrived at during the conference

  9. Cytokine vaccination: neutralising IL-1alpha autoantibodies induced by immunisation with homologous IL-1alpha

    DEFF Research Database (Denmark)

    Svenson, M; Hansen, M B; Thomsen, Allan Randrup

    2000-01-01

    High-affinity IgG autoantibodies (aAb) to IL-1alpha are among the most frequently found aAb to cytokines in humans. To establish an animal model with aAb to IL-1alpha, we immunised mice with recombinant murine IL-1alpha. Unprimed and Bacille Calmette-Guérin (BCG)-primed BALB/cA mice were vaccinat...

  10. Liquid-scintillation alpha-detection techniques

    International Nuclear Information System (INIS)

    McKlveen, J.W.; McDowell, W.J.

    1983-01-01

    Accurate, quantitative determinations of alpha-emitting nuclides by conventional plate-counting methods are difficult because of sample self-absorption problems in counting and because of non-reproducible losses in conventional sample separation methods. Liquid scintillation alpha spectrometry offers an attractive with no sample self-absorption or geometry problems and with 100% counting efficiency. Sample preparation may include extraction of the alpha emitter of interest by a specific organic-phase-soluble compound directly into the liquid scintillation counting medium. Detection electronics use energy and pulse-shape discrimination to yield alpha spectra without beta and gamma background interference. Specific procedures have been developed for gross alpha, uranium, plutonium, thorium, and polonium assay. Possibilities for a large number of other applications exist. Accuracy and reproducibility are typically in the 1% range. Backgrounds on the order of 0.01 cpm are readily achievable. The paper will present an overview of liquid scintillation alpha counting techniques and some of the results achieved for specific applications

  11. Structures of G [alpha [superscript i1

    Energy Technology Data Exchange (ETDEWEB)

    Johnston, Christopher A.; Willard, Francis S.; Jezyk, Mark R.; Fredericks, Zoey; Bodor, Erik T.; Jones, Miller B.; Blaesius, Rainer; Watts, Val J.; Harden, T. Kendall; Sondek, John; Ramer, J. Kevin; Siderovski, David P. (Karo Bio); (UNC); (Purdue)

    2010-07-19

    Heterotrimeric G proteins are molecular switches that regulate numerous signaling pathways involved in cellular physiology. This characteristic is achieved by the adoption of two principal states: an inactive, GDP bound state and an active, GTP bound state. Under basal conditions, G proteins exist in the inactive, GDP bound state; thus, nucleotide exchange is crucial to the onset of signaling. Despite our understanding of G protein signaling pathways, the mechanism of nucleotide exchange remains elusive. We employed phage display technology to identify nucleotide state-dependent G{alpha} binding peptides. Herein, we report a GDP-selective G{alpha} binding peptide, KB-752, that enhances spontaneous nucleotide exchange of G{alpha}{sub i} subunits. Structural determination of the G{alpha}{sub i1}/peptide complex reveals unique changes in the G{alpha} switch regions predicted to enhance nucleotide exchange by creating a GDP dissociation route. Our results cast light onto a potential mechanism by which G{alpha} subunits adopt a conformation suitable for nucleotide exchange.

  12. Current status and perspectives in alpha radioimmunotherapy.

    Science.gov (United States)

    Chérel, M; Davodeau, F; Kraeber-Bodéré, F; Chatal, J F

    2006-12-01

    Systemic administration of radiolabeled antibody directed against tumor antigens in radioimmunotherapy (RIT) enables to specifically target the cancer cells and to destroy them. So far, this strategy has proven its efficiency in the treatment of some hematological cancers with antibodies labeled with beta emitting radionuclides. In the last 2 decades, availability of short half life alpha emitters prompted to consider their use in RIT. Contrary to beta particles, alpha particles have a short path length and display a high lineic energy transfer. Those physical characteristics open new fields of clinical applications complementary to beta-RIT. To date, alpha-RIT is still at a preclinical stage of development: the radiolabeling methods need to be optimized to ensure in vivo stability of the radiopharmaceuticals. Some radionuclides have complex decay schemes with daughters emitting further alpha particles whose toxicity needs to be investigated. The modalities of administration of radiolabeled antibodies in animal models require also to be improved for delivering higher doses to tumor targets. A comprehensive analysis of the specific events occurring at cell or tissue level in response to alpha irradiation would be of great interest in order to define the best therapeutic association for residual disease or consolidation treatments. This approach has been proven to be efficient in increasing antitumor response either by using high doses with organ protection (kidney, bone marrow) or by a synergistic effect between alpha-RIT and associated treatments, such as chemotherapy.

  13. Systemic Targeted Alpha Radiotherapy for Cancer

    Directory of Open Access Journals (Sweden)

    Allan B. J.

    2013-09-01

    Full Text Available Background: The fundamental principles of internal targeted alpha therapy for cancer were established many decades ago.The high linear energy transfer (LET of alpha radiation to the targeted cancer cellscauses double strand breaks in DNA. At the same time, the short range radiation spares adjacent normal tissues. This targeted approach complements conventional external beam radiotherapy and chemotherapy. Such therapies fail on several fronts, such as lack of control of some primary cancers (e.g.glioblastoma multiformeand to inhibit the development of lethal metastatic cancer after successful treatment of the primary cancer. Objective: This review charts the developing role of systemic high LET, internal radiation therapy. Method: Targeted alpha therapy is a rapidly advancing experimental therapy that holds promise to deliver high cytotoxicity to targeted cancer cells. Initially thought to be indicated for leukemia and micrometastases, there is now evidence that solid tumors can also be regressed. Results: Alpha therapy may be molecular or physiological in its targeting. Alpha emitting radioisotopes such as Bi-212, Bi-213, At-211 and Ac-225 are used to label monoclonal antibodies or proteins that target specifc cancer cells. Alternatively, Radium-233 is used for palliative therapy of breast and prostate cancers because of its bone seeking properties. Conclusion: Preclinical studies and clinical trials of alpha therapy are discussed for leukemia, lymphoma, melanoma, glioblastoma multiforme, bone metastases, ovarian cancer, pancreatic cancer and other cancers.

  14. Antitumor Effects In Vitro and In Vivo and Mechanisms of Protection against Melanoma B16F10-Nex2 Cells By Fastuosain, a Cysteine Proteinase from Bromelia fastuosa

    Directory of Open Access Journals (Sweden)

    Carla A. Guimarães-Ferreira

    2007-09-01

    Full Text Available In the present work, the antitumor effect of fastuosain, a cysteine proteinase from Bromelia fastuosa, was investigated. In the intravenous model of lung colonization in C57BI/6 mice, fastuosain and bromelain injected intraperitoneally were protective, very few nodules of B16F10-Nex2 melanoma cells were detected. Tumor cells treated with fastuosain showed reduced expression of CD44 and decreased invasion through Matrigel, lost their cytoplasmic extensions and substrate adherence, became round and detached, forming strongly bound cell clusters in suspension. Peritoneal cells recruited and activated by fastuosain treatment (mainly monocytic cells and lymphocytes migrated to the lung, where pulmonary melanoma metastases grew. Adoptive transference of peritoneal cells recruited by fastuosain had no protective effect against lung metastases in recipient mice. Treatment of green fluorescent protein -chimeric animals with fastuosain did not change the number of cells that migrated to the lung, compared to PBSinjected control mice, but the number of positive major histocompatibility complex class II cells increased with fastuosain treatment. Murine antibodies against fastuosain, bromelain, cathepsins B and L crossreacted in ELISA and recognized surface and cytoplasmic components expressed on B16F10-Nex2 cells. Anti-fastuosain antibodies were cytotoxic/lytic to B16F10-Nex2 cells. Antitumor effects of fastuosain involve mainly the direct effect of the enzyme and elicitation of protective antibodies.

  15. The role of secretory leukocyte proteinase inhibitor and elafin (elastase-specific inhibitor/skin-derived antileukoprotease as alarm antiproteinases in inflammatory lung disease

    Directory of Open Access Journals (Sweden)

    Sallenave Jean-Michel

    2000-08-01

    Full Text Available Abstract Secretory leukocyte proteinase inhibitor and elafin are two low-molecular-mass elastase inhibitors that are mainly synthesized locally at mucosal sites. It is thought that their physicochemical properties allow them to efficiently inhibit target enzymes, such as neutrophil elastase, released into the interstitium. Historically, in the lung, these inhibitors were first purified from secretions of patients with chronic obstructive pulmonary disease and cystic fibrosis. This suggested that they might be important in controlling excessive neutrophil elastase release in these pathologies. They are upregulated by 'alarm signals' such as bacterial lipopolysaccharides, and cytokines such as interleukin-1 and tumor necrosis factor and have been shown to be active against Gram-positive and Gram-negative bacteria, so that they have joined the growing list of antimicrobial 'defensin-like' peptides produced by the lung. Their site of synthesis and presumed functions make them very attractive candidates as potential therapeutic agents under conditions in which the excessive release of elastase by neutrophils might be detrimental. Because of its natural tropism for the lung, the use of adenovirus-mediated gene transfer is extremely promising in such applications.

  16. High-resolution X-ray study of the effects of deuteration on crystal growth and the crystal structure of proteinase K.

    Science.gov (United States)

    Chatake, Toshiyuki; Ishikawa, Takuya; Yanagisawa, Yasuhide; Yamada, Taro; Tanaka, Ichiro; Fujiwara, Satoru; Morimoro, Yukio

    2011-11-01

    Deuteration of macromolecules is an important technique in neutron protein crystallography. Solvent deuteration of protein crystals is carried out by replacing water (H(2)O) with heavy water (D(2)O) prior to neutron diffraction experiments in order to diminish background noise. The effects of solvent deuteration on the crystallization of proteinase K (PK) with polyethylene glycol as a precipitant were investigated using high-resolution X-ray crystallography. In previous studies, eight NO(3)(-) anions were included in the PK crystal unit cell grown in NaNO(3) solution. In this study, however, the PK crystal structure did not contain NO(3)(-) anions; consequently, distortions of amino acids arising from the presence of NO(3)(-) anions were avoided in the present crystal structures. High-resolution (1.1 Å) X-ray diffraction studies showed that the degradation of PK crystals induced by solvent deuteration was so small that this degradation would be negligible for the purpose of neutron protein crystallography experiments at medium resolution. Comparison of the nonhydrogen structures of nondeuterated and deuterated crystal structures demonstrated very small structural differences. Moreover, a positive correlation between the root-mean-squared differences and B factors indicated that no systematic difference existed. © 2011 International Union of Crystallography. All rights reserved.

  17. [Proteinase activity in Candida albicans strains isolated from the oral cavity of immunocompromised patients, with oral candidiasis and in healthy subjects].

    Science.gov (United States)

    Hernández-Solís, Sandra E; Rueda-Gordillo, Florencio; Rojas-Herrera, Rafael A

    2014-01-01

    Candida albicans has a variety of virulence factors, including secreted aspartyl proteases, which are determinant factors in the pathogenesis of this yeast in immunocompromised patients. Proteinase activity was identified in C. albicans strains isolated from the oral cavity of immunocompromised patients with cancer, diabetes and HIV+, with oral candidiasis and in healthy subjects. Two hundred and fifty C. albicans strains were analyzed, distributed in 5 different groups: patients with cancer, diabetes, HIV+, with oral candidiasis and healthy subjects. Proteolytic activity was identified in 46% of the strains from cancer patients, 54% from HIV+ patients, 60% from diabetics, 70% from oral candidiasis patients, and 42% from healthy subjects. Activity was higher in strains from immunocompromised and oral candidiasis patients than in healthy subjects. Differences were observed between the candidiasis-healthy, candidiasis-HIV+, and diabetic-healthy groups. No differences were observed between the oral candidiasis, diabetes and cancer patients, between the diabetes and HIV+ patients, or between the cancer patients, HIV+ patients and healthy subjects. The present results suggest that although secreted aspartyl proteases are important in the pathogenesis of C. albicans, their activity depends on host conditions. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  18. Anti-proteinase 3 antibodies in diffuse systemic sclerosis (SSc with normotensive renal impairment: is it suggestive for an overlapping between SSc and idiopathic vasculitis?

    Directory of Open Access Journals (Sweden)

    V. Campanella

    2011-09-01

    Full Text Available Objective. To test the prevalence of anti-neutrophil cytoplasmic antibodies (ANCA in systemic sclerosis (SSc and to verify a possible association of ANCA with normotensive renal involvement in SSc. Patients and methods: 51 patients affected by SSc, 35 with diffuse scleroderma (dSSc and 16 with limited scleroderma (lSSc, were tested for ANCA by indirect immunofluorescence (IIF on human ethanol and formalin-acetone-fixed granulocytes (before and after DNase treatment, by conventional enzyme linked immuno-sorbent assay (ELISA and by capture-ELISA. Results. Six out of 51 selected SSc patients had ANCA by IIF (11.7% and five presented a perinuclear/nuclear atypical ANCA pattern. In all cases we only found anti-proteinase3 (aPR3 antibodies. All ANCA positive patients had diffuse form of SSc (17.1%, all were anti-Scl70 positive (aScl70, five patients had proteinuria, three had microscopic haematuria. All ANCA positive patients were normotensive with normal renin plasma levels, the mean erythrocyte sedimentation rate (ESR was higher in this group compared to the other SSc patients. Conclusions. Our study shows that aPR3 is not rare in dSSc. According to the clinical and serological findings and to the recent literature, we can hypothesise that when ANCA are found in SSc, an overlapping of scleroderma with systemic necrotizing vasculitis should be suspected.

  19. Proteinase-activated receptor 2 and disease biomarkers in cerebrospinal fluid in cases with autopsy-confirmed prion diseases and other neurodegenerative diseases.

    Science.gov (United States)

    Rohan, Zdenek; Smetakova, Magdalena; Kukal, Jaromir; Rusina, Robert; Matej, Radoslav

    2015-03-31

    Proteinase-activated receptor 2 (PAR-2) has been shown to promote both neurotoxic and neuroprotective effects. Similarly, other routinely used nonspecific markers of neuronal damage can be found in cerebrospinal fluid (CSF) and can be used as biomarkers for different neurodegenerative disorders. Using enzyme-linked immunosorbent assays and western blotting we assessed PAR-2, total-tau, phospho-tau, beta-amyloid levels, and protein 14-3-3 in the CSF of former patients who had undergone a neuropathological autopsy after death and who had been definitively diagnosed with a prion or other neurodegenerative disease. We did not find any significant correlation between levels of PAR-2 and other biomarkers, nor did we find any differences in PAR-2 levels between prion diseases and other neurodegenerative conditions. However, we confirmed that very high total-tau levels were significantly associated with definitive prion diagnoses and exhibited greater sensitivity and specificity than protein 14-3-3, which is routinely used as a marker. Our study showed that PAR-2, in CSF, was not specifically altered in prion diseases compared to other neurodegenerative conditions. Our results also confirmed that very high total-tau protein CSF levels were significantly associated with a definitive Creutzfeldt-Jakob disease (CJD) diagnosis and should be routinely tested as a diagnostic marker. Observed individual variability in CSF biomarkers provide invaluable feedback from neuropathological examinations even in "clinically certain" cases.

  20. Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis.

    Science.gov (United States)

    Shahbazi, Mehdi; Zahedifard, Farnaz; Taheri, Tahereh; Taslimi, Yasaman; Jamshidi, Shahram; Shirian, Sadegh; Mahdavi, Niousha; Hassankhani, Mehdi; Daneshbod, Yahya; Zarkesh-Esfahani, Sayyed Hamid; Papadopoulou, Barbara; Rafati, Sima

    2015-01-01

    Canine Visceral Leishmaniasis (CVL) is a major veterinary and public health problem caused by Leishmania infantum (L. infantum) in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE) and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL.

  1. Purification and characterization of a thermostable keratinolytic serine alkaline proteinase from Streptomyces sp. strain AB1 with high stability in organic solvents.

    Science.gov (United States)

    Jaouadi, Bassem; Abdelmalek, Badis; Fodil, Djamila; Ferradji, Fatma Zohra; Rekik, Hatem; Zaraî, Nedia; Bejar, Samir

    2010-11-01

    A keratinolytic alkaline proteinase (KERAB) was isolated from Streptomyces sp. strain AB1. Based on MALDI-TOF mass spectrometry analysis, the purified enzyme is a monomer with a molecular mass of 29850.17Da. The NH(2)-terminal sequence of the enzyme was determined to be TQANPPSWGLDDIDQTAL. This keratinase was completely inhibited by phenylmethanesulfonyl fluoride (PMSF) and diiodopropyl fluorophosphates (DIFP), which suggests that it belongs to the serine protease family. Using keratin azure as a substrate, the optimum pH and temperature values for keratinase activity were pH 11.5 and 75 degrees C, respectively. This keratinase was stable between 30 and 60 degrees C and pH 4 and 11 for 4 and 96 h, respectively, and thermoactivity and thermostability were enhanced in the presence of 5 mM Mg(2+). Its catalytic efficiency was higher than those of SAPB-L31I/T33S/N99Y, nattokinase and subtilisin Carlsberg. KERAB exhibited stability to detergents and high resistance against organic solvents and was able to degrade feathers completely. These properties make KERAB a potential candidate for future applications in detergent formulations, dehairing during leather processing, and non-aqueous peptide biocatalysis. Copyright 2010 Elsevier Ltd. All rights reserved.

  2. Cross-talk between toll-like receptor 4 (TLR4) and proteinase-activated receptor 2 (PAR(2) ) is involved in vascular function.

    Science.gov (United States)

    Bucci, M; Vellecco, V; Harrington, L; Brancaleone, V; Roviezzo, F; Mattace Raso, G; Ianaro, A; Lungarella, G; De Palma, R; Meli, R; Cirino, G

    2013-01-01

    Proteinase-activated receptors (PARs) and toll-like receptors (TLRs) are involved in innate immune responses. The aim of this study was to evaluate the possible cross-talk between PAR(2) and TLR4 in vessels in physiological condition and how it varies following stimulation of TLR4 by using in vivo and ex vivo models. Thoracic aortas were harvested from both naïve and endotoxaemic rats for in vitro studies. Arterial blood pressure was monitored in anaesthetized rats in vivo. LPS was used as a TLR4 agonist while PAR(2) activating peptide (AP) was used as a PAR(2) agonist. Aortas harvested from TLR4(-/-) mice were also used to characterize the PAR(2) response. PAR(2) , but not TLR4, expression was enhanced in aortas of endotoxaemic rats. PAR(2) AP-induced vasorelaxation was increased in aortic rings of LPS-treated rats. TLR4 inhibitors, curcumine and resveratrol, reduced PAR(2) AP-induced vasorelaxation and PAR(2) AP-induced hypotension in both naïve and endotoxaemic rats. Finally, in aortic rings from TLR4(-/-) mice, the expression of PAR(2) was reduced and the PAR(2) AP-induced vasodilatation impaired compared with those from wild-type mice and both resveratrol and curcumine were ineffective. Cross-talk between PAR(2) and TLR4 contributes to vascular homeostasis. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

  3. Cross-talk between toll-like receptor 4 (TLR4) and proteinase-activated receptor 2 (PAR2) is involved in vascular function

    Science.gov (United States)

    Bucci, M; Vellecco, V; Harrington, L; Brancaleone, V; Roviezzo, F; Mattace Raso, G; Ianaro, A; Lungarella, G; De Palma, R; Meli, R; Cirino, G

    2013-01-01

    Background and Purpose Proteinase-activated receptors (PARs) and toll-like receptors (TLRs) are involved in innate immune responses. The aim of this study was to evaluate the possible cross-talk between PAR2 and TLR4 in vessels in physiological condition and how it varies following stimulation of TLR4 by using in vivo and ex vivo models. Experimental Approach Thoracic aortas were harvested from both naïve and endotoxaemic rats for in vitro studies. Arterial blood pressure was monitored in anaesthetized rats in vivo. LPS was used as a TLR4 agonist while PAR2 activating peptide (AP) was used as a PAR2 agonist. Aortas harvested from TLR4–/– mice were also used to characterize the PAR2 response. Key Results PAR2, but not TLR4, expression was enhanced in aortas of endotoxaemic rats. PAR2AP-induced vasorelaxation was increased in aortic rings of LPS-treated rats. TLR4 inhibitors, curcumine and resveratrol, reduced PAR2AP-induced vasorelaxation and PAR2AP-induced hypotension in both naïve and endotoxaemic rats. Finally, in aortic rings from TLR4–/– mice, the expression of PAR2 was reduced and the PAR2AP-induced vasodilatation impaired compared with those from wild-type mice and both resveratrol and curcumine were ineffective. Conclusions and Implications Cross-talk between PAR2 and TLR4 contributes to vascular homeostasis. PMID:22957757

  4. Cytokine vaccination: neutralising IL-1alpha autoantibodies induced by immunisation with homologous IL-1alpha

    DEFF Research Database (Denmark)

    Svenson, M; Hansen, M B; Thomsen, Allan Randrup

    2000-01-01

    High-affinity IgG autoantibodies (aAb) to IL-1alpha are among the most frequently found aAb to cytokines in humans. To establish an animal model with aAb to IL-1alpha, we immunised mice with recombinant murine IL-1alpha. Unprimed and Bacille Calmette-Guérin (BCG)-primed BALB/cA mice were vaccinated...... in mice by vaccination with recombinant murine IL-1alpha conjugated to PPD. Studies of the effects of IL-1alpha aAb in such animals may help clarify the importance of naturally occurring IL-1alpha aAb in humans and permit the evaluation of future therapies with cytokine aAb in patients...... with IL-1alpha coupled to purified protein derivative of tuberculin (PPD). Both unprimed and primed animals developed IgG aAb to IL-1alpha. These aAb persisted at high levels more than 100 days after vaccination and did not cross-react with murine IL-1beta. The induced anti-IL-1alpha aAb inhibited binding...

  5. Alpha 1-blockers vs 5 alpha-reductase inhibitors in benign prostatic hyperplasia. A comparative review

    DEFF Research Database (Denmark)

    Andersen, J T

    1995-01-01

    During recent years, pharmacological treatment of symptomatic benign prostatic hyperplasia (BPH) has become the primary treatment choice for an increasing number of patients. The 2 principal drug classes employed are alpha 1-blockers and 5 alpha-reductase inhibitors. Current information from...... of patients who will respond well to alpha 1-blockers have yet to be identified, and data concerning the long term effects of these drugs are not yet available. 5 alpha-Reductase inhibitors have a slow onset of effect, but treatment leads to improvement in symptoms, reduction of the size of the prostate gland...... or unwilling to undergo surgical resection of the prostate will benefit from such therapy....

  6. Selectivity of the imidazoline alpha-adrenoceptor agonists (oxymetazoline and cirazoline) for human cloned alpha 1-adrenoceptor subtypes.

    Science.gov (United States)

    Horie, K.; Obika, K.; Foglar, R.; Tsujimoto, G.

    1995-01-01

    1. To investigate the structure-activity relationships of alpha-adrenoceptor agonists for the alpha 1-adrenoceptor subtypes, we have compared the imidazoline class of compounds, oxymetazoline and cirazoline, with the phenethylamine, noradrenaline, in their affinities and also in their intrinsic activities in Chinese hamster ovary (CHO) cells stably expressing the cloned human alpha 1-adrenoceptor subtypes (alpha 1a-, alpha 1b-, and alpha 1d-subtypes). 2. Radioligand binding studies with [125I]-HEAT showed that cirazoline and oxymetazoline had higher affinities at alpha 1a-subtype than at alpha 1b- and alpha 1d-subtypes, while noradrenaline had higher affinity at the alpha 1d-subtype than at alpha 1a- and alpha 1b-subtypes. 3. In functional studies, cirazoline caused transients of cytosolic Ca2+ concentrations ([Ca2+]i response) in a concentration-dependent manner and developed a maximal response similar to that to noradrenaline in CHO cells expressing the alpha 1a-subtype, while it acted as a partial agonist at alpha 1b- and alpha 1d-adrenoceptors. Oxymetazoline, on the other hand, was a weak agonist at alpha 1a-adrenoceptors, and has no intrinsic activity at the other subtypes. 4. Using the phenoxybenzamine inactivation method, the relationships between receptor occupancy and noradrenaline-induced [Ca2+]i response for alpha 1a- and alpha 1d-subtypes were found to be linear, whereas it was moderately hyperbolic for the alpha 1b-subtype, indicating the absence of receptor reserves in CHO cells expressing alpha 1a- and alpha 1d-subtypes while there exists a small receptor reserve for CHO cells expressing the alpha 1b-subtype.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8564227

  7. Anti-IL-1alpha autoantibodies in early rheumatoid arthritis

    DEFF Research Database (Denmark)

    Forslind, K; Svensson, Birte; Svenson, M

    2001-01-01

    To investigate the potential predictive value of autoantibodies against IL1-alpha (anti-IL-1alpha) in patients with early rheumatoid arthritis (RA).......To investigate the potential predictive value of autoantibodies against IL1-alpha (anti-IL-1alpha) in patients with early rheumatoid arthritis (RA)....

  8. The nitric oxide-donating pravastatin derivative, NCX 6550 [(1S-[1alpha(betaS*, deltaS*), 2alpha, 6alpha, 8beta-(R*), 8a alpha

    Science.gov (United States)

    Dever, G; Spickett, C M; Kennedy, S; Rush, C; Tennant, G; Monopoli, A; Wainwright, C L

    2007-01-01

    Statins possess anti-inflammatory effects that may contribute to their ability to slow atherogenesis, whereas nitric oxide (NO) also influences inflammatory cell adhesion. This study aimed to determine whether a novel NO-donating pravastatin derivative, NCX 6550 [(1S-[1alpha(betaS*,deltaS*),2alpha,6alpha,8beta-(R*),8a alpha

  9. Measurement of radium micro-precipitates using alpha spectrometry and total alpha counting methods

    International Nuclear Information System (INIS)

    Hosseini, T.; Fathivand, A.A.

    2004-01-01

    Background: This study consists of two parts. The first part deals with both qualitative and quantitative analysis of 226 Ra suing alpha spectrometry measurement method. In the second part, the percentage of radioactive equilibrium between 226 Radium and its daughter products were determined by alpha spectrometry and total alpha measurement system after elapsed time of 15 days from precipitation. Materials and methods: Twelve 226 Radium samples as Barium-Radium Sulfate in form of micro-precipitates on millipore and What man 42 filters were prepared. An alpha spectrometer with surface barrier detector and a total alpha measurement system consists of scintillation crystal assembly Zinc Silver were used for counting. Results: The minimum detection limit of alpha spectrometry and total alpha counting for 226 Radium measurements in samples for counting time equal to 10000 seconds, were found to be 3.7 mBq and 15.8 mBq respectively. Results from total alpha counting showed that radioactive equilibrium between 226 Radium and its daughter products reached to about 92%± 3.5, where as, in the case of alpha spectrometry radioactive equilibrium, it was destroyed due to vacuum during counting the sample. Also in case of alpha spectrometry, the optimum sample to detector distance, was found to be 0.5 centimeter. Conclusion: From this study it was concluded that micro-precipitation can be used as a proper method for sample preparation and alpha spectrometry due to its lower detection limit to measure low concentration of 224 Radium and 226 Radium in these precipitates, prepared from different samples. Besides it is not time consuming and sources can be measured immediately after sample preparation

  10. Alpha-particle fluence in radiobiological experiments.

    Science.gov (United States)

    Nikezic, Dragoslav; Yu, Kwan Ngok

    2017-03-01

    Two methods were proposed for determining alpha-particle fluence for radiobiological experiments. The first involved calculating the probabilities of hitting the target for alpha particles emitted from a source through Monte Carlo simulations, which when multiplied by the activity of the source gave the fluence at the target. The second relied on the number of chemically etched alpha-particle tracks developed on a solid-state nuclear track detector (SSNTD) that was irradiated by an alpha-particle source. The etching efficiencies (defined as percentages of latent tracks created by alpha particles from the source that could develop to become visible tracks upon chemical etching) were computed through Monte Carlo simulations, which when multiplied by the experimentally counted number of visible tracks would also give the fluence at the target. We studied alpha particles with an energy of 5.486 MeV emitted from an 241Am source, and considered the alpha-particle tracks developed on polyallyldiglycol carbonate film, which is a common SSNTD. Our results showed that the etching efficiencies were equal to one for source-film distances of from 0.6 to 3.5 cm for a circular film of radius of 1 cm, and for source-film distances of from 1 to 3 cm for circular film of radius of 2 cm. For circular film with a radius of 3 cm, the etching efficiencies never reached 1. On the other hand, the hit probability decreased monotonically with increase in the source-target distance, and fell to zero when the source-target distance was larger than the particle range in air. © The Author 2016. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

  11. Cancer radioimmunotherapy with alpha-emitting nuclides

    Energy Technology Data Exchange (ETDEWEB)

    Couturier, Olivier [INSERM U 601, Nantes (France); Place Alexis Ricordeau, Department of Nuclear Medicine, Nantes, Cedex (France); Supiot, Stephane; Degraef-Mougin, Marie; Faivre-Chauvet, Alain; Carlier, Thomas; Chatal, Jean-Francois; Davodeau, Francois; Cherel, Michel [INSERM U 601, Nantes (France)

    2005-04-01

    In lymphoid malignancies and in certain solid cancers such as medullary thyroid carcinoma, somewhat mixed success has been achieved when applying radioimmunotherapy (RIT) with {beta}-emitters for the treatment of refractory cases. The development of novel RIT with {alpha}-emitters has created new opportunities and theoretical advantages due to the high linear energy transfer (LET) and the short path length in biological tissue of {alpha}-particles. These physical properties offer the prospect of achieving selective tumoural cell killing. Thus, RIT with {alpha}-emitters appears particularly suited for the elimination of circulating single cells or cell clusters or for the treatment of micrometastases at an early stage. However, to avoid non-specific irradiation of healthy tissues, it is necessary to identify accessible tumoural targets easily and rapidly. For this purpose, a small number of {alpha}-emitters have been investigated, among which only a few have been used for in vivo preclinical studies. Another problem is the availability and cost of these radionuclides; for instance, the low cost and the development of a reliable actinium-225/bismuth-213 generator were probably determining elements in the choice of bismuth-213 in the only human trial of RIT with an {alpha}-emitter. This article reviews the literature concerning monoclonal antibodies radiolabelled with {alpha}-emitters that have been developed for possible RIT in cancer patients. The principal radio-immunoconjugates are considered, starting with physical and chemical properties of {alpha}-emitters, their mode of production, the possibilities and difficulties of labelling, in vitro studies and finally, when available, in vivo preclinical and clinical studies. (orig.)

  12. Cancer radioimmunotherapy with alpha-emitting nuclides.

    Science.gov (United States)

    Couturier, Olivier; Supiot, Stéphane; Degraef-Mougin, Marie; Faivre-Chauvet, Alain; Carlier, Thomas; Chatal, Jean-François; Davodeau, François; Cherel, Michel

    2005-05-01

    In lymphoid malignancies and in certain solid cancers such as medullary thyroid carcinoma, somewhat mixed success has been achieved when applying radioimmunotherapy (RIT) with beta-emitters for the treatment of refractory cases. The development of novel RIT with alpha-emitters has created new opportunities and theoretical advantages due to the high linear energy transfer (LET) and the short path length in biological tissue of alpha-particles. These physical properties offer the prospect of achieving selective tumoural cell killing. Thus, RIT with alpha-emitters appears particularly suited for the elimination of circulating single cells or cell clusters or for the treatment of micrometastases at an early stage. However, to avoid non-specific irradiation of healthy tissues, it is necessary to identify accessible tumoural targets easily and rapidly. For this purpose, a small number of alpha-emitters have been investigated, among which only a few have been used for in vivo preclinical studies. Another problem is the availability and cost of these radionuclides; for instance, the low cost and the development of a reliable actinium-225/bismuth-213 generator were probably determining elements in the choice of bismuth-213 in the only human trial of RIT with an alpha-emitter. This article reviews the literature concerning monoclonal antibodies radiolabelled with alpha-emitters that have been developed for possible RIT in cancer patients. The principal radio-immunoconjugates are considered, starting with physical and chemical properties of alpha-emitters, their mode of production, the possibilities and difficulties of labelling, in vitro studies and finally, when available, in vivo preclinical and clinical studies.

  13. Pharmacologic specificity of alpha-2 adrenergic receptor subtypes

    Energy Technology Data Exchange (ETDEWEB)

    Petrash, A.; Bylund, D.

    1986-03-01

    The authors have defined alpha-2 adrenergic receptor subtypes in human and rat tissues using prazosin as a subtype selective drug. Prazosin has a lower affinity (250 nM) at alpha-2A receptor and a higher affinity (5 nM) at alpha-2B receptors. In order to determine if other adrenergic drugs are selective for one or the other subtypes, the authors performed (/sup 3/H)yohimbine inhibition experiments with various adrenergic drugs in tissues containing alpha-2A, alpha-2B or both subtypes. Oxymetazoline, WB4101 and yohimbine were found to be 80-, 20- and 10-fold more potent at alpha-2A receptors than at alpha-2B receptors. Phentolamine, adazoxan, (+)- and (-)-mianserin, clonidine, (+)-butaclamol, (-)- and (+)-norepinephrine, epinephrine, dopamine and thioridazine were found to have equal affinities for the two subtypes. These results further validate the subdivision of alpha-2 adrenergic receptors into alpha-2A and alpha-2B subtypes.

  14. Alpha 1 B- but not alpha 1 A-adrenoceptors mediate inositol phosphate generation

    NARCIS (Netherlands)

    Michel, M. C.; Hanft, G.; Gross, G.

    1990-01-01

    We used novel highly subtype-selective antagonists to study whether alpha 1A- and/or alpha 1B-adrenoceptors mediate the stimulation of inositol phosphate generation by noradrenaline in rat cerebral cortex. Phentolamine (10 microM) and prazosin (100 nM) completely abolished the stimulated inositol

  15. Role of macrophage inflammatory protein-1 alpha (MIP-1 alpha) in acute lung injury in rats

    DEFF Research Database (Denmark)

    Shanley, T P; Schmal, H; Friedl, H P

    1995-01-01

    The role of macrophage inflammatory protein-1 alpha (MIP-1 alpha) in the pathogenesis of acute lung injury in rats after intrapulmonary deposition of IgG immune complexes or intratracheal administration of LPS has been assessed. Critical to these studies was the cloning and functional expression...

  16. Alpha-in-air monitor for continuous monitoring based on alpha to beta ratio

    International Nuclear Information System (INIS)

    Somayaji, K.S.; Venkataramani, R.; Swaminathan, N.; Pushparaja

    1997-01-01

    Measurement of long-lived alpha activity collected on a filter paper in continuous air monitoring of ambient working environment is difficult due to interference from much larger concentrations of short-lived alpha emitting daughter products of 222 Rn and 220 Rn. However, the ratio between the natural alpha and beta activity is approximately constant and this constancy of the ratio is used to discriminate against short-lived natural radioactivity in continuous air monitoring. Detection system was specially designed for the purpose of simultaneous counting of alpha and beta activity deposited on the filter paper during continuous monitoring. The activity ratios were calculated and plotted against the monitoring duration up to about six hours. Monitoring was carried out in three facilities with different ventilation conditions. Presence of any long-lived alpha contamination on the filter paper results in increase in the alpha to beta ratio. Long-lived 239 Pu contamination of about 16 DAC.h could be detected after about 45 minutes of commencement of the sampling. The experimental results using prototype units have shown that the approach of using alpha to beta activity ratio method to detect long-lived alpha activity in the presence of short-lived natural activity is satisfactory. (author)

  17. PLE CATALYZED HYDROLYZES OF ALPHA-SUBSTITUTED ALPHA-HYDROXY ESTERS - THE INFLUENCE OF THE SUBSTITUENTS

    NARCIS (Netherlands)

    MOORLAG, H; KELLOGG, RM

    1991-01-01

    The enzymatic hydrolyses of a variety of alpha-substituted mandelic and lactic esters using pig liver esterase (PLE) have been investigated. High to moderate enantioselectivity was found for various alpha-substituted mandelic esters, whereas PLE showed low to no enantioselectivity for

  18. Increased voluntary exercise in mice deficient for tumour necrosis factor-alpha and lymphotoxin-alpha.

    NARCIS (Netherlands)

    Netea, M.G.; Kullberg, B.J.; Vonk, A.G.; Verschueren, I.; Joosten, L.A.B.; Meer, J.W.M. van der

    2007-01-01

    BACKGROUND: The endogenous mediators playing a role in the sensing of fatigue and cessation of exercise are yet to be characterized. We hypothesized that proinflammatory cytokines, in particular tumour necrosis factor-alpha (TNFalpha) and lymphotoxin-alpha (LT) transmit signals leading to fatigue.

  19. Alpha-particle elastic scattering on [sup 16]O in the four [alpha]-particle model

    Energy Technology Data Exchange (ETDEWEB)

    Li Qingrun (CCAST (World Lab.), Beijing (China) Inst. of High Energy Physics, Academia Sinica, Beijing (China)); Yang Yongxu (Dept. of Physics, Guangxi Normal Univ., Guilin (China))

    1993-08-23

    A folding potential describing the alpha-particle scattering on [sup 16]O is constructed based on the four [alpha]-particle model of the nucleus [sup 16]O. This folding potential provides a good description of the experimental data covering a broad energy range. (orig.)

  20. Consistent Measurements of $\\alpha_{s}$ from Precise Oriented Event Shape Distributions

    CERN Document Server

    Abreu, P.; Adye, T.; Adzic, P.; Albrecht, Z.; Alderweireld, T.; Alekseev, G.D.; Alemany, R.; Allmendinger, T.; Allport, P.P.; Almehed, S.; Amaldi, U.; Amapane, N.; Amato, S.; Anassontzis, E.G.; Andersson, P.; Andreazza, A.; Andringa, S.; Antilogus, P.; Apel, W.D.; Arnoud, Y.; Asman, B.; Augustin, J.E.; Augustinus, A.; Baillon, P.; Bambade, P.; Barao, F.; Barbiellini, G.; Barbier, R.; Bardin, D.Yu.; Barker, G.J.; Baroncelli, A.; Battaglia, M.; Baubillier, M.; Becks, K.H.; Begalli, M.; Behrmann, A.; Beilliere, P.; Belokopytov, Yu.; Benekos, N.C.; Benvenuti, A.C.; Berat, C.; Berggren, M.; Bertini, D.; Bertrand, D.; Besancon, M.; Bigi, M.; Bilenky, Mikhail S.; Bizouard, M.A.; Bloch, D.; Blom, H.M.; Bonesini, M.; Bonivento, W.; Boonekamp, M.; Booth, P.S.L.; Borgland, A.W.; Borisov, G.; Bosio, C.; Botner, O.; Boudinov, E.; Bouquet, B.; Bourdarios, C.; Bowcock, T.J.V.; Boyko, I.; Bozovic, I.; Bozzo, M.; Branchini, P.; Brenke, T.; Brenner, R.A.; Bruckman, P.; Brunet, J.M.; Bugge, L.; Buran, T.; Burgsmuller, T.; Buschbeck, B.; Buschmann, P.; Cabrera, S.; Caccia, M.; Calvi, M.; Camporesi, T.; Canale, V.; Carena, F.; Carroll, L.; Caso, C.; Castillo Gimenez, M.V.; Cattai, A.; Cavallo, F.R.; Chabaud, V.; Charpentier, P.; Chaussard, L.; Checchia, P.; Chelkov, G.A.; Chierici, R.; Chochula, P.; Chorowicz, V.; Chudoba, J.; Cieslik, K.; Collins, P.; Contri, R.; Cortina, E.; Cosme, G.; Cossutti, F.; Cowell, J.H.; Crawley, H.B.; Crennell, D.; Crepe-Renaudin, Sabine; Crosetti, G.; Cuevas Maestro, J.; Czellar, S.; Davenport, M.; Da Silva, W.; Deghorain, A.; Della Ricca, G.; Delpierre, P.; Demaria, N.; De Angelis, A.; De Boer, W.; De Clercq, C.; De Lotto, B.; De Min, A.; De Paula, L.; Dijkstra, H.; Di Ciaccio, L.; Dolbeau, J.; Doroba, K.; Dracos, M.; Drees, J.; Dris, M.; Duperrin, A.; Durand, J.D.; Eigen, G.; Ekelof, T.; Ekspong, G.; Ellert, M.; Elsing, M.; Engel, J.P.; Erzen, B.; Espirito Santo, M.C.; Fanourakis, G.; Fassouliotis, D.; Fayot, J.; Feindt, M.; Ferrari, P.; Ferrer, A.; Ferrer-Ribas, E.; Ferro, F.; Fichet, S.; Firestone, A.; Flagmeyer, U.; Foeth, H.; Fokitis, E.; Fontanelli, F.; Franek, B.; Frodesen, A.G.; Fruhwirth, R.; Fulda-Quenzer, F.; Fuster, J.; Galloni, A.; Gamba, D.; Gamblin, S.; Gandelman, M.; Garcia, C.; Gaspar, C.; Gaspar, M.; Gasparini, U.; Gavillet, P.; Gazis, Evangelos; Gele, D.; Ghodbane, N.; Gil Botella, Ines; Glege, F.; Gokieli, R.; Golob, B.; Gomez-Ceballos, G.; Goncalves, P.; Gonzalez Caballero, I.; Gopal, G.; Gorn, L.; Gracco, V.; Grahl, J.; Graziani, E.; Green, C.; Grimm, H.J.; Gris, P.; Grosdidier, G.; Grzelak, K.; Gunther, M.; Guy, J.; Hahn, F.; Hahn, S.; Haider, S.; Hallgren, A.; Hamacher, K.; Hansen, J.; Harris, F.J.; Hedberg, V.; Heising, S.; Hernandez, J.J.; Herquet, P.; Herr, H.; Hessing, T.L.; Heuser, J.M.; Higon, E.; Holmgren, S.O.; Holt, P.J.; Hoorelbeke, S.; Houlden, M.; Hrubec, J.; Huet, K.; Hughes, G.J.; Hultqvist, K.; Jackson, John Neil; Jacobsson, R.; Jalocha, P.; Janik, R.; Jarlskog, C.; Jarlskog, G.; Jarry, P.; Jean-Marie, B.; Johansson, Erik Karl; Jonsson, P.; Joram, C.; Juillot, P.; Kapusta, Frederic; Karafasoulis, K.; Katsanevas, S.; Katsoufis, E.C.; Keranen, R.; Kersevan, B.P.; Khomenko, B.A.; Khovansky, N.N.; Kiiskinen, A.; King, B.; Kinvig, A.; Kjaer, N.J.; Klapp, O.; Klein, Hansjorg; Kluit, P.; Kokkinias, P.; Koratzinos, M.; Kourkoumelis, C.; Kuznetsov, O.; Krammer, M.; Kriznic, E.; Krumshtein, Z.; Kubinec, P.; Kurowska, J.; Kurvinen, K.; Lamsa, J.W.; Lane, D.W.; Langefeld, P.; Laugier, J.P.; Lauhakangas, R.; Leder, G.; Ledroit, Fabienne; Lefebure, V.; Leinonen, L.; Leisos, A.; Leitner, R.; Lemonne, J.; Lenzen, G.; Lepeltier, V.; Lesiak, T.; Lethuillier, M.; Libby, J.; Liko, D.; Lipniacka, A.; Lippi, I.; Lorstad, B.; Loken, J.G.; Lopes, J.H.; Lopez, J.M.; Lopez-Fernandez, R.; Loukas, D.; Lutz, P.; Lyons, L.; MacNaughton, J.; Mahon, J.R.; Maio, A.; Malek, A.; Malmgren, T.G.M.; Maltezos, S.; Malychev, V.; Mandl, F.; Marco, J.; Marco, R.; Marechal, B.; Margoni, M.; Marin, J.C.; Mariotti, C.; Markou, A.; Martinez-Rivero, C.; Martinez-Vidal, F.; Marti i Garcia, S.; Masik, J.; Mastroyiannopoulos, N.; Matorras, F.; Matteuzzi, C.; Matthiae, G.; Mazzucato, F.; Mazzucato, M.; McCubbin, M.; McKay, R.; McNulty, R.; McPherson, G.; Meroni, C.; Meyer, W.T.; Migliore, E.; Mirabito, L.; Mitaroff, W.A.; Mjornmark, U.; Moa, T.; Moch, M.; Moller, Rasmus; Monig, Klaus; Monge, M.R.; Moreau, X.; Morettini, P.; Morton, G.; Muller, U.; Munich, K.; Mulders, M.; Mulet-Marquis, C.; Muresan, R.; Murray, W.J.; Muryn, B.; Myatt, G.; Myklebust, T.; Naraghi, F.; Nassiakou, M.; Navarria, F.L.; Navas, Sergio; Nawrocki, K.; Negri, P.; Nemecek, S.; Neufeld, N.; Nicolaidou, R.; Nielsen, B.S.; Niezurawski, P.; Nikolenko, M.; Nomokonov, V.; Normand, A.; Nygren, A.; Olshevsky, A.G.; Onofre, A.; Orava, R.; Orazi, G.; Osterberg, K.; Ouraou, A.; Paganoni, M.; Paiano, S.; Pain, R.; Paiva, R.; Palacios, J.; Palka, H.; Papadopoulou, T.D.; Papageorgiou, K.; Pape, L.; Parkes, C.; Parodi, F.; Parzefall, U.; Passeri, A.; Passon, O.; Pegoraro, M.; Peralta, L.; Pernicka, M.; Perrotta, A.; Petridou, C.; Petrolini, A.; Phillips, H.T.; Pierre, F.; Pimenta, M.; Piotto, E.; Podobnik, T.; Pol, M.E.; Polok, G.; Poropat, P.; Pozdnyakov, V.; Privitera, P.; Pukhaeva, N.; Pullia, A.; Radojicic, D.; Ragazzi, S.; Rahmani, H.; Ratoff, P.N.; Read, Alexander L.; Rebecchi, P.; Redaelli, Nicola Giuseppe; Regler, M.; Reid, D.; Reinhardt, R.; Renton, P.B.; Resvanis, L.K.; Richard, F.; Ridky, J.; Rinaudo, G.; Rodrigo, German; Rohne, O.; Romero, A.; Ronchese, P.; Rosenberg, E.I.; Rosinsky, P.; Roudeau, P.; Rovelli, T.; Royon, C.; Ruhlmann-Kleider, V.; Ruiz, A.; Saarikko, H.; Sacquin, Y.; Sadovsky, A.; Sajot, G.; Salt, J.; Sampsonidis, D.; Sannino, M.; Schneider, H.; Schwemling, P.; Schwering, B.; Schwickerath, U.; Schyns, M.A.E.; Scuri, Fabrizio; Seager, P.; Sedykh, Yu.; Segar, A.M.; Sekulin, R.; Shellard, R.C.; Sheridan, A.; Siebel, M.; Simard, L.; Simonetto, F.; Sisakian, A.N.; Smadja, G.; Smirnova, O.; Smith, G.R.; Sopczak, A.; Sosnowski, R.; Spassoff, T.; Spiriti, E.; Sponholz, P.; Squarcia, S.; Stanescu, C.; Stanic, S.; Stevenson, K.; Stocchi, A.; Strauss, J.; Strub, R.; Stugu, B.; Szczekowski, M.; Szeptycka, M.; Tabarelli, T.; Tegenfeldt, F.; Terranova, F.; Thomas, J.; Timmermans, Jan; Tinti, N.; Tkachev, L.G.; Todorova, S.; Tomaradze, A.; Tome, B.; Tonazzo, A.; Tortora, L.; Transtromer, G.; Treille, D.; Tristram, G.; Trochimczuk, M.; Troncon, C.; Tsirou, A.; Turluer, M.L.; Tyapkin, I.A.; Tzamarias, S.; Ullaland, O.; Valenti, G.; Vallazza, E.; Vander Velde, C.; Van Apeldoorn, G.W.; Van Dam, Piet; Van Doninck, Walter; Van Eldik, J.; Van Lysebetten, A.; Van Remortel, N.; Van Vulpen, I.; Vassilopoulos, N.; Vegni, G.; Ventura, L.; Venus, W.; Verbeure, F.; Verlato, M.; Vertogradov, L.S.; Verzi, V.; Vilanova, D.; Vitale, L.; Vodopianov, A.S.; Vollmer, C.; Voulgaris, G.; Vrba, V.; Wahlen, H.; Walck, C.; Weiser, C.; Wicke, D.; Wickens, J.H.; Wilkinson, G.R.; Winter, M.; Witek, M.; Wolf, G.; Yi, J.; Zalewska, A.; Zalewski, P.; Zavrtanik, D.; Zevgolatakos, E.; Zimine, N.I.; Zucchelli, G.C.; Zumerle, G.

    2000-01-01

    An updated analysis using about 1.5 million events recorded at $\\sqrt{s} =M_Z$ with the DELPHI detector in 1994 is presented. Eighteen infrared and collinear safe event shape observables are measured as a function of the polar angle of the thrust axis. The data are compared to theoretical calculations in${\\cal O} (\\alpha_s^2)$ including the event orientation. A combined fit of $\\alpha_s$ and of the renormalization scale $x_{\\mu}$ in $\\cal O(\\alpha_s^2$)yields an excellent description of the high statistics data. The weighted average from 18 observables including quark mass effects and correlations is $\\alpha_s(M_Z^2) = 0.1174 \\pm 0.0026$. The final result, derived from the jet cone energy fraction, the observable with the smallest theoretical and experimental uncertainty, is $\\alpha_s(M_Z^2) = =0:1180 0:0006(exp:) 0:0013(hadr:) 0:0008(scale) 0:0007(mass). Further studies include an s determination using theoretical predictions in the next-to-leading log approximation (NLLA), matched NLLA and O(\\alpha^{2}_{s})...

  1. Activity monitoring of alpha-bearing wastes

    International Nuclear Information System (INIS)

    Birkhoff, G.; Bondar, L.

    1980-01-01

    The paper aims at the survey on the actual situation in activity monitoring of alpha-bearing wastes. Homogeneous materials such as liquid-, gaseous- and homogeneous solid wastes are amenable to destructive analyses of representative samples. Available destructive analyses methods are sensitive and precise enough to cope with all requirements in alpha-waste monitoring. The more difficult problems are encountered with alpha-contaminated solids, when representative sampling is not practicable. Non-destructive analysis techniques are applied for monitoring this category of solid wastes. The techniques for nondestructive analysis of alpha-bearing wastes are based on the detection of gamma and/or neutron-emission of actinides. Principles and a theory of non-destructive radiometric assay of plutonium contaminated solid waste streams are explained. Guidelines for the calibration of instruments and interpretation of experimental data are given. Current theoretical and experimental development work in this problem area is reviewed. Evaluations concerning capabilities and limitations of monitoring systems for alpha-bearing solid wastes are very complex and out of the scope of this paper

  2. Role of Frontal Alpha Oscillations in Creativity

    Science.gov (United States)

    Lustenberger, Caroline; Boyle, Michael R.; Foulser, A. Alban; Mellin, Juliann M.; Fröhlich, Flavio

    2015-01-01

    Creativity, the ability to produce innovative ideas, is a key higher-order cognitive function that is poorly understood. At the level of macroscopic cortical network dynamics, recent EEG data suggests that cortical oscillations in the alpha frequency band (8 – 12 Hz) are correlated with creative thinking. However, whether alpha oscillations play a fundamental role in creativity has remained unknown. Here we show that creativity is increased by enhancing alpha power using 10 Hz transcranial alternating current stimulation (10Hz-tACS) of the frontal cortex. In a study of 20 healthy participants with a randomized, balanced cross-over design, we found a significant improvement of 7.4% in the Creativity Index measured by the Torrance Test of Creative Thinking, a comprehensive and most frequently used assay of creative potential and strengths. In a second similar study with 20 subjects, 40Hz-tACS was used in instead of 10Hz-tACS to rule out a general “electrical stimulation” effect. No significant change in the Creativity Index was found for such frontal gamma stimulation. Our results suggest that alpha activity in frontal brain areas is selectively involved in creativity; this enhancement represents the first demonstration of specific neuronal dynamics that drive creativity and can be modulated by non-invasive brain stimulation. Our findings agree with the model that alpha recruitment increases with internal processing demands and is involved in inhibitory top-down control, which is an important requirement for creative ideation. PMID:25913062

  3. Purification and characterization of alpha-amylase from rat pancreatic acinar carcinoma. Comparison with pancreatic alpha-amylase.

    OpenAIRE

    Reddy, M K; Heda, G D; Reddy, J K

    1987-01-01

    alpha-Amylase was purified to apparent homogeneity from normal pancreas and a transplantable pancreatic acinar carcinoma of the rat by affinity chromatography on alpha-glucohydrolase inhibitor (alpha-GHI) bound to aminohexyl-Sepharose 4B. Recovery was 95-100% for both pancreas and tumour alpha-amylases. They were monomeric proteins, with Mr approx. 54000 on SDS/polyacrylamide-gel electrophoresis. Isoelectric focusing of both normal and tumour alpha-amylases resolved each into two major isoenz...

  4. Opposing actions of the progesterone metabolites, 5alpha-dihydroprogesterone (5alphaP) and 3alpha-dihydroprogesterone (3alphaHP) on mitosis, apoptosis, and expression of Bcl-2, Bax and p21 in human breast cell lines.

    Science.gov (United States)

    Wiebe, John P; Beausoleil, Michel; Zhang, Guihua; Cialacu, Valentin

    2010-01-01

    Previous studies have shown that breast tissues and breast cell lines convert progesterone (P) to 5alpha-dihydroprogesterone (5alphaP) and 3alpha-dihydroprogesterone (3alphaHP) and that 3alphaHP suppresses, whereas 5alphaP promotes, cell proliferation and detachment. The objectives of the current studies were to determine if the 5alphaP- and 3alphaHP-induced changes in cell numbers are due to altered rates of mitosis and/or apoptosis, and if 3alphaHP and 5alphaP act on tumorigenic and non-tumorigenic cells, regardless of estrogen (E) and P receptor status. The studies were conducted on tumorigenic (MCF-7, MDA-MB-231, T47D) and non-tumorigenic (MCF-10A) human breast cell lines, employing several methods to assess the effects of the hormones on cell proliferation, mitosis, apoptosis and expression of Bcl-2, Bax and p21. In all four cell lines, 5alphaP increased, whereas 3alphaHP decreased cell numbers, [(3)H]thymidine uptake and mitotic index. Apoptosis was stimulated by 3alphaHP and suppressed by 5alphaP. 5alphaP resulted in increases in Bcl-2/Bax ratio, indicating decreased apoptosis; 3alphaHP resulted in decreases in Bcl-2/Bax ratio, indicating increased apoptosis. The effects of either 3alphaHP or 5alphaP on cell numbers, [(3)H]thymidine uptake, mitosis, apoptosis, and Bcl-2/Bax ratio, were abrogated when cells were treated simultaneously with both hormones. The expression of p21 was increased by 3alphaHP, and was unaffected by 5alphaP. The results provide the first evidence that 5alphaP stimulates mitosis and suppresses apoptosis, whereas 3alphaHP inhibits mitosis and stimulates apoptosis. The opposing effects of 5alphaP and 3alphaHP were observed in all four breast cell lines examined and the data suggest that all breast cancers (estrogen-responsive and unresponsive) might be suppressed by blocking 5alphaP formation and/or increasing 3alphaHP. The findings further support the hypothesis that progesterone metabolites are key regulatory hormones and that changes

  5. Low energy alphas in the drift detector

    International Nuclear Information System (INIS)

    Snowden-Ifft, D.P.; Lawson, T.; Spooner, N.J.C.; Villaume, N.

    2004-01-01

    The Directional Recoil Identification From Tracks project is a US-UK endeavor to build and operate a low pressure negative ion TPC (NITPC) to search for weakly interacting massive particles (WIMPs) thought to make up the dark matter in our Galaxy. Low energy (∼10 keV) alpha events from U and Th decays within the walls and wires of the detector can enter the active volume of the detector and be confused for WIMP interactions. This paper presents data on and a model of low energy alphas in a NITPC operated at 40 Torr CS 2 with the aim of understanding and removing this potentially serious background. A comparison of the data to this model reveals good agreement with range predictions of SRIM2000 and allows us to calculate the energy dissipation per ion pair, W=19.0±0.5 eV for low energy alphas in CS 2

  6. Liver replacement for alpha1-antitrypsin deficiency

    Science.gov (United States)

    Putnam, Charles W.; Porter, Kendrick A.; Peters, Robert L.; Ashcavai, Mary; Redeker, Allan G.; Starzl, Thomas E.

    2010-01-01

    A 16-year-old girl with advanced cirrhosis and severe alpha1-antitrypsin deficiency of the homozygous PiZZ phenotype was treated by orthotopic liver transplantation. After replacement of the liver with a homograft from a donor with the normal PiMM phenotype, the alpha1-antitrypsin concentration in the recipient’s serum rose to normal; it had the PiMM phenotype. Two and a third years later, chronic rejection necessitated retransplantation. Insertion of a homograft from a heterozygous PiMZ donar was followed by the identification of that phenotype in the recipient’s serum. Neither liver graft developed the alpha1-antitrypsin glycoprotein deposits seen with the deficiency state. These observations confirm that this hepatic- based inborn error metabolism is metabolically cured by liver replacement. PMID:320694

  7. Investigation of arginine A-specific cysteine proteinase gene expression profiling in clinical Porphyromonas gingivalis isolates against photokilling action of the photo-activated disinfection.

    Science.gov (United States)

    Pourhajibagher, Maryam; Ghorbanzadeh, Roghayeh; Bahador, Abbas

    2018-02-01

    Porphyromonas gingivalis is a significant root canal pathogen capable of causing endodontic infections, which during their treatment may receive sub-lethal doses of photo-activated disinfection (sPAD). As sPAD can influence microbial virulence, this study was designed to evaluate the effect of sPAD on gene expression level of arginine A-specific cysteine proteinase (rgpA), as one of the underlying virulence factors involved in the development of endodontic infection via P. gingivalis strains. To find out the sPAD against 16 clinical isolates of PAD-resistant P. gingivalis that were isolated in vivo, we used toluidine blue O (TBO), methylene blue (MB), and indocyanine green (ICG) as the photosensitizers, which were excited with specific wavelength of light in vitro. Quantitative real-time PCR (qRT-PCR) was then applied to monitor gene expression of rgpA in P. gingivalis isolates to characterize its virulence agent and understand the effect of sPAD on its pathogenicity. Maximal sPAD that could not decrease the count of P. gingivalis isolates were 6.25, 15.6, and 25 μg/mL at fluencies of 171.87, 15.6, and 93.75 J/cm 2 for TBO, ICG, and MB, respectively. ICG-sPAD could suppress the rgpA gene expression about 14-fold, while MB and TBO-mediated sPAD could cause the attenuation of rgpA expression about 4.9- and 11.6-fold, respectively. ICG-sPAD with the maximum ability to reduce rgpA gene expression compared with other photosensitizers can be an appropriate candidate for the treatment of endodontic infections.

  8. Serum and fecal canine α1-proteinase inhibitor concentrations reflect the severity of intestinal crypt abscesses and/or lacteal dilation in dogs.

    Science.gov (United States)

    Heilmann, Romy M; Parnell, Nolie K; Grützner, Niels; Mansell, Joanne; Berghoff, Nora; Schellenberg, Stefan; Reusch, Claudia E; Suchodolski, Jan S; Steiner, Jörg M

    2016-01-01

    Gastrointestinal (GI) protein loss, due to lymphangiectasia or chronic inflammation, can be challenging to diagnose. This study evaluated the diagnostic accuracy of serum and fecal canine α1-proteinase inhibitor (cα1PI) concentrations to detect crypt abscesses and/or lacteal dilation in dogs. Serum and fecal cα1PI concentrations were measured in 120 dogs undergoing GI tissue biopsies, and were compared between dogs with and without crypt abscesses/lacteal dilation. Sensitivity and specificity were calculated for dichotomous outcomes. Serial serum cα1PI concentrations were also evaluated in 12 healthy corticosteroid-treated dogs. Serum cα1PI and albumin concentrations were significantly lower in dogs with crypt abscesses and/or lacteal dilation than in those without (both P <0.001), and more severe lesions were associated with lower serum cα1PI concentrations, higher 3 days-mean fecal cα1PI concentrations, and lower serum/fecal cα1PI ratios. Serum and fecal cα1PI, and their ratios, distinguished dogs with moderate or severe GI crypt abscesses/lacteal dilation from dogs with only mild or none such lesions with moderate sensitivity (56-92%) and specificity (67-81%). Serum cα1PI concentrations increased during corticosteroid administration. We conclude that serum and fecal α1PI concentrations reflect the severity of intestinal crypt abscesses/lacteal dilation in dogs. Due to its specificity for the GI tract, measurement of fecal cα1PI appears to be superior to serum cα1PI for diagnosing GI protein loss in dogs. In addition, the serum/fecal cα1PI ratio has an improved accuracy in hypoalbuminemic dogs, but serum cα1PI concentrations should be carefully interpreted in corticosteroid-treated dogs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Cysteine proteinases regulate chloroplast protein content and composition in tobacco leaves: a model for dynamic interactions with ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) vesicular bodies.

    Science.gov (United States)

    Prins, Anneke; van Heerden, Philippus D R; Olmos, Enrique; Kunert, Karl J; Foyer, Christine H

    2008-01-01

    The roles of cysteine proteinases (CP) in leaf protein accumulation and composition were investigated in transgenic tobacco (Nicotiana tabacum L.) plants expressing the rice cystatin, OC-1. The OC-1 protein was present in the cytosol, chloroplasts, and vacuole of the leaves of OC-1 expressing (OCE) plants. Changes in leaf protein composition and turnover caused by OC-1-dependent inhibition of CP activity were assessed in 8-week-old plants using proteomic analysis. Seven hundred and sixty-five soluble proteins were detected in the controls compared to 860 proteins in the OCE leaves. A cyclophilin, a histone, a peptidyl-prolyl cis-trans isomerase, and two ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activase isoforms were markedly altered in abundance in the OCE leaves. The senescence-related decline in photosynthesis and Rubisco activity was delayed in the OCE leaves. Similarly, OCE leaves maintained higher leaf Rubisco activities and protein than controls following dark chilling. Immunogold labelling studies with specific antibodies showed that Rubisco was present in Rubisco vesicular bodies (RVB) as well as in the chloroplasts of leaves from 8-week-old control and OCE plants. Western blot analysis of plants at 14 weeks after both genotypes had flowered revealed large increases in the amount of Rubisco protein in the OCE leaves compared to controls. These results demonstrate that CPs are involved in Rubisco turnover in leaves under optimal and stress conditions and that extra-plastidic RVB bodies are present even in young source leaves. Furthermore, these data form the basis for a new model of Rubisco protein turnover involving CPs and RVBs.

  10. House Dust Mites Induce Production of Endothelin-1 and Matrix Metalloproteinase-9 in Keratinocytes via Proteinase-Activated Receptor-2 Activation.

    Science.gov (United States)

    Yamada, Yoshihito; Matsumoto, Tatsumi

    2017-01-01

    Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by skin barrier dysfunction and abnormal immune response. House dust mites (HDM) are a major source of allergens, some of which have cysteine and serine protease activities. Keratinocytes stimulated by HDM-derived proteases have been suggested to contribute to the pathogenesis of AD by producing various cytokines. However, whether keratinocytes contribute to the induction of pruritus in AD, especially by producing pruritus-related mediators upon stimulation with HDM-derived proteases, has not been fully elucidated. We examined whether the production of endothelin-1 (ET-1), matrix metalloproteinase (MMP)-2, and MMP-9 in keratinocytes can be induced by stimulation with Dermatophagoides farinae extracts, and if so, whether pretreatment with a protease inhibitor or proteinase-activated receptor-2 (PAR-2) antagonist affects the production of these mediators in keratinocytes. Although MMP-2 levels were undetectable in the culture supernatants, the production of ET-1 and MMP-9 was increased upon stimulation with HDM extracts in a concentration- and time-dependent manner and suppressed by pretreatment of HDM extracts with serine protease inhibitor, but not with cysteine protease inhibitor. Mite-derived serine proteases also induced ET-1 and MMP-9 production in a concentration- and time-dependent manner. Moreover, pretreatment with a PAR-2 antagonist inhibited the production of ET-1 and MMP-9 in keratinocytes. These results suggest that the activation of PAR-2 on keratinocytes by HDM-derived serine proteases induces the production of ET-1 and MMP-9, and may contribute to the induction of pruritus in AD. © 2017 S. Karger AG, Basel.

  11. Variability in proteinase-antiproteinase balance, nutritional status, and quality of life in stable chronic obstructive pulmonary disease due to tobacco and nontobacco etiology

    Directory of Open Access Journals (Sweden)

    Anant Mohan

    2016-01-01

    Full Text Available Context: Although the role of proteinase/antiproteinase imbalance in chronic obstructive pulmonary disease (COPD due to tobacco is well established, information in COPD due to nontobacco etiology is sparse. Aims: To assess the variability in metalloproteinase activity in COPD related to tobacco and nontobacco causes. Settings and Design: This is a hospital-based, prospective, observational study. Subjects and Methods: Serum matrix metalloproteinase-9 (MMP-9 and tissue inhibitor of metalloproteinases-1 (TIMP-1 were estimated in 200 subjects divided equally into four groups, i.e. COPD in tobacco smokers, COPD in nonsmokers but with exposure to biomass-related indoor air pollution, smokers without COPD, and nonsmoking healthy controls. Anthropometric skinfold measurements, quality of life (QOL using St. George Respiratory Questionnaire, and exercise capacity using the 6-min walk test (6-MWT were carried out. Groups were compared using analysis of variance and Kruskal–Wallis plus Mann–Whitney U-test to assess differences between groups. The Chi-square and Fisher's exact tests were used to evaluate associations among categorical variables. Spearman's rank correlation was calculated to assess the correlation between data. Results: Patients with COPD due to either tobacco or nontobacco etiology were older, more malnourished, had worse QOL, and poorer exercise capacity compared to non-COPD subjects. Triceps, subscapular, and suprailiac skinfold thicknesses were less in smokers with COPD than biomass-related COPD. MMP-9 and TIMP-1 levels were similar across all groups. TIMP-1 significantly correlated with 6-MWT among all groups. Conclusions: The protease-antiprotease balance in COPD is similar irrespective of the presence or absence of tobacco exposure but is related to poor exercise capacity.

  12. Mechanisms for proteinase-activated receptor 1-triggered prostaglandin E2 generation in mouse osteoblastic MC3T3-E1 cells.

    Science.gov (United States)

    Maeda, Yuma; Sekiguchi, Fumiko; Yamanaka, Rumi; Sugimoto, Ryo; Yamasoba, Daichi; Tomita, Shiori; Nishikawa, Hiroyuki; Kawabata, Atsufumi

    2015-02-01

    We analyzed signaling mechanisms for prostaglandin E2 (PGE2) production following activation of proteinase-activated receptor-1 (PAR1), a thrombin receptor, in preosteoblastic MC3T3-E1 cells. PAR1 stimulation caused PGE2 release, an effect suppressed by inhibitors of COX-1, COX-2, iPLA2, cPLA2, MAP kinases (MAPKs), Src, EGF receptor (EGFR) tyrosine kinase (EGFR-TK) and matrix metalloproteinase (MMP), but not by an intracellular Ca2+ chelator or inhibitors of PI3 kinase, protein kinase C (PKC) and NF-κB. PAR1 activation induced phosphorylation of MAPKs and upregulation of COX-2. The phosphorylation of p38 MAPK was suppressed by inhibitors of Src and EGFR-TK. The COX-2 upregulation was dependent on ERK, p38, EGFR-TK, Src, and COX-2 itself. PAR1 activation also induced MEK-dependent phosphorylation of cAMP response element binding protein (CREB). All inhibitors of EP1, EP2, EP3 and EP4 receptors suppressed the PAR1-triggered PGE2 release. Exogenously applied PGE2 facilitated PAR1-triggered COX-2 upregulation, but it alone had no effect. Together, the PAR1-mediated PGE2 production in MC3T3-E1 cells appears to involve iPLA2 and cPLA2 for arachidonic acid release, and the MEK/ERK/CREB and Src/MMP/EGFR/p38 pathways for COX-2 upregulation, which is facilitated by endogenous PGE2 formed by COX-2. These signaling mechanisms might underlie the role of the thrombin/PAR1/PGE2 system in the early stage of the bone healing.

  13. Lactobacillus bulgaricus proteinase expressed in Lactococcus lactis is a powerful carrier for cell wall-associated and secreted bovine beta-lactoglobulin fusion proteins.

    Science.gov (United States)

    Bernasconi, Eric; Germond, Jacques-Edouard; Delley, Michèle; Fritsché, Rodolphe; Corthésy, Blaise

    2002-06-01

    Lactic acid bacteria have a good potential as agents for the delivery of heterologous proteins to the gastrointestinal mucosa and thus for the reequilibration of inappropriate immune responses to food antigens. Bovine beta-lactoglobulin (BLG) is considered a major allergen in cow's milk allergy. We have designed recombinant Lactococcus lactis expressing either full-length BLG or BLG-derived octapeptide T6 (IDALNENK) as fusions with Lactobacillus bulgaricus extracellular proteinase (PrtB). In addition to constructs encoding full-length PrtB for the targeting of heterologous proteins to the cell surface, we generated vectors aiming at the release into the medium of truncated PrtB derivatives lacking 100 (PrtB partial differential, PrtB partial differential-BLG, and PrtB partial differential-T6) or 807 (PrtBdelta) C-terminal amino acids. Expression of recombinant products was confirmed using either anti-PrtB, anti-BLG, or anti-peptide T6 antiserum. All forms of the full-length and truncated recombinant products were efficiently translocated, irrespective of the presence of eucaryotic BLG sequences in the fusion proteins. L. lactis expressing PrtB partial differential-BLG yielded up to 170 microg per 10(9) CFU in the culture supernatant and 9 microg per 10(9) CFU at the bacterial cell surface within 14 h. Therefore, protein fusions relying on the use of PrtB gene products are adequate for concomitant cell surface display and secretion by recombinant L. lactis and thus may ensure maximal bioavailability of the eucaryotic antigen in the gut-associated lymphoid tissue.

  14. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    International Nuclear Information System (INIS)

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun; Xiao, Shaobo

    2010-01-01

    Research highlights: → FMDV L pro inhibits poly(I:C)-induced IFN-α1/β mRNA expression. → L pro inhibits MDA5-mediated activation of the IFN-α1/β promoter. → L pro significantly reduced the transcription of multiple IRF-responsive genes. → L pro inhibits IFN-α1/β promoter activation by decreasing IRF-3/7 in protein levels. → The ability to process eIF-4G of L pro is not necessary to inhibit IFN-α1/β activation. -- Abstract: The leader proteinase (L pro ) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-β (IFN-β) antagonist that disrupts the integrity of transcription factor nuclear factor κB (NF-κB). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-α1/β expression caused by L pro was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-α/β. Furthermore, overexpression of L pro significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L pro mutants indicated that the ability to process eIF-4G of L pro is not required for suppressing dsRNA-induced activation of the IFN-α1/β promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-κB, L pro also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  15. The region approximately between amino acids 81 and 137 of proteinase K-resistant PrPSc is critical for the infectivity of the Chandler prion strain.

    Science.gov (United States)

    Shindoh, Ryo; Kim, Chan-Lan; Song, Chang-Hyun; Hasebe, Rie; Horiuchi, Motohiro

    2009-04-01

    Although the major component of the prion is believed to be the oligomer of PrP(Sc), little information is available concerning regions on the PrP(Sc) molecule that affect prion infectivity. During the analysis of PrP(Sc) molecules from various prion strains, we found that PrP(Sc) of the Chandler strain showed a unique property in the conformational-stability assay, and this property appeared to be useful for studying the relationship between regions of the PrP(Sc) molecule and prion infectivity. Thus, we analyzed PrP(Sc) of the Chandler strain in detail and analyzed the infectivities of the N-terminally denatured and truncated forms of proteinase K-resistant PrP. The N-terminal region of PrP(Sc) of the Chandler strain showed region-dependent resistance to guanidine hydrochloride (GdnHCl) treatment. The region approximately between amino acids (aa) 81 and 137 began to be denatured by treatment with 1.5 M GdnHCl. Within this stretch, the region comprising approximately aa 81 to 90 was denatured almost completely by 2 M GdnHCl. Furthermore, the region approximately between aa 90 and 137 was denatured completely by 3 M GdnHCl. However, the C-terminal region thereafter was extremely resistant to the GdnHCl treatment. This property was not observed in PrP(Sc) molecules of other prion strains. Denaturation of the region between aa 81 and 137 by 3 M GdnHCl significantly prolonged the incubation periods in mice compared to that for the untreated control. More strikingly, the denaturation and removal of this region nearly abolished the infectivity. This finding suggests that the conformation of the region between aa 81 and 137 of the Chandler strain PrP(Sc) molecule is directly associated with prion infectivity.

  16. Advances in alpha air monitoring instrumentation

    International Nuclear Information System (INIS)

    Valentine, A.M.

    1979-01-01

    Early detection of airborne radioactive material is important for effective protection of workers, particularly at facilities handling high toxicity alpha-emitting radionuclides such as 238 Pu and 239 Pu. Variable levels of ever present and naturally occurring alpha-emitting radon daughters made gross alpha air monitoring systems unreliable for live-time monitoring of airborne plutonium. Hence, good live-time monitoring was not possible until an improved alpha air monitoring instrument capable of monitoring select plutonium radionuclides of concern was developed and marketed by nuclear instrumentation companies in the USA during the late 1960s and early 1970s. This monitoring instrument utilized a solid-state silicon-diffused junction type of detector and pulse-height analyser system to achieve a radon-daughter rejection capability. This rejection capability has resulted in an improved sensitivity for specific alpha-emitting radionuclides such as 238 Pu, 239 Pu and 241 Am. Current models of this instrument are capable of detecting 40 dis/min (0.65 Bq) of plutonium alpha activity. This activity is about equal to the integrated activity workers would be exposed to if they worked for eight hours in an airborne concentration equal to the maximum permissible concentration for transportable 239 Pu. The Los Alamos Scientific Laboratory began using these instruments in its plutonium research laboratories when they became available in the early 1970s. Approximately 300 are currently in service, and their use has resulted in much improved protection of plutonium workers because accidental airborne releases can now be detected soon after they occur. General design features of these instruments are discussed together with actual operational experience and field calibration procedures. (author)

  17. Parallel Genetic Algorithm for Alpha Spectra Fitting

    Science.gov (United States)

    García-Orellana, Carlos J.; Rubio-Montero, Pilar; González-Velasco, Horacio

    2005-01-01

    We present a performance study of alpha-particle spectra fitting using parallel Genetic Algorithm (GA). The method uses a two-step approach. In the first step we run parallel GA to find an initial solution for the second step, in which we use Levenberg-Marquardt (LM) method for a precise final fit. GA is a high resources-demanding method, so we use a Beowulf cluster for parallel simulation. The relationship between simulation time (and parallel efficiency) and processors number is studied using several alpha spectra, with the aim of obtaining a method to estimate the optimal processors number that must be used in a simulation.

  18. Enteric alpha defensins in norm and pathology

    Directory of Open Access Journals (Sweden)

    Lisitsyn Nikolai A

    2012-01-01

    Full Text Available Abstract Microbes living in the mammalian gut exist in constant contact with immunity system that prevents infection and maintains homeostasis. Enteric alpha defensins play an important role in regulation of bacterial colonization of the gut, as well as in activation of pro- and anti-inflammatory responses of the adaptive immune system cells in lamina propria. This review summarizes currently available data on functions of mammalian enteric alpha defensins in the immune defense and changes in their secretion in intestinal inflammatory diseases and cancer.

  19. Clustering of Lyman-Alpha Emitters galaxies

    Science.gov (United States)

    Francke, Harold

    2009-06-01

    Galaxy clustering properties have been studied for decades to constrain cosmological parameters and have today, with large datasets of high-redshift sources piling up, become a powerful tool to discriminate and characterize primeval galaxies. In the last years, several Lyman-Alpha Emitter (LAE) galaxy samples have been gathered, which are big, uniform and compact enough to allow clustering analysis. Here we present a summary of the discussion session on the clustering properties of LAEs at the "Understanding Lyman-Alpha Emitters" conference.

  20. Diffusion of cobalt in alpha zirconium

    International Nuclear Information System (INIS)

    Ghosh, U.; Pruthi, D.D.; Anand, M.S.

    1979-01-01

    The diffusion of 60 Co in alpha zirconium has been studied in the temperature range of 873-1123 K using the sectioning technique. The diffusion parameters obey the following Arrhenius equation:- D = (1.132 +- 0.22)10 -4 exp((-136.43 +- 3.50 KJ)/RT)m 2 s -1 . The diffusivity in this phase is as high as that observed in β-phase. It is also about five orders of magnitude higher when compared to the self diffusion values. The high diffusivity of cobalt in alpha zirconium has been explained on the basis of an interstitial mechanism. (auth.)

  1. Radiological hazards of alpha-contaminated waste

    International Nuclear Information System (INIS)

    Rodgers, J.C.

    1982-01-01

    The radiological hazards of alpha-contaminated wastes are discussed in this overview in terms of two components of hazard: radiobiological hazard, and radioecological hazard. Radiobiological hazard refers to human uptake of alpha-emitters by inhalation and ingestion, and the resultant dose to critical organs of the body. Radioecological hazard refers to the processes of release from buried wastes, transport in the environment, and translocation to man through the food chain. Besides detailing the sources and magnitude of hazards, this brief review identifies the uncertainties in their estimation, and implications for the regulatory process

  2. The murine alpha B-crystallin/small heat shock protein enhancer: identification of alpha BE-1, alpha BE-2, alpha BE-3, and MRF control elements.

    OpenAIRE

    Gopal-Srivastava, R; Piatigorsky, J

    1993-01-01

    The murine alpha B-crystallin gene (a member of the small heat shock protein family) is expressed constitutively at high levels in the lens and at lower levels in many other tissues, including skeletal muscle. We have previously used the herpes simplex virus thymidine kinase promoter fused to the human growth hormone gene to identify an alpha B-crystallin enhancer at positions -427 to -259 that has high activity in muscle and low activity in lens cell lines. In the study reported here, we per...

  3. Fan-less long range alpha detector

    Science.gov (United States)

    MacArthur, Duncan W.; Bounds, John A.

    1994-01-01

    A fan-less long range alpha detector which operates by using an electrical field between a signal plane and the surface or substance to be monitored for air ions created by collisions with alpha radiation. Without a fan, the detector can operate without the possibility of spreading dust and potential contamination into the atmosphere. A guard plane between the signal plane and the electrically conductive enclosure and maintained at the same voltage as the signal plane, reduces leakage currents. The detector can easily monitor soil, or other solid or liquid surfaces.

  4. Recoil-alpha-fission and recoil-alpha-alpha-fission events observed in the reaction Ca-48 + Am-243

    NARCIS (Netherlands)

    Forsberg, U.; Rudolph, D.; Andersson, L. -L.; Nitto, A. Di; Düllmann, Ch E.; Gates, J. M.; Golubev, P.; Gregorich, K. E.; Gross, C. J.; Herzberg, R. -D.; Hessberger, F. P.; Khuyagbaatar, J.; Kratz, J. V.; Rykaczewski, K.; Sarmiento, L. G.; Schädel, M.; Yakushev, A.; Åberg, S.; Ackermann, D.; Block, M.; Brand, H.; Carlsson, B. G.; Cox, D.; Derkx, X.; Dobaczewski, J.; Eberhardt, K.; Even, J.; Fahlander, C.; Gerl, J.; Jäger, E.; Kindler, B.; Krier, J.; Kojouharov, I.; Kurz, N.; Lommel, B.; Mistry, A.; Mokry, C.; Nazarewicz, W.; Nitsche, H.; Omtvedt, J. P.; Papadakis, P.; Ragnarsson, I.; Runke, J.; Schaffner, H.; Schausten, B.; Shi, Y.; Thörle-Pospiech, P.; Torres, T.; Traut, T.; Trautmann, N.; Türler, A.; Ward, A.; Ward, D. E.; Wiehl, N.

    2016-01-01

    Products of the fusion-evaporation reaction Ca-48 + Am-243 were studied with the TASISpec set-up at the gas-filled separator TASCA at the GSI Helmholtzzentrum f\\"ur Schwerionenforschung. Amongst the detected thirty correlated alpha-decay chains associated with the production of element Z=115, two

  5. Alpha 1-adrenoceptor subtypes in the rat ventricular muscle.

    Science.gov (United States)

    Kinami, J; Tsuchihashi, H; Baba, S; Mano, F; Maruyama, K; Nagatomo, T

    1992-02-01

    Scatchard analyses of [3H]prazosin binding in rat ventricular muscle membranes showed biphasic curves, which identified alpha 1High- and alpha 1Low-affinity sites. The alpha 1High-affinity site was completely inhibited by 1 microM phenoxybenzamine. The displacement potencies of alpha 1-adrenergic antagonists were characterized by [3H]prazosin binding to alpha 1High- and alpha 1Low-affinity sites in the absence and presence of 1 microM phenoxybenzamine. The affinities of most chemicals for alpha 1Low-affinity sites were significantly lower than those for alpha 1High-affinity sites, but WB-4101 (2-(2,6-dimethoxy-phenoxyethyl)aminomethyl-1,4-benzodioxane), arotinolol, cinanserin, nifedipine, and p-aminoclonidine had the same affinities for both alpha 1Low- and alpha 1High-affinity sites. These results show that two alpha 1-adrenoceptor subtypes, alpha 1High- and alpha 1Low-affinity, are present in the rat heart, and that there are physical variations in alpha 1-adrenoceptor binding sites, based on their selectivity to antagonists.

  6. Six HLA-D region alpha-chain genes on human chromosome 6: polymorphisms and associations of DC alpha-related sequences with DR types.

    OpenAIRE

    Spielman, R S; Lee, J; Bodmer, W F; Bodmer, J G; Trowsdale, J

    1984-01-01

    Analysis of cosmid clones containing genes related to the HLA-DR alpha chain calls for at least six HLA-D region alpha-chain coding sequences in man; namely, DR alpha, DC alpha, DX alpha (very closely related to DC alpha), SB alpha 1, SB alpha 2 (two closely linked genes on the same cosmid clones), and DZ alpha. The first four genes have been described previously. SB alpha 2 and DZ alpha are recently identified genes, characterized by their unique and, from a limited study, nonpolymorphic ban...

  7. Phenomenological Alpha-Alpha Interactions and its Application on 16O Nucleus

    International Nuclear Information System (INIS)

    Qandil, O.S.A.A.

    2013-01-01

    The thesis includes the study of alpha-alpha interactions and apply them to the nucleus of the 16 O by using Hartree-Fock method for bosons. The used potentials are Ali-Bodmer potential, Modified Woods- Saxon potential, Fish-Bone I (FB-I) and Fish-Bone II (FB-II) potential. Also, the properties of nuclear structure of 16 O nucleus are calculated such as binding energy, form factor and root mean square radius. The results we have obtained were compared with previous results in the same field of research topic. This work requires knowledge of different kinds of potentials and the difference between the phenomenological and microscopic interactions and research in Hartree-Fock method and nuclear models, especially shell model. The content of the present thesis can be summarized as the following: The first Chapter, (Introduction), presents detailed informations about the nuclear structure, clustering, alpha clustering and nuclear models especially the shell model. Also, a review of previous researches in the research topic ( phenomenological alpha- alpha interactions). The second chapter, (Alpha-Alpha interactions), many types of phenomenological alpha-alpha interactions and its various forms are displayed, including what has been used in the thesis.The third chapter, (Derivation of Hartree-Fock Method for Bosons), illustrates the theoretical calculations which have been used in the thesis, the derivation of the Hartree-Fock method for bosons, the variational method and the derivation of binding energy, form factor and root mean square radius equations. the fourth chapter, (Numerical Results and Discussion), includes the present theoretical results of the nuclear structure of 16 O nucleus using the previous four potentials, and comparing our present results with the experimental and previous theoretical ones.

  8. Thermochemical Stability of alpha-Amino-alpha-carbonylmethyl Radicals and Their Resonance As Measured by ESR.

    Science.gov (United States)

    Welle, Frank M.; Beckhaus, Hans-Dieter; Rüchardt, Christoph

    1997-02-07

    ESR spectra of the captodative alpha-amino-alpha-carbonylmethyl radicals 8 have been recorded. No coalescence temperature for the rotation of the two NMe groups was found at temperatures below the decomposition temperature of the radicals. From known coalescence temperatures and rotational barriers of substituted methyl radicals the rotational barrier of >/=17 kcal mol(-)(1) was estimated for the (*)C-N bond in the radicals 8. Enthalpies DeltaH(diss) and entropies DeltaS(diss) of the homolytic dissociation of 7a,c,d into 8a,c,d have been obtained from equilibrium measurements by ESR. By correcting for substituent interaction enthalpies in 7 (steric and geminal), a radical stabilization enthalpy RSE = -20.7 +/- 1.0 kcal mol(-)(1) was obtained for 8. By addition of the known RSEs of dialkylamino- and carbonyl groups, a RSE = -9.9 kcal mol(-)(1) is predicted for 8. The difference between the experimental and predicted values of 10.8 kcal mol(-)(1) is attributed to a synergistic captodative substituent effect. A linear correlation between the radical stabilization enthalpies of the radicals 8 and of other mono- and disubstituted alkyl radicals and their ESR aH(alpha) coupling constants was found. According to this correlation the reduction of aH(alpha) by 1 G corresponds to an increase in RSE of 1.57 kcal mol(-)(1). The large resonance of the captodative alpha-amino-alpha-carbonylmethyl radicals 3, expressed by their high RSE, their small aH(alpha) coupling constant, and their high rotational barrier, can be rationalized by a strong interaction between the alpha-amino and the alpha-carbonyl groups similar to that in amides and expressed in the resonance structures 6.

  9. Isolation and characterization of a cDNA from flowers of Cynara cardunculus encoding cyprosin (an aspartic proteinase) and its use to study the organ-specific expression of cyprosin.

    Science.gov (United States)

    Cordeiro, M C; Xue, Z T; Pietrzak, M; Pais, M S; Brodelius, P E

    1994-03-01

    Poly(A)+ RNA isolated from flower buds of Cynara cardunculus has been used to prepare a cDNA library. Screening of the cDNA after expression of cloned DNA with antibodies raised against the large subunit of cyprosin 3 resulted in the isolation of six positive clones. One of these clones (cypro1s; a 1.7 kb Eco RI fragment) codes for cyprosin. The nucleotide sequence contain a 1419 bp open reading frame coding for 473 amino acids (aa) including a putative full-length mature protein (440 aa) and a partial prosequence (33 aa). Cypro1s contains a 162 bp 3' non-coding region followed by a poly(A) tail. The deduced amino acid sequence shows high homology to other plant aspartic proteinases. The homology to mammalian and microbial aspartic proteinases is somewhat lower. Plant aspartic proteinases contain an insert of around 100 aa. We are modelling where this plant-specific insert will appear in the structure of cyprosin. Using cypro1s as a probe in northern blot analysis, the expression of cyprosin in developing flowers and other tissues has been studied. The signal on the northern blot increased for RNA samples from early (flower buds 6 mm in length) to later stages of floral development (flower buds up to 40 mm in length). In late stages of floral development (open flowers 50 mm in length and styles from such flowers) no hybridization signal was visualized showing that the synthesis of mRNA encoding the cyprosin starts in early stages of floral development and switches off at maturation of the flower.(ABSTRACT TRUNCATED AT 250 WORDS)

  10. Production of pi /sup 0/ at large transverse momentum in alpha alpha and alpha p collisions at the CERN Intersecting Storage Rings

    CERN Document Server

    Karabarbounis, A; Fields, T; Filippas-Tassos, A; Fokitis, E; Goldberg, M; Kourkoumelis, C; Lissauer, D; Mannelli, I; Molzon, W; Mouzourakis, P; Palmer, R B; Rahm, David Charles; Rehak, P; Resvanis, L K; Stumer, I; Trakkas, C; Willis, W

    1981-01-01

    Inclusive pi /sup 0/ production has been measured at the CERN Intersecting Storage Rings in alpha alpha and alpha p collisions near 90 degrees , for p/sub T/ between 2 and 5 GeV/c. The differential cross sections show a slower exponential fall-off with p/sub T/ than has been observed in pp collisions at the corresponding nucleon- nucleon centre-of-mass energies at large p/sub T/. The ratio of the pi /sup 0/ production cross sections for alpha alpha collisions to those for pp collisions is observed to be larger than 16. (8 refs).

  11. A survey of the alpha-nucleon interaction

    International Nuclear Information System (INIS)

    Ali, S.; Ahmad, A.A.Z.; Ferdous, N.

    1984-10-01

    A survey of the alpha-nucleon interaction is made. The experimental work on angular distributions of differential scattering cross-sections and polarizations in proton-alpha and neutron-alpha scattering is described. The phenomenological approach which includes the study of both local and non-local potentials reproducing the experimental alpha-nucleon scattering data, is discussed. Basic studies of the alpha-nucleon interaction attempting to build an interaction between an alpha particle and a nucleon from first principles are then described. A critical discussion of the results with some concluding remarks suggesting the direction for further investigation is made. (author)

  12. Production of {alpha}-glucosidases by Bacillus sp. strains

    Energy Technology Data Exchange (ETDEWEB)

    Castro, G.R. [Univ. Nacional de Tucuman, Facultad de Bioquimica, Quimica y Farmacia, Catedra de Microbiologia Superior, PROIMI-MIRCEN, Tucuman (Argentina); Baigori, M.D. [Univ. Nacional de Tucuman, Facultad de Bioquimica, Quimica y Farmacia, Catedra de Microbiologia Superior, PROIMI-MIRCEN, Tucuman (Argentina); Sineriz, F. [Univ. Nacional de Tucuman, Facultad de Bioquimica, Quimica y Farmacia, Catedra de Microbiologia Superior, PROIMI-MIRCEN, Tucuman (Argentina)

    1995-12-31

    {alpha}-Glucosidase was detected in four wild-type amylolytic production strains belonging to the Bacillus genus. The strains showed {alpha}-glucosidase activity in extracellular and membrane-bound fractions. Kinetic studies of the {alpha}-glucosidase synthesis in the batch cultures of four strains of the Bacillus genus showed two profiles: partially and totally growth-linked synthesis. The presence of different activities and production profiles of {alpha}-glucosidase in the strains at high or low glucose concentrations in the medium would indicate that {alpha}-glucosidase may have a role in the regulation of the metabolism of {alpha}-polysaccharides. (orig.)

  13. Remarks on tilde g_{alpha}-irresolute maps

    Directory of Open Access Journals (Sweden)

    Nirmala Rebecca Paul

    2011-12-01

    Full Text Available Only a few of the class of generalized closed sets form a topology. The class of tilde g_{alpha}-closed sets is one among them. The aim of this paper is to introduce the different notions of irresolute function using tilde g_{alpha}-closed sets and study some of their basic properties.We also study the relation between strongly tilde g_{alpha}- continuous and perfectly eg-continuous functions. We also introduce tilde g_{alpha}-compact and ilde g_{alpha}-connectedspaces and study their properties using tilde g_{alpha}-continuous and eg-irresolute functions.

  14. Neuropsychiatric Complications Associated with Interferon - Alpha ...

    African Journals Online (AJOL)

    Several adverse effects have been associated with interferon alpha 2b treatment and neuropsychiatric effects have also been commonly reported. Psychosis and mood disorders have been described in the literature. This case report is of a 30 year old man with malignant melanoma stage 3a who was receiving adjuvant ...

  15. Alpha -2b treatment of Malignant Melanoma

    African Journals Online (AJOL)

    Introduction. Interferon preparations such as interferon alpha are widely used in the treatment of a number of non malignant and malignant conditions including malignant melanoma where it used. Mania has been observed in patients undergoing interferon treatment especially after significant dose- reduction or treatment ...

  16. Laboratory system for alpha particle spectroscopy

    International Nuclear Information System (INIS)

    Dean, J.R.; Chiu, N.W.

    1987-03-01

    An automated alpha particle spectroscopy system has beeen designed and fabricated. It consists of two major components, the automatic sample changer and the controller/data acquisition unit. It is capable of unattended analysis of ten samples for up to 65,000 seconds per sample

  17. Biomarkers of Alpha Particle Radiation Exposure

    Science.gov (United States)

    2014-04-01

    work towards the identification of gene-based biomarkers of alpha-particle radiation exposure. Peripheral blood mononuclear cells (PBMN) isolated from...manipulation et l’exposition au rayonnement ionisant chez les humains . CSSP-2012-CD-1117 and CSSP-2012-CD-1114 iii Table of contents...19 Acknowledgements This work was supported by the Centre for

  18. Superparamagnetic relaxation in alpha-Fe particles

    DEFF Research Database (Denmark)

    Bødker, Franz; Mørup, Steen; Pedersen, Michael Stanley

    1998-01-01

    The superparamagnetic relaxation time of carbon-supported alpha-Fe particles with an average size of 3.0 Mm has been studied over a large temperature range by the use of Mossbauer spectroscopy combined with AC and DC magnetization measurements. It is found that the relaxation time varies with tem...

  19. Extraction of molybdenum VI by alpha benzoinoxime

    International Nuclear Information System (INIS)

    Achache, M.; Meklati, M.

    1990-06-01

    The concentration of molybdenum, was studied using alpha benzoinoxime dissolved in chloroform. Several acids and salt at different levels of concentration were investigated as well as other parameters such as (mixing time, extractant to metal ratio, temperature etc.) The molybdenum stippling was also studied in alkaline medium with the subsequent recovery of the extractant and solvent

  20. Syndecan-4 associates with alpha-actinin

    DEFF Research Database (Denmark)

    Greene, Daniel K; Tumova, Sarka; Couchman, John R

    2002-01-01

    during the formation of focal adhesions. To date, a direct link between syndecan-4 and the cytoskeleton has remained elusive. We now demonstrate by Triton X-100 extraction immunoprecipitation and in vitro binding assays that the focal adhesion component alpha-actinin interacts with syndecan-4 in a beta...

  1. Genetics Home Reference: alpha-1 antitrypsin deficiency

    Science.gov (United States)

    ... rapid heartbeat upon standing. Affected individuals often develop emphysema, which is a lung disease caused by damage to the small air ... exposure to tobacco smoke accelerates the appearance of emphysema symptoms and damage to the lungs. About 10 percent of infants with alpha-1 ...

  2. Diagnosis of foetal membrane ruptures: Placental alpha ...

    African Journals Online (AJOL)

    AbstRACt. Context: Pre‑labour rupture of membranes (PROM) is a common obstetric complication which presents a diagnostic challenge, especially in equivocal cases. Standard methods of diagnosis are limited by high false positives and negatives. This study compared the accuracy of a biomarker placental alpha ...

  3. Evaluation of microcrystalline cellulose modifed from alpha ...

    African Journals Online (AJOL)

    Alpha cellulose was obtained from Costus afer and part of it was modified to microcrystalline cellulose (CAMCC). The physicochemical properties of the microcrystalline cellulose were determined and compared with those of commercial microcrystalline cellulose (Avicel 101). The swelling capacity, hydration capacity, loss ...

  4. Physicochemical and powder properties of alpha- and ...

    African Journals Online (AJOL)

    Cob alpha-celluloses (CAC) was extracted from maize cobs by defibering, delignification and bleaching; then subjected to acid hydrolysis to obtain Cob- microcrystalline-cellulose (CMCC). Their physicochemical properties were evaluated and compared with those of Avicel®, a commercial variety of microcrystalline ...

  5. Comparison of alpha decay and alpha transfer reactions in the lead region. [R matrix theory, absolute alpha widths, cross sections, 93 MeV

    Energy Technology Data Exchange (ETDEWEB)

    DeVries, R.M.

    1978-01-01

    Data were taken for five transitions in the lead region allowing a quantitative comparison with corresponding alpha-decay data via R-matrix theory using the same target + alpha nuclear potential. Good agreement between the absolute reduced widths determined from the two sets of data suggests that in transfer reactions, as in alpha decay, an alpha particle in its ground state is transferred in a one-step process. In a separate analysis, elastic and total reaction cross sections for the systems ..cap alpha.. + /sup 208/Pb, /sup 209/Bi were analyzed to obtain a limited set of potentials which, in turn, were used to calculate absolute alpha widths. Existing shell model calculations give ..gamma../sub ..cap alpha..//sup 2/ values three orders of magnitude smaller.

  6. Involvement of the clock gene Rev-erb alpha in the regulation of glucagon secretion in pancreatic alpha-cells.

    Directory of Open Access Journals (Sweden)

    Elaine Vieira

    Full Text Available Disruption of pancreatic clock genes impairs pancreatic beta-cell function, leading to the onset of diabetes. Despite the importance of pancreatic alpha-cells in the regulation of glucose homeostasis and in diabetes pathophysiology, nothing is known about the role of clock genes in these cells. Here, we identify the clock gene Rev-erb alpha as a new intracellular regulator of glucagon secretion. Rev-erb alpha down-regulation by siRNA (60-70% inhibition in alphaTC1-9 cells inhibited low-glucose induced glucagon secretion (p<0.05 and led to a decrease in key genes of the exocytotic machinery. The Rev-erb alpha agonist GSK4112 increased glucagon secretion (1.6 fold and intracellular calcium signals in alphaTC1-9 cells and mouse primary alpha-cells, whereas the Rev-erb alpha antagonist SR8278 produced the opposite effect. At 0.5 mM glucose, alphaTC1-9 cells exhibited intrinsic circadian Rev-erb alpha expression oscillations that were inhibited by 11 mM glucose. In mouse primary alpha-cells, glucose induced similar effects (p<0.001. High glucose inhibited key genes controlled by AMPK such as Nampt, Sirt1 and PGC-1 alpha in alphaTC1-9 cells (p<0.05. AMPK activation by metformin completely reversed the inhibitory effect of glucose on Nampt-Sirt1-PGC-1 alpha and Rev-erb alpha. Nampt inhibition decreased Sirt1, PGC-1 alpha and Rev-erb alpha mRNA expression (p<0.01 and glucagon release (p<0.05. These findings identify Rev-erb alpha as a new intracellular regulator of glucagon secretion via AMPK/Nampt/Sirt1 pathway.

  7. Nicotine enhances expression of the alpha 3, alpha 4, alpha 5, and alpha 7 nicotinic receptors modulating calcium metabolism and regulating adhesion and motility of respiratory epithelial cells.

    Science.gov (United States)

    Zia, S; Ndoye, A; Nguyen, V T; Grando, S A

    1997-09-01

    The purpose of this study was to investigate the possibility of direct toxic effects of nicotine (Nic) on human bronchial epithelial cells (BEC) suggested by our previous findings of functional nicotinic acetylcholine receptors (nAChRs) in the epithelial cells lining mucocutaneous membranes. We now demonstrate for the first time that human and murine BEC both in vivo and in vitro express functional nAChRs, and that classic alpha 3, alpha 4, alpha 5 and alpha 7 subunits can contribute to formation of these acetylcholine-gated ion channels. In human bronchial and mouse lung tissues, and in cultures of human BEC, the nAChRs were visualized by subunit-specific antibodies on the cell membranes, particularly at the sites of cell-to-cell contacts. The epithelial cells of submucosal glands abundantly expressed alpha 7 nAChRs. Smoking significantly (p epithelial nAChRs apparently involve regulation of cell-to-cell communications, adhesion and motility, because Mec caused rapid and profound changes in these cell functions which were reversible by Nic. An over exposure of BEC to Nic, however, produced an antagonist-like effect, suggesting that the pathobiological effects of Nic toxicity might result from both activation of nAChR channels and nAChR desensitization. We conclude that medical consequences of smoking can be mediated by direct toxic effects of inhaled Nic on the respiratory tissues wherein Nic specifically binds to and activates the nicotinic ion channels present on the cell surfaces of BEC. We believe that outside the neural system Nic interferes with functioning of non-neuronal cholinergic networks by displacing from nAChR its natural ligand acetylcholine which acts as a local hormone or cytokine in a variety of non-neuronal locations.

  8. Chromaticity separation and the alpha response.

    Science.gov (United States)

    Haigh, S M; Cooper, N R; Wilkins, A J

    2018-01-08

    Chromatic gratings can be uncomfortable to view and can evoke a large haemodynamic response. Both the discomfort and the amplitude of the haemodynamic response increase monotonically with the perceptual difference in the colour of the component bars of the grating, as registered by the separation in their chromaticity in the CIE 1976 UCS diagram. Individuals with photosensitive epilepsy exhibit epileptiform EEG activity in response to flickering light of alternate colours. The probability of the epileptiform response again increases monotonically with the separation of the colours in the CIE UCS diagram. We investigated whether alpha power, which is known to reflect the excitation of large populations of neurons, is similarly affected by the separation in chromaticity. Chromatic square-wave gratings with bars that differed in CIE UCS chromaticity were presented, together with a central fixation cross. In 18 non-clinical participants, alpha responses were recorded over the visual cortex (O1, Oz, O2, PO3, POz, PO4, P1, P2) and compared to responses in prefrontal cortex (F1, F2). Gratings comprised bars of two alternate colours that either had a small difference in chromaticity (mean CIE UCS separation of 0.03), a medium difference (mean separation of 0.19), or a large difference (mean separation of 0.43). The colour pairs had chromaticities that lay on the red-green, red-blue, or blue-green borders of the screen gamut. Regardless of the hue, the larger the separation in chromaticity, the greater the alpha desynchronization and the lower the alpha power (p = 0.004), but only in posterior electrodes (p < 0.001). Together this indicates that differences in colour evoke a cortical excitation that increases monotonically with the colour difference. In this respect the alpha response resembles the haemodynamic response. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. The measurement of $\\alpha_s$ from event shapes with the DELPHI detector at the highest LEP energies

    CERN Document Server

    Abdallah, J; Adam, W; Adzic, P; Albrecht, T; Alderweireld, T; Alemany-Fernandez, R; Allmendinger, T; Allport, P P; Amaldi, Ugo; Amapane, N; Amato, S; Anashkin, E; Andreazza, A; Andringa, S; Anjos, N; Antilogus, P; Apel, W D; Arnoud, Y; Ask, S; Åsman, B; Augustin, J E; Augustinus, A; Baillon, Paul; Ballestrero, A; Bambade, P; Barbier, R; Bardin, Dimitri Yuri; Barker, G; Baroncelli, A; Battaglia, Marco; Baubillier, M; Becks, K H; Begalli, M; Behrmann, A; Ben-Haim, E; Benekos, N C; Benvenuti, Alberto C; Bérat, C; Berggren, M; Berntzon, L; Bertrand, D; Besançon, M; Besson, N; Bloch, D; Blom, M; Bluj, M; Bonesini, M; Boonekamp, M; Booth, P S L; Borisov, G; Botner, O; Bouquet, B; Bowcock, T J V; Boyko, I; Bracko, M; Brenner, R; Brodet, E; Brückman, P; Brunet, J M; Bugge, L; Buschmann, P; Calvi, M; Camporesi, T; Canale, V; Carena, F; Castro, N; Cavallo, F R; Chapkin, M M; Charpentier, P; Checchia, P; Chierici, R; Shlyapnikov, P; Chudoba, J; Chung, S U; Cieslik, K; Collins, P; Contri, R; Cosme, G; Cossutti, F; Costa, M J; Crennell, D J; Cuevas-Maestro, J; D'Hondt, J; Dalmau, J; Da Silva, T; Da Silva, W; Della Ricca, G; De Angelis, A; de Boer, Wim; De Clercq, C; De Lotto, B; De Maria, N; De Min, A; De Paula, L S; Di Ciaccio, Lucia; Di Simone, A; Doroba, K; Drees, J; Dris, M; Eigen, G; Ekelöf, T J C; Ellert, M; Elsing, M; Espirito-Santo, M C; Fanourakis, G K; Fassouliotis, D; Feindt, M; Fernández, J; Ferrer, A; Ferro, F; Flagmeyer, U; Föth, H; Fokitis, E; Fulda-Quenzer, F; Fuster, J A; Gandelman, M; García, C; Gavillet, P; Gazis, E N; Gokieli, R; Golob, B; Gómez-Ceballos, G; Gonçalves, P; Graziani, E; Grosdidier, G; Grzelak, K; Guy, J; Haag, C; Hallgren, A; Hamacher, K; Hamilton, K; Haug, S; Hauler, F; Hedberg, V; Hennecke, M; Herr, H; Hoffman, J; Holmgren, S O; Holt, P J; Houlden, M A; Hultqvist, K; Jackson, J N; Jarlskog, G; Jarry, P; Jeans, D; Johansson, E K; Johansson, P D; Jonsson, P; Joram, C; Jungermann, L; Kapusta, F; Katsanevas, S; Katsoufis, E C; Kernel, G; Kersevan, Borut P; Kerzel, U; Kiiskinen, A P; King, B T; Kjaer, N J; Kluit, P; Kokkinias, P; Kourkoumelis, C; Kuznetsov, O; Krumshtein, Z; Kucharczyk, M; Lamsa, J; Leder, G; Ledroit, F; Leinonen, L; Leitner, R; Lemonne, J; Lepeltier, V; Lesiak, T; Liebig, W; Liko, D; Lipniacka, A; Lopes, J H; López, J M; Loukas, D; Lutz, P; Lyons, L; MacNaughton, J; Malek, A; Maltezos, S; Mandl, F; Marco, J; Marco, R; Maréchal, B; Margoni, M; Marin, J C; Mariotti, C; Markou, A; Martínez-Rivero, C; Masik, J; Mastroyiannopoulos, N; Matorras, F; Matteuzzi, C; Mazzucato, F; Mazzucato, M; McNulty, R; Meroni, C; Migliore, E; Mitaroff, W A; Mjörnmark, U; Moa, T; Moch, M; Mönig, K; Monge, R; Montenegro, J; Moraes, D; Moreno, S; Morettini, P; Müller, U; Münich, K; Mulders, M; Mundim, L M; Murray, W; Muryn, B; Myatt, Gerald; Myklebust, T; Nassiakou, M; Navarria, Francesco Luigi; Nawrocki, K; Nicolaidou, R; Nikolenko, M; Oblakowska-Mucha, A; Obraztsov, V F; Olshevskii, A G; Onofre, A; Orava, Risto; Österberg, K; Ouraou, A; Oyanguren, A; Paganoni, M; Paiano, S; Palacios, J P; Palka, H; Papadopoulou, T D; Pape, L; Parkes, C; Parodi, F; Parzefall, U; Passeri, A; Passon, O; Peralta, L; Perepelitsa, V F; Perrotta, A; Petrolini, A; Piedra, J; Pieri, L; Pierre, F; Pimenta, M; Piotto, E; Podobnik, T; Poireau, V; Pol, M E; Polok, G; Poropat, P; Pozdnyakov, V; Pukhaeva, N; Pullia, Antonio; Rames, J; Ramler, L; Read, A; Rebecchi, P; Rehn, J; Reid, D; Reinhardt, R; Renton, P B; Richard, F; Rídky, J; Rivero, M; Rodríguez, D; Romero, A; Ronchese, P; Roudeau, Patrick; Rovelli, T; Ruhlmann-Kleider, V; Ryabtchikov, D; Sadovskii, A; Salmi, L; Salt, J; Savoy-Navarro, A; Schwickerath, U; Segar, A; Sekulin, R L; Siebel, M; Sissakian, A N; Smadja, G; Smirnova, O G; Sokolov, A; Sopczak, A; Sosnowski, R; Spassoff, Tz; Stanitzki, M; Stocchi, A; Strauss, J; Stugu, B; Szczekowski, M; Szeptycka, M; Szumlak, T; Tabarelli de Fatis, T; Taffard, A C; Tegenfeldt, F; Timmermans, J; Tkatchev, L G; Tobin, M; Todorovova, S; Tomé, B; Tonazzo, A; Tortosa, P; Travnicek, P; Treille, D; Tristram, G; Trochimczuk, M; Troncon, C; Turluer, M L; Tyapkin, I A; Tyapkin, P; Tzamarias, S; Uvarov, V; Valenti, G; van Dam, P; Van Eldik, J; Van Lysebetten, A; Van Remortel, N; Van Vulpen, I B; Vegni, G; Veloso, F; Venus, W A; Verdier, P; Verzi, V; Vilanova, D; Vitale, L; Vrba, V; Wahlen, H; Washbrook, A J; Weiser, C; Wicke, D; Wickens, J H; Wilkinson, G; Winter, M; Witek, M; Yushchenko, O P; Zalewska-Bak, A; Zalewski, Piotr; Zavrtanik, D; Zhuravlov, V; Zimin, N I; Zinchenko, A I; Zupan, M

    2004-01-01

    Hadronic event shape distributions are determined from data in e+e- collisions between 183 and 207 GeV. From these the strong coupling alpha_s is extracted in O(alpha_s^2), NLLA and matched O(alpha_s^2)+NLLA theory. Hadronisation corrections evaluated with fragmentation model generators as well as an analytical power ansatz are applied. Comparing these measurements to those obtained at and around M_Z allows a combined measurement of alpha_s from all DELPHI data and a test of the energy dependence of the strong coupling.

  10. Muramidase, alpha-1 antitrypsin, alpha-1 antichymotrypsin, and S-100 protein immunoreactivity in giant cell lesions.

    Science.gov (United States)

    Regezi, J A; Zarbo, R J; Lloyd, R V

    1987-01-01

    A spectrum of giant cell lesions was evaluated for muramidase, alpha-1 antitrypsin, alpha-1 antichymotrypsin, and S-100 protein immunoreactivity using an avidin-biotin-complex immunoperoxidase method. Peripheral giant cell granuloma, central giant cell granuloma, giant cell tumor, osteitis fibrosa cystica, cherubism, and giant cell tumor of tendon sheath showed similar patterns of reactivity. Granulomatous inflammatory lesions stained more intensely for muramidase than did noninflammatory lesions. Alpha-1-antichymotrypsin was a slightly better marker of giant cell lesions than was alpha-1-antitrypsin. Positive S-100 protein staining in half the lesions was thought to be due to the presence of Langerhans cells. The results supported the belief that giant cell lesions of bone and tendon sheath are differentiated toward cells of the mononuclear-phagocyte system and that multinucleated giant cells are derived from macrophages.

  11. Measurement of total alpha activity in water; Messung der Gesamt-Alpha-Aktivitaet in Wasser

    Energy Technology Data Exchange (ETDEWEB)

    Eikenberg, Jost [Paul Scherrer Institut (PSI), Villigen (Switzerland). Abt. fuer Strahlenschutz und Sicherheit; Florschuetz, Bernd [Hessisches Landesamt fuer Umwelt und Geologie, Kassel (Germany). Dezernat 15 - Strahlenschutz; Salvamoser, Josef [Institut fuer Angewandte Isotopen-, Gas- und Umweltuntersuchungen, Woerthsee (Germany); Steinkopff, Thomas [Deutscher Wetterdienst, Offenbach am Main (Germany); Wilhelm, Christoph [Karlsruher Institut fuer Technologie (KIT), Eggenstein-Leopoldshafen (Germany). Sicherheitsmanagement - Analytische Labore; Wisser, Sascha [FCI, Mainz (Germany)

    2014-04-01

    The article describes the measurement of the total alpha activity in an (evaporated) liquid sample, and the various sample preparation methods for measurements with proportional counters or LSC. (orig.)

  12. Synthesis and coupling reactions of alpha,alpha-dialkylated amino acids with nucleobase side chains.

    OpenAIRE

    Azumaya, I; Aebi, R; Kubik, S; Rebek, J

    1995-01-01

    Several di- and tripeptides containing protected purine (adenine) and pyrimidine (thymine) residues on their side chains were synthesized. The parent amino acids alpha, alpha-dialkylated in a symmetrical manner. An effective coupling procedure was developed for these sterically hindered amino acids: the fluoren-9-ylmethyloxycarbonyl-protected amino acid was dehydrated to its oxazolinone form, which was coupled in good yields with amino esters in hot tetrachloroethane.

  13. Catalytic enantioselective Michael addition reactions of alpha-nitroesters to alpha,beta-unsaturated ketones

    NARCIS (Netherlands)

    Keller, E; Veldman, N; Spek, AL; Feringa, BL

    1997-01-01

    Enantioselective Michael additions of alpha-nitroesters 2a-d with alpha,beta-unsaturated ketones were carried out in the presence of a catalytic amount of chiral Al-Li-(R,R')-2,2'-dihydroxy-1,1'-binaphthyl ('AlLiBINOL') complex prepared in situ from LiAlH4 and 2.45 equiv. of (R,R')-BINOL. The

  14. CAMEX-4 NOAA LYMAN-ALPHA HYGROMETER V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The CAMEX-4 NOAA Lyman-Alpha Hygrometer dataset was collected by the NOAA Lyman-alpha Total Water Hygrometer, which was flown during the fourth field campaign in the...

  15. Alpha-(trifluoromethyl)amine derivatives via nucleophilic trifluoromethylation of nitrones.

    Science.gov (United States)

    Nelson, D W; Owens, J; Hiraldo, D

    2001-04-20

    (Trifluoromethyl)trimethylsilane (TMSCF(3)) reacts with nitrones to afford alpha-(trifluoromethyl)hydroxylamines protected as O-trimethylsilyl ethers. Potassium t-butoxide initiates the nucleophilic trifluoromethylation. The reaction works best with alpha,N-diaryl nitrones, and the conditions are compatible with a range of substituents on the aryl groups. Acidic deprotection of the nitrone/TMSCF(3) adducts generates alpha-(trifluoromethyl)hydroxylamines. Catalytic hydrogenation of the adducts produces alpha-(trifluoromethyl)amines. Nitrone/TMSCF(3) adducts with strong electron-withdrawing groups on the alpha-aryl ring or heterocyclic alpha-aryl groups undergo an elimination/addition sequence to generate alpha,alpha-bis(trifluoromethyl)amines. Nitrones with alkyl groups bound directly to the 1,3-dipolar moiety fail to react with TMSCF(3), but trifluoromethylation of beta,gamma-unsaturated nitrones followed by reduction of the double bond can circumvent this limitation.

  16. Plasma Ubiquinone, Alpha-Tocopherol and Cholesterol in Man

    DEFF Research Database (Denmark)

    Karlsson, Jan; Diamant, Bertil; Edlund, Per Olof

    1992-01-01

    Farmakologi, Coenzyme Q10, free cholesterol, vitamin E, antioxidants, Alpha-Tocopherol, vitamin Q, plasma, LDL-particle......Farmakologi, Coenzyme Q10, free cholesterol, vitamin E, antioxidants, Alpha-Tocopherol, vitamin Q, plasma, LDL-particle...

  17. Estimation of Time-Varying Autoregressive Symmetric Alpha Stable

    Data.gov (United States)

    National Aeronautics and Space Administration — In the last decade alpha-stable distributions have become a standard model for impulsive data. Especially the linear symmetric alpha-stable processes have found...

  18. Interferon alpha for chronic hepatitis D.

    Science.gov (United States)

    Abbas, Zaigham; Khan, Muhammad Arsalan; Salih, Mohammad; Jafri, Wasim

    2011-12-07

    Hepatitis D virus is a small defective RNA virus that requires the presence of hepatitis B virus infection to infect a person. Hepatitis D is a difficult-to-treat infection. Several clinical trials have been published on the efficacy of interferon alpha for hepatitis D virus (HDV) infection. However, there are few randomised trials evaluating the effects of interferon alpha, and it is difficult to judge any benefit of this intervention from the individual trials. To evaluate the beneficial and harmful effects of interferon alpha for patients with chronic hepatitis D. We identified relevant for the review randomised clinical trials by electronic searches in the Cochrane Hepato-Biliary Group Controlled Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL) in The Cochrane Library, MEDLINE, EMBASE, and Science Citation Index Expanded until May 2011. We also checked the bibliographic references of identified randomised trials, textbooks, and review articles in order to find randomised trials not identified by the electronic searches. Randomised clinical trials evaluating interferon alpha versus placebo or no intervention for patients with chronic hepatitis D infection. Two authors assessed the trials and extracted data on mortality, virologic, biochemical, and histological response as well as adverse events at end of treatment and six months or more after completing treatment. The analyses were performed using the intention-to-treat principle including all randomised participants irrespective of follow-up. Drop-outs, withdrawals, and non-compliance were considered as treatment failures. Data were analysed with fixed- and random-effects models. Reported results were based on fixed-effect model except in cases where statistical significance varied between the two models. Six randomised trials fulfilled the inclusion criteria. Two hundred and one randomised participants (male = 174) were included. The risk of bias in all the included trials was high

  19. Silicon vertex detector upgrade in the ALPHA experiment

    CERN Document Server

    Amole, C; Ashkezari, M.D; Baquero-Ruiz, M; Bertsche, W; Burrows, C; Butler, E; Capra, A; Cesar, C.L; Chapman, S; Charlton, M; Deller, A; Eriksson, S; Fajans, J; Friesen, T; Fujiwara, M.C; Gill, D.R; Gutierrez, A; Hangst, J.S; Hardy, W.N; Hayden, M.E; Humphries, A.J; Isaac, C.A; Jonsell, S; Kurchaninov, L; Little, A; Madsen, N; McKenna, J.T.K; Menary, S; Napoli, S.C; Nolan, P; Olchanski, K; Olin, A; Povilus, A; Pusa, P; Rasmussen, C.Ø; Robicheaux, F; Sacramento, R.L; Sampson, J.A; Sarid, E; Seddon, D; Silveira, D.M; So, C; Stracka, S; Tharp, T; Thompson, R.I; Thornhill, J; Tooley, M.P; Van Der Werf, D.P; Wells, D

    2013-01-01

    The Silicon Vertex Detector (SVD) is the main diagnostic tool in the ALPHA-experiment. It provides precise spatial and timing information of antiproton (antihydrogen) annihilation events (vertices), and most importantly, the SVD is capable of directly identifying and analysing single annihilation events, thereby forming the basis of ALPHA ' s analysis. This paper describes the ALPHA SVD and its upgrade, installed in the ALPHA ' s new neutral atom trap.

  20. Expression of alpha-amylase in Bacillus licheniformis.

    OpenAIRE

    Rothstein, D M; Devlin, P E; Cate, R L

    1986-01-01

    In Bacillus licheniformis, alpha-amylase production varied more than 100-fold depending on the presence or absence of a catabolite-repressing carbon source in the growth medium. alpha-Amylase was produced during the growth phase and not at the onset of the stationary phase. Induction of alpha-amylase correlated with synthesis of mRNA initiating at the promoter of the alpha-amylase gene.